Dallas, TX, United States

The University of Texas Southwestern Medical Center is one of the leading medical education and biomedical research institutions in the United States. UT Southwestern is located in Southwestern Medical District, a 231-acre campus in Dallas incorporating UT Southwestern Medical School, UT Southwestern Graduate School of Biomedical science, UT Southwestern School of Health Professions, and four affiliated hospitals: Parkland Memorial Hospital, Children's Medical Center, Zale Lipshy University Hospital, and St. Paul University Hospital. It also has programs with affiliated hospitals at several sites in Dallas, Richardson, Fort Worth, Waco, Austin, and Wichita Falls. Wikipedia.


Time filter

Source Type

Patent
Philips and University of Texas Southwestern Medical Center | Date: 2015-03-31

A method of operating a magnetic resonance imaging system (10) with regard to acquiring multiple-phase dynamic contrast-enhanced magnetic resonance images, the method comprising steps of acquiring (48) a first set of magnetic resonance image data (x_(pre)) prior to administering a contrast agent to the subject of interest (20), by employing a water/fat magnetic resonance signal separation technique, determining (52) a first image of the spatial distribution of fat (I_(pre)) of at least the portion of the subject of interest (20), acquiring (50) at least a second set of magnetic resonance image data (x_(2)) of at least the portion of the subject of interest (20) after administering the contrast agent to the subject of interest (20), by employing a water/fat magnetic resonance signal separation technique, determining (54) at least a second image of the spatial distribution of fat (I_(2)^(ph)) of at least the portion of the subject of interest (20), applying (56) an image registration method to the second image of the spatial distribution of fat with reference to the first image of the spatial distribution of fat (I_(pre)) for correcting a potential motion of the subject of interest (20); and a magnetic resonance imaging system (10) having a control unit (26) that is configured to carry out steps (56-64) of such a method; and a software module (44) for carrying out such a method, wherein the method steps (56-64) to be conducted are converted into a program code that is implementable in a memory unit (30) and is executable by a processor unit (32) of the magnetic resonance imaging system (10).


Maity P.,University of Texas Southwestern Medical Center | Pemberton R.P.,University of California at Davis | Tantillo D.J.,University of California at Davis | Tambar U.K.,University of Texas Southwestern Medical Center
Journal of the American Chemical Society | Year: 2013

Although the aromatic aza-Claisen rearrangement is a general strategy for accessing substituted aromatic amines, there are no highly enantioselective examples of this process. We report the first Brønsted acid catalyzed enantioselective indole aza-Claisen rearrangement for the synthesis of chiral 3-amino-2-substituted indoles. We present evidence for an arene CH-O interaction as a source of activation and stereoinduction, which is an unprecedented phenomenon in enantioselective Brønsted acid catalysis. The products of this reaction can be transformed into 3-aminooxindoles, which are prevalent in many biologically active small molecules. © 2013 American Chemical Society.


Boroughs L.K.,Children's Medical Center Dallas | Deberardinis R.J.,Children's Medical Center Dallas | Deberardinis R.J.,University of Texas Southwestern Medical Center
Nature Cell Biology | Year: 2015

Activation of oncogenes and loss of tumour suppressors promote metabolic reprogramming in cancer, resulting in enhanced nutrient uptake to supply energetic and biosynthetic pathways. However, nutrient limitations within solid tumours may require that malignant cells exhibit metabolic flexibility to sustain growth and survival. Here, we highlight these adaptive mechanisms and also discuss emerging approaches to probe tumour metabolism in vivo and their potential to expand the metabolic repertoire of malignant cells even further. © 2015 Macmillan Publishers Limited.


North C.S.,University of Texas Southwestern Medical Center | Oliver J.,University of Texas at Dallas
American Journal of Public Health | Year: 2012

Objectives. Using a comprehensive disaster model, we examined predictors of posttraumatic stress disorder (PTSD) in combined data from 10 different disasters. Methods. The combined sample included data from 811 directly exposed survivors of 10 disasters between 1987 and 1995. We used consistent methods across all 10 disaster samples, including full diagnostic assessment. Results. In multivariate analyses, predictors of PTSD were female gender, younger age, Hispanic ethnicity, less education, ever-married status, predisaster psychopathology, disaster injury, and witnessing injury or death; exposure through death or injury to friends or family members and witnessing the disaster aftermath did not confer additional PTSD risk. Intentionally caused disasters associated with PTSD in bivariate analysis did not independently predict PTSD in multivariate analysis. Avoidance and numbing symptoms represented a PTSD marker. Conclusions. Despite confirming some previous research findings, we found no associations between PTSD and disaster typology. Prospective research is needed to determine whether early avoidance and numbing symptoms identify individuals likely to develop PTSD later. Our findings may help identify at-risk populations for treatment research.


Krawczyk D.C.,University of Texas at Dallas | Krawczyk D.C.,University of Texas Southwestern Medical Center | D'Esposito M.,University of California at Berkeley
Human Brain Mapping | Year: 2013

Cognitive performance is affected by motivation. Few studies, however, have investigated the neural mechanisms of the influence of motivation through potential monetary punishment on working memory. We employed functional MRI during a delayed recognition task that manipulated top-down control demands with added monetary incentives to some trials in the form of potential losses of bonus money. Behavioral performance on the task was influenced by loss-threatening incentives in the form of faster and more accurate performance. As shown previously, we found enhancement of activity for relevant stimuli occurs throughout all task periods (e.g., stimulus encoding, maintenance, and response) in both prefrontal and visual association cortex. Further, these activation patterns were enhanced for trials with possible monetary loss relative to nonincentive trials. During the incentive cue, the amygdala and striatum showed significantly greater activation when money was at a possible loss on the trial. We also evaluated patterns of functional connectivity between regions responsive to monetary consequences and prefrontal areas responsive to the task. This analysis revealed greater delay period connectivity between and the left insula and prefrontal cortex with possible monetary loss relative to nonincentive trials. Overall, these results reveal that incentive motivation can modulate performance on working memory tasks through top-down signals via amplification of activity within prefrontal and visual association regions selective to processing the perceptual inputs of the stimuli to be remembered. Hum Brain Mapp , 2013. © 2011 Wiley Periodicals, Inc. Copyright © 2011 Wiley Periodicals, Inc..


Younger S.T.,University of Texas Southwestern Medical Center | Corey D.R.,University of Texas Southwestern Medical Center
Nucleic Acids Research | Year: 2011

Synthetic small duplex RNAs that are fully complementary to gene promoters can silence transcription in mammalian cells. microRNAs (miRNAs) are endogenous small regulatory RNAs that sequence specifically regulate gene expression. We have developed a computational method to identify potential miRNA target sites within gene promoters. Ten candidate miRNAs predicted to target the human progesterone receptor (PR) gene promoter were tested for their ability to modulate gene expression. Several miRNA mimics inhibited PR gene expression and miR-423-5p, which targets a highly conserved region of the PR promoter, was chosen for detailed analysis. Chromatin immunoprecipitation revealed that the miR-423-5p mimic decreased RNA polymerase II occupancy and increased histone H3 lysine 9 dimethylation (H3K9me2) at the PR promoter, indicative of chromatin-level silencing. Transcriptional silencing was transient, independent of DNA methylation, and associated with recruitment of Argonaute 2 (AGO2) to a non-coding RNA (ncRNA) transcript that overlaps the PR gene promoter. The miR-423-5p mimic also silenced expression of immunoglobulin superfamily member 1 (IGSF1), an additional gene with a predicted target site within its promoter. While additional investigations of endogenous miRNA function will be necessary, these observations suggest that recognition of gene promoters by miRNAs may be a natural and general mechanism for regulating gene transcription. © 2011 The Author(s).


Zhou C.,University of Texas at Dallas | Long M.,University of Texas Southwestern Medical Center | Qin Y.,University of Texas at Dallas | Sun X.,University of Texas Southwestern Medical Center | Zheng J.,University of Texas at Dallas
Angewandte Chemie - International Edition | Year: 2011

Going to waste: Renal clearance of 2-nm glutathione-coated luminescent gold nanoparticles (GS-AuNPs) was more than 10 to 100 times better than that of the similarly sized nonluminescent AuNPs coated by bis(p-sulfonatophenyl) phenylphosphine and cysteine. Urinary excretion of the particles was imaged with X-ray computed tomography (see picture of a mouse after injection of GS-AuNPs) and luminescence techniques in real time. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Roland P.S.,University of Texas Southwestern Medical Center | Tobey E.,University of Texas at Dallas
Cell | Year: 2013

For their work on the development of the modern cochlear implant, which bestows hearing to individuals with profound deafness, Ingeborg Hochmair, Graeme Clark, and Blake Wilson are the 2013 recipients of the Lasker∼DeBakey Clinical Medical Research Award. © 2013 Elsevier Inc.


Flores G.,University of Texas Southwestern Medical Center | Flores G.,Children's Medical Center Dallas | Lin H.,University of Texas Southwestern Medical Center
American Journal of Clinical Nutrition | Year: 2013

Background: Childhood overweight is a substantial public-health problem, but little is known about predictors of early childhood overweight. Objective: We aimed to identify factors - alone and in combination - that predict kindergarten overweight. Design: We analyzed nationally representative data from the Early Childhood Longitudinal Study-Birth Cohort, a longitudinal cohort study of 6800 children followed from birth through kindergarten. Multivariable logistic regression and recursive partitioning analysis (RPA) were performed to identify individual and clusters of parental, prenatal/pregnancy, infant, and toddler factors predicting kindergarten overweight. The main outcome was kindergarten overweight [body mass index (BMI) ≥85th percentile, which includes obesity]. Results: The prevalence of kindergarten overweight was 32%. By using combinations (derived from 131 factors) of a weight-for-length or BMI ≥85th percentile at earlier ages, race/ethnicity, a maternal gestational diabetes history, birth weight, and ages at solid-food introduction and the child pulling to a stand, the RPA identified 6 groups with a particularly high prevalence of kindergarten overweight (56-100%) and 2 groups with a particularly low prevalence (11-15%). An especially high prevalence was noted for children with a ≥85th BMI percentile at preschool age (77%) and in children with a ≥85th BMI percentile at 2 y old, for white children whose mother had gestational diabetes (100%), and for minority children with a birth weight <2695.5 g and who pulled themselves to a stand at <7.5 mo old (89%). Conclusion: Clusters of parental, prenatal/pregnancy, infant, and toddler factors can be used to predict which children are at particularly high and low risk of becoming overweight kindergartners. © 2013 American Society for Nutrition.


Willis B.L.,Cooper Institute | Ayers C.R.,University of Texas Southwestern Medical Center
Circulation | Year: 2012

Background-Family history of coronary heart disease (CHD) has been well studied as an independent risk factor for CHD events in the short term (<10 years). However, data are sparse on the association between family history and risk for CHD across long-term follow-up. Methods and Results-We included 49 255 men from the Cooper Center Longitudinal Study. Premature family history of CHD was defined as the presence of angina, myocardial infarction, angioplasty, or bypass surgery in a relative <50 years of age. Cause-specific mortality was obtained from the National Death Index. The association between premature family history and cardiovascular disease (CVD) or CHD death was compared across 3 unique follow-up periods (0-10, >10-20, and >20 years). Lifetime risk was estimated by use of a modified survival analytic technique adjusted for competing risk with non-CVD death as the competing event. After 811 708 person-years of follow-up, there were 919 CHD deaths and 1456 CVD deaths. After adjustment for traditional risk factors, premature family history was associated with CHD mortality >10 to 20 years (1.59; 95% confidence interval, 1.14-2.22) and >20 years (1.43; 95% confidence interval, 1.05-1.95) with wider confidence intervals at 0 to 10 years (1.32; 95% confidence interval, 0.76-2.31). Similar findings were observed for CVD mortality. Compared with men without a family history of coronary artery disease, premature family history was associated with an 50% higher lifetime risk for both CHD and CVD mortality (13.7% versus 8.9% and 21% versus 14.1%, respectively). Conclusion-Premature family history was associated with a persistent increase in both CHD and CVD mortality risk across long-term follow-up, resulting in significantly higher lifetime risk estimates. © 2012 American Heart Association, Inc.


Huang H.,University of Texas Southwestern Medical Center | Gundapuneedi T.,University of Texas Southwestern Medical Center | Rao U.,Meharry Medical College | Rao U.,Vanderbilt University
Neuropsychopharmacology | Year: 2012

Childhood maltreatment has been known to produce long-lasting impairments in behavioral, cognitive and social functioning, but their underlying mechanisms are not well-understood. A better understanding of their underlying mechanisms will aid in developing effective preventive interventions. Nineteen adolescent volunteers with no personal history of a psychiatric illness, but who were exposed to maltreatment during childhood, and 13 adolescent volunteers with no personal or family history of a psychiatric disorder (controls) underwent diffusion tensor imaging (DTI) studies. The participants were then followed longitudinally at 6-month intervals for up to 5 years to determine the onset of mood and substance use disorders. The associations among fractional anisotropy (FA) values obtained from the DTI scans at baseline and psychopathology at follow-up were examined. At baseline, adolescents exposed to childhood maltreatment had significantly lower FA values in the left and right superior longitudinal fasciculi, right cingulum bundle projecting to the hippocampus, left inferior fronto-occipital fasciculus, and splenium of the corpus callosum compared with controls. Adolescents who developed major depressive disorder at follow-up had significantly lower FA values in the superior longitudinal fasciculi and the right cingulum-hippocampal projection compared with their counterparts who did not develop the illness. Adolescents who developed substance use disorder during follow-up had significantly lower FA values in the right cingulum-hippocampal projection than their counterparts without the disorder. These preliminary results suggest that white matter disruptions observed in adolescents exposed to childhood maltreatment may be associated with increased vulnerability to psychopathology, specifically depressive and substance use disorders. © 2012 American College of Neuropsychopharmacology.


Kim H.S.,University of Texas Southwestern Medical Center | Minna J.D.,University of Texas at Dallas | Minna J.D.,University of Texas Southwestern Medical Center | White M.A.,University of Texas Southwestern Medical Center
Cell | Year: 2013

Genome-wide association studies (GWASs) have unraveled a large number of cancer risk alleles. Understanding how these allelic variants predispose to disease is a major bottleneck confronting translational application. In this issue, Li and colleagues combine GWASs with The Cancer Genome Atlas (TCGA) to disambiguate the contributions of germline and somatic variants to tumorigenic gene expression programs. They find that close to half of the known risk alleles for estrogen receptor (ER)-positive breast cancer are expression quantitative trait loci (eQTLs) acting upon major determinants of gene expression in tumors. © 2013 Elsevier Inc.


Krawczyk D.C.,University of Texas at Dallas | Krawczyk D.C.,University of Texas Southwestern Medical Center
Brain Research | Year: 2012

There has been a growing interest in understanding the complex cognitive processes that give rise to human reasoning. This review focuses on the cognitive and neural characteristics of relational reasoning and analogy performance. Initially relational reasoning studies that have investigated the neural basis of abstract reasoning with an emphasis on the prefrontal cortex are described. Next studies of analogical reasoning are reviewed with insights from neuropsychological and neuroimaging studies. Additionally, studies of cognitive components in analogical reasoning are described. This review draws together insights from numerous studies and concludes that prefrontal areas exhibit domain independence in relational reasoning, while posterior areas within the temporal, parietal, and occipital lobes show evidence of domain dependence in reasoning. Lastly, future directions in the study of relational reasoning are discussed. © 2010 Elsevier B.V. All rights reserved.


Ryu K.W.,University of Texas at Dallas | Ryu K.W.,University of Texas Southwestern Medical Center | Kim D.-S.,University of Texas at Dallas | Kraus W.L.,University of Texas at Dallas
Chemical Reviews | Year: 2015

The diverse subcellular locations and functions of poly(ADP-ribose) (PAR) and the members of the poly(ADPribose) Polymerase (PARP) family provide many opportunities to impact molecular processes and biological outcomes. The functions of the enzymatically active PARP family members are intimately tied to the nicotinamide adenine dinucleotide (NAD+) biosynthetic pathways, which provide a ready supply of ADPR units for catalysis and targeting, and underlie some of the functional interplay observed with other NAD+-utilizing enzymes. PARP proteins utilize NAD+ as a donor of ADP-ribose units and transfer these units to their target proteins. ADP-ribose transfer occurs at the catalytic domain of PARPs, which contains a donor s site with a PARP signature motif that binds NAD+ and an acceptor site where ADP-ribose chains are extended. Another layer of biological process that PAR modulates is protein degradation through ubiquitylation. greater understanding of the physiology and pathophysiology of PARPs will help us to target them more effectively, using PARP inhibitors as therapeutics.


Hefetz-Sela S.,University of Texas at Dallas | Scherer P.E.,University of Texas at Dallas | Scherer P.E.,University of Texas Southwestern Medical Center
Pharmacology and Therapeutics | Year: 2013

The prevalence of obesity has increased dramatically in recent decades, reaching epidemic proportions. It is becoming clear that obesity is associated not only with type 2 diabetes mellitus and cardiovascular disease, but also with multiple types of cancer. Obesity is characterized by impaired adipose tissue function, leading to adipocyte hypertrophy, inflammation, hypoxia and induced angiogenesis, extracellular matrix remodeling and fibrosis as well as additional stress responses. While epidemiological data indicate that obesity is a well-established risk factor for certain malignancies, the molecular mechanisms underlying the link between obesity and cancer are still poorly understood. Recent data implicates systemic and paracrine factors secreted from adipose tissue during the obese state, promoting cancer development and progression. Here, we focus on the obesity-associated adipose tissue remodeling that may not only lead to metabolic complications, but also to a permissive pro-tumorigenic environment. Particular attention is given to the local pro-tumorigenic effects derived from adipocytes that present an important part of the tumor microenvironment of at least some cancers, in an attempt to describe the nature of the major players of the adipocyte-cancer cell crosstalk that dictates to a large extent tumor progression. © 2013 Elsevier Inc.


Asterholm I.W.,University of Texas at Dallas | Scherer P.E.,University of Texas at Dallas | Scherer P.E.,University of Texas Southwestern Medical Center
American Journal of Pathology | Year: 2010

Metabolically healthy individuals effectively adapt to changes in nutritional state. Here, we focus on the effects of the adipocyte-derived secretory molecule adiponectin on adipose tissue in mouse models with genetically altered adiponectin levels. We found that higher adiponectin levels increased sensitivity to the lipolytic effects of adrenergic receptor agonists. In parallel, adiponectin-overexpressing mice also display enhanced clearance of circulating fatty acids and increased expansion of subcutaneous adipose tissue with chronic high fat diet (HFD) feeding. These adaptive changes to the HFD were associated with increased mitochondrial density in adipocytes, smaller adipocyte size, and a general transcriptional up-regulation of factors involved in lipid storage through efficient esterification of free fatty acids. The physiological response to adiponectin overexpression resembles in many ways the effects of chronic exposure to β3-adrenergic agonist treatment, which also results in improvements in insulin sensitivity. In addition, using a novel computed tomography-based method for measurements of hepatic lipids , we resolved the temporal events taking place in the liver in response to acute HFD exposure in both wild-type and adiponectin-overexpressing mice. Increased levels of adiponectin potently protect against HFD-induced hepatic lipid accumulation and preserve insulin sensitivity. Given these profound effects of adiponectin, we propose that adiponectin is a factor that increases the metabolic flexibility of adipose tissue, enhancing its ability to maintain proper function under metabolically challenging conditions. Copyright © American Society for Investigative Pathology.


Kroemer G.,French Institute of Health and Medical Research | Kroemer G.,Institute Gustave Roussy | Kroemer G.,University of Paris Descartes | Marino G.,French Institute of Health and Medical Research | And 2 more authors.
Molecular Cell | Year: 2010

Autophagy is a tightly regulated pathway involving the lysosomal degradation of cytoplasmic organelles or cytosolic components. This pathway can be stimulated by multiple forms of cellular stress, including nutrient or growth factor deprivation, hypoxia, reactive oxygen species, DNA damage, protein aggregates, damaged organelles, or intracellular pathogens. Both specific, stimulus-dependent and more general, stimulus-independent signaling pathways are activated to coordinate different phases of autophagy. Autophagy can be integrated with other cellular stress responses through parallel stimulation of autophagy and other stress responses by specific stress stimuli, through dual regulation of autophagy and other stress responses by multifunctional stress signaling molecules, and/or through mutual control of autophagy and other stress responses. Thus, autophagy is a cell biological process that is a central component of the integrated stress response. © 2010 Elsevier Inc.


Bird J.A.,University of Texas Southwestern Medical Center | Crain M.,Children's Medical Center Dallas | Varshney P.,Dell
Journal of Pediatrics | Year: 2015

Objective To determine the utility of food allergy panel testing among patients referred to a pediatric food allergy center. Study design Retrospective chart review of all new patients seen between September 2011 and December 2012 by 1 provider in a tertiary referral pediatric food allergy center. A cost analysis was performed to calculate the estimated cost of evaluation for patients who have received a food allergy panel. Results Of 797 new patient encounters, 284 (35%) patients had received a food allergy panel. Only 90 (32.8%) individuals had a history warranting evaluation for food allergy; 126 individuals were avoiding a food baased on recommendations from the referring provider and 112 (88.9%) were able to re-introduce at least 1 food into their diet. The positive predictive value of food allergy panel testing in this unselected population was 2.2%. The estimated cost of evaluation for this population was $79 412. Conclusions Food allergy panel testing often results in misdiagnosis of food allergy, overly restrictive dietary avoidance, and an unnecessary economic burden on the health system. Copyright © 2015 Elsevier Inc. All rights reserved.


Doern C.D.,University of Texas Southwestern Medical Center
Journal of Clinical Microbiology | Year: 2013

Invasive fungal infections are a significant cause of morbidity and mortality in patients who receive immunosuppressive therapy, such as solid organ and hematopoietic stem cell transplant (HSCT) recipients. Many of the fungi associated with these infections are angioinvasive and are best diagnosed by visualizing the organism in or culturing the organism from deep tissue. However, obtaining such tissue often requires an invasive procedure. Many HSCT recipients are thrombocytopenic, making such procedure too risky because of potential bleeding complications. Additionally, positive blood cultures are rare for patients with angioinvasive fungal infections, making this diagnostic strategy of little value. Undiagnosed fungal infections in these patient populations are a significant cause of mortality. Prophylactic use of antifungal agents, such as the echinocandins, during periods of neutropenia or graft-versus-host disease may prevent some fungal infections but increase the risk for others. Detection of fungal antigens in body fluids, including cryptococcus capsular polysaccharide, histoplasma antigen, galactomannan, and-D-glucan, is viewed as being clinically useful for at least the presumptive diagnosis of invasive fungal infections.-D-Glucan is an attractive antigen in that it is found in a broad range of fungal agents, including the commonly encountered agents Candida spp., Aspergillus spp., and Pneumocystis jirovecii. Cross-reactions with certain hemodialysis filters, beta-lactam antimicrobials, and immunoglobulins, which raise concerns about false-positive tests, have also been described. As a result, the use of this testing must be closely monitored. In this point-counterpoint, we have asked Elitza Theel, who directs the Infectious Disease Serology Laboratory at the Mayo Clinic, to address why she believes that this test has value in the diagnosis of invasive fungal infections. We have asked Christopher Doern, Director of Clinical Microbiology at Children's Medical Center of Dallas, why he questions the clinical value of-D-glucan testing. Copyright © 2013, American Society for Microbiology. All Rights Reserved.


Sherry A.D.,University of Texas Southwestern Medical Center | Sherry A.D.,University of Texas at Dallas | Wu Y.,University of Texas Southwestern Medical Center
Current Opinion in Chemical Biology | Year: 2013

The rate of water exchange in lanthanide complexes is often overlooked as an important parameter in the design of responsive MR imaging agents. Most often, the number of inner-sphere water coordination sites or the rotational mobility of the complex is considered as the central theme while water exchange is either assumed to be " fast enough" or entirely ignored. On the contrary, relaxation and shift theories predict that water exchange rates may indeed be the key parameter one should consider in any new molecular design. In this short review, the impact of water exchange rates on three classes of lanthanide-based MRI contrast agents, T1-based relaxation agents, T2 exchange line-broadening agents and chemical exchange saturation transfer (CEST) agents, is illustrated and discussed. © 2013 Elsevier Ltd.


News Article | April 20, 2016
Site: www.sciencenews.org

Don’t blame lab mice for shortfalls in their ability to mimic human immune systems — blame their upbringing. Mice with more experience fighting pathogens have immune system reactions more like humans’, conclude two studies published online April 20. “Dirty” mice bought from pet stores or caught in the wild have more humanlike immune systems than clean lab mice do, researchers report in Nature. And in Cell Host & Microbe, scientists find that infecting lab mice with a series of viruses and parasites alters their immune responses to be similar to those of dirty mice and humans. In recent years, scientists have debated whether mice are adequate stand-ins for humans. Some say mice are poor substitutes, and that money should instead be spent on bolstering human studies (SN: 3/23/13, p. 10). Others look at the same data and conclude that mice do a pretty good job of representing humans (SN: 9/20/14, p. 14). Plus, many important studies could not be done with humans, so mice are a necessity. But even mouse fans recognize there is room for improvement. “All science is an approximation of the real situation,” says immunologist Andrew Macpherson of the University Hospital of Bern, Switzerland, who relies on mice models. “I don’t think anybody doubts that the models don’t always accurately recapitulate what is happening in humans.” The new papers show where mice fall short and suggest ways to improve their ability to mimic people, he says. Lab mice’s immune system responses “really do look different” from that of humans’, says immunologist David Masopust, coauthor of both studies. Masopust, of the University of Minnesota in Minneapolis, and colleagues wondered whether those dissimilarities are due to irreconcilable differences in the genetic makeup of mice and humans or if the environment plays a role. His group counted immune cells in blood from adult lab mice, adult humans and human umbilical cords. Of special interest were “memory CD8+ T cells,” which cull body cells that are infected with viruses or bacteria or that have become cancerous. Lab mice and human infants have few of these memory cells, while adult humans have a plethora. That indicates that lab mice have inexperienced immune systems, much like human babies. The finding, “is one of those things that once you know it, it’s incredibly obvious,” says E. John Wherry, an immunologist at the University of Pennsylvania. “Mice are like humans raised in a bubble.” Masopust agrees. “They live a preposterously hygienic existence.” Even mice with severe immune deficiencies can thrive in immaculately clean labs. Ultraclean lab mice can’t emulate the sort of history most human immune systems experience, says Tiffany Reese, a viral immunologist at the University of Texas Southwestern Medical Center in Dallas. Adults carry an average of eight to 12 chronic viruses, such as Epstein-Barr virus (the cause of mononucleosis). Worm parasites infect about 2 billion people worldwide. And by adulthood, people have usually fought off multiple colds, flus and other infections. Masopust’s team found that the memory T cell profiles of wild and pet-store mice more closely resembled that of adult humans than lab mice’s did. Housing lab mice next to pet-store mice for a month caused their immune system to change, making the lab mice resemble the dirty mice, the researchers reported in Nature. In discrepancies between studies of lab mice and humans, “the mouse may not be at fault,” Masopust says. “It’s the way that they are cared for.” An experienced immune system not only looks different, it also works differently from an inexperienced one, Reese and colleagues report in Cell Host & Microbe. Reese infected lab mice with two types of herpesviruses, gave them the flu and inoculated them with an intestinal parasite. She then compared how uninfected mice reacted to a yellow fever vaccine with how chronically infected mice reacted. Uninfected mice made more antibodies against the vaccine. The result might help explain why some vaccines that look promising in animal studies don’t pan out in human trials. Controlled infections may increase understanding of how pathogens interact with each other, with friendly microbes that live in the body and with the host’s immune system, says Reese’s coauthor Herbert Virgin, a viral immunologist at Washington University School of Medicine in St. Louis. Researchers have a bias that mice are not humans, says Virgin, “But I think that’s too simplistic a view. We shouldn’t be asking whether the mouse is a perfect model for humans, but whether we can make the mouse emulate more closely the basic nature of human physiology.”


News Article | September 14, 2016
Site: www.nature.com

The subjects were Th-Cre knock-in heterozygous male mice backcrossed more than 20 times to the C57BL/6 strain (Thtm1(cre)Te; EM:00254)51 (behavioural and anatomical studies, n = 71 mice), Th-Cre transgenic heterozygous male mice on a mixed C57BL/6 and CD1 background (Tg(Th-cre)1Tmd)52 (for ex vivo hippocampal electrophysiology, n = 42), and C57BL/6 male mice (Charles River; n = 7). They were >8 weeks old at the start of the experiments, several of which continued for many months, and assigned to groups randomly. All mice were given water ad libidum, kept under a 12 h light/dark cycle (lights on 7:00); given food ad libidum in unit recording studies but food-restricted for event arena training (85% of free-feeding weight monitored daily throughout the study, after behavioural training). Behavioural testing was performed during the light phase of the cycle, and all critical tests were conducted ‘blind’. All procedures were overseen by the University of Edinburgh Ethical Review Committee, compliant with the UK Animals (Scientific Procedures) Act 1986 and with the European Communities Council Directive of 24 November 1986 (86/609/EEC) legislation governing the maintenance of laboratory animals and their use in scientific experiments; and with guidelines of the Animal Welfare Committee of Hokkaido University; were approved by the animal care and use committee (IACUC) at the University of Texas Southwestern Medical Center and comply with federal regulations set forth by the National Institutes of Health. The Cre-inducible AAV were obtained from the University of North Carolina (UNC) Vector Core Facilities. The viral concentration was 8.0 × 1012 particles ml–1 for AAV5/EF1a-DIO-hChR2(H134R)–eYFP (ChR2–eYFP), 3.2 × 1012 particles ml–1 for AAV5/EF1a-DIO-eArch3.0–eYFP (eArch3.0–eYFP), 4.0 × 1012 particles ml–1 for AAV5/EF1a-DIO-eNpHR3.0–eYFP (eNpHR3.0–eYFP), 4.0 × 1012 particles ml–1 for AAV5/EF1a-DIO–eYFP (eYFP control), and 1 × 1012 to 7 × 1012 particles ml–1 for AAV2/EF1a-DIO-hChR2(H134R)–eYFP (for ex vivo hippocampal electrophysiology). Virus were subdivided into aliquots stored at −80 °C until use. Anaesthesia was induced using isoflurane (induction, 5%; maintenance, 1–2%; air-flow, 1 l min−1). The animals were placed in the stereotactic frame (Kopf Instruments). For viral or retrobead (Lumafluor) injection, a small hole was drilled into the skull over the target site. The virus (0.75–1 μl) or retrobead solution (0.1 μl) was then injected at 0.1 μl min−1 into the target site using a Nanofil syringe (WPI) and UMP3 pump (WPI) mounted directly on the stereotactic frame. After each injection, the needle was kept in place for 10 min to ensure proper diffusion of the virus. Animals recovered on a heating pad until normal behaviour resumed. All experiments involving viral constructs were performed at least 3 weeks after surgery to allow for sufficient expression. Viral infusion coordinates were VTA (from bregma53: anterior–posterior (AP), −3.50 mm; mediolateral (ML), 0.50 mm; and dorsal–ventral (DV) from the dura, −4.40 mm) and LC (AP, −5.45 mm; ML, 1.20 mm; and DV, −3.65 mm). Event arena pharmacological experiment (Fig. 1). Bilateral 26-gauge microinjection steel guide cannulae (2.5 mm length, 3.0 mm distance between cannulae; Plastics One) with stylets that protruded 0.5 mm below the end of the cannula (33 gauge, Plastics One) were implanted into the dorsal hippocampus. The cannula implantation coordinates were (AP, −2.10 mm; ML, ± 1.50 mm; and DV, −2.00 mm). Extracellular recording during novelty exploration (Fig. 2). the Cre-inducible AAV5 ChR2–eYFP virus (1 μl) was unilaterally injected into VTA or LC as mentioned above. Four jeweller’s screws were then placed in the skull and the ground wire was attached to one of the skull screws. The microdrive implantation coordinates were VTA (AP, −3.52 mm; ML, 0.48 mm; and DV, −4.00 mm) and LC (AP, −5.45 mm; ML, 1.00 mm; and DV, −2.80 mm). Adhesive cement (C&B metabond, Parkell) and dental acrylic were then sculpted around the microdrive. Tract tracing experiment (Fig. 3 and Extended Data Fig. 5). For anterograde tracing, the Cre-inducible AAV5 eYFP virus (1 μl) was unilaterally injected into VTA or LC as described above. For retrograde tracing, retrobeads (0.1 μl) were unilaterally injected into CA1 (AP, −2.18 mm; ML, 1.18 mm; and DV, −1.36 mm), CA3 (AP, −2.18 mm; ML, 2.68 mm; and DV, −2.05 mm) and DG (AP, −2.18 mm; ML, 1.36 mm; and DV, −1.82 mm). Event arena experiment with optogenetics (Fig. 4). Arterial oxygen saturation, heart rate and breath rate were monitored by MouseOx instrument (STARR Life Science). The Cre-inducible AAV5 virus (ChR2–eYFP or eYFP, 1 μl per side) was injected bilaterally into VTA and LC according to the procedure described above. A dual ferrule optical fibre implant (0.22 numerical aperture, 200 μm core diameter; Doric Lenses) was implanted vertically into VTA (AP, −3.40 mm; ML, ± 0.50 mm; and DV, −4.30 mm). Subsequently, a two-ferrule optical fibre implant (0.22 numerical aperture, 200 μm; Doric Lenses) was implanted into LC at −30° angle to coronal plane (AP, −5.45 mm; ML, ± 0.90 mm; and DV, −3.00 mm). Additionally, bilateral 26-gauge steel guide cannulae (4.0 mm length, 3.0 mm distance between cannulae) with stylets (33 gauge) that protruded 0.5 mm below the end of the cannula was inserted into the dorsal hippocampus at a 40° angle to coronal plane (AP, −2.10 mm; ML, ± 1.50 mm; and DV, −2.00 mm). Event arena experiment with pharmacological inactivation (Fig. 6). The Cre-inducible AAV5 virus (eArch3.0–eYFP or eYFP, 1 μl per side) was injected bilaterally into LC, and a two-ferrule optical fibre implant was then implanted into LC at −30° angle to coronal plane as described above. Additionally, bilateral 26-gauge steel guide cannulae (4.8 mm length, 1.0 mm distance between cannulae) with stylets (33 gauge) that protruded 0.5 mm below the end of the cannula was inserted into VTA (AP, −3.40 mm; ML, ± 0.50 mm; and DV, −4.40 mm). Ex vivo hippocampal electrophysiology (Fig. 5). The Cre-inducible AAV2 ChR2–eYFP virus (0.75–1 μl per side) was bilaterally injected into LC (AP, −5.45 mm; ML, ± 0.90 mm; and DV, −3.00 mm) over a 10–15 min period using a borosilicate glass electrode (10–15 MΩ) pulled with a horizontal pipet puller (P-97, Sutter Instrument) and a picospritzer (Parker) timed by a Master-8 pulse stimulator (A.M.P.I.). After each injection, the pipette was kept in place for 5 min to ensure proper diffusion of the virus. Postoperative analgesia. Carprofen (0.08 ml kg−1 body weight), or buprenorphine (0.1 mg kg−1 body weight), was administered by subcutaneous injection at the end of all surgical procedures. All mice were allowed a recovery period of at least 7 days for them to regain their pre-surgery weights before electrophysiological or behavioural testing. Everyday memory was tested in an event arena: a square open field (120 cm wide × 120 cm long) with walls (35 cm high) made out of transparent Plexiglas (Fig. 1a) with four adjacent start boxes (black Plexiglas). The name ‘event arena’ derives from it being an arena in which ‘events’ happen (for example, finding food)13. The floor of the arena, arranged in a 5 × 5 grid, was covered with ~2 cm of sawdust and had two intramaze landmarks (a white metal cube located at row 3, column 2, and a black rubber flash light at row 3, column 4). The Plexiglas sandwells, in which food reward was potentially available, could be fitted into any of the 23 remaining sandwell positions (positions occupied by internal cues were excluded). The mice had access to the arena and sandwells when the startbox door was opened in any trials. Light levels, checked every day, were 25–35 lx. Data were recorded using custom-made LabVIEW software (National Instruments), using the image from camera placed above the arena. For novelty exploration, a square Plexiglas open field with transparent walls (70 cm wide × 70 cm long × 30 cm high) was placed in the middle of the event arena (Fig. 1b). To maintain the novelty of the environment, a wide range of floor substrates (dried leaves, shredded paper, feathers, acrylic pompoms, corks, lolly sticks, Lego blocks, pipe cleaners, shredded straws and sea shells) that covered the floor of the box were used, as in a previously published study that used rats13. Sample size, randomization, blinding and replication. Sample size was determined based on variability in pilot data. A distinctive feature of event arena tasks is that most but not all comparisons are ‘within-subject’ design in which every single subject is exposed to every single treatment, including the control treatment. This typically reduces the number of subjects required for statistical significance and avoids issues associated with randomization. All non-rewarded probe tests (see below) were analysed blind and, being conducted against a stable performance background, were typically conducted twice or three times (internal replication). Averaging data helped reduce variability. Shaping and habituation. After handling, habituation (eight sessions) involved training mice to dig in the sandwells to retrieve food (a half of cereal ‘Cheerios’) and carry it to the start box. Everyday training protocol. The goal in each daily session was to encode the changing daily location of a rewarded sandwell encountered during two consecutive sample trials (two retrievals of buried food in each trial), and then, 10 min later, return to that same location during the choice trial (Fig. 1a). The choice trial was a retrieval test that involved a rewarded sandwell in a location that matched the sample location (the ‘correct’ location; win-stay rule) and four non-rewarded sandwells placed in other locations around the arena (the ‘incorrect’ locations). Training sessions were conducted daily (5–7 sessions per week) using 16 different sandwell configurations with rewarded sandwell positions counterbalanced between mice (Extended Data Fig. 1a). We calculated a performance index, using the formula performance index = 100 − [100 × (errors/4)], based on the number of errors made during the choice trial (an error being defined as digging at an incorrect sandwell). The value expected on the basis of chance was 50% (two errors). With each behavioural cohort, we began conducting critical memory probe tests once mice reached average performance index of 75% (equivalent to one error, computed as average of last five training sessions). This typically happened within 35 training sessions, but sometimes additional training was needed after surgical procedures. Memory probe tests. The primary data measures of the study were derived from ‘memory tests’ performed as ‘probe’ tests, defined as sessions in which none of the sandwells contained any accessible food pellets. The mice were cued with a food pellet in the start box, and then allowed to search for the correct sandwell for 60 s from the first dig of any sandwell. After 60 s, the experimenter quietly entered the room and buried pellets in the correct sandwell, allowing the mouse to retrieve them (one by one as in training, this limited extinction). Dig time at each sandwells was measured, and the relative proportion of time at the correct and incorrect sandwells was calculated. The value expected on the basis of chance was 20%. Probe tests were always separated by at least two sessions of regular training. In the ‘reward magnitude’ probe tests (Fig. 1b and Extended Data Fig. 1d), animals retrieved either two pellets (low reward) or eight pellets (high reward) during memory encoding. Novelty exploration. The mice underwent seven sessions of habituation to the box with sawdust placed in the event arena. For post-encoding unexpected environmental novelty, the mouse was placed into the centre of the box lined with a novel floor surface and allowed to explore freely for 5 min. Microinfusion/injection of drugs. To help reduce any stress, all drugs were infused in the home cages. The stylets in the guide cannulae were replaced by a double infusion cannula (33 gauge, Plastics One) connected to two 5-μl microsyringes (WPI) in a microinfusion pump (Native Instruments) via flexible plastic tubing (C232CS, Plastics One) filled with Fluorinert (3 M). The tips of infusion cannulae projected 0.5 mm below the tip of the guide cannulae. For intra-hippocampal microinjection, 0.5 μl of drug per cannula was infused at 0.2 μl min–1 (2.5 min). Infusion cannulas were left in place for a further 2.5 min before being replaced with stylets to aid drug absorption. For intra-VTA microinjection, 0.3 μl was injected at a rate of 0.3 μl min–1 (1 min) followed by 1 min of waiting. The mice were habituated to the experimental procedure of injection and to vehicle injection before the drug test to minimize the potential novelty of the procedure. Mice received drug injection 20 min (hippocampal microinfusions and intraperitoneal injection of clonidine) or 3 min (VTA microinfusions) before the novelty exploration. Drug concentrations. For microinfusions, the concentrations used were 21.1 mM (6.25 μg μl–1) for β-adrenoceptor antagonist propranolol ((S)-(−)-propranolol hydrochloride, 295.80 g mol–1; Sigma-Aldrich), 3.1 mM (1 μg μl–1) for D /D receptor antagonist SCH23390 (SCH 23390 hydrochloride, 324.24 g mol–1; Tocris) and 2% w/v for voltage-gated sodium channel blocker lidocaine (lidocaine hydrochloride monohydrate, 288.81 g mol–1; Sigma-Aldrich). α -Adrenoceptor agonist clonidine (clonidine hydrochloride, 266.55 g mol–1; Sigma-Aldrich) was administered intraperitoneally at a dose of 50 μg kg–1 of body weight. We used 0.9% NaCl in H O (saline) as a vehicle and for control infusions. Both vehicle and drug solutions were stored in 100 μl aliquots at −20 °C until use. The mice were extensively habituated to the experimental procedure of photostimulation and to flickering blue light for several weeks before the optogenetic photostimulation test to minimize the potential novelty of the procedure. Laser stimulation, consisting of 20 5-ms pulses of 473-nm light at 25 Hz, delivered every 5 s (average stimulation rate 4 Hz) for the duration of 5 min (Fig. 4b), was performed in home cages using two blue solid-state diode pumped lasers (18–19 mW, Laser 2000) connected to either a dual fibre optic patch cord (for VTA; 0.22 numerical aperture, 200 μm core diameter; Doric Lenses) or two single fibre optic patch cords (for LC; 0.22 numerical aperture, 200 μm, Doric Lenses). Both lasers were synchronously controlled using custom-built LabVIEW software. Apparatus and light stimulation. The behavioural apparatus consisted of a rectangular wooden box with three compartments: a screening chamber with space for the home cage, as well as ‘familiar’ and ‘novel’ chambers (both 30 cm wide × 70 cm long). The apparatus was surrounded by black curtains and light level was kept at 25–35 lx. The floor of the familiar environment was covered in sawdust (the floor substrate of the event arena), and the floor of the novel environment was covered in one of the floor substrates used as novelty in the event arena experiments. Each floor substrate was used only once for each mouse. Unit activity was recorded extracellularly using the implanted custom-built screw-driven microdrive consisting of a 200 μm optic fibre surrounded by four tetrodes (an ‘optetrode’) that protruded 400–800 μm beyond the fibre tip54. Signals were fed through a 16 channel unity gain headstage amplifier (Axona), band-pass filtered at 300–5,000 Hz, amplified 1,000–40,000 times, digitized at 50 kHz and stored for subsequent analysis. Spike capturing was done on-line using amplitude threshold. Recorded neurons were identified as TH+ using Cre-dependent ChR2 expression and low-frequency light stimulation55. Laser stimulation was performed using a blue solid-state diode pumped laser (473 nm, Laser 2000) connected to a fibre optic patch cord (0.22 numerical aperture, 200 μm core diameter), and controlled with the data capturing software (Axona). Epochs of 60 light pulses (1 Hz, 5 ms pulse duration) at different light intensities (0.1–20 mW) were then administered and each tetrode was screened for light-evoked spikes. Spikes were classified as ‘light-evoked’ if their latency from the onset of the light pulse was between 0 and 15 ms, all other spikes being classified as ‘spontaneous’. Units were classified as light-responsive if (a) a cell fired a light-evoked spike in response to more than one third of light pulses, (b) the shape of the mean light-evoked waveform of a unit closely resembled the spontaneous waveform of the same unit. Units with basal firing rates above 20 Hz were excluded from this analysis because of intrinsically high probability of spiking within the 15 ms window after the light pulse. Novelty exploration. Two weeks after surgery, the implanted mice underwent 5 days of habituation to the experimental apparatus and the familiar environment. Following habituation, mice underwent daily screening trials for light-responsive neurons in the home cage. If no light-responsive cells were found, the mouse was allowed to explore the familiar environment for 5 min and was then unplugged. If one or more recording channels showed a light-responsive unit, the mouse was subjected to an ‘exploration’ trial after a 10 min delay (Fig. 2b). The mouse might first be placed in the novel environment for 5 min, followed by 25 min in the home cage and 5 min in the familiar environment; alternatively, the order of novel and familiar exploration was reversed (in a counterbalanced manner). Baseline recording (5 min) was performed in the home cage between two exploration trials. These particular time delays were chosen to mimic timing used in the event arena experiments, where there was a 30 min delay between memory encoding and novelty exploration. The light-sensitive unit was again confirmed in home cage with 1 Hz light stimulation after open field exploration. The microdrive was advanced by ~40 μm at the end of daily session to ensure that recordings are made from a different population of neurons. Recording and analysis. Recorded spikes were clustered using Klusterkwik 1.5 unsupervised clustering algorithm (http://klusta-team.github.io/klustakwik/) on the basis of their energy and first principal component of the waveform. Clusters were then corrected manually using Klusters spike sorting software (http://neurosuite.sourceforge.net/), on the basis of several additional parameters (width of waveform, amplitude, time at peak, auto- and cross-correlograms). Data were analysed using Matlab R2012a (MathWorks). Firing patterns were characterized in terms of firing rate, rate of burst events and firing rate of spikes within bursts. Bursts were defined, using classic criteria56, as trains of two or more spikes with an interspike interval of less than 80 ms, followed by an interspike interval of more than 160 ms. For comparison of novelty modulation in VTA-TH+ and LC-TH+ neurons, firing rates of individual neurons in the novel and familiar environments were binned in 10-s bins and normalized to the average home cage firing rate of all identified neurons in respective brain areas. For additional analysis of the novelty modulation, binned firing rates of individual neurons in the novel environment were z-scored to their respective firing rates in the familiar environment. Under deep pentobarbital anaesthesia (100 mg kg–1 of body weight, intraperitoneally), the mice were fixed transcardially with 4% paraformaldehyde in 0.1 M sodium phosphate buffer, pH 7.2, post-fixed in the same fixative for 24 h, and placed in 30% sucrose in phosphate buffer. Sections of fixed brains (30 μm in thickness) were prepared using a freezing microtome (SM2000R, Leica Microsystems) for immunohistochemistry. Immunofluorescence. Antibodies used included: goat anti-GFP57, guinea-pig anti-NET58 and mouse anti-TH (AB152, Millipore). eYFP was visualized by anti-GFP immunostaining. All immunohistochemical incubations were done at room temperature. Sections were incubated successively with 10% normal donkey serum for 20 min, a mixture of primary antibodies overnight (1 μg ml–1), and a mixture of Alexa Fluor 488-, Cy3- or Cy5-labelled species-specific secondary antibodies (Invitrogen; Jackson ImmunoResearch) for 2 h at a dilution of 1:200. Fluorescent in situ hybridization. Brains were freshly obtained under deep diethyl ether anaesthesia and immediately frozen in powdered dry ice. Fresh-frozen sections (20 μm) were cut on a cryostat (CM1900, Leica Microsystems). All sections were mounted on silane-coated glass slides. Mouse cDNA fragments of TH (bases 1–1025; GenBank accession number AY855842), and NET (bases 124–814, MMU76306) were subcloned into the pBluescript II plasmid vector. Digoxigenin- or fluorescein-labelled cRNA probes were transcribed in vitro for fluorescent in situ hybridization59. Image acquisition and data analysis. Images were taken with a confocal laser-scanning microscope (FV1200, Olympus) equipped with diode laser lines, and UPlanSApo (20×, numerical aperture 0.75) and PlanApoN (60×, numerical aperture 1.4, oil-immersion) objective lenses (Olympus). To avoid cross talk between multiple fluorophores, Alexa Fluor 488, Cy3, and Alexa Fluor 647 fluorescent signals were acquired sequentially using the 473, 559, and 647 nm excitation laser lines, respectively. All images show single optical sections. For quantification of anterograde tracing, we obtained images with a 20× objective and then created images of the entire hippocampus with a Fluoview image stitching software (Olympus). For analysis, the separate colour components were converted to greyscale, and the area of eYFP- and TH-positive elements were measured with Integrated Morphometry Analysis module (MetaMorph software, Molecular Devices). Hippocampal slice preparation. Thick coronal slices (300 μm) containing the hippocampus were cut from Th-Cre transgenic mice52 expressing AAV2 ChR2–eYFP in LC (8–12 weeks old) in low light conditions to prevent unwanted ChR2 activation. Animals were anaesthetized under 1.5–2% isoflurane, and the brains removed and blocked following rapid decapitation. Hippocampal slices were prepared using a vibratome (VT 1000S, Leica Microsystems) in ice cold N-methyl-d-Glucosamine (NMDG) ringer solution (in mM): 5 NaCl, 57 NMDG, 37.5 Na-pyruvate, 12.5 Na-lactate, 5 Na-ascorbate, 2.5 KCl, 1.25 NaH PO , 25 NaHCO , 25 glucose, 10 MgSO ·7H O, 0.5 CaCl ·2H O, the pH was set between 7.3 and 7.4 using 12 N HCl, the osmolarity was adjusted as needed to 315 mOsm using glucose and the solution was bubbled with 95% O and 5% CO gas. Slices were maintained in NMDG ringer at room temperature for no longer than 15 min and then transferred to artificial cerebrospinal fluid (in mM): 125 NaCl, 2.5 KCl, 1.25 NaH PO , 1.3 MgCl , 2 CaCl , 25 NaHCO and 25 dextrose continuously bubbled with 95% O and 5% CO gas, where they were kept up to 6 h, protected from light, for experimentation. One slice per animal was used. Ex vivo whole-cell recordings. Slices were transferred to a submersion recording chamber and were perfused with artificial cerebrospinal fluid at a rate of 1–2 ml min−1 at 26–29 °C. EPSC recordings were performed with GABA receptor antagonist picrotoxin (602.58 g mol–1; Sigma-Aldrich) at a concentration of 50 μM and D receptor antagonist eticlopride (eticlopride hydrochloride, 377.31 g mol–1; Sigma-Aldrich) at a concentration of 100 nM in the bath. A borosilicate glass electrode (3–5 MΩ), pulled with a horizontal pipet puller (P-97), was filled with Cs-methanesulfonate pipet solution (in mM): 110 CsMeSO , 15 CsCl, 8 NaCl, 2 EGTA, 10 HEPES, 3 QX-314, 2 ATP and 0.3 GTP adjusted to 295 mOsm and pH 7.3. Whole-cell pyramidal cell recordings from area CA1 were acquired using a combination of visualized and blind patch techniques. Cells were held at −60 mV using a multiclamp 700B amplifier (Molecular Devices). A bipolar stimulating electrode (FHC) was placed in the stratum radiatum region of dorsal CA1 within 100–200 μm of the recording electrode and stimulation (delivered at a rate of 0.2 Hz) was set to elicit current responses of 50–150 pA. Data were acquired and analysed automatically using P-Clamp 10 (Molecular Devices). Recordings were discarded if the series resistance varied by more than 20% or if the initial holding current exceeded 70 pA. Following a 5 min baseline acquisition, hippocampal slices were given a 470-nm light stimulus (consisting of three trains delivered every 2 min, of 60 5-ms pulses, applied at 18 Hz (Extended Data Fig. 7b)) through the 40× objective lenses. Following the light stimulus, baseline stimulation resumed. Ex vivo field recordings. Slices were transferred to a submersion chamber and were perfused with artificial cerebrospinal fluid at a rate of ~2 ml min−1 at 29–31 °C. Field recordings from the stratum radiatum of dorsal CA1 were acquired using a borosilicate glass electrode (1–3 MΩ) filled with artificial cerebrospinal fluid. A bipolar stimulating electrode was also placed in the stratum radiatum of CA1 within 100–200 μm of the recording electrode and stimulation (one stimulus every 30 s) was set to elicit a fEPSP slope that was ~50% of the maximum value. A stable 15 min baseline was obtained, followed by 10 more min of baseline stimulation with or without simultaneous optogenetic stimulation of LC-TH+ axons. Photostimulation consisting of four trains, at a 1 s interval, consisting of four 10-ms pulses of 470-nm light at 16 Hz, delivered every 30 s for the duration of 10 min (Extended Data Fig. 7c), was applied through the 10× objective (directly before the Schaffer collateral stimulus). After the 25 min baseline, a weak theta-burst tetanus was applied consisting of four trains, at a 100 ms interval, consisting of three pulses at 50 Hz (12 pulses in total). Baseline stimulation then resumed as described above for 45 min. Every two traces were averaged to reduce variability. Data were acquired and analysed automatically using P-Clamp 10. Ex vivo pharmacology. Where indicated, the following drugs were bath applied: α-adrenoceptor antagonist prazosin (prazosin hydrochloride, 419.86 g mol–1; Sigma-Aldrich) at a concentration of 30 μM, β-adrenoceptor antagonist propranolol ((S)-(−)-propranolol hydrochloride, 295.80 g mol–1; Sigma-Aldrich) at a concentration of 30 μM, D /D receptor antagonist SCH 39166 (SCH 39166 hydrochloride, 394.73 g mol–1; Tocris) at a concentration of 100 nM (intracellular recordings, n = 2) and D /D receptor antagonist SCH 23390 (SCH 23390 hydrochloride, 324.24 g mol–1; Tocris) at a concentration of 100 nM (intracellular recordings, n = 3) or 1 μM (extracellular recordings). Anaesthesia was induced using isoflurane for LC optrode recording as described above or with 4.8% chloral hydrate for VTA recording (induction: 480 mg kg–1 body weight, intraperitoneally; maintenance: 120 mg kg–1 body weight, intraperitoneally). Recordings were made using a 125 μm 1 MΩ tungsten electrode (A-M systems). This electrode was connected to a differential AC amplifier (A-M Systems), signals were band-pass filtered at 300 Hz to 5 kHz, amplified 10,000 times and digitized at 20 kHz. Spiking activity was defined as spikes that exceed five standard deviations from the mean value of the baseline signal (1 min before laser stimulation (LC) or drug infusion (VTA)). Multi-unit activity in each trace was then normalized to the pre-stimulation (LC) or pre-infusion (VTA) baseline. LC optrode recording. The tungsten electrode was coupled to an optic fibre (0.22 numerical aperture, 200 μm core diameter) (an ‘optrode’). This optrode was positioned above LC (AP, −5.45 mm; ML, 1.00 mm; and DV, −2.80 mm), and was then gradually lowered in 50 μm increments until multi-unit activity was observed. Laser stimulation was performed using a green solid-state diode pumped laser (532 nm, Laser 2000) with 10–20 mW output from the fibre. For quantification of eArch3.0 recordings, baseline was measured over 30 s before the start of illumination (Pre), level of inhibition was measured over 5 min light on period (LC-on), rebound activity was measured over 1 min after the end of illumination (rebound) and post-inhibition baseline was measured 4–5 min after the end of illumination (Post). VTA recording with pharmacology. The drug cannula (33 gauge, Plastics One) was positioned in the VTA at 14° angle to sagittal plane (AP, −3.52 mm; ML, 0.48 mm; and DV, −4.40 mm) and the recording electrode was positioned vertically at the boundary of the VTA. For intra-VTA microinjection of lidocaine, infusion parameters were the same as those used in the behavioural experiment as described above. For quantification, pre-infusion baseline was measured over 30 s before the start of infusion (Pre), level of inhibition was measured 3–8 min after the start of infusion (Lid)—the period that corresponds to novelty exploration, and post-inhibition baseline was measured 17–18 min after the start of injection (Post). Statistical analyses were performed using SPSS version 19 (IBM). All data are expressed as mean ± s.e.m. Statistical significance was always determined by ANOVAs, before orthogonal comparisons where possible or Tukey’s HSD tests as appropriate to correct for multiple comparisons, paired t-tests and one-sample t-tests. All statistical tests were two-tailed. Analysis of probe test performance was done on the basis on the ‘% correct dig time’ score. In the pharmacological inactivation experiment (Fig. 6), three animals that persistently failed to show any novelty-induced memory enhancement in all control conditions (all 24-h probe test scores (% correct dig) in the ‘novelty with vehicle’ condition at below chance level, that is, <20%) were eliminated from the whole data set before statistical analysis (leaving n = 15). The rationale for this was that it was not possible to measure the impact of pharmacological manipulations on the novelty effect in animals that are not susceptible to it, the existence of the novelty effect having been established in the first cohort of mice (Fig. 1b). All source data for the preparation of graphs and statistical analysis are presented online. All other relevant data that support the conclusions of the study are available from the authors on request.


News Article | March 16, 2016
Site: www.nature.com

All animal studies were performed with the approval of the Institutional Animal Care Committees of Sun Yat-sen University, the University of California San Diego, West China Hospital, and the University of Texas Southwestern Medical Center. The eyeball was enucleated from a one-month-old New Zealand white rabbit and washed with PBS (containing antibiotics) three times. After the cornea and iris were removed, a small cut was made in the posterior capsule of the lens; the capsule with attached epithelium was removed and cut into 1 × 1 mm2 pieces. The pieces of epithelium were cultured in minimum essential media supplemented with 20% FBS, NEAA, and 50 μg ml−1 gentamicin. A 17-week-old human fetal eyeball was purchased from Advanced Bioscience Resources, Inc. (San Francisco, California). Post-mortem human eyes were obtained from San Diego Eye Bank. The human LECs were cultured according to the same methods as above. For in vitro differentiation, LECs were cultured on Matrigel-coated six-well plates or eight-well chambers. Lentoid body was formed after 21 days in minimum essential media supplemented with NEAA, 1% FBS, 100 ng ml−1 FGF2, and 5 μg ml−1 insulin. Images of lentoid tissue were obtained using a Leica M205FA stereo microscope. Membrane-tomato/membrane-green (mTmG)-targeted ROSAmTmG mice were purchased from the Jackson Laboratory (Bar Harbour, ME; stock no. 7576) and maintained as homozygotes. P0-3.9-GFPcre mice expressing an eGFP–Cre recombinase fusion protein under the control of the Pax6 lens ectoderm enhancer and the Pax6 P0 promoter26 were maintained in a FVB/N background. Lineage-tracing experiments were performed by crossing the homozygous ROSAmTmG reporter mouse strain with the P0-3.9-GFPcre deleter strain. Eyes were dissected at P1, P14, and P30 and fixed overnight in 4% formaldehyde. Tissues were then incubated in 10% sucrose and embedded in OCT for cryo-sectioning. Frozen sections were washed in PBS and imaged on a Zeiss Axio Imager fluorescence microscope. Bmi1fl/fl mice were generated as previously described27. Nestin-cre mice28 were obtained from the Jackson Laboratory. For BrdU pulses, mice were injected with 100 mg kg−1 BrdU (Sigma) dissolved in PBS, then maintained on drinking water that contained 1 mg ml−1 BrdU until sacrifice. For gene expression studies, lenses of Pax6P0-3.9-GFPcre mice were dissected under a dissecting microscope. Lens capsular bag was opened from the posterior surface by making three crisscross incisions. The capsular bag was opens and lens material extruded. GFP-positive LECs in the mid-anterior capsular area were separated mechanically from GFP-negative LECs in the remaining capsular areas under a fluorescence microscope. RNA was isolated using RNeasy Mini Kit (Qiagen). To image cataracts, mice were anaesthetized with Avertin, and one drop of 1% Mydriacyl (Alcon) was administered per eye. Eyes were immediately visualized in vivo using a light microscope. For histology, mice were perfused with heparinized saline followed by 4% paraformaldehyde (PFA) in PBS. Dissected eyes were fixed in 4% PFA overnight, embedded in paraffin, and sectioned by the UT Southwestern Molecular Pathology core facility. For BrdU staining, slides were deparaffinized, and subjected to heat-mediated antigen retrieval (in 10 mM sodium citrate, pH 6.0). Slides were stained with primary mouse anti-BrdU (Caltag, MD5000, 1:200) overnight at 4 °C. Slides were subsequently stained with Alexa Fluor 555-conjugated goat anti-mouse IgG1 secondary antibody (Life Technologies, 1:500) and 1 mg ml−1 DAPI (1:500) for 1 h at room temperature. The number of BrdU-labelled cells was divided by the total number of DAPI+ cells in a single layer of LECs. Lentiviral shRNA targeting the human BMI1 gene (NCBI Reference Sequence: NM_005180.8) was purchased from Origene (TL314462), ShRNA targeting sequences were as follow: 5′-AATGCCATATTGGTATATGACATAACAGG-3′ and 5′-GTAAGAATCAGATGGCATTATGCTTGTTG-3′. Two shRNAs were used separately, and a non-effective 29-mer scrambled shRNA was used as a control. Lentiviral shRNA particles were prepared using shRNA lentiviral packaging kit (Origene, TR30022). Viruses were harvested at 48 h and 72 h post-transfection. LECs were cultured on Matrigel-coated 3.5-mm dishes with lentoid formation medium for 30 days. Cells were washed twice with ice-cold PBS, and lysed in RIPA lysis buffer with PMSF. Protein concentration was determined by BCA protein assay kit. Thirty micrograms of total protein lysate was loaded onto 10% SDS–PAGE gel and then transferred to a PVDF membrane (Millipore) at 70 V for 2 h. The membrane was probed with the following primary antibody at 4 °C overnight: anti-αA-crystallin (sc-22389, Santa Cruz), anti-β-crystallin (sc-48335, Santa Cruz), anti-γ-crystallin (sc-22415, Santa Cruz) and anti-β-actin (sc-47778, Santa Cruz), and then incubated with HRP-conjugated anti-rabbit, anti-mouse, or anti-goat secondary antibody for 1 h at room temperature. The immunodetection was visualized using a blot imaging system (Fluor Chem Q, Protein Simple) with ECL buffer (Millipore). New Zealand white rabbits (n = 29, four rabbits died from systemic infections unrelated to surgery. The remaining 25 rabbits were used to assess regeneration), and long-tailed macaques (Macaca fascicularis) monkeys (n = 6) underwent minimally invasive capsulorhexis surgery. Only the left eye of each animal was used for experiments. Slit-lamp biomicroscopy and photography were performed at different time points to monitor lens regeneration. Rabbits were euthanized at day 1, day 7, and one month after surgery, and the treated eyes were enucleated. The lenses were harvested for histologic analysis using haematoxylin and eosin staining. For the macaques, enucleation of the treated eye was performed 4 months post-surgery and the lenses were harvested for the same histologic examinations. The eyes were fixed, paraffin-embedded, and sectioned at 5 μm through the cornea, pupil, and optic nerve with the lens in situ. RNA was isolated from rabbit LECs, mature lens fibre cells and LECs in P0-3.9-GFPcre mice using an RNeasy Mini Kit (Qiagen) and subjected to on-column DNase digestion. cDNA was synthesized using a Superscript III reverse transcriptase kit according to the manufacturer’s instructions (Invitrogen). Quantitative PCR was performed via 40 cycle amplification using gene-specific primers (Supplementary Table 1) and Power SYBR Green PCR Master Mix on a 7500 Real-Time PCR System (Applied Biosystems). Measurements were performed in triplicate and normalized to endogenous GAPDH levels. The relative fold change in expression was calculated using the ΔΔC method (C values <30). Rabbit LECs were fixed in 4% PFA for 20 min, then permeabilized with 0.3% Triton X-100-PBS for 10 min and blocked in PBS solution containing 5% BSA, followed by an overnight incubation in primary antibodies at 4 °C. After three washes in PBS, cells were incubated with secondary antibody for 1 h in room temperature. Cell nuclei were counterstained with DAPI. The following antibodies were used: goat anti-Sox2 polyclonal antibody (Santa Cruz), rabbit anti-PAX6 polyclonal antibody (PRB-278P, Covance), mouse anti-Bmi1 antibody (ab14389, Abcam), and mouse anti-Ki67 monoclonal antibody (550609, BD Sciences). The secondary antibodies, Alexa Fluor 488- or 568-conjugated anti-mouse or anti-rabbit IgG (Invitrogen), were used at a dilution of 1:500. Images were obtained using an Olympus FV1000 confocal microscope. We used BrdU labelling to identify and quantify proliferating LECs from human cadaver eyes. Whole-mount human lens capsules were pulsed with BrdU and then stained with an antibody against BrdU to determine the distribution and density of proliferating LECs. In brief, within 12–24 h after death, lenses from post-mortem donor eyes were obtained from the Eye Bank of Zhongshan Ophthalmic Center in Guangzhou, China. Twelve lenses in total from six donors were used for the experiment. A small puncture injury was made on the anterior surface of a post-mortem human lens using a 30-gauge needle. The lenses were cultured at 37 °C in Dulbecco modified Eagle medium (DMEM) supplemented with 10% FBS in a humidified incubator with 5% CO . The contralateral lens from the same donor was treated under the same conditions but did not receive a puncture injury and was used as a control. To label the proliferating LECs, both groups of lenses were incubated in 100 μg ml−1 BrdU (Sigma-Aldrich) 24 h after the puncture injury. The lens was then removed from the capsular bag, and the lens capsules were fixed in 4% formaldehyde and subjected to BrdU staining using a standard immunohistochemistry protocol according to the manufacturer’s instructions (CST, Boston, Massachusetts). Images were taken using a Carl Zeiss microscope (Jena, Germany). This study was approved by the institutional review board of the Zhongshan Ophthalmic Center (ZOC). Informed written consent was obtained from the parents or guardians of the infants before enrolment, and the tenets of the Declaration of Helsinki were followed throughout the study. The study was conducted in accordance with an international guideline and protocol for visual function measurements in paediatric cataract surgery and a protocol of the Childhood Cataract Program of the Chinese Ministry of Health (CCPMOH) and had an independent data and safety monitoring board of ZOC-CCPMOH. The current standard-of-care treatment for paediatric cataract involves removal of the cataractous lens through a relatively large opening using anterior continuous curvilinear capsulorhexis (ACCC, about 6 mm in diameter, Extended Data Fig. 1), followed by cataract extraction and artificial lens implantation or placement of postoperative aphakic eyeglasses/contact lens in paediatric cataract patients younger than two years. Some patients underwent additional posterior continuous curvilinear capsulorhexis (PCCC) and anterior vitrectomy. We established a new capsulorhexis surgery method to facilitate lens regeneration (Fig. 3a). First, we decreased the size of the capsulorhexis opening to 1.0–1.5 mm in diameter. This results in a minimal wound of about 1.2 mm2 in area, which is only about 4.3% the size of the wound created by the current method. Second, we moved the location of the capsulorhexis to the peripheral area of the lens instead of the central area. A 0.9 mm phacoemulsification probe was used to remove the lens contents and/or cortical opacities. These changes provide significant advantages. First, it considerably reduces the size of the injury, which resulted in a lower incidence of inflammation and much faster healing. Second, it moves the wound scar away from the central visual axis to the periphery, leading to improved visual axis transparency. Third, it preserves a nearly intact transparent lens capsule and layer of LECs, which have regenerative potential and are critically required for the regeneration of a natural lens. The clinical trial is an open label, randomized controlled trial in a study population of paediatric cataract patients (age: 0–2 years). Except the trial participants, all other parties (care providers, outcome assessors) were blinded to treatment allocation. A clinical trial consort flowchart is listed in the Extended Data Fig. 8a. Paediatric patients were enrolled accordingly inclusion and exclusion criteria below (ClinicalTrials.gov identifier: NCT01844258). Inclusion criteria were the following: infants were ≤24 months old, and diagnosed with bilateral uncomplicated congenital cataract with an intact non-fibrotic capsular bag. Exclusion criteria included preoperative intraocular pressure (IOP) >21 mm Hg, premature birth, family history of ocular disease, ocular trauma, or other abnormalities, such as microcornea, persistent hyperplastic primary vitreous, rubella, or Lowe syndrome. In total, twelve paediatric cataract patients (24 eyes) received the new minimally invasive lens surgery (Table 1). Twenty-five paediatric cataract patients (50 eyes in total) were enrolled as the control group to receive the current standard surgical treatment (Extended Data Fig. 8a). Bilateral eye surgeries of the same patient were conducted during the same operation session. We defined the incidence of corneal oedema as a >5% increase in central corneal thickness one week post-surgery, and the incidence of severe anterior chamber inflammation as a Flare value >10 evaluated by Pentacam system (OCULUS, Germany) and Laser flare meter (KOWA FM-600, Japan). Early-onset ocular hypertension was identified as IOP >21 mm Hg by Tonopen (Reichert, Seefeld, Germany) within one month after surgery. Macular oedema was identified by fundus OCT (iVue, Optovue, Germany) as an increase in central macular thickness >10% one week post-surgery. When indicated, VAO, defined by visual decline and the degree to which the fundus was obscured, was treated with YAG laser capsulotomy at follow-up. Compared to infants operated on using our new surgical technique, infants who received the traditional technique had a higher incidence of anterior chamber inflammation one week after surgery, early-onset ocular hypertension, and increased VAO (Table 1). However, in the group treated with our new method, a transparent regenerated biconvex lens was found in 100% of eyes three months after surgery, while no regenerated biconvex lenses formed in the group treated with the standard technique. In addition, 100% of the capsular openings healed within one month after surgery in the experimental group, but no capsular openings healed in the control group. Testing equipment included a set of Teller Acuity Cards (Vistech Consultants, Dayton, Ohio). The set of cards consists of 15 cards with gratings ranging in spatial frequency from 0.32 to 38 cycles per cm, in half-octave steps, and one blank grey card. A 4-mm peephole in each card allows the tester to view the child’s face through the card during testing. Test distance was kept constant by use of an aid to measure the distance from the child’s eyes to the card throughout testing. For 38 cm, the aid was the distance measured from the tester’s elbow to a specific knuckle on the tester’s hand, and for 55 cm, the aid was the length (55 cm) of the Teller Acuity Card. Testers were instructed to hold the cards without wrapping their fingers around the front side of the card, as this may attract the child’s attention. Testers presented the cards directly in front of the child and observed the child either over the top of the card or through the peephole in the card. During each acuity test, a masked visual acuity examiner was aware that the gratings were arranged in order from lower to higher spatial frequencies in half-octave steps, but were masked to the absolute spatial frequency of the grating on each card. The subset of spatial frequencies used for each test was selected according to a pseudorandom order from among three possible subsets of spatial frequencies for the subject’s age group. All three subsets for each age group included spatial frequencies known to be well above the threshold for that age group. To keep the visual acuity examiner masked to the absolute spatial frequency, the visual acuity examiner was not permitted to look at the front of the card to confirm the location of the grating. Instead, the visual acuity examiner asked an assistant to confirm the location of the grating on the card, after the visual acuity examiner had shown a card to the subject enough times to assess whether or not the subject could detect the grating. A clinical examiner was masked to the acuity results and the assigned patient group. Acuity was scored as the spatial frequency of the finest grating and was converted to log values before data analysis. We used a handheld auto-refractometer (PlusoptiX A09, OptiMed, Sydney, Australia) to evaluate the function of the regenerated lenses according to the manufacturer’s methods. Descriptive statistics was provided for the primary and secondary endpoints measured by intervention groups at each time point. Mean and standard deviation was reported for continuous variables and count and percentage is reported for categorical variables. To assess whether the primary outcome, decimal acuity, was significantly improved within each group, we performed the pre-post comparison between decimal acuity measured at baseline and study endpoint using paired t-tests. Normality of the data was checked and non-parametric alternatives, Wilcoxon signed-rank test is considered if the assumption was severely violated. To evaluate whether the mean response profiles in two groups were similar, we used the linear mixed-effect model taking account for the within-subjects correlation. The baseline decimal acuity was not adjusted by the model due to the homogeneity of this measurement as shown in the summary statistics. As the standard-of-care approach requires laser surgery at 3 months while the novel treatment does not, we fit two models using before and after laser surgery data, separately, to demonstrate the superiority of the novel approach. In each model, the outcome is the decimal acuity measured at four time points: baseline, 1 week, 3 months (before or after laser surgery) and 6 months; time (baseline as the reference level), treatment assignment and their interaction are the fixed effects; and patient is the random effect. Significant associations are identified using likelihood ratio test (LRT) by comparing models with and without a fixed effect. A linear mixed-effect model is fit again by dropping out the insignificant fixed effect until the final model is selected. A contrast test is performed when necessary. For the secondary aim, we compared the proportions of each condition of complications between two groups. We assumed the occurrence of complications for eyes from the same patient were independent. The mean difference and its 95% confidence interval was reported. A two-proportion z-test was used with the nonparametric χ2 test as alternative if the normality assumption was violated. All tests were two-sided and a P value less than 0.05 is considered to be statistically significant. Accommodative response was measured by an open-field autorefractor (SRW-5001K; Shin-Nippon, Tokyo, Japan), which allows targets to be viewed at any distance. The paediatric patients were positioned for autorefractor measurement with assistance from their parents. The patients were guided to fixate binocularly at a near target (33 cm, 5 × 5 array of smiley faces of N10 size) and a far target (3 m, 5 × 5 array of smiley faces of N10 size) by a trained and certified investigator or study coordinator. The measurements from non-cycloplegic autorefraction were performed three times at each target distance by the same trained and certified investigator throughout the study, in order to maintain accuracy and consistency throughout the trial. Measurements were taken in the same quiet environment with consistent room illumination to diminish the influence of distracting factors and to maintain subjects’ concentration. The spherical equivalent refractive value (SER) was recorded for each measurement and the mean value was calculated for evaluation of an accommodative response. The value of accommodative response was the difference between SER values for the near and the far target. We also used dynamic retinoscopy to measure the infants’ accommodation29, 30, 31. In brief, we recorded a lens dioptre value using retinoscopy when a patient was guided to fixate on a target 3 m away. Then another lens dioptre value was recorded when the target was moved closer, at a distance of 33 cm from the eyes. The difference between these two measurements was used to evaluate lens accommodative power.


News Article | November 11, 2015
Site: www.nature.com

Melanoma specimens were obtained with informed consent from all patients according to protocols approved by the Institutional Review Boards of the University of Michigan Medical School (IRBMED approvals HUM00050754 and HUM00050085; see ref. 27) and the University of Texas Southwestern Medical Center. Tumours were dissociated in Sterile Closed System Tissue Grinders (SKS Science) in enzymatic digestion medium containing 200 U ml−1 collagenase IV (Worthington) for 20 min at 37 °C. DNase (50–100 U ml−1) was added to reduce clumping of cells during digestion. Cells were filtered with a 40- μm cell strainer to obtain a single-cell suspensions. All melanomas used in this study stably expressed DsRed and luciferase so that the melanoma cells could be unambiguously distinguished from mouse cells by flow cytometry and by bioluminescence imaging. When isolated by flow cytometry, cells were also stained with antibodies against mouse CD45 (30-F11-APC, eBiosciences), mouse CD31 (390-APC, Biolegend), Ter119 (TER-119-APC, eBiosciences) and human HLA-A, -B, -C (G46-2.6-FITC, BD Biosciences) to select live human melanoma cells and to exclude contaminating mouse endothelial and haematopoietic cells. Live human melanoma cells were thus isolated by flow cytometry by sorting cells that were positive for DsRed and HLA and negative for mouse CD45, Ter119 and CD31. All antibody labelling was performed for 20 min on ice, followed by washing and centrifugation. Before flow cytometric analysis, cells were re-suspended in staining medium (L15 medium containing bovine serum albumin (1 mg ml−1), 1% penicillin/streptomycin, and 10 mM HEPES, pH 7.4) containing 4′,6-diamidino-2-phenylindole (DAPI; 5 μg ml−1; Sigma) to eliminate dead cells from sorts and analyses. Sorts and analyses were performed using a FACSAria flow cytometer (Becton Dickinson). After sorting, an aliquot of sorted melanoma cells was always reanalysed to check for purity, which was usually greater than 95%. For analysis of circulating melanoma cells, blood was collected from each mouse by cardiac puncture with a syringe pretreated with citrate-dextrose solution (Sigma). Red blood cells were precipitated by Ficoll sedimentation according to the manufacturer’s instructions (Ficoll Paque Plus, GE Healthcare). Remaining cells were washed with Hanks’ balanced salt solution (Invitrogen) before antibody staining and flow cytometric analysis. For limiting dilution analysis, cells for each mouse were sorted into individual wells of 96-well V-bottomed plates containing staining medium and loaded into syringes directly from the well (one well into one syringe into one mouse). After sorting, cells were counted and resuspended in staining medium with 25% high-protein Matrigel (product 354248; BD Biosciences). Subcutaneous injections were performed into the right flank of NOD.CB17-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice (Jackson Laboratory) in a final volume of 50 μl. Each mouse was transplanted with 100 melanoma cells unless otherwise specified. Tumour formation was evaluated regularly by palpation of the injection site, and the subcutaneous tumours were measured every 10 days until any tumour in the mouse cohort reached 2.5 cm in its largest diameter. Mice were monitored daily for signs of distress and euthanized when they exhibited distress according to a standard body condition score or within 24 h of their tumours reaching 2.5 cm in largest diameter, whichever came first. We adhered to this limit in all experiments. Organs were analysed visually and by bioluminescence imaging (see details below) for presence of macrometastases and micrometastases. These experiments were performed according to protocols approved by the Institutional Animal Care and Use Committee at the University of Texas Southwestern Medical Center (protocol 2011-0118). Intravenous injections were done by injecting cells into the tail vein of NSG mice in 100 μl of staining medium. For intrasplenic injections the mice were anaesthetized with isoflourane, then the left flank was shaved and disinfected with an ethanol wipe and iodine swab. An incision was made into the intraperitoneal cavity. The spleen was exposed with forceps and cells were injected slowly in a 40 μl volume of staining medium. The peritoneum was then sutured and skin was closed with clips. Mice were injected with buprenex before surgery and then again 12 h after surgery. A bicistronic lentiviral construct carrying dsRed2 and luciferase (dsRed2-P2A-Luc) was generated (for bioluminescence imaging) and cloned into the FUW lentivrial expression construct. The primers that were used for generating this construct were: dsRed2 forward, 5′-CGACTCTAGAGGATCCatggatagcactgagaacgtc-3′ (capital letters indicate homology to FUW backbone); dsRed2 reverse, 5′-TCCACGTCTCCAGC CTGCTTCAGCAGGCTGAAGTTAGTAGCTCCGCTTCCctggaacaggtggtggc-3′ (capital letters indicate P2A sequences); luciferase forward, 5′-GCCTGCTGAAGCAGGCTGGAGACGTGGAGGAGAACCCTGGACCTGGATCCatggaagacgccaaaaacataaag-3′ (capital letters indicate P2A sequences) and luciferase reverse, 5′-GCTTGATATCGAATTCttacacggcgatctttccgc-3′ (capital letters indicate homology to FUW backbone). All constructs were generated using the In-Fusion HD cloning system (Clontech) and sequence verified. For virus production, 0.9 μg of the appropriate plasmid and 1 μg of helper plasmids (0.4 μg pMD2G and 0.6 μg of psPAX2) were transfected into 293T cells using polyjet (Signagen) according to the manufacturer’s instructions. Replication incompetent viral supernatants were collected 48 h after transfection and filtered through a 0.45- μm filter. Approximately 300,000 freshly dissociated melanoma cells were infected with viral supernatants supplemented with 10 μg ml−1 poybrene (Sigma) for 4 h. Cells were then washed twice with staining medium, and about 25,000 cells (a mixture of infected and non-infected cells) were suspended in staining medium with 25% high-protein Matrigel (product 354248; BD Biosciences) then injected subcutaneously into NSG mice. After growing to 1–2 cm in diameter, tumours were excised and dissociated into single-cell suspensions, and luciferase-dsRed+ or green fluorescent protein (GFP)+ cells were collected by flow cytometry for injection into secondary recipients. Metastasis was monitored by bioluminescence imaging in secondary recipients. All shRNAs were expressed from a pGIPZ miRNA-based construct with TurboGFP from GE Dharmacon. For ALDH1L2, the following GE Dharmacon shRNA clones were used: V2LHS_30207, V2LHS_30209. For MTHFD1 the following GE Dharmacon shRNA clones were used: V2LHS_216208 and V2LHS_196832. Mice were injected with 100 luciferase-dsRed+ cells on the right flank and monitored until tumour diameters approached 2.5 cm, at which point they were imaged along with an uninjected control mouse using an IVIS Imaging System 200 Series (Caliper Life Sciences) with Living Image software. Mice were injected intraperitoneally with 100 μl of PBS containing d-luciferin monopotassium salt (40 μg ml−1) (Biosynth) 5 min before imaging, followed by general anaesthesia 2 min before imaging. After imaging of the whole mouse, the mice were euthanized and individual organs were surgically removed and quickly imaged. The exposure time of images ranged from 10 to 60 s depending on signal intensity. The bioluminescence signal was quantified with ‘region of interest’ measurement tools in Living Image (Perkin Elmer) software. After imaging, tumours and organs were fixed in 10% neutral-buffered formalin for histopathology. For live imaging, mice were imaged once a month, and whole body bioluminescence was quantified using Living Image Software (Perkin Elmer). Mice were euthanized by cervical dislocation. Subcutaneous tumours and metastatic nodules were dissected, immediately homogenized in 80% methanol chilled with dry ice (Honeywell), vortexed vigorously, and metabolites were extracted overnight at −80 °C. The following day, samples were centrifuged at 13,000g for 15 min at 4 °C, the supernatant was collected, and metabolites from the pellet were re-extracted with 80% methanol at −80 °C for 4 h. After centrifugation, both supernatants were pooled and lyophilized using a SpeedVac (Thermo). To inhibit spontaneous oxidation, samples were extracted with 80% methanol containing 0.1% formic acid in some experiments47. Dried metabolites were reconstituted in 0.03% formic acid in water, vortexed and centrifuged, then the supernatant was analysed using liquid chromatography-tandem mass spectrometry (LC–MS/MS). A Nexera Ultra High Performance Liquid Chromatograph (UHPLC) system (Shimadzu) was used for liquid chromatography, with a Polar-RP HPLC column (150 × 2 mm, 4 μm, 80 Å, Phenomenex) and the following gradient: 0–3 min 100% mobile phase A; 3–15 min 100–0% A; 15–17 min 0% A; 17–18 min 0–100% A; 18–23 min 100% A. Mobile phase A was 0.03% formic acid in water. Mobile phase B was 0.03% formic acid in acetonitrile. The flow rate was 0.5 ml min−1 and the column temperature was 35 °C. A triple quadrupole mass spectrometer (AB Sciex QTRAP 5500) was used for metabolite detection as previously described48. Chromatogram peak areas were integrated using Multiquant (AB Sciex). To measure GSH and GSSG levels, some metabolite extractions were performed with 0.1% formic acid in 80% methanol, to inhibit spontaneous GSH oxidation. To calculate GSH and GSSG amounts, a standard curve was prepared by adding known quantities of GSH and GSSG to tumour metabolite extracts. Mice were injected intraperitoneally with 2 g kg−1 body mass of uniformly 13C-labelled glucose (Cambridge Isotopes) and were analysed 15, 30 and 60 min later. Mice were fasted for 14 h before the injection. In most experiments, subcutaneous tumours and metastatic nodules were surgically excised and homogenized in ice cold 50% methanol for GC–MS and in 80% dry ice-cold methanol for LC–MS analysis. Metabolites were extracted with three freeze-thaw cycles in liquid nitrogen. Supernatant was collected after a 15 min centrifugation at 13,000g at 4 °C and lyophilized. Metabolites were derivatized with trimethylsilyl (TMS) at 42 °C for 30 min for GC–MS analysis. 13C-enrichment analysis was performed by GC–MS as previously described48. For LC–MS analysis, lyophilized samples were resuspended in either 0.03% formic acid in water or in 5 mM ammonium acetate in water depending on the method of analysis. For 13C-enrichment analysis of lactate, serine and glycine by LC–MS/MS, we used the liquid chromatography procedure described above for LC–MS/MS metabolomics analysis with the following modifications: the liquid chromatography gradient was 0–3 min 100% mobile phase A; 3–15 min 100–0% A; 15–17 min 0% A; 17–17.5 min 0–100% A; 17.5–20 min 100% A. For analysis of 3-PG, the liquid chromatography conditions were: mobile phase A, 5 mM ammonium acetate in water and mobile phase B, 5 mM ammonium acetate in acetonitrile, and a Fusion-RP HPLC column (150 × 2 mm, 4 μm, 80 Å, Phenomenex). The liquid chromatography gradient was: 0–3 min 100% mobile phase A; 3–9 min 100–0% A; 9–11 min 0% A; 11–12 min 0–100% A; 12–15 min 100% A. For metabolite detection a triple quadrupole mass spectrometer (AB Sciex QTRAP 5500) was used on multiple reaction monitoring mode as previously described, with some modifications48. The following transitions were used: positive mode: serine 106.1/60 (M + 1: 107.1/60 and 107.1/61, M + 2: 108.1/61 and 108.1/62, M + 3: 109.1/62), glycine 76/30 (M + 1 77/30 and 77/31, M + 2 78/31); negative mode: lactate 89/43 (M + 1 90/43 and 90/44, M + 2 91/44 and 91/45, M + 3 92/45), 3-PG 185/79 (M + 1 186/79, M + 2 187/79, M + 3 188/79) and 185/97 (M + 1 186/97, M + 2 187/97, M + 3 188/97). Unlabelled tissue was used as a negative control to confirm isotopic labelling in specific transitions. Melanomas were generally dissociated enzymatically as described above. Equal numbers of dissociated cells (500,000–2,000,000) from subcutaneous tumours, Ficoll-depleted blood, or metastatic nodules were loaded with dyes to assess mitochondrial mass, mitochondrial membrane potential, and ROS levels. The dyes that were used to assess these parameters were all obtained from Life Technologies. We stained the dissociated cells for 20–45 min at 37 °C with 5 μM Mitotracker Green, Mitotracker DeepRed, CellROX Green, or CellROX DeepRed in HBSS-free (Ca2+- and Mg2+-free) to assess mitochondrial mass, mitochondrial membrane potential, mitochondrial and cytoplasmic ROS, respectively. For each indicator, staining intensity per cell was assessed by flow cytometry in live human melanoma cells (positive for human HLA and dsRed and negative for DAPI and mouse CD45/CD31/Ter119). All animal experiments were performed according to protocols approved by the Institutional Animal Care and Use Committee at the University of Texas Southwestern Medical Center (protocol 2011-0118). Unless otherwise stated, 100 freshly dissociated melanoma cells were injected subcutaneously into the right flanks of NSG mice. When tumours became palpable, in some experiments mice were injected subcutaneously with NAC (Sigma, 200 mg kg−1 day−1 in 200 μl PBS, pH 7.4) or PBS as a control. Mice were injected with their last NAC dose 10 min before being euthanized for end-point analysis. In experiments where mice received NAC via the drinking water, NAC was dissolved in PBS at 1 mg ml−1 and the water was changed every 2 days. In other experiments methotrexate (Tocris, 1.25 mg kg−1 day−1 in 100 μl PBS) was injected intraperitoneally 5 days per week. Mice that received methotrexate were simultaneously administered thymidine (Sigma, 3 mg per mouse per day in 100 μl PBS) and hypoxanthine (Sigma, 750 μg per mouse per day in 100 μl PBS) to prevent suppression of nucleotide biosynthesis. Tumour growth was monitored weekly with a caliper. Experiments were terminated when any tumour in the cohort reached 2.5 cm in size. At the end of experiments, blood was collected by cardiac puncture. Organs were analysed for micrometastases and macrometastases by bioluminescence imaging and visual inspection. Subcutaneous tumours or metastatic nodules were surgically excised as quickly as possible after euthanizing the mice then melanoma cells were mechanically dissociated and NADPH and NADP+ were measured using NADPH/NADP Glo-Assay (Promega) following the manufactures instructions. Luminescence was measured using a using a FLUOstar Omega plate reader (BMG Labtech). Values were normalized to protein concentration, measured using a bicinchoninic acid (BCA) protein assay (Thermo). Tissue lysates were prepared in Kontes tubes with disposable pestles using RIPA Buffer (Cell Signaling Technology) supplemented with phenylmethylsulfonyl fluoride (Sigma), and protease and phosphatase inhibitor cocktails (Roche). The BCA protein assay (Thermo) was used to quantify protein concentrations. Equal amounts of protein (15–30 μg) were separated on 4–20% Tris Glycine SDS gels (BioRad) and transferred to polyvinylidene difluoride membranes (BioRad). Membranes were blocked for 30 min at room temperature with 5% milk in TBS supplemented with 0.1% Tween20 (TBST) then incubated with primary antibodies overnight at 4 °C. After incubating with horseradish peroxidase conjugated secondary antibodies (Cell Signaling Technology), membranes were developed using SuperSignal West Pico or Femto chemiluminescence reagents (Thermo). Blots were stripped with 1% SDS, 25 mM glycine, pH 2, before re-probing. The following primary antibodies were used for western blot analyses: ALDH1L2 (LifeSpan Bio; LS-C178510), DHFR (LifeSpan Bio; LS-C138829), MTHFR (LifeSpan Bio; LS-C157974), SHMT1 (Cell Signaling; 12612S), SHMT2 (Cell Signaling; 12762S), MTHFD1 (ProteinTech; 10794-1-AP), MTHFD2 (ProtenTech; 12270-1-AP) and aActin (Abcam, ab8227). Tissues were fixed in 4% paraformaldehyde for 12 h at 4 °C, and then transferred to 30% sucrose for 24 h for cryoprotection. Tissues were then frozen in OCT. Sections (10 μm) were permeabilized in PBS with 0.2% Triton (PBT), three times for 5 min each, and blocked in 5% goat serum in PBT for 30 min at room temperature. Sections were then stained with primary antibodies overnight: ALDH1L2 (LS-C178510, LifeSpan Bio; 1:50) and S100 (Z0311, Dako, 1:500). The next day, sections were washed in PBS with 0.2% Triton and stained with secondary goat anti-rabbit antibody (Invitrogen) at 1:500 for 30 min in the dark at room temperature. Sections were washed with PBT with DAPI (1:1,000) and mounted for imaging. No statistical methods were used to predetermine sample size. The data in most figure panels reflect several independent experiments performed on different days using melanomas derived from several patients. Variation is always indicated using standard deviation. For analysis of statistical significance, we first tested whether there was homogeneity of variation across treatments (as required for ANOVA) using Levene’s test, or when only two conditions were compared, using the F-test. In cases where the variation significantly differed among treatments, the data were log -transformed. If the data contained zero values, 1/2 of the smallest non-zero value was added to all measurements before log transformation. If the data contained negative values, all measurements were log-modulus transformed (L(x) = sign(x) × log(|x| + 1)). In the rare cases when the transformed data continued to exhibit variation that significantly differed among treatments, we used a non-parametric Kruskal–Wallis test or a non-parametric Mann–Whitney test to assess the significance of differences among populations and treatments. Usually, variation did not significantly differ among treatments. Under those circumstances, two-tailed Student’s t-tests were used to test the significance of differences between two treatments. When more than two treatments were compared, a one-way ANOVA followed by Dunnett’s multiple comparisons tests were performed. A two-way ANOVA followed by Dunnett’s multiple comparisons tests were used in cases where more than two groups were compared with repeated measures. Hierarchical clustering was performed using Euclidean distance in Metaboanalyst49. Mouse cages were randomized between treatments in all in vivo experiments (mice within the same cage had to be part of the same treatment). No blinding was used in any experiment. In all xenograft assays we injected 4–8-week-old NSG mice, 5 mice per treatment. Both male and female mice were used. For long-term assays, we injected 10 mice per treatment to account for non-melanoma related deaths (NSG mice are susceptible to death from opportunistic infections). When mice died before the end of experiments due to opportunistic infections the data from those mice were excluded. There were only two experiments in which this occurred. In Fig. 1c, d, 0–4 mice per melanoma line were found dead owing to an opportunistic bacterial infection before termination of the experiment and were excluded from the reported results. In Fig. 2b, 0–3 mice per melanoma line were found dead owing to opportunistic infections, before the first imaging time point after transplantation. These mice were excluded from the reported results.


News Article | February 23, 2017
Site: phys.org

The discovery, published on February 23, 2017 in the journal Cell, reveals new details about the evolution of sex. The protein acts as a nearly universal, biochemical "key" that enables two cell membranes to become one, resulting in the combination of genetic material—a necessary step for sexual reproduction. New details about the protein's function could help fight parasitic diseases, such as malaria, and boost efforts to control insect pests. "Our findings show that nature has a limited number of ways it can cause cells to fuse together into a single cell," said William Snell, a senior author of the study and a research professor in the University of Maryland Department of Cell Biology and Molecular Genetics. Snell joined UMD in June 2016, but performed the majority of the work at his previous institution, the University of Texas Southwestern Medical Center. "A protein that first made sex possible—and is still used for sexual reproduction in many of Earth's organisms—is identical to the protein used by dengue and Zika viruses to enter human cells," Snell said. "This protein must have really put the spice in the primordial soup." Snell and his colleagues studied the protein, called HAP2, in the single-celled green alga Chlamydomonas reinhardtii. HAP2 is common among single-celled protozoans, plants and arthropods—although it is not found in fungi or vertebrates such as humans. Prior results from Snell and his collaborators, as well as other research groups, indicated that HAP2 is necessary for sex cell fusion in the organisms that possess the protein. But the precise mechanism remained unclear. For the current study, Snell and his colleagues at UT Southwestern used sophisticated computer analysis tools to compare the amino acid sequence of Chlamydomonas HAP2 with that of known viral fusion proteins. The results suggested a striking degree of similarity, especially in a region called the "fusion loop" that enables the viral proteins to successfully invade a cell. If HAP2 functioned like a viral fusion protein, Snell reasoned, then disrupting HAP2's fusion loop should block its ability to fuse sex cells. Sure enough, when Snell's team changed just a single amino acid in the fusion loop of Chlamydomonas HAP2, the protein entirely lost its function. The sex cells were able to stick together—a process that depends on other proteins—but they were not able to complete the final fusion of their cell membranes. Similarly, the cells could not fuse when the researchers introduced an antibody that covered up the HAP2 fusion loop. "We were thrilled with these results, because they supported our new model of HAP2 function," Snell said. "But we needed to visualize the three-dimensional structure of the HAP2 protein to be sure it was similar to viral fusion proteins." Snell reached out to Felix Rey, a structural biologist at the Pasteur Institute in Paris who specializes in viruses. Coincidentally, Rey and his colleagues had just determined the structure of Chlamydomonas HAP2 using X-ray crystallography. Rey's results demonstrated that, indeed, HAP2 was functionally identical to dengue and Zika viral fusion proteins. "The HAP2 protein from Chlamydomonas is folded in an identical fashion to the viral proteins," Rey said, referring to the molecular folding that creates the three-dimensional structure of all proteins from a simple chain of amino acids. "The resemblance is unmistakable." HAP2 appears to be necessary for cell fusion in a wide variety of organisms, including disease-causing protozoans, invasive plants and destructive insect pests. So far, every known version of HAP2 shares the one critical amino acid in the fusion loop region. As such, HAP2 could provide a promising target for vaccines, therapies and other control methods. Snell is particularly encouraged by the possibility of controlling malaria, which is caused by the single-celled protozoan Plasmodium falciparum. "Developing a vaccine that blocks the fusion of Plasmodium sex cells would be a huge step forward," Snell said, noting that Plasmodium has a complex life cycle that depends on both mosquito and human hosts. "Our findings strongly suggest new strategies to target Plasmodium HAP2 that could disrupt the mosquito-borne stage of the Plasmodium life cycle." Explore further: Sperm-egg fusion proteins have same structure as those used by Zika and other viruses More information: The research paper, "The ancient gamete fusogen HAP2 is a eukaryotic class II fusion protein," Juliette Fedry, Yanjie Liu, Gerard Péhau-Arnaudet, Jimin Pei, Wenhao Li, M. Alejandra Tortorici, Francois Traincard, Annalisa Meola, Gerard Bricogne, Nick Grishin, William J. Snell, Félix A. Rey and Thomas Krey, was published February 23, 2017 in the journal Cell.


News Article | February 23, 2017
Site: www.sciencenews.org

Like many living things, a cancer cell cannot survive without oxygen. When young and tiny, a malignancy nestles inside a bed of blood vessels that keep it fed. As the mass grows, however, its demand for oxygen outpaces supply. Pockets within the tumor become deprived and send emergency signals for new vessel growth, a process called angiogenesis. In the 1990s, a popular cancer-fighting theory proposed interfering with angiogenesis to starve tumors to death. One magazine writer in 2000 called the strategy “the most important single insight about cancer of the past 50 years.” It made such intuitive sense. Rakesh Jain viewed angiogenesis through a different lens. Trained as an engineer, not a biologist, Jain was studying tumor vasculature during the height of excitement about drugs that could impede vessel growth. He was bothered by the fact that capillaries that arise in the tumor aren’t normal; they’re gnarled and porous, incapable of effective blood flow in the same way a leaky pipe is lousy at delivering water. The expanding tumor squeezes smaller vessels, making them even less able to transport blood. “The mantra was, ‘Let’s starve tumors,’ ” recalls Jain, director of the Edwin L. Steele Laboratories for Tumor Biology at Harvard Medical School. “I said, ‘No, we need to do the opposite.’ ” In 2001, he published a commentary in Nature Medicine predicting that angiogenesis inhibitors would not entirely shrivel the tumor. Instead, he argued, starving tumors might make them harder to treat. “I was sticking my neck out and saying this is not a good thing to do,” he says. “I had tremendous resistance.” Time has proved him right. Once they came on the market, anti-angiogenesis drugs were not the boon doctors had hoped for. Most disturbing, some patients saw their tumors shrink, only to have the disease return with renewed vengeance. Today, more than a decade after the introduction of the first tumor-starving drug, researchers have a far greater understanding of the role of oxygen deprivation in cancer. Instead of slowing tumors, hypoxia appears to trigger a metabolic panic that can drive growth, drug resistance and metastasis. Rescue proteins called hypoxia-inducible factors, or HIFs, open a bag of tricks so tumors can adapt and outrun the body’s defenses. But there’s now reason for hope: Recent insights into the effects of oxygen deprivation in cancer are sparking new ideas and providing the blueprint for treatments that could short-circuit a cancer’s ability to survive and spread, and help make existing drugs more effective. While the idea of starving cancer made sense, the approach may have underestimated the strength and complexity of a tumor’s resilience. Since oxygen is essential for so much of life, nature equips cells with elaborate safeguards that kick in when the oxygen-rich blood supply dwindles — whether the cells are part of a tumor or part of a muscle straining for one last push of strength. When oxygen levels drop, newly minted proteins stampede throughout the cell, turning on a frenzy of chemical reactions that offer protection from the crisis. Cancer cells distort this natural coping mechanism for their own means. Growing new vessels is just one move in an elaborate strategy. Many changes accompany hypoxia, including: The malignant cells loosen from each other and become less adhesive, ready to break free; tendrils of collagen, a natural binding substance, form and start to reach out to nearby vessels; and proteins appear on the cell surface to pump out lactic acid, a product of the tumor’s switch from primarily aerobic to anaerobic respiration. Researchers now think stopping enough of these and other changes could cripple the cancer. Much of the research focuses on the proteins that are among the first to deploy when a cell senses a danger of asphyxiation. “At zero oxygen, the cell can’t survive,” says Daniele Gilkes of Johns Hopkins University School of Medicine. “Inside a tumor you will see these regions of necrosis,” or dead cells. But those cells that are low on oxygen but still alive will produce new proteins: Key among them are HIF-1 and HIF-2. Both are transcription factors — they help transcribe DNA instructions into RNA. Under normal conditions, the genes that make HIF proteins are mostly silent. Once HIF proteins are made, they turn on genes — Gilkes estimates there are hundreds — that enable cells to live when oxygen concentrations are low. Gilkes’ target of choice is HIF-1. It is not only a first responder, but the protein also appears to be key to cancer’s spread. Tumors with high levels of HIF-1, particularly when concentrated at the invasive outer edge of the mass, are more likely to become metastatic, invading other parts of the body. The reverse is also true: Human tumors transplanted into mice that genetically can’t produce HIF-1 are less likely to spread. The reasons are complicated, Gilkes says, but she considers one thing really interesting: HIF-1 is involved with a lot of enzymes in collagen formation. The collagen appears to provide a means of escape. Last year, in a review in the International Journal of Molecular Sciences, Gilkes described genes, found by her lab group and others, that breast tumors activate to degrade the surrounding environment. In turn, the tumor wraps itself in a stringy web of collagen. As the collagen forms, the strands stretch outward from the tumor and latch onto nearby vessels. “We think cancer cells will find this collagen and use it to migrate and glide.” She calls them “collagen highways.” Her laboratory captured video of human tumor cells migrating along a fibrous strand. “To see them move is really scary.” Once they’ve broken from their home tumor, many types of cancer, including prostate and breast cancers, commonly move into bones. This is no coincidence, Gilkes says. Bones lack the dense thickets of blood vessels that run through soft tissues. That means cancer cells migrating from a hypoxic environment, and therefore already trained for low oxygen, would find hospitable surroundings in the bone. Her lab group is now looking for ways to block collagen formation to close the travel lanes and perhaps keep the cancer from spreading. She and others are also working to find a way to inhibit HIF-1 directly, but so far those efforts have proved challenging. HIF-1’s accomplice, HIF-2, may be a more available target. HIF-2 is a molecule made of two parts that clamp onto DNA to trigger production of other proteins that make tumors tougher to kill. In 2009, structural biologists at University of Texas Southwestern Medical Center in Dallas discovered that the HIF-2 protein had a large cavity. “Usually proteins don’t have holes inside them,” says James Brugarolas, leader of UT Southwestern’s kidney cancer program. With the discovery, researchers began working on a way to use the gap as a foothold for drugs. Now in development, the experimental drug PT2399 slips inside HIF-2 and effectively breaks the molecule in two. Brugarolas and colleagues from six other institutions and the biotech company Peloton Therapeutics Inc. in Dallas published results of the first animal tests of the compound in Nature in November. In mice with implanted grafts of human kidney tumors, PT2399 split HIF-2 and slowed growth in 56 percent of tumors — better than a standard treatment. Brugarolas hypothesizes that the drug worked only about half the time because the other half of tumors relied more heavily on HIF-1. A similar HIF-2–busting drug is now in Phase I safety testing in humans, described in June in Chicago at the annual meeting of the American Society of Clinical Oncology. While Phase I studies are not designed to test whether the treatment works, the drug showed few side effects among 51 people with advanced kidney cancer who took the drug at ever-increasing doses. The patients had already been through multiple types of treatments, one as many as seven. After taking the drug, 16 patients experienced a slowing in disease progression, three more had a partial response and one a complete reversal. Given the dearth of treatments for advanced kidney cancer, Brugarolas says, “this is a big deal.” Still more molecules throw a lifeline to hypoxic tumors in ways that scientists are just beginning to understand. In 2008, pathologist David Cheresh and colleagues at the University of California, San Diego announced a curious discovery in Nature: Depriving cells of vascular endothelial growth factor, or VEGF — the key protein responsible for new vessel growth in a tumor and the main target of drugs that block angiogenesis — could actually make tumors more aggressive. His team went on to discover the same was true for another popular class of drugs, which work by depriving a tumor cell of nutrients in the same way anti-angiogenesis drugs limit oxygen. The drugs, called EGFR inhibitors, were capable of doing the opposite of what was expected: They could make tumors stronger. Cheresh believes that hypoxia — and other stresses of low blood supply, like nutrient deprivation — inflict a wound on the tumor. When normal tissues sustain an injury (like a cut), they immediately enter a period of healing and regeneration. The bleeding stops and the skin grows back. Low oxygen delivers a blow to tumor cells, sending them into a similar state of rejuvenation, he says. “They’re now prepared to survive not only the hypoxia, but everything else thrown at it.” In 2014, Cheresh published his take on why this occurs, at least in some cases, in Nature Cell Biology. He and his team described a molecule called avb3 found on the surface of drug-resistant tumors that appears to reprogram tumor cells into a stem cell–like state. As embers of the original tumor that are often impermeable to treatment, these stem cell–like cells can lie quietly for a time and then reignite. The discovery of avb3 has redefined how Cheresh thinks about resistance. He no longer believes that tumors defy chemotherapy in the way bacteria overcome antibiotics, with only the strongest cells surviving and then roaring back to become dominant. “The tumor cells are adapting, changing in real time,” Cheresh says. In short, his data suggest that when EGFR inhibitors deprive a cell of nutrients, some cells survive not because they are naturally tougher, but because the appearance of avb3 transforms them into drug-resistant stem cells. The good news is that laboratory tests suggest an experimental drug might block this reprogramming, and it may even prevent chemotherapy resistance. A clinical trial will soon begin that combines usual cancer treatment with this avb3-disabling drug, in a one-two punch aimed at reversing or delaying resistance so the treatment can do its job. There are still more ways tumors withstand low oxygen. They start eating leftovers. HIF-1 triggers a switch from oxygen-based aerobic respiration to anaerobic respiration using pyruvate, a product of glucose breaking down. The strategy works in the short term; it’s the reason your muscles keep pumping for a time, even when you’re gasping for air on the last few yards of a sprint. Problem is, anaerobic respiration leaves a trail of lactic acid. A lot of it. “Lactic acid buildup leads to a precipitous drop in pH inside of the tumor,” says Shoukat Dedhar of the BC Cancer Research Centre in Vancouver. To compensate, HIF-1 deploys a fleet of proteins that remove the acid so it won’t accumulate and burn up the cell. Dedhar’s laboratory didn’t start out studying hypoxia. “We had tumors that were readily metastatic and genetically related tumors that couldn’t metastasize,” he says. Those tumors that easily spread were producing HIF-1, along with products from other genes. Searching for the functions of those genes, his group and others found two proteins important in pH balance. The first, MCT-4, acts like a molecular sump pump, bailing out lactic acid. But it’s not enough to normalize the pH, Dedhar says. That job goes to the second protein, carbonic anhydrase 9, or CAIX. “Its job is simply to convert carbon dioxide to bicarbonate, which then neutralizes the acid,” he says. In March 2016, in a review in Frontiers in Cell and Developmental Biology, Dedhar and colleagues described how to improve cancer treatment by taking away some of the tools for hypoxia survival — that is, keeping the cell from neutralizing acid — while simultaneously giving drugs that boost the immune system. His team has developed new compounds that specifically block CAIX. Since CAIX is almost exclusively produced in tumor cells, CAIX inhibitors should theoretically have few side effects. A Phase I safety trial is testing possible drugs now. Harvard’s Jain is still making the case for bathing the tumors in oxygen, giving them more blood, not less. This could keep the tumor from becoming hypoxic and throwing up a new series of defenses, including a flood of angiogenesis-promoting proteins, which produce tormented circulation. When he proposed that concept in 2001, “I thought abnormal vessels were bad,” he says. “I now think they are worse.” His idea is to make tumor vasculature more normal, using the very drugs that he was concerned about almost two decades ago. His research suggests that giving anti-angiogenesis drugs in modest doses will keep the vessels from becoming abnormal, making them less tortured and more capable of normal blood flow (SN: 10/5/13, p. 20). He believes the restored oxygen not only shuts down the hypoxic response that gives the cancer a survival advantage, but also serves as a conduit for chemotherapy drugs and immune cells to penetrate deeper into the tumor. Oxygen is also necessary for radiation to work. His latest experiments take the concept of more oxygen, not less, even further. He combined two chemotherapy drugs with losartan, a generic medicine used to control blood pressure. The result, reported in Nature Communications in 2013, was a delay in pancreatic and breast tumor growth in mice. Another experiment from Jain and colleagues, published in 2016 in Translational Oncology, had similar results. “We are finding every therapy works better when we do this,” he says. A clinical trial is now under way at Massachusetts General Hospital testing whether giving losartan during radiation and chemotherapy will improve results for pancreatic cancer patients. The concept still remains unproven, but Jain has reason for optimism. And he is no longer in the scientific minority. Last May, he received the National Medal of Science from President Barack Obama, who commended Jain for “groundbreaking discoveries of principles leading to the development and novel use of drugs for treatment of cancer.” Jain hopes to see the day, not long in the future, when hypoxic tumors are defeated by giving them the very thing they need the most. This article appears in the March 4, 2017, issue of Science News with the headline, "Deflating cancer: New approaches to low oxygen may thwart tumors."


The International Association of HealthCare Professionals is pleased to welcome Revathi P. Naadimuthu, MD, Ophthalmologist, to their prestigious organization with her upcoming publication in the Leading Physicians of the World. She is a highly trained and qualified ophthalmologist with an extensive expertise in all facets of her work. Dr. Revathi P. Naadimuthu has been in practice for more than 12 years and is currently serving patients at Freehold Ophthalmology LLC in Freehold, New Jersey. Additionally, she is affiliated with CentraState Medical Center and the Ocean Medical Center. Dr. Revathi P. Naadimuthu graduated with her Medical Degree in 2004 at the University of Medicine and Dentistry of New Jersey Robert Wood Johnson Medical School. She then went on to undertake an Ophthalmology residency at New York College, before her fellowship in Cornea and External Disease was completed at the University of Texas Southwestern Medical Center, Dallas. Dr. Naadimuthu is certified by the American Board of Ophthalmology, and is an expert in surgical ophthalmology, clinical ophthalmology, and conditions affecting the cornea. To keep up to date with the latest advances in her field, Dr. Naadimuthu maintains a professional membership with the American Academy of Ophthalmology, and lectures on ophthalmology annually. Dr. Naadimuthu attributes her success to the support and encouragement she has had from her family, as well as her passion for ophthalmology. Learn more about Dr. Naadimuthu by reading her upcoming publication in The Leading Physicians of the World. FindaTopDoc.com is a hub for all things medicine, featuring detailed descriptions of medical professionals across all areas of expertise, and information on thousands of healthcare topics.  Each month, millions of patients use FindaTopDoc to find a doctor nearby and instantly book an appointment online or create a review.  FindaTopDoc.com features each doctor’s full professional biography highlighting their achievements, experience, patient reviews and areas of expertise.  A leading provider of valuable health information that helps empower patient and doctor alike, FindaTopDoc enables readers to live a happier and healthier life.  For more information about FindaTopDoc, visit http://www.findatopdoc.com


News Article | February 23, 2017
Site: www.eurekalert.org

Researchers determine that a protein required for sperm-egg fusion is identical to a protein viruses use to invade host cells; discovery could help fight parasitic diseases like malaria Sexual reproduction and viral infections actually have a lot in common. According to new research, both processes rely on a single protein that enables the seamless fusion of two cells, such as a sperm cell and egg cell, or the fusion of a virus with a cell membrane. The protein is widespread among viruses, single-celled protozoans, and many plants and arthropods, suggesting that the protein evolved very early in the history of life on Earth. The discovery, published on February 23, 2017 in the journal Cell, reveals new details about the evolution of sex. The protein acts as a nearly universal, biochemical "key" that enables two cell membranes to become one, resulting in the combination of genetic material--a necessary step for sexual reproduction. New details about the protein's function could help fight parasitic diseases, such as malaria, and boost efforts to control insect pests. "Our findings show that nature has a limited number of ways it can cause cells to fuse together into a single cell," said William Snell, a senior author of the study and a research professor in the University of Maryland Department of Cell Biology and Molecular Genetics. Snell joined UMD in June 2016, but performed the majority of the work at his previous institution, the University of Texas Southwestern Medical Center. "A protein that first made sex possible -- and is still used for sexual reproduction in many of Earth's organisms -- is identical to the protein used by dengue and Zika viruses to enter human cells," Snell said. "This protein must have really put the spice in the primordial soup." Snell and his colleagues studied the protein, called HAP2, in the single-celled green alga Chlamydomonas reinhardtii. HAP2 is common among single-celled protozoans, plants and arthropods -- although it is not found in fungi or vertebrates such as humans. Prior results from Snell and his collaborators, as well as other research groups, indicated that HAP2 is necessary for sex cell fusion in the organisms that possess the protein. But the precise mechanism remained unclear. For the current study, Snell and his colleagues at UT Southwestern used sophisticated computer analysis tools to compare the amino acid sequence of Chlamydomonas HAP2 with that of known viral fusion proteins. The results suggested a striking degree of similarity, especially in a region called the "fusion loop" that enables the viral proteins to successfully invade a cell. If HAP2 functioned like a viral fusion protein, Snell reasoned, then disrupting HAP2's fusion loop should block its ability to fuse sex cells. Sure enough, when Snell's team changed just a single amino acid in the fusion loop of Chlamydomonas HAP2, the protein entirely lost its function. The sex cells were able to stick together -- a process that depends on other proteins--but they were not able to complete the final fusion of their cell membranes. Similarly, the cells could not fuse when the researchers introduced an antibody that covered up the HAP2 fusion loop. "We were thrilled with these results, because they supported our new model of HAP2 function," Snell said. "But we needed to visualize the three-dimensional structure of the HAP2 protein to be sure it was similar to viral fusion proteins." Snell reached out to Felix Rey, a structural biologist at the Pasteur Institute in Paris who specializes in viruses. Coincidentally, Rey and his colleagues had just determined the structure of Chlamydomonas HAP2 using X-ray crystallography. Rey's results demonstrated that, indeed, HAP2 was functionally identical to dengue and Zika viral fusion proteins. "The HAP2 protein from Chlamydomonas is folded in an identical fashion to the viral proteins," Rey said, referring to the molecular folding that creates the three-dimensional structure of all proteins from a simple chain of amino acids. "The resemblance is unmistakable." HAP2 appears to be necessary for cell fusion in a wide variety of organisms, including disease-causing protozoans, invasive plants and destructive insect pests. So far, every known version of HAP2 shares the one critical amino acid in the fusion loop region. As such, HAP2 could provide a promising target for vaccines, therapies and other control methods. Snell is particularly encouraged by the possibility of controlling malaria, which is caused by the single-celled protozoan Plasmodium falciparum. "Developing a vaccine that blocks the fusion of Plasmodium sex cells would be a huge step forward," Snell said, noting that Plasmodium has a complex life cycle that depends on both mosquito and human hosts. "Our findings strongly suggest new strategies to target Plasmodium HAP2 that could disrupt the mosquito-borne stage of the Plasmodium life cycle." In addition to Snell and Rey, co-authors of the study include: Juliette Fedry, Gerard Péhau-Arnaudet, M. Alejandra Tortorici, Francois Traincard and Annalisa Meola (Pasteur Institute); Yanjie Liu, Jimin Pei, Wenhao Li and Nick Grishin (UT Southwestern); Gerard Bricogne (Global Phasing, Ltd.); and Thomas Krey (Pasteur Institute, Hannover Medical School and German Center for Infection Research). The research paper, "The ancient gamete fusogen HAP2 is a eukaryotic class II fusion protein," Juliette Fedry, Yanjie Liu, Gerard Péhau-Arnaudet, Jimin Pei, Wenhao Li, M. Alejandra Tortorici, Francois Traincard, Annalisa Meola, Gerard Bricogne, Nick Grishin, William J. Snell, Félix A. Rey and Thomas Krey, was published February 23, 2017 in the journal Cell. This work was supported by the United States National Institutes of Health (Award Nos. GM56778 and GM094575), the Welch Foundation (Award No. I-1505), the European Research Council, the Pasteur Institute and the French National Center for Scientific Research. The content of this article does not necessarily reflect the views of these organizations. University of Maryland College of Computer, Mathematical, and Natural Sciences 2300 Symons Hall College Park, MD 20742 http://www. @UMDscience About the College of Computer, Mathematical, and Natural Sciences The College of Computer, Mathematical, and Natural Sciences at the University of Maryland educates more than 7,000 future scientific leaders in its undergraduate and graduate programs each year. The college's 10 departments and more than a dozen interdisciplinary research centers foster scientific discovery with annual sponsored research funding exceeding $150 million.


News Article | November 10, 2016
Site: www.eurekalert.org

Neoadjuvant endocrine therapy - designed to reduce the size of breast tumors before surgical removal - appears to be as effective as neoadjuvant chemotherapy for patients with localized, estrogen-receptor (ER)-positive breast cancer with considerably fewer side effects. The study conducted by a Massachusetts General Hospital (MGH) Cancer Center research team appears in the current print issue of JAMA Oncology and was published online earlier this year. "Estrogen-receptor-positive tumors are generally highly receptive to endocrine therapy with drugs such as tamoxifen and aromatase inhibitors. But while endocrine therapy is the most important component of adjuvant or postsurgical therapy, use of neoadjuvant endocrine therapy has been low in the U.S.," says Aditya Bardia, MD, MPH, of the MGH Cancer Center, an assistant professor of Medicine at Harvard Medical School and senior author of the study. "Since the chemotherapy more commonly used in this situation might not be the best option for patients with these tumors, we conducted a comprehensive, systematic review and meta-analysis to evaluate rigorously the existing scientific evidence." The authors note that previous studies of neoadjuvant endocrine therapy have been small and as a result had limited statistical power. Their search for prospective, randomized clinical trials of neoadjuvant therapy for ER-positive breast cancer turned up 20 studies involving almost 3,500 patients. In addition to comparing the results of neoadjuvant endocrine and chemotherapy, the researchers also analyzed whether the different types of endocrine therapy - drugs like tamoxifen, which block signaling at the estrogen receptor, or aromatase inhibitors, which inhibit estrogen production - changed treatment results. Overall, treatment outcomes were similar in patients treated with neoadjuvant chemotherapy and those receiving neoadjuvant endocrine therapy, but patients receiving chemotherapy had significantly greater toxic side effects. Comparing the results of different endocrine therapies revealed that neoadjuvant treatment with aromatase inhibitors was significantly more effective than treatment with tamoxifen-like drugs. "Endocrine therapy is an approved option for neoadjuvant treatment of localized estrogen-receptor-positive breast cancer, so there's no reason our findings cannot be applied to treatment right now," says Laura Spring, MD, the lead author and a senior oncology fellow at the MGH Cancer Center. "With the spurt in development of new targeted therapies, particularly CDK4/6 inhibitors, more research is needed to look at combining endocrine therapy with those drugs for neoadjuvant treatment." Such a study combining endocrine therapy with a CDK 4/6 inhibitor is currently underway at the MGH and other sites, and information about the trial is available at https:/ . Additional co-authors of the JAMA Oncology report are Kerry Reynolds, MD, Michelle Gadd, MD, Leif Ellisen, MD, PhD, Steven Isakoff, MD, and Beverly Moy, MD, MGH Cancer Center; and Arjun Gupta, MD, University of Texas Southwestern Medical Center. Support for the study includes a Jerry Younger Grant for Clinical and Translational Breast Cancer Research and National Institutes of Health grants 5K12 CA087723-13 and 5T32 CA071345-19. Massachusetts General Hospital, founded in 1811, is the original and largest teaching hospital of Harvard Medical School. The MGH Research Institute conducts the largest hospital-based research program in the nation, with an annual research budget of more than $800 million and major research centers in HIV/AIDS, cardiovascular research, cancer, computational and integrative biology, cutaneous biology, human genetics, medical imaging, neurodegenerative disorders, regenerative medicine, reproductive biology, systems biology, photomedicine and transplantation biology. The MGH topped the 2015 Nature Index list of health care organizations publishing in leading scientific journals, earned the prestigious 2015 Foster G. McGaw Prize for Excellence in Community Service. In August 2016 the MGH was once again named to the Honor Roll in the U.S. News & World Report list of "America's Best Hospitals."


News Article | November 9, 2015
Site: www.techtimes.com

Russian billionaire and Silicon Valley investor Yuri Milner, 53, gave out $21 million to seven Breakthrough Prize awardees Sunday, Nov. 8. The awards were given to scientists to honor their contributions to the fields of life sciences, math and physics. Among the recipients of the three-year-old award-giving body were six scientists, a huge group of researchers and a high school student. Each of the awaredees received $3 million. The seven recepients of the 2016 Breakthrough Prize Awards include Karl Deisseroth from Stanford University and Edward Boyden from Massachusetts Institute of Technology for their discovery of optogenetics, which involves the use of light signals to control brain activity and treat Parkinson's disease. Another recipient is Helen Hobbs from the University of Texas Southwestern Medical Center, who presented genetic ideas pertaining to the unsuccessful cholesterol-clearing up mechanisms of the body. Her insights will help experts develop new ways to prevent liver and heart diseases. John Hardy from the University College London, was also honored for his discovery of genetic changes in the protein plaque that accumulates in the brain during the early phase of Alzheimer's disease. The last awardee in the Life Sciences category is Svante Pääbo, the director of the Max Planck Institute for Evolutionary Anthropology, who worked on ancient genomes and DNA that shed a light on the possible links of modern humans to Neanderthals and other long-lost relatives. In the field of math, Ian Agol from the University of California at Berkeley received the award for doing the final touches on a topology theory that started in 1982. In Physics, the $3 million reward would be divided to 1,377 scientists for taking a part in a scientific collaboration that aims to reveal the mystery behind neutrinos, a typical yet elusive particle that besets all existence. The participants will receive about $2,300 each. A high school student from North Royalton, Ohio, named Ryan Chester was the youngest recipient and was awarded due to his excellent video entitled "Some Cool Ways of Looking at the Special Theory of Relativity." Aside from the seven major awards, Breakthrough Prize also gave out New Horizons Prizes for individual experts, whose scientific investigations and staff are still in the initial stages of the research. The winners were awarded during a black-tie event held at NASA's Ames Research Center in Mountain View. Animator Seth MacFarlane was the host while Grammy winner Pharell Williams performed and Hollywood celebrities Hilary Swank and Russell Crowe were presenters. "I would love to give $3 million to each one, but we're not there yet," said Milner. He added that he came up with such event to celebrate the careers of scientific experts in a glamorous way, just like how television shows and movies are recognized.


News Article | February 17, 2017
Site: www.eurekalert.org

Brooklyn, NY - A new test using peripheral vision reaction time could lead to earlier diagnosis and more effective treatment of mild traumatic brain injury, often referred to as a concussion, according to Peter J. Bergold, PhD, professor of physiology and pharmacology at SUNY Downstate Medical Center and corresponding author of a study newly published online by the Journal of Neurotrauma. While most patients with mild traumatic brain injury or concussion fully recover, a significant number do not, and earlier diagnosis could lead to better management of patients at risk for developing persistent symptoms, according to Dr. Bergold and his co-authors. Lingering symptoms may include loss of concentration and/or memory, confusion, anxiety, headaches, irritability, noise and light sensitivity, dizziness, and fatigue. "Mild traumatic brain injury is currently diagnosed with subjective clinical assessments," says Dr. Bergold. "The potential utility of the peripheral vision reaction test is clear because it is an objective, inexpensive, and rapid test that identifies mild traumatic brain injury patients who have a more severe underlying injury." Dr. Bergold's co-authors include colleagues from the University of Texas Southwestern Medical Center; The University of Texas at Dallas; Washington University; the National Institute of Neurological Disorders and Stroke; the Uniformed Services University of the Health Sciences; and SUNY Downstate. The article published by the Journal of Neurotrauma is titled "Measurement of Peripheral Vision Reaction Time Identifies White Matter Disruption in Patients with Mild Traumatic Brain Injury" and is available online at: http://online. . The research leading to the results published by the Journal of Neurotrauma was funded by an award from the United States Army Medical Research and Materiel Command, award number W81XWH1011061. Support was also received by the Center for Neuroscience and Regenerative Medicine. The United States Army Medical Research and Materiel Command had no input into the design and conduct of the study; the collection, management, analysis, or interpretation of the data; or the preparation, review, or approval of the manuscript. The content is solely the responsibility of the authors and does not necessarily represent the official views of the funding sources. SUNY Downstate Medical Center, founded in 1860, was the first medical school in the United States to bring teaching out of the lecture hall and to the patient's bedside. A center of innovation and excellence in research and clinical service delivery, SUNY Downstate Medical Center comprises a College of Medicine, College of Nursing, College of Health Related Professions, a School of Graduate Studies, a School of Public Health, University Hospital of Brooklyn, and a multifaceted biotechnology initiative including the Downstate Biotechnology Incubator and BioBAT for early-stage and more mature companies, respectively. SUNY Downstate ranks twelfth nationally in the number of alumni who are on the faculty of American medical schools. More physicians practicing in New York City have graduated from SUNY Downstate than from any other medical school. For more information, visit http://www. .


News Article | November 9, 2015
Site: news.yahoo.com

SAN FRANCISCO (Reuters) - The following is a list of winners of the Breakthrough Prizes, worth $3 million each, announced on Sunday in Mountain View, California. Karl Deisseroth, Investigator of the Howard Hughes Medical Institute and the D.H. Chen Professor of Bioengineering and of Psychiatry at Stanford University, and Ed Boyden, Professor of Biological Engineering and Brain and Cognitive Sciences at the MIT Media Lab and the MIT McGovern Institute The two will each win a separate $3 million prize for developing and implementing optogenetics – the programming of neurons to express light-activated ion channels and pumps, so that their electrical activity can be controlled by light, potentially opening a new path to treatments for Parkinson’s, depression, Alzheimer’s and blindness. Helen Hobbs, Investigator of the Howard Hughes Medical Institute and a Professor of Internal Medicine and Molecular Genetics at the University of Texas Southwestern Medical Center Hobbs discovered human genetic variants that alter the levels and distribution of cholesterol and other lipids, inspiring new approaches to the prevention of cardiovascular and liver disease. Hardy discovered mutations in the Amyloid Precursor Protein gene (APP) that cause early onset Alzheimer’s. Svante Pääbo, Director at the Max Planck Institute for Evolutionary Anthropology Pääbo pioneered the sequencing of ancient DNA and ancient genomes, illuminating the origins of modern humans and our relationships to extinct relatives such as Neanderthals. Ian Agol, Associate Professor Department of Mathematics at University of California, Berkeley Agol contributed low dimensional topology and geometric group theory, including work on the solutions of the tameness, virtual Haken and virtual fibering conjectures. The prize will be shared among 1370 physicists representing five international teams, who will share $1 million, and their seven team leaders, who will share $2 million. The teams built technically challenging experiments in underground caves to trap the neutrino, and thereby confirmed the theory of neutrino oscillation. Super-Kamiokande, led by Takaaki Kajita of the Institute for the Physics and Mathematics of the Universe in Tokyo and Director at Institute of Cosmic Ray Research, and Yoichiro Suzuki, Daya Bay, led by Yifang Wang of the Institute for High Energy Physics and Kam-Biu Luk, Professor, Department of Physics University of California, at Berkeley K2K and T2K, led by Koichiro Nishikawa of K2K – from KEK to Kamioka – Long-Baseline Neutrino Oscillation Experiment SNO, led by Arthur B. McDonald, Director of the Sudbury Neutrino Observatory Institute Ryan Chester, a high-school senior from North Royalton High School in the U.S. state of Ohio, won $250,000 in scholarships for his video explaining the theory of relativity. An additional $100,000 went to his school for a science lab, plus $50,000 to his teacher Richard Nestoff.


News Article | November 9, 2015
Site: news.yahoo.com

SAN FRANCISCO (Reuters) - The following is a list of winners of the Breakthrough Prizes, worth $3 million each, announced on Sunday in Mountain View, California. Karl Deisseroth, Investigator of the Howard Hughes Medical Institute and the D.H. Chen Professor of Bioengineering and of Psychiatry at Stanford University, and Ed Boyden, Professor of Biological Engineering and Brain and Cognitive Sciences at the MIT Media Lab and the MIT McGovern Institute The two will each win a separate $3 million prize for developing and implementing optogenetics – the programming of neurons to express light-activated ion channels and pumps, so that their electrical activity can be controlled by light, potentially opening a new path to treatments for Parkinson’s, depression, Alzheimer’s and blindness. Helen Hobbs, Investigator of the Howard Hughes Medical Institute and a Professor of Internal Medicine and Molecular Genetics at the University of Texas Southwestern Medical Center Hobbs discovered human genetic variants that alter the levels and distribution of cholesterol and other lipids, inspiring new approaches to the prevention of cardiovascular and liver disease. Hardy discovered mutations in the Amyloid Precursor Protein gene (APP) that cause early onset Alzheimer’s. Svante Pääbo, Director at the Max Planck Institute for Evolutionary Anthropology Pääbo pioneered the sequencing of ancient DNA and ancient genomes, illuminating the origins of modern humans and our relationships to extinct relatives such as Neanderthals. Ian Agol, Associate Professor Department of Mathematics at University of California, Berkeley Agol contributed low dimensional topology and geometric group theory, including work on the solutions of the tameness, virtual Haken and virtual fibering conjectures. The prize will be shared among 1370 physicists representing five international teams, who will share $1 million, and their seven team leaders, who will share $2 million. The teams built technically challenging experiments in underground caves to trap the neutrino, and thereby confirmed the theory of neutrino oscillation. Super-Kamiokande, led by Takaaki Kajita of the Institute for the Physics and Mathematics of the Universe in Tokyo and Director at Institute of Cosmic Ray Research, and Yoichiro Suzuki, Daya Bay, led by Yifang Wang of the Institute for High Energy Physics and Kam-Biu Luk, Professor, Department of Physics University of California, at Berkeley K2K and T2K, led by Koichiro Nishikawa of K2K – from KEK to Kamioka – Long-Baseline Neutrino Oscillation Experiment SNO, led by Arthur B. McDonald, Director of the Sudbury Neutrino Observatory Institute


News Article | March 31, 2016
Site: www.nanotech-now.com

Abstract: Bacteria are the most abundant form of life on Earth, and they are capable of living in diverse habitats ranging from the surface of rocks to the insides of our intestines. Over millennia, these adaptable little organisms have evolved a variety of specialized mechanisms to move themselves through their particular environments. In two recent Caltech studies, researchers used a state-of-the-art imaging technique to capture, for the first time, three-dimensional views of this tiny complicated machinery in bacteria. Credit: Science/AAAS "Bacteria are widely considered to be 'simple' cells; however, this assumption is a reflection of our limitations, not theirs," says Grant Jensen, a professor of biophysics and biology at Caltech and an investigator with the Howard Hughes Medical Institute (HHMI). "In the past, we simply didn't have technology that could reveal the full glory of the nanomachines--huge complexes comprising many copies of a dozen or more unique proteins--that carry out sophisticated functions." Jensen and his colleagues used a technique called electron cryotomography to study the complexity of these cell motility nanomachines. The technique allows them to capture 3-D images of intact cells at macromolecular resolution--specifically, with a resolution that ranges from 2 to 5 nanometers (for comparison, a whole cell can be several thousand nanometers in diameter). First, the cells are instantaneously frozen so that water molecules do not have time to rearrange to form ice crystals; this locks the cells in place without damaging their structure. Then, using a transmission electron microscope, the researchers image the cells from different angles, producing a series of 2-D images that--like a computed tomography, or CT, scan--can be digitally reconstructed into a 3-D picture of the cell's structures. Jensen's laboratory is one of only a few in the entire world that can do this type of imaging. In a paper published in the March 11 issue of the journal Science, the Caltech team used this technique to analyze the cell motility machinery that involves a structure called the type IVa pilus machine (T4PM). This mechanism allows a bacterium to move through its environment in much the same way that Spider-Man travels between skyscrapers; the T4PM assembles a long fiber (the pilus) that attaches to a surface like a grappling hook and subsequently retracts, thus pulling the cell forward. Although this method of movement is used by many types of bacteria, including several human pathogens, Jensen and his team used electron cryotomography to visualize this cell motility mechanism in intact Myxococcus xanthus--a type of soil bacterium. The researchers found that the structure is made up of several parts, including a pore on the outer membrane of the cell, four interconnected ring structures, and a stemlike structure. By systematically imaging mutants, each of which lacked one of the 10 T4PM core components, and comparing these mutants with normal M. xanthus cells, they mapped the locations of all 10 T4PM core components, providing insights into pilus assembly, structure, and function. "In this study, we revealed the beautiful complexity of this machine that may be the strongest motor known in nature. The machine lets M. xanthus, a predatory bacterium, move across a field to form a 'wolf pack' with other M. xanthus cells, and hunt together for other bacteria on which to prey," Jensen says. Another way that bacteria move about their environment is by employing a flagellum--a long whiplike structure that extends outward from the cell. The flagellum is spun by cellular machinery, creating a sort of propeller that motors the bacterium through a substrate. However, cells that must push through the thick mucus of the intestine, for example, need more powerful versions of these motors, compared to cells that only need enough propeller power to travel through a pool of water. In a second paper, published in the online early edition of the Proceedings of the National Academy of Sciences (PNAS) on March 14, Jensen and his colleagues again used electron cryotomography to study the differences between these heavy-duty and light-duty versions of the bacterial propeller. The 3-D images they captured showed that the varying levels of propeller power among several different species of bacteria can be explained by structural differences in these tiny motors. In order for the flagellum to act as a propeller, structures in the cell's motor must apply torque--the force needed to cause an object to rotate--to the flagellum. The researchers found that the high-power motors have additional torque-generating protein complexes that are found at a relatively wide radius from the flagellum. This extra distance provides greater leverage to rotate the flagellum, thus generating greater torque. The strength of the cell's motor was directly correlated with the number of these torque-generating complexes in the cell. "These two studies establish a technique for solving the complete structures of large macromolecular complexes in situ, or inside intact cells," Jensen says. "Other structure determination methods, such as X-ray crystallography, require complexes to be purified out of cells, resulting in loss of components and possible contamination. On the other hand, traditional 2-D imaging alone doesn't let you see where individual protein pieces fit in the complete structure. Our electron cryotomography technique is a good solution because it can be used to look at the whole cell, providing a complete picture of the architecture and location of these structures." ### The work involving the type IVa pilus machinery was published in a Science paper titled "Architecture of the type IVa pilus machine." First author Yi-Wei Chang is a research scientist at Caltech; additional coauthors include collaborators from the Max Planck Institute for Terrestrial Microbiology, in Marburg, Germany, and from the University of Utah. The study was funded by the National Institutes of Health (NIH), HHMI, the Max Planck Society, and the Deutsche Forschungsgemeinschaft. Work involving the flagellum machinery was published in a PNAS paper titled "Diverse high-torque bacterial flagellar motors assemble wider stator rings using a conserved protein scaffold." Additional coauthors include collaborators from Imperial College London; the University of Texas Southwestern Medical Center; and the University of Wisconsin-Madison. The study was supported by funding from the UK's Biotechnology and Biological Sciences Research Council and from HHMI and NIH. For more information, please click If you have a comment, please us. Issuers of news releases, not 7th Wave, Inc. or Nanotechnology Now, are solely responsible for the accuracy of the content.


News Article | October 3, 2016
Site: www.scientificcomputing.com

Today, the National Science Foundation (NSF) announced $10 million in awards to 10 "Big Data Spokes" projects to initiate research on specific topics identified by the Big Data Regional Innovation Hubs (BD Hubs). Project topics range from precision agriculture to personalized education. The data spokes reflect the unique priorities and capabilities of the four BD Hubs, which represent consortia from the Midwest, Northeast, South and West of the country. "The BD Spokes advance the goals and regional priorities of each BD Hub, fusing the strengths of a range of institutions and investigators and applying them to problems that affect the communities and populations within their regions," said Jim Kurose, assistant director of NSF's Computer and Information Science and Engineering Directorate. "We are pleased to be making this substantial investment today to accelerate the nation's big data R&D innovation ecosystem." NSF is also making available an additional $1 million toward planning efforts and Early-Concept Grants for Exploratory Research (EAGER) awards in support of the nation's big data innovation ecosystem. In March 2012, the Administration launched the Big Data Research and Development Initiative to improve the ability to solve some of the nation's most pressing challenges by extracting knowledge and insights from large, complex collections of digital data. The BD Hubs, announced last year, are one way NSF is addressing this need by fostering multi-sector collaborations among academia, industry and government, and bringing together a wide range of stakeholders to solve regional challenges. Like the BD Hubs, the BD Spokes will take on a convening and coordinating role, as opposed to directly conducting research. Each will gather important stakeholders, engage end users and solution providers, and form multi-disciplinary teams to tackle questions no single field can solve alone. However, unlike the BD Hubs, which aim to span the full range of data-driven challenges and solutions in a geographic region, each BD Spoke will have a specific, goal-driven mission. An example of the types of activities the awards will support includes an effort, led by a team from the Massachusetts Institute of Technology (MIT), Brown University and Drexel University, to develop a data licensing approach and automated platform that will allow individuals and organizations to share data. The platform will ensure that data sharing conforms to the imposed licensing restrictions. Partners on the project include Elsevier, Intel, Microsoft Research, Oracle, Rhode Island Hospital and Thomson Reuters. "The Big Data Hub and Big Data Spokes have been a great way to connect those of us working in Big Data in the Northeast," said Samuel Madden, professor of Electrical Engineering & Computer Science at MIT and a principal investigator on the project. "As a computer scientist focused on software for sharing data, I've been able to connect to a diverse group of researchers and leaders interested in a wide range of broader issues, ranging from hardware infrastructure to software architectures to the legal, ethical, and societal implications of data sharing." Another project, led by Gari Clifford, associate professor of Biomedical Informatics at Emory University, will investigate how to use data from diverse sources, including fitness trackers and environmental monitors, to improve patient care. As its first pilot, the project will focus on African-Americans and Latinos diagnosed with cardiovascular disease. Partners include Amazon, Emory Critical Care Center, Cerner, Relus Technologies and the University of Texas Southwestern Medical Center. Among the planning grants being announced today, Robin Gandhi from the University of Nebraska Omaha will investigate how Big Data can be used to improve America's infrastructure, starting with sensors on bridges. Abani Patra from the University at Buffalo will lead a collaboration to spur innovations in the energy sector using Big Data. The full list of planning projects is as follows: The BD Hubs and Spokes programs are part of a larger effort at NSF to advance data science and engineering. In Fiscal Year 2017, NSF will invest more than $110 million in Big Data research.


News Article | October 3, 2016
Site: www.scientificcomputing.com

Today, the National Science Foundation (NSF) announced $10 million in awards to 10 "Big Data Spokes" projects to initiate research on specific topics identified by the Big Data Regional Innovation Hubs (BD Hubs). Project topics range from precision agriculture to personalized education. The data spokes reflect the unique priorities and capabilities of the four BD Hubs, which represent consortia from the Midwest, Northeast, South and West of the country. "The BD Spokes advance the goals and regional priorities of each BD Hub, fusing the strengths of a range of institutions and investigators and applying them to problems that affect the communities and populations within their regions," said Jim Kurose, assistant director of NSF's Computer and Information Science and Engineering Directorate. "We are pleased to be making this substantial investment today to accelerate the nation's big data R&D innovation ecosystem." NSF is also making available an additional $1 million toward planning efforts and Early-Concept Grants for Exploratory Research (EAGER) awards in support of the nation's big data innovation ecosystem. In March 2012, the Administration launched the Big Data Research and Development Initiative to improve the ability to solve some of the nation's most pressing challenges by extracting knowledge and insights from large, complex collections of digital data. The BD Hubs, announced last year, are one way NSF is addressing this need by fostering multi-sector collaborations among academia, industry and government, and bringing together a wide range of stakeholders to solve regional challenges. Like the BD Hubs, the BD Spokes will take on a convening and coordinating role, as opposed to directly conducting research. Each will gather important stakeholders, engage end users and solution providers, and form multi-disciplinary teams to tackle questions no single field can solve alone. However, unlike the BD Hubs, which aim to span the full range of data-driven challenges and solutions in a geographic region, each BD Spoke will have a specific, goal-driven mission. An example of the types of activities the awards will support includes an effort, led by a team from the Massachusetts Institute of Technology (MIT), Brown University and Drexel University, to develop a data licensing approach and automated platform that will allow individuals and organizations to share data. The platform will ensure that data sharing conforms to the imposed licensing restrictions. Partners on the project include Elsevier, Intel, Microsoft Research, Oracle, Rhode Island Hospital and Thomson Reuters. "The Big Data Hub and Big Data Spokes have been a great way to connect those of us working in Big Data in the Northeast," said Samuel Madden, professor of Electrical Engineering & Computer Science at MIT and a principal investigator on the project. "As a computer scientist focused on software for sharing data, I've been able to connect to a diverse group of researchers and leaders interested in a wide range of broader issues, ranging from hardware infrastructure to software architectures to the legal, ethical, and societal implications of data sharing." Another project, led by Gari Clifford, associate professor of Biomedical Informatics at Emory University, will investigate how to use data from diverse sources, including fitness trackers and environmental monitors, to improve patient care. As its first pilot, the project will focus on African-Americans and Latinos diagnosed with cardiovascular disease. Partners include Amazon, Emory Critical Care Center, Cerner, Relus Technologies and the University of Texas Southwestern Medical Center. Among the planning grants being announced today, Robin Gandhi from the University of Nebraska Omaha will investigate how Big Data can be used to improve America's infrastructure, starting with sensors on bridges. Abani Patra from the University at Buffalo will lead a collaboration to spur innovations in the energy sector using Big Data. The full list of planning projects is as follows: The BD Hubs and Spokes programs are part of a larger effort at NSF to advance data science and engineering. In Fiscal Year 2017, NSF will invest more than $110 million in Big Data research.


Miller W.L.,University of California at San Francisco | Auchus R.J.,University of Texas Southwestern Medical Center
Endocrine Reviews | Year: 2011

Steroidogenesis entails processes by which cholesterol is converted to biologically active steroid hormones. Whereas most endocrine texts discuss adrenal, ovarian, testicular, placental, and other steroidogenic processes in a gland-specific fashion, steroidogenesis is better understood as a single process that is repeated in each gland with cell-type-specific variations on a single theme. Thus, understanding steroidogenesis is rooted in an understanding of the biochemistry of the various steroidogenic enzymes and cofactors and the genes that encode them. The first and rate-limiting step in steroidogenesis is the conversion of cholesterol to pregnenolone by a single enzyme, P450scc (CYP11A1), but this enzymatically complex step is subject to multiple regulatory mechanisms, yielding finely tuned quantitative regulation. Qualitative regulation determining the type of steroid to be produced is mediated by many enzymes and cofactors. Steroidogenic enzymes fall into two groups: cytochrome P450 enzymes and hydroxysteroid dehydrogenases. A cytochrome P450 may be either type 1 (in mitochondria) or type 2 (in endoplasmic reticulum), and a hydroxysteroid dehydrogenase may belong to either the aldo-keto reductase or short-chain dehydrogenase/reductase families. The activities of these enzymes are modulated by posttranslational modifications and by cofactors, especially electron-donating redox partners. The elucidation of the precise roles of these various enzymes and cofactors has been greatly facilitated by identifying the genetic bases of rare disorders of steroidogenesis. Some enzymes not principally involved in steroidogenesis may also catalyze extraglandular steroidogenesis, modulating the phenotype expected to result from some mutations. Understanding steroidogenesis is of fundamental importance to understanding disorders of sexual differentiation, reproduction, fertility, hypertension, obesity, and physiological homeostasis. Copyright © 2011 by The Endocrine Society.


Cohen I.G.,Harvard University | Amarasingham R.,University of Texas Southwestern Medical Center | Lo B.,University of California at San Francisco
Health Affairs | Year: 2014

Predictive analytics, or the use of electronic algorithms to forecast future events in real time, makes it possible to harness the power of big data to improve the health of patients and lower the cost of health care. However, this opportunity raises policy, ethical, and legal challenges. In this article we analyze the major challenges to implementing predictive analytics in health care settings and make broad recommendations for overcoming challenges raised in the four phases of the life cycle of a predictive analytics model: acquiring data to build the model, building and validating it, testing it in real-world settings, and disseminating and using it more broadly. For instance, we recommend that model developers implement governance structures that include patients and other stakeholders starting in the earliest phases of development. In addition, developers should be allowed to use already collected patient data without explicit consent, provided that they comply with federal regulations regarding research on human subjects and the privacy of health information. © 2014 by Project HOPE - The People-to-People Health Foundation.


Lu M.,National Institute of Allergy and Infectious Diseases | Varley A.W.,University of Texas Southwestern Medical Center | Munford R.S.,National Institute of Allergy and Infectious Diseases
PLoS Pathogens | Year: 2013

Measures that bolster the resolution phase of infectious diseases may offer new opportunities for improving outcome. Here we show that inactivation of microbial lipopolysaccharides (LPS) can be required for animals to recover from the innate immune tolerance that follows exposure to Gram-negative bacteria. When wildtype mice are exposed to small parenteral doses of LPS or Gram-negative bacteria, their macrophages become reprogrammed (tolerant) for a few days before they resume normal function. Mice that are unable to inactivate LPS, in contrast, remain tolerant for several months; during this time they respond sluggishly to Gram-negative bacterial challenge, with high mortality. We show here that prolonged macrophage reprogramming is maintained in vivo by the persistence of stimulatory LPS molecules within the cells' in vivo environment, where naïve cells can acquire LPS via cell-cell contact or from the extracellular fluid. The findings provide strong evidence that inactivation of a stimulatory microbial molecule can be required for animals to regain immune homeostasis following parenteral exposure to bacteria. Measures that disable microbial molecules might enhance resolution of tissue inflammation and help restore innate defenses in individuals recovering from many different infectious diseases.


Skaug B.,University of Texas Southwestern Medical Center | Chen J.,University of Texas Southwestern Medical Center | Du F.,University of Texas Southwestern Medical Center | He J.,University of North Carolina at Chapel Hill | And 2 more authors.
Molecular Cell | Year: 2011

A20 is a potent anti-inflammatory protein that inhibits NF-κB, and A20 dysfunction is associated with autoimmunity and B cell lymphoma. A20 harbors a deubiquitination enzyme domain and can employ multiple mechanisms to antagonize ubiquitination upstream of NEMO, a regulatory subunit of the IκB kinase complex (IKK). However, direct evidence of IKK inhibition by A20 is lacking, and the inhibitory mechanism remains poorly understood. Here we show that A20 can directly impair IKK activation without deubiquitination or impairment of ubiquitination enzymes. We find that polyubiquitin binding by A20, which is largely dependent on A20's seventh zinc-finger motif (ZnF7), induces specific binding to NEMO. Remarkably, this ubiquitin-induced recruitment of A20 to NEMO is sufficient to block IKK phosphorylation by its upstream kinase TAK1. Our results suggest a noncatalytic mechanism of IKK inhibition by A20 and a means by which polyubiquitin chains can specify a signaling outcome. © 2011 Elsevier Inc.


Montalbano A.P.,University of Texas Southwestern Medical Center | Hawgood S.,University of California at San Francisco | Mendelson C.R.,University of Texas Southwestern Medical Center
Endocrinology | Year: 2013

Previously we obtained compelling evidence that the fetus provides a critical signal for the initiation of term labor through developmental induction of surfactant protein (SP)-A expression by the fetal lung and secretion into amniotic fluid (AF). We proposed that interactions of AF macrophage(Mφ) Toll-like receptors (TLRs) with SP-A, at term, or bacterial components, at preterm, result in their activation and migration to the pregnant uterus. Herein the timing of labor in wild-type (WT) C57BL/6 mice was compared with mice homozygous null for TLR2, SP-A, SP-D, or doubly deficient in SP-A and SP-D. Interestingly, TLR2-/- females manifested a significant (P<0.001) delay in timing of labor compared with WT as well as reduced expression of the myometrial contraction-associated protein (CAP) gene, connexin-43, and Mφ marker, F4/80, at 18.5 d postcoitum (dpc). Whereas in first pregnancies, SP-A-/-, SP-D-/-, and SP-A/D-/- females delivered at term (∼19.5 dpc), in second pregnancies, parturition was delayed by approximately 12 h in SP-A-/- (P = 0.07) and in SP-A/D-/- (P <0.001) females. Myometrium of SP-A/D-/- females expressed significantly lower levels of IL-1β, IL-6, and CAP genes, connexin-43, and oxytocin receptor at 18.5 dpc compared with WT. F4/80 + AF Mφs from TLR2-/- and SP-A/D-/- mice expressed significantly lower levels of both proinflammatory and antiinflammatory activation markers (e.g. IL-1β, IL-6, ARG1, YM1) compared with gestation-matched WT AF Mφs. These novel findings suggest that the pulmonary collectins acting via TLR2 serve a modulatory role in the timing of labor; their relative impact may be dependent on parity. Copyright © 2013 by The Endocrine Society.


Ku C.-J.,University of Texas Southwestern Medical Center | Wang Y.,University of Texas Southwestern Medical Center | Weiner O.D.,University of California at San Francisco | Altschuler S.J.,University of Texas Southwestern Medical Center | Wu L.F.,University of Texas Southwestern Medical Center
Cell | Year: 2012

How complex signaling networks shape highly coordinated, multistep cellular responses is poorly understood. Here, we made use of a network-perturbation approach to investigate causal influences, or "crosstalk," among signaling modules involved in the cytoskeletal response of neutrophils to chemoattractant. We quantified the intensity and polarity of cytoskeletal marker proteins over time to characterize stereotyped cellular responses. Analyzing the effects of network disruptions revealed that, not only does crosstalk evolve rapidly during polarization, but also that intensity and polarity responses are influenced by different patterns of crosstalk. Interestingly, persistent crosstalk is arranged in a surprisingly simple circuit: a linear cascade from front to back to microtubules influences intensities, and a feed-forward network in the reverse direction influences polarity. Our approach provided a rational strategy for decomposing a complex, dynamically evolving signaling system and revealed evolving paths of causal influence that shape the neutrophil polarization response. © 2012 Elsevier Inc.


Houk A.R.,University of California at San Francisco | Jilkine A.,University of Texas Southwestern Medical Center | Jilkine A.,University of Arizona | Mejean C.O.,Yale University | And 6 more authors.
Cell | Year: 2012

Little is known about how neutrophils and other cells establish a single zone of actin assembly during migration. A widespread assumption is that the leading edge prevents formation of additional fronts by generating long-range diffusible inhibitors or by sequestering essential polarity components. We use morphological perturbations, cell-severing experiments, and computational simulations to show that diffusion-based mechanisms are not sufficient for long-range inhibition by the pseudopod. Instead, plasma membrane tension could serve as a long-range inhibitor in neutrophils. We find that membrane tension doubles during leading-edge protrusion, and increasing tension is sufficient for long-range inhibition of actin assembly and Rac activation. Furthermore, reducing membrane tension causes uniform actin assembly. We suggest that tension, rather than diffusible molecules generated or sequestered at the leading edge, is the dominant source of long-range inhibition that constrains the spread of the existing front and prevents the formation of secondary fronts. Video Abstract: © 2012 Elsevier Inc.


Villasenor A.,University of California at San Francisco | Cleaver O.,University of Texas Southwestern Medical Center
Seminars in Cell and Developmental Biology | Year: 2012

Growth and development of embryonic organs goes hand in hand with development of the vascular system. Blood vessels have been known for centuries to supply nutrients and oxygen to all cell types in an organism, however, they have more recently been shown to provide specific cues required for the formation and functionality of a number of tissues. Here, we review the role of blood vessels during pancreas formation, from early specification of the initial pancreatic bud, to its growth and maturation. The overarching theme that emerges from the many studies carried out in the past decade is that the vasculature likely plays diverse and changing roles during pancreas organogenesis. Blood vessels are required for endocrine specification at the onset of pancreatic budding, while only a few days later, blood vessels suppress pancreatic branching and exocrine differentiation. In this review, we summarize our understanding to date about the crosstalk between the pancreas and its vasculature, and we provide a perspective on the promises and challenges of the field. © 2012 .


Yu C.-H.,Leiden University | Yu C.-H.,University of Texas Southwestern Medical Center | Teulade-Fichou M.-P.,Leiden University | Olsthoorn R.C.L.,Leiden University | Olsthoorn R.C.L.,University Paris - Sud
Nucleic Acids Research | Year: 2014

Guanine-rich sequences can fold into four-stranded structures of stacked guanine-tetrads, so-called G-quadruplexes (G4). These unique motifs have been extensively studied on the DNA level; however, exploration of the biological roles of G4s at the RNA level is just emerging. Here we show that G4 RNA when introduced within coding regions are capable of stimulating -1 ribosomal frameshifting (-1 FS) in vitro and in cultured cells. Systematic manipulation of the loop length between each G-tract revealed that the -1 FS efficiency positively correlates with G4 stability. Addition of a G4-stabilizing ligand, PhenDC3, resulted in higher -1 FS. Further, we demonstrated that the G4s can stimulate +1 FS and stop codon readthrough as well. These results suggest a potentially novel translational gene regulation mechanism mediated by G4 RNA. © 2013 The Author(s) 2013. Published by Oxford University Press.


Manzanillo P.S.,University of California at San Francisco | Shiloh M.U.,University of Texas Southwestern Medical Center | Portnoy D.A.,University of California at Berkeley | Cox J.S.,University of California at San Francisco
Cell Host and Microbe | Year: 2012

Cytosolic bacterial pathogens activate the cytosolic surveillance pathway (CSP) and induce innate immune responses, but how the host detects vacuolar pathogens like Mycobacterium tuberculosis is poorly understood. We show that M. tuberculosis also initiates the CSP upon macrophage infection via limited perforation of the phagosome membrane mediated by the ESX-1 secretion system. Although the bacterium remains within the phagosome, this permeabilization results in phagosomal and cytoplasmic mixing and allows extracellular mycobacterial DNA to access host cytosolic receptors, thus blurring the distinction between "vacuolar" and "cytosolic" pathogens. Activation of cytosolic receptors induces signaling through the Sting/Tbk1/Irf3 axis, resulting in IFN-β production. Surprisingly, Irf3-/- mice, which cannot respond to cytosolic DNA, are resistant to long-term M. tuberculosis infection, suggesting that the CSP promotes M. tuberculosis infection. Thus, cytosolic sensing of mycobacterial DNA plays a key role in M. tuberculosis pathogenesis and likely contributes to the high type I IFN signature in tuberculosis. © 2012 Elsevier Inc.


Liu H.,University of Texas Southwestern Medical Center | Rankin S.,Oklahoma Medical Research Foundation | Yu H.,University of Texas Southwestern Medical Center
Nature Cell Biology | Year: 2013

Timely dissolution of sister-chromatid cohesion in mitosis ensures accurate chromosome segregation to guard against aneuploidy and tumorigenesis. The complex of shugoshin and protein phosphatase 2A (SGO1-PP2A) protects cohesin at centromeres from premature removal by mitotic kinases and WAPL in prophase. Here we address the regulation and mechanism of human SGO1 in centromeric cohesion protection, and show that cyclin-dependent kinase (CDK)-mediated, mitosis-specific phosphorylation of SGO1 activates its cohesion-protection function and enables its direct binding to cohesin. The phospho-SGO1-bound cohesin complex contains PP2A, PDS5 and hypophosphorylated sororin, but lacks WAPL. Expression of non-phosphorylatable sororin bypasses the requirement for SGO1-PP2A in centromeric cohesion. Thus, mitotic phosphorylation of SGO1 targets SGO1-PP2A to cohesin, promotes dephosphorylation of PDS5-bound sororin and protects centromeric cohesin from WAPL. PP2A-orchestrated, site-selective dephosphorylation of cohesin and its regulators underlies centromeric cohesion protection. © 2013 Macmillan Publishers Limited. All rights reserved.


Chalasani N.P.,Indiana University | Hayashi P.H.,University of North Carolina at Chapel Hill | Bonkovsky H.L.,CarolinasHealthCare System | Navarro V.J.,Care Network | And 2 more authors.
American Journal of Gastroenterology | Year: 2014

Idiosyncratic drug-induced liver injury (DILI) is a rare adverse drug reaction and it can lead to jaundice, liver failure, or even death. Antimicrobials and herbal and dietary supplements are among the most common therapeutic classes to cause DILI in the Western world. DILI is a diagnosis of exclusion and thus careful history taking and thorough work-up for competing etiologies are essential for its timely diagnosis. In this ACG Clinical Guideline, the authors present an evidence-based approach to diagnosis and management of DILI with special emphasis on DILI due to herbal and dietary supplements and DILI occurring in individuals with underlying liver disease. © 2014 by the American College of Gastroenterology.


News Article | February 24, 2017
Site: www.businesswire.com

BURLINGTON, Mass.--(BUSINESS WIRE)--ArQule, Inc. (Nasdaq: ARQL) today announced it will report financial results for the fourth quarter and full year 2016 before the market opens on Tuesday, March 7, 2017. The Company will hold a conference call and webcast on the same day at 9:00 a.m. ET to discuss these results and provide a general business update. The live webcast can be accessed in the “Investors & Media” section of our website, www.arqule.com, under “Events & Presentations." You may also listen to the call by dialing (877) 868-1831 within the U.S. or (914) 495-8595 outside the U.S. A replay will be available two hours after the completion of the call and can be accessed in the “Investors & Media” section of our website, www.arqule.com, under “Events and Presentations." ArQule is a biopharmaceutical company engaged in the research and development of targeted therapeutics to treat cancers and rare diseases. Our mission is to discover, develop and commercialize novel small molecule drugs in areas of high unmet need that will dramatically extend and improve the lives of our patients. Our clinical-stage pipeline consists of five drug candidates, all of which are in targeted, biomarker-defined patient populations, making ArQule a leader among companies our size in precision medicine. ArQule’s most advanced product, in phase 3 clinical development, is tivantinib (ARQ 197), an oral, selective inhibitor of the c-MET receptor tyrosine kinase, for second-line treatment of MET-overexpressing hepatocellular carcinoma in partnership with Daiichi Sankyo in the West and Kyowa Hakko Kirin in Asia. ArQule’s proprietary pipeline includes: ARQ 087, a multi-kinase inhibitor designed to preferentially inhibit the fibroblast growth factor receptor (FGFR) family, in phase 2 for iCCA and in phase 1b for multiple oncology indications; ARQ 092, a selective inhibitor of the AKT serine/threonine kinase, in phase 1 for multiple oncology indications as well as ultra-rare Proteus syndrome, in partnership with the National Institutes of Health (NIH); ARQ 751, a next generation AKT inhibitor, in phase 1 for patients with AKT1 and PI3K mutations; and ARQ 761, a β-lapachone analog being evaluated as a promoter of NQO1-mediated programmed cancer cell necrosis, in phase 1/2 in multiple oncology indications in partnership with the University of Texas Southwestern Medical Center. In addition, we have advanced ARQ 531, an investigational, orally bioavailable, potent and reversible inhibitor of both wild type and C481S-mutant BTK, through toxicology testing and plan to file an Investigational New Drug Application in early 2017. ArQule’s current discovery efforts are focused on the identification and development of novel kinase inhibitors, leveraging the Company’s proprietary library of compounds. You can follow us on Twitter and LinkedIn.


BURLINGTON, Mass.--(BUSINESS WIRE)--ArQule, Inc. (Nasdaq: ARQL) today announced that preclinical data was presented on Bruton's tyrosine kinase (BTK) inhibitor, ARQ 531, in a poster presentation by The Ohio State University at the American Society of Hematology (ASH) Annual Meeting. The presentation highlighted preclinical studies of ARQ 531 in Chronic Lymphocytic Leukemia (CLL). ARQ 531 is an investigational, orally bioavailable, potent and reversible BTK inhibitor. Title: The Bruton's Tyrosine Kinase (BTK) Inhibitor ARQ 531 Effectively Inhibits Wild Type and C481S Mutant BTK and Is Superior to Ibrutinib in a Mouse Model of Chronic Lymphocytic Leukemia The presentation can be viewed at https://www.arqule.com/wp-content/uploads/ASH-2016-ARQ-531-in-CLL.pdf. "Irreversible kinase inhibitors directed at BTK have really changed the landscape of CLL but at extended follow up, we are beginning to see a subset of high risk patients who are relapsing,” said Dr. Jennifer Woyach, M.D., of The Ohio State University College of Medicine. “Small molecules that target BTK that are not dependent upon the C481 site represent an exciting option for future clinical trials. We are excited to be working with ArQule on this project and look forward to initiating the first in man study with ARQ 531." “We began our BTK discovery program in 2011 which ultimately lead to the selection of ARQ 531, a potent reversible inhibitor of both wild type and mutant BTK,” said Dr. Giovanni Abbadessa, M.D., PhD., Vice President of Clinical Development, Translational Medicine and Medical Affairs at ArQule. “With the recent emergence in 2015 of BTK resistance we concentrated our efforts in this growing CLL patient population. We are pleased to be working with The Ohio State University to finish preclinical studies on this exciting program. We remain on track to file an IND application early next year.” ARQ 531 is an investigational, orally bioavailable, potent and reversible Bruton’s tyrosine kinase (BTK) inhibitor. Biochemical and cellular studies have shown that ARQ 531inhibits both the wild type and C481S-mutant forms of BTK. The C481S mutation is a known emerging resistance mechanism for first generation irreversible BTK inhibitors. ARQ 531 has high oral bioavailability as well as good ADME, pharmacokinetic and metabolic properties. The company plans to file an IND for ARQ 531 in early 2017. BTK is a therapeutic target that has been clinically proven to inhibit B-cell receptor signaling in blood cancers. ArQule is a biopharmaceutical company engaged in the research and development of targeted therapeutics to treat cancers and rare diseases. Our mission is to discover, develop and commercialize novel small molecule drugs in areas of high unmet need that will dramatically extend and improve the lives of our patients. Our clinical-stage pipeline consists of five drug candidates, all of which are in targeted, biomarker-defined patient populations, making ArQule a leader among companies our size in precision medicine. ArQule’s lead product, in phase 3 clinical development, is tivantinib (ARQ 197), an oral, selective inhibitor of the c-MET receptor tyrosine kinase, for second-line treatment of hepatocellular carcinoma in partnership with Daiichi Sankyo in the West and Kyowa Hakko Kirin in Asia. ArQule’s proprietary pipeline includes: ARQ 087, a multi-kinase inhibitor designed to preferentially inhibit the fibroblast growth factor receptor (FGFR) family, in phase 2 for iCCA and in phase 1b for multiple oncology indications; ARQ 092, a selective inhibitor of the AKT serine/threonine kinase, in phase 1 for multiple oncology indications as well as ultra-rare Proteus syndrome, in partnership with the National Institutes of Health (NIH); ARQ 751, a next generation AKT inhibitor, in phase 1 for patients with AKT1 and PI3K mutations; and ARQ 761, a β-lapachone analog being evaluated as a promoter of NQO1-mediated programmed cancer cell necrosis, in phase 1/2 in multiple oncology indications in partnership with the University of Texas Southwestern Medical Center. In addition, we have advanced ARQ 531, an investigational, orally bioavailable, potent and reversible inhibitor of both wild type and C481S-mutant BTK, into toxicology testing and plan to file an Investigational New Drug Application in early 2017. ArQule’s current discovery efforts are focused on the identification and development of novel kinase inhibitors, leveraging the Company’s proprietary library of compounds. You can follow us on Twitter and LinkedIn. This press release contains forward-looking statements regarding preclinical experiments and planned clinical trials with ARQ 531. These statements are based on the Company’s current beliefs and expectations, and are subject to risks and uncertainties that could cause actual results to differ materially. Positive information about pre-clinical results does not ensure that clinical trials will be successful. For example, ARQ 531 may not demonstrate promising therapeutic effect in man; in addition, and ARQ 531 may not exhibit an adequate safety profile in ongoing toxicology testing and may not demonstrate appropriate safety in planned, current or later stage or larger scale clinical trials as a result of known or as yet unanticipated side effects. The results achieved in later stage trials may not be sufficient to meet applicable regulatory standards or to justify further development. Problems or delays may arise during clinical trials or in the course of developing, testing or manufacturing these compounds that could lead the Company to discontinue development. Even if later stage clinical trials are successful, unexpected concerns may arise from subsequent analysis of data or from additional data. Obstacles may arise or issues may be identified in connection with review of clinical data with regulatory authorities. Regulatory authorities may disagree with the Company’s view of the data or require additional data or information or additional studies. Drug development involves a high degree of risk. Only a small number of research and development programs result in the commercialization of a product. Furthermore, ArQule may not have the financial or human resources to successfully pursue drug discovery in the future. For more detailed information on the risks and uncertainties associated with the Company’s drug development and other activities, see the Company’s periodic reports filed with the Securities and Exchange Commission. The Company does not undertake any obligation to publicly update any forward-looking statements.


ArQule to host investor conference call on February 17, 2017 at 8:30 A.M. ET BURLINGTON, Mass. and TOKYO and MUNICH and PARSIPPANY, N.J., Feb. 17, 2017 /PRNewswire/ -- ArQule, Inc. (Nasdaq: ARQL) and Daiichi Sankyo today announced that the METIV-HCC phase 3 study of tivantinib in hepatocellular carcinoma (HCC) did not meet its primary endpoint of improving overall survival. METIV-HCC is a biomarker-selected, double-blind, placebo-controlled, randomized phase 3 study evaluating tivantinib (2:1) versus best supportive care in patients with MET-overexpressing, inoperable HCC intolerant to or previously-treated with systemic therapy. A total of 340 patients with MET-overexpressing HCC analyzed by a validated immunohistochemical assay were randomized in the intent-to-treat population for efficacy analysis. The primary endpoint of the study is overall survival. Secondary endpoints include progression-free survival and safety. Full results from the trial will be presented at an upcoming scientific forum. "HCC is a disease with high unmet need, especially in the second-line setting, so these results are disappointing for the patients as well as the investigators and the companies," said Paolo Pucci, Chief Executive Officer of ArQule. "Despite the negative outcome of this study, we remain committed to applying rigorous science to unmet needs for patients with cancer," said Antoine Yver, MD, MSc, Executive Vice President and Global Head, Oncology Research and Development, Daiichi Sankyo. "We would like to take this opportunity to thank all of the investigators, and especially the patients, for their participation in this study." The ArQule investor conference call can be accessed in the "Investors and Media" section of ArQule's website, www.arqule.com, under "Events and Presentations." You may also listen to the call by dialing (877) 868-1831 within the U.S. or (914) 495-8595 outside the U.S. and using the passcode 74015633. A replay will be available two hours after the completion of the call and can be accessed in the "Investors and Media" section of our website, www.arqule.com, under "Events and Presentations." About Hepatocellular Carcinoma (HCC) Liver cancer is the sixth most common cancer globally with 782,000 new cases in 2012 and is the second most common cause of cancer-related death with 745,000 deaths in 2012.1 HCC accounts for about 90 percent of primary liver cancers.2 Cirrhosis, chronic hepatitis B and C and smoking are recognized worldwide as factors increasing the risk of HCC.2 About Tivantinib (ARQ 197) ArQule and Daiichi Sankyo have a licensing, co-development and co-commercialization agreement for tivantinib in the U.S., Europe, South America and the rest of the world, excluding Japan, China (including Hong Kong), South Korea and Taiwan. About ArQule ArQule is a biopharmaceutical company engaged in the research and development of targeted therapeutics to treat cancers and rare diseases. Our mission is to discover, develop and commercialize novel small molecule drugs in areas of high unmet need that will dramatically extend and improve the lives of our patients. Our clinical-stage pipeline consists of five drug candidates, all of which are in targeted, biomarker-defined patient populations, making ArQule a leader among companies our size in precision medicine. ArQule's lead product, in phase 3 clinical development, is tivantinib (ARQ 197), an oral, selective inhibitor of the c-MET receptor tyrosine kinase, for second-line treatment of patients with MET-overexpressing hepatocellular carcinoma in partnership with Daiichi Sankyo in the West and Kyowa Hakko Kirin in Asia. ArQule's proprietary pipeline includes: ARQ 087, a multi-kinase inhibitor designed to preferentially inhibit the fibroblast growth factor receptor (FGFR) family, in phase 2 for iCCA and in phase 1b for multiple oncology indications; ARQ 092, a selective inhibitor of the AKT serine/threonine kinase, in phase 1 for multiple oncology indications as well as ultra-rare Proteus syndrome, in partnership with the National Institutes of Health (NIH); ARQ 751, a next generation AKT inhibitor, in phase 1 for patients with AKT1 and PI3K mutations; and ARQ 761, a β-lapachone analog being evaluated as a promoter of NQO1-mediated programmed cancer cell necrosis, in phase 1/2 in multiple oncology indications in partnership with the University of Texas Southwestern Medical Center. In addition, we have advanced ARQ 531, an investigational, orally bioavailable, potent and reversible inhibitor of both wild type and C481S-mutant BTK, into toxicology testing and plan to file an Investigational New Drug Application in early 2017. ArQule's current discovery efforts are focused on the identification and development of novel kinase inhibitors, leveraging the Company's proprietary library of compounds. You can follow us on Twitter and LinkedIn. About Daiichi Sankyo Cancer Enterprise The vision of Daiichi Sankyo Cancer Enterprise is to leverage our world-class, innovative science and push beyond traditional thinking in order to create meaningful treatments for patients with cancer. We are dedicated to transforming science into value for patients, and this sense of obligation informs everything we do. Anchored by our Antibody Drug Conjugate (ADC) and Acute Myeloid Leukemia (AML) franchises, our cancer pipeline includes more than 20 small molecules, monoclonal antibodies and ADCs stemming from our powerful research engines: our two laboratories for biologic/immuno-oncology and small molecules in Japan, and Plexxikon Inc., our small molecule structure-guided R&D center in Berkeley, CA. Compounds in development include: quizartinib, an oral FLT3 inhibitor, for FLT3-ITD+ AML; DS-8201, a HER2-targeting ADC, for HER2-expressing breast or gastric cancer or other HER2-expressing solid tumors; pexidartinib, an oral CSF-1R inhibitor, for tenosynovial giant cell tumor (TGCT), which is also being explored in a range of solid tumors in combination with the anti-PD1 immunotherapy, pembrolizumab; and tivantinib, an oral MET inhibitor, for second-line treatment of patients with MET-overexpressing hepatocellular carcinoma in partnership with ArQule, Inc. About Daiichi Sankyo Daiichi Sankyo Group is dedicated to the creation and supply of innovative pharmaceutical products to address diversified, unmet medical needs of patients in both mature and emerging markets. With over 100 years of scientific expertise and a presence in more than 20 countries, Daiichi Sankyo and its 16,000 employees around the world draw upon a rich legacy of innovation and a robust pipeline of promising new medicines to help people. In addition to a strong portfolio of medicines for hypertension and thrombotic disorders, under the Group's 2025 Vision to become a "Global Pharma Innovator with a Competitive Advantage in Oncology," Daiichi Sankyo research and development is primarily focused on bringing forth novel therapies in oncology, including immuno-oncology, with additional focus on new horizon areas, such as pain management, neurodegenerative diseases, heart and kidney diseases, and other rare diseases. For more information, please visit: www.daiichisankyo.com. Daiichi Sankyo, Inc., headquartered in Parsippany, New Jersey, is a member of the Daiichi Sankyo Group. For more information on Daiichi Sankyo, Inc., please visit: www.dsi.com. This press release contains forward-looking statements regarding the Company's clinical trials with tivantinib (ARQ 197). These statements are based on the Company's current beliefs and expectations, and are subject to risks and uncertainties that could cause actual results to differ materially. Positive information about pre-clinical and early stage clinical trial results does not ensure that later stage or larger scale clinical trials will be successful. For example, tivantinib may not demonstrate promising therapeutic effect or appropriate safety profiles in current or later stage or larger scale clinical trials as a result of known or as yet unanticipated side effects. The results achieved in later stage trials may not be sufficient to meet applicable regulatory standards or to justify further development. Problems or delays may arise prior to the initiation of planned clinical trials, during clinical trials or in the course of developing, testing or manufacturing that could lead the Company or its partners and collaborators to fail to initiate or to discontinue development. Even if later stage clinical trials are successful, unexpected concerns may arise from subsequent analysis of data or from additional data. Obstacles may arise or issues may be identified in connection with review of clinical data with regulatory authorities. Regulatory authorities may disagree with the Company's view of the data or require additional data or information or additional studies. In addition, the planned timing of initiation and completion of clinical trials for tivantinib is subject to the ability of the Company as well as Daiichi Sankyo, Inc., our development partner for tivantinib, and Kyowa Hakko Kirin, a licensee of tivantinib, to enroll patients, enter into agreements with clinical trial sites and investigators, and overcome technical hurdles and other issues related to the conduct of the trials for which each of them is responsible. There is a risk that these issues may not be successfully resolved. In addition, we and our partners are utilizing companion diagnostic tests to identify MET-overexpressing patients in the METIV-HCC, JET-HCC and other trials. We may encounter difficulties in developing and obtaining approval for companion diagnostics, including issues relating to selectivity/specificity, analytical validation, reproducibility, or clinical validation. Any delay or failure by our collaborators or us to develop or obtain regulatory approval of the companion diagnostics could delay or prevent approval of our product candidates. Drug development involves a high degree of risk. Only a small number of research and development programs result in the commercialization of a product. Positive pre-clinical data may not be supported in later stages of development. Furthermore, ArQule may not have the financial or human resources to successfully pursue drug discovery in the future. Moreover, with respect to partnered programs, even if certain compounds show initial promise, Daiichi Sankyo or Kyowa Hakko Kirin may decide not to license or continue to develop them, as the case may be. In addition, Daiichi Sankyo and Kyowa Hakko Kirin have certain rights to unilaterally terminate their agreements with ArQule. If either company were to do so, the Company might not be able to complete development and commercialization of the applicable licensed products on its own. For more detailed information on the risks and uncertainties associated with the Company's drug development and other activities, see the Company's periodic reports filed with the Securities and Exchange Commission. The Company does not undertake any obligation to publicly update any forward-looking statements.


News Article | December 15, 2015
Site: www.biosciencetechnology.com

A team of researchers at UC San Francisco has devised a new approach for early stage drug discovery that uses techniques from the world of computer vision in combination with a powerful new tool: a lineage of genetically modified cancer cells in a dish that change their appearance when treated with drugs targeting common disease pathways. The researchers hope to employ these cells -- in combination with algorithms akin to those used by face-recognition software -- to quickly organize the hundreds of thousands of molecules in institutional compound libraries according to their biological function, greatly facilitating the search for new and better drugs for cancer and other diseases. For this reason, they dubbed the cells ORACLs (Optimized Reporter cell lines for Annotating Compound Libraries). A paper describing the new approach was published online Dec. 14 in the journal Nature Biotechnology. Co-senior authors Steven Altschuler, Ph.D., and Lani Wu, Ph.D., are professors of pharmaceutical chemistry in the UCSF School of Pharmacy and conducted the initial research in this area while at the University of Texas Southwestern Medical Center. The pair envision making drug discovery more like the field of genomics, where researchers have spent decades annotating large genomic databases as more and more functions are discovered for individual genes and regulatory elements. As a result of this community effort, researchers can now rapidly identify particular genes of interest for more targeted investigations. Using ORACLs to functionally annotate existing compound libraries could have similar benefits, enabling researchers to quickly identify drugs that are clinically relevant for specific disease pathways, and significantly streamlining the way scientists use the shared drug discovery facilities, or cores, which are common in academic research. “Drug screening cores are very busy – they run continuously,” Wu said. “The problem is that screening results generally cannot be reused. When you have a new biological target you want to hit with a drug, you have to go and screen the whole compound library again.” Researchers commonly screen for effective drugs using “reporter cells,” which are carefully designed to change their behavior or appearance in response to a successful treatment. But the creation of an ORACL – a single reporter cell line capable of distinguishing between multiple common drug classes – required a different approach. “We didn't know how to design such a versatile reporter cell based on our existing biological knowledge,” said Chien-Hsiang “Charles” Hsu, a graduate student in the Altschuler and Wu lab and co-lead author on the new study. “So we thought, why not just randomly tag proteins and screen the cells with drugs to see which line could produce the response that fit our goal?" The researchers created an assortment of 93 cell lines, based on an existing line of lung cancer cells, by tagging randomly selected genes with a fluorescent marker. They treated cells from each of these lineages with a panel of 30 compounds belonging to six commonly used classes of cancer drugs, then analyzed images of the cells with custom algorithms. As the team had hoped, different drugs caused cells to change their shape and pattern of fluorescence in distinct ways that enabled the software to deduce which type of drug had been applied. As anticipated, some cell lines were more informative than others: To the researchers’ delight, one cell line proved capable of distinguishing between the six drug classes with 94 percent accuracy. Some of the more subtle aspects of the cells’ characteristic responses to different drug types required the computer algorithms to detect, but others were obvious under the microscope: DNA damaging drugs caused cells to swell, while HDAC inhibitors produced a spiky appearance. MTOR inhibitors produced dim fluorescence throughout the cell, while treatment with hsp90 inhibitors resulted in fluorescent speckles within the cytoplasm. The researchers had found their ORACL. To test their ORACL cells, the team screened through more than 10,000 small molecules of unknown function from several institutional compound libraries. Their analysis identified 106 molecules whose effects on the cells matched those of the “training” drugs that had been used to initially select the ORACL, and further experiments confirmed that at least 90 of these matches affected the same biological pathway as the training drug. To the researchers’ surprise, ORACL cells also produced characteristic responses to a slew of additional compounds, some of which turned out to belong to drug classes not included in the original training set, including ER and Aurora Kinase inhibitors, glucocorticoid steroids, ATPase inhibitors, and dihihydrofolate reductase inhibitors, as well as a dozen other groups of molecules whose shared biological function is still not known. “What’s amazing,” said Altschuler, “is that we were able to do one screen, one time, and fish out molecules that were in many different diverse classes at once.” The researchers are currently scaling up their screens to enable rapid annotation of compound libraries containing hundreds of thousands molecules. They hope that ORACLs will be adopted to identify new compounds that hit biological pathways where current drugs have too many side effects or miss certain patient populations. They could find compounds that affect pathways affected by few known drugs or even drugs whose effects seem far from any known biological pathway. “These are really early steps in drug discovery,” Altschuler said. “We hope finding more high quality compounds will make all the later steps more efficient. Currently the process takes billions of dollars over many, many years. Wouldn't it be nice to make that easier?” Major funding for the research was provided by the National Institutes of Health.


News Article | October 8, 2016
Site: www.techtimes.com

Strokes, injuries and even neurological diseases such as Alzheimer's, Parkinson's or Huntington's seem to follow the same chain of events, in spite of their distinct triggers. New research has found the end of the chain, responsible for the fatal step through carving up a cell's DNA. The conclusions of the research create new possibilities in the development of drugs aimed at treating and even preventing the process. The study, published on Oct. 7 in Science, documents the experiments conducted inside lab-grown cells, signed by Ted Dawson, director of the Institute for Cell Engineering at the Johns Hopkins University School of Medicine, and Valina Dawson, professor of neurology. Cell death occurs because of the toxic and/or stressful insults, and a scientific way to stop its activity would represent a huge step in the fight against neurodegenerative disorders, as well as any other form of cell injury. The shared mechanisms of these diseases share a programmed brain cell death the team named parthanatos, which is an enzyme involved in almost all forms of cellular injuries. Other scientific studies showed that a protein named mitochondrial apoptosis-inducing factor (AIF) sometimes leaves its normal place in the cellular mitochondria moving toward the nucleus, triggering the carving up of the genome housed in the nucleus. The process is responsible for cell death. However, AIF cannot cut DNA alone. The endonuclease G promotes DNA degradation cooperating with AIF but doesn't play an important role in the chromatinolysis that is PARP-dependend, therefore not being responsible for cell death. But Yingfei Wang, assistant professor at the University of Texas Southwestern Medical Center, tested which proteins interact most powerfully with the AIF by screening other human proteins. Using custom molecules known as small interfering RNAs, she tried to stop the formation of proteins in human cells grown in her laboratory to test the hypothesis of preventing cell death. Of the total number of proteins, the macrophage migration inhibitory factor (MIF) was the one found responsible for the interaction. Although MIF's capacity to affect DNA was only clearly linked to stroke. Disabling the MIF gene in mice reduced the number of strokes. The paper also mentioned that some chemical compounds blocking the action of MIF in the cells grown in the lab protected them from parthanatos. According to the scientists, they also plan on testing them on animals to maximize the efficiency. "We're interested in finding out whether MIF is also involved in Parkinson's, Alzheimer's and other neurodegenerative diseases," explained Dawson. Blocking the interaction of the two or the entire nuclease activity of MIF would open a major line of therapeutic opportunities for patients of various diseases. © 2017 Tech Times, All rights reserved. Do not reproduce without permission.


News Article | October 7, 2016
Site: www.chromatographytechniques.com

Despite their different triggers, the same molecular chain of events appears to be responsible for brain cell death from strokes, injuries and even such neurodegenerative diseases as Alzheimer's. Now, researchers at Johns Hopkins say they have pinpointed the protein at the end of that chain of events, one that delivers the fatal strike by carving up a cell's DNA. The find, they say, potentially opens up a new avenue for the development of drugs to prevent, stop or weaken the process. A report on the research appears in the Oct. 7 issue of the journal Science. The new experiments, conducted in laboratory-grown cells, build on earlier work by research partners Ted Dawson, M.D., Ph.D., now director of the Institute for Cell Engineering at the Johns Hopkins University School of Medicine, and Valina Dawson, Ph.D., professor of neurology. Their research groups found that despite their very different causes and symptoms, injury, stroke, Alzheimer's disease, Parkinson's disease and the rare, fatal genetic disorder Huntington's disease have a shared mechanism of a distinct form of "programmed" brain cell death they named parthanatos after the personification of death in Greek mythology and PARP, an enzyme involved in the process. "I can't overemphasize what an important form of cell death it is; it plays a role in almost all forms of cellular injury," Dawson says. His and Valina Dawson's research groups have spent years delineating each of the links in the parthanatos chain of events and the roles of the proteins involved. The current study, they say, has completed the chain. From previous studies, the researchers knew that when a protein called mitochondrial apoptosis-inducing factor, or AIF, leaves its usual place in the energy-producing mitochondria of the cell and moves to the nucleus, it sparks the carving up of the genome housed in the nucleus and leads to cell death. But AIF itself, they say, can't cut DNA. So then-postdoctoral fellow Yingfei Wang, Ph.D., now an assistant professor at the University of Texas Southwestern Medical Center, used a protein chip to screen thousands of human proteins to find those that interacted most strongly with AIF. Working with the 160 candidates she uncovered, she then used custom molecules called small interfering RNAs to stop each of those proteins' manufacture, one by one, in lab-grown human cells to see if doing so would prevent cell death. One of the 160 proteins, known as macrophage migration inhibitory factor (MIF), was a winner. "We found that AIF binds to MIF and carries it into the nucleus, where MIF chops up DNA," Dawson says. "We think that's the final execution step in parthanatos." The group reports that in work to be published, it also identified a few chemical compounds that block MIF's action in the lab-grown cells, protecting them from parthanatos. Dawson says they plan to test these in animals, and modify them to maximize their safety and effectiveness. He cautions that while parthanatos is known to cause cell death in many brain conditions, MIF's ability to chop up DNA has so far only been definitively linked with stroke -- when the MIF gene was disabled in mice, the damage caused by a stroke was dramatically reduced. "We're interested in finding out whether MIF is also involved in Parkinson's, Alzheimer's and other neurodegenerative diseases," he says. If so, and if an inhibitor of MIF proves successful in testing, it could have implications for treating many conditions, he says.


BURLINGTON, Mass., TOKYO, MUNICH, & PARSIPPANY, N.J.--(BUSINESS WIRE)--ArQule, Inc. (Nasdaq: ARQL) and Daiichi Sankyo today announced that the METIV-HCC phase 3 study of tivantinib in hepatocellular carcinoma (HCC) did not meet its primary endpoint of improving overall survival. METIV-HCC is a biomarker-selected, double-blind, placebo-controlled, randomized phase 3 study evaluating tivantinib (2:1) versus best supportive care in patients with MET-overexpressing, inoperable HCC intolerant to or previously-treated with systemic therapy. A total of 340 patients with MET-overexpressing HCC analyzed by a validated immunohistochemical assay were randomized in the intent-to-treat population for efficacy analysis. The primary endpoint of the study is overall survival. Secondary endpoints include progression-free survival and safety. Full results from the trial will be presented at an upcoming scientific forum. “HCC is a disease with high unmet need, especially in the second-line setting, so these results are disappointing for the patients as well as the investigators and the companies,” said Paolo Pucci, Chief Executive Officer of ArQule. “Despite the negative outcome of this study, we remain committed to applying rigorous science to unmet needs for patients with cancer,” said Antoine Yver, MD, MSc, Executive Vice President and Global Head, Oncology Research and Development, Daiichi Sankyo. “We would like to take this opportunity to thank all of the investigators, and especially the patients, for their participation in this study.” The ArQule investor conference call can be accessed in the “Investors and Media” section of ArQule’s website, www.arqule.com, under “Events and Presentations.” You may also listen to the call by dialing (877) 868-1831 within the U.S. or (914) 495-8595 outside the U.S. and using the passcode 74015633. A replay will be available two hours after the completion of the call and can be accessed in the “Investors and Media” section of our website, www.arqule.com, under “Events and Presentations.” About Hepatocellular Carcinoma (HCC) Liver cancer is the sixth most common cancer globally with 782,000 new cases in 2012 and is the second most common cause of cancer-related death with 745,000 deaths in 2012.1 HCC accounts for about 90 percent of primary liver cancers.2 Cirrhosis, chronic hepatitis B and C and smoking are recognized worldwide as factors increasing the risk of HCC.2 About Tivantinib (ARQ 197) ArQule and Daiichi Sankyo have a licensing, co-development and co-commercialization agreement for tivantinib in the U.S., Europe, South America and the rest of the world, excluding Japan, China (including Hong Kong), South Korea and Taiwan. About ArQule ArQule is a biopharmaceutical company engaged in the research and development of targeted therapeutics to treat cancers and rare diseases. Our mission is to discover, develop and commercialize novel small molecule drugs in areas of high unmet need that will dramatically extend and improve the lives of our patients. Our clinical-stage pipeline consists of five drug candidates, all of which are in targeted, biomarker-defined patient populations, making ArQule a leader among companies our size in precision medicine. ArQule’s lead product, in phase 3 clinical development, is tivantinib (ARQ 197), an oral, selective inhibitor of the c-MET receptor tyrosine kinase, for second-line treatment of patients with MET-overexpressing hepatocellular carcinoma in partnership with Daiichi Sankyo in the West and Kyowa Hakko Kirin in Asia. ArQule’s proprietary pipeline includes: ARQ 087, a multi-kinase inhibitor designed to preferentially inhibit the fibroblast growth factor receptor (FGFR) family, in phase 2 for iCCA and in phase 1b for multiple oncology indications; ARQ 092, a selective inhibitor of the AKT serine/threonine kinase, in phase 1 for multiple oncology indications as well as ultra-rare Proteus syndrome, in partnership with the National Institutes of Health (NIH); ARQ 751, a next generation AKT inhibitor, in phase 1 for patients with AKT1 and PI3K mutations; and ARQ 761, a β-lapachone analog being evaluated as a promoter of NQO1-mediated programmed cancer cell necrosis, in phase 1/2 in multiple oncology indications in partnership with the University of Texas Southwestern Medical Center. In addition, we have advanced ARQ 531, an investigational, orally bioavailable, potent and reversible inhibitor of both wild type and C481S-mutant BTK, into toxicology testing and plan to file an Investigational New Drug Application in early 2017. ArQule’s current discovery efforts are focused on the identification and development of novel kinase inhibitors, leveraging the Company’s proprietary library of compounds. You can follow us on Twitter and LinkedIn. About Daiichi Sankyo Cancer Enterprise The vision of Daiichi Sankyo Cancer Enterprise is to leverage our world-class, innovative science and push beyond traditional thinking in order to create meaningful treatments for patients with cancer. We are dedicated to transforming science into value for patients, and this sense of obligation informs everything we do. Anchored by our Antibody Drug Conjugate (ADC) and Acute Myeloid Leukemia (AML) franchises, our cancer pipeline includes more than 20 small molecules, monoclonal antibodies and ADCs stemming from our powerful research engines: our two laboratories for biologic/immuno-oncology and small molecules in Japan, and Plexxikon Inc., our small molecule structure-guided R&D center in Berkeley, CA. Compounds in development include: quizartinib, an oral FLT3 inhibitor, for FLT3-ITD+ AML; DS-8201, a HER2-targeting ADC, for HER2-expressing breast or gastric cancer or other HER2-expressing solid tumors; pexidartinib, an oral CSF-1R inhibitor, for tenosynovial giant cell tumor (TGCT), which is also being explored in a range of solid tumors in combination with the anti-PD1 immunotherapy, pembrolizumab; and tivantinib, an oral MET inhibitor, for second-line treatment of patients with MET-overexpressing hepatocellular carcinoma in partnership with ArQule, Inc. About Daiichi Sankyo Daiichi Sankyo Group is dedicated to the creation and supply of innovative pharmaceutical products to address diversified, unmet medical needs of patients in both mature and emerging markets. With over 100 years of scientific expertise and a presence in more than 20 countries, Daiichi Sankyo and its 16,000 employees around the world draw upon a rich legacy of innovation and a robust pipeline of promising new medicines to help people. In addition to a strong portfolio of medicines for hypertension and thrombotic disorders, under the Group’s 2025 Vision to become a “Global Pharma Innovator with a Competitive Advantage in Oncology,” Daiichi Sankyo research and development is primarily focused on bringing forth novel therapies in oncology, including immuno-oncology, with additional focus on new horizon areas, such as pain management, neurodegenerative diseases, heart and kidney diseases, and other rare diseases. For more information, please visit: www.daiichisankyo.com. Daiichi Sankyo, Inc., headquartered in Parsippany, New Jersey, is a member of the Daiichi Sankyo Group. For more information on Daiichi Sankyo, Inc., please visit: www.dsi.com. This press release contains forward-looking statements regarding the Company's clinical trials with tivantinib (ARQ 197). These statements are based on the Company's current beliefs and expectations, and are subject to risks and uncertainties that could cause actual results to differ materially. Positive information about pre-clinical and early stage clinical trial results does not ensure that later stage or larger scale clinical trials will be successful. For example, tivantinib may not demonstrate promising therapeutic effect or appropriate safety profiles in current or later stage or larger scale clinical trials as a result of known or as yet unanticipated side effects. The results achieved in later stage trials may not be sufficient to meet applicable regulatory standards or to justify further development. Problems or delays may arise prior to the initiation of planned clinical trials, during clinical trials or in the course of developing, testing or manufacturing that could lead the Company or its partners and collaborators to fail to initiate or to discontinue development. Even if later stage clinical trials are successful, unexpected concerns may arise from subsequent analysis of data or from additional data. Obstacles may arise or issues may be identified in connection with review of clinical data with regulatory authorities. Regulatory authorities may disagree with the Company's view of the data or require additional data or information or additional studies. In addition, the planned timing of initiation and completion of clinical trials for tivantinib is subject to the ability of the Company as well as Daiichi Sankyo, Inc., our development partner for tivantinib, and Kyowa Hakko Kirin, a licensee of tivantinib, to enroll patients, enter into agreements with clinical trial sites and investigators, and overcome technical hurdles and other issues related to the conduct of the trials for which each of them is responsible. There is a risk that these issues may not be successfully resolved. In addition, we and our partners are utilizing companion diagnostic tests to identify MET-overexpressing patients in the METIV-HCC, JET-HCC and other trials. We may encounter difficulties in developing and obtaining approval for companion diagnostics, including issues relating to selectivity/specificity, analytical validation, reproducibility, or clinical validation. Any delay or failure by our collaborators or us to develop or obtain regulatory approval of the companion diagnostics could delay or prevent approval of our product candidates. Drug development involves a high degree of risk. Only a small number of research and development programs result in the commercialization of a product. Positive pre-clinical data may not be supported in later stages of development. Furthermore, ArQule may not have the financial or human resources to successfully pursue drug discovery in the future. Moreover, with respect to partnered programs, even if certain compounds show initial promise, Daiichi Sankyo or Kyowa Hakko Kirin may decide not to license or continue to develop them, as the case may be. In addition, Daiichi Sankyo and Kyowa Hakko Kirin have certain rights to unilaterally terminate their agreements with ArQule. If either company were to do so, the Company might not be able to complete development and commercialization of the applicable licensed products on its own. For more detailed information on the risks and uncertainties associated with the Company's drug development and other activities, see the Company's periodic reports filed with the Securities and Exchange Commission. The Company does not undertake any obligation to publicly update any forward-looking statements.


News Article | November 4, 2015
Site: www.nature.com

What are some of the biggest obstacles in bringing big data into drug discovery? Certainly we'll benefit from integrating large data sets, but it is imperative that this is not uncoupled from biological investigation. One of the challenges in pharma, at a time of increased externalization and partnering for research, is how to retain deep biological insight and connect that to the interrogation of these large data sets. One without the other is misguided. As we move from the lab into the clinic, it is useful to study large, longitudinal clinical data sets. But again, interrogating those data without clinical insight is not very meaningful. The big prizes will go to those who connect the large clinical data sets with an abundance of preclinical data, and bring all that together. I don't think anybody has got that right yet. In the academic world, the two new drugs to treat high cholesterol that target the protein PCSK9 are a great example of making the connection. Helen Hobbs of the University of Texas Southwestern Medical Center and her partners connected formidable genetics, biological understanding and chemical insight to lead to the important new medicines. In the pharma world, Genentech seems to have made a long-term investment in biology and linked that to clinical data to treat the right person with the right drug — witness Herceptin for patients with breast cancer. How are drug companies figuring out where to place their bets? Given the attention deficit disorder and externalization of research in pharma, and the ever-increasing demands of venture capital and other financial markets in drug discovery, sometimes we are seeing elements of risk aversion and a herd effect. There must be 20 companies looking for the next PD-1 [a cancer immunotherapy target] right now. But we are also seeing examples of pharma companies making very big, bold decisions — going after certain bespoke immune therapies, for example, without any hugely compelling evidence that this approach will work. I think there is an argument for companies externalizing and partnering on research, so that they are not missing something at the bleeding edge of discovery. At the same time, those decisions of when and where you jump in, with enough confidence, are truly challenging. What is a good example of a tough decision? Look at microbiome research. Everyone is really excited about the microbiome. It is tantalizing that all these billions of bacteria in our gut, skin and everywhere else influence disease and how we respond to drugs. We have seen associations between particular bacterial flora and disease states, but when will we be comfortable enough to invest substantially in making medicines to alter someone's microbiome? I don't know the answer to that. To date, most of the data set out to define the diverse repertoire of microbes are from small numbers of individuals. Conducting evaluations from larger cohorts of subjects is daunting, and long-term clinical data on these cohorts have been limited. Finally, we lack the robust tests that would let us modify the microbiome in a durable way and have a meaningful impact on the disease process. Why does Sanford Burnham Prebys combine basic research with drug discovery? Pharma often has a disconnect between the applied research in making medicines and the deep biological insight and ongoing experimentation that informs it. We have 80 people here from pharma who have made making medicines their whole career. Our principal investigators can work hand-in-hand with drug discoverers, and that enables us to pursue research and go after targets that nobody else will work on because they are too uncertain. Building up the big-data element creates a unique situation in this work. For example, when we start thinking about autoimmunity, about what happens when, with which T cells and B cells, how do we start disentangling those complexities? This is where big-data bioinformatics can be hugely useful. That to me underscores more than ever the need both to analyse large data sets and to stay connected to researchers who understand all the moving pieces with a view from a little higher up. Is it difficult to find research staff with skills in gathering and analysing big data? Yes, we struggle with this. Part of the challenge is training and funding individuals who are sophisticated enough to pursue that. They also tend to be gobbled up by potentially more lucrative fields outside the life sciences. We have been searching hard to attract and recruit the next wave of systems biologists and other people who can analyse large amounts of data. You want to have a critical mass of people doing that together, and it's really tough. There are people who generate data and people who analyse data, but there are few who do both really well. I believe the winners are the ones who can pull it all together.


The International Association of HealthCare Professionals is pleased to welcome Dr. Michael Sutker, MD to their prestigious organization with his upcoming publication in The Leading Physicians of the World. Dr. Sutker is a General and Bariatric Surgeon holding over two years of experience and an extensive expertise in all facets of his work. Dr. Michael Sutker is currently serving patients his private practice, Michael Sutker, M.D., P.A in Dallas, Texas, and is also affiliated with Medical City Dallas Hospital, where he is the Medical Director of Robotic Surgery and the Chair of the Advanced Clinical Advisory Board. Dr. Sutker’s career in medicine began in 2008, when he graduated with his Medical Degree from the University of Texas Southwestern Medical Center. He completed his residency in General Surgery at the same institution. He then went on to undertake his fellowship training in Minimally Invasive and Bariatric Surgery at the University of California-San Francisco. To keep up to date with the latest advances and developments in his field, Dr. Sutker maintains professional memberships with the Society of American Gastrointestinal and Endoscopic Surgeons, the American College of Surgeons, the American Medical Association, the Texas Medical Association, and the Dallas County Medical Society. He attributes his great success to his training, his availability, and his hard work. When Dr. Sutker is not assisting patients, he enjoys spending time with his family. Learn more about Dr. Sutker by reading his upcoming publication in The Leading Physicians of the World. FindaTopDoc.com is a hub for all things medicine, featuring detailed descriptions of medical professionals across all areas of expertise, and information on thousands of healthcare topics.  Each month, millions of patients use FindaTopDoc to find a doctor nearby and instantly book an appointment online or create a review.  FindaTopDoc.com features each doctor’s full professional biography highlighting their achievements, experience, patient reviews and areas of expertise.  A leading provider of valuable health information that helps empower patient and doctor alike, FindaTopDoc enables readers to live a happier and healthier life.  For more information about FindaTopDoc, visit http://www.findatopdoc.com


News Article | November 2, 2016
Site: www.sciencedaily.com

The first unbiased genetic screen for sleep defects in mice has yielded two interesting mutants, Sleepy, which sleeps excessively, and Dreamless, which lacks rapid eye movement (REM) sleep. The findings are the first step towards discovering the biochemistry that controls the switch from wakefulness to sleep, the researchers say. Although sleep is essential for life and we spend one third of our lives doing it, the function of sleep, and the physiology that regulates it, remain longstanding mysteries in biology. Researchers have hypothesized that there is a substance that builds up when we are awake, and then has to be discharged or recovered while we are sleeping. "But we still don't understand those processes," said HHMI Investigator Joseph Takahashi at the University of Texas Southwestern Medical Center in Dallas. To address these issues, Takahashi and former HHMI Investigator Masashi Yanagisawa, now Professor in the International Institute for Integrative Sleep Medicine at the University of Tsukuba in Japan, decided to take an unbiased exploratory approach. Instead of beginning with a hypothesis about specific genes that might be involved, the researchers introduced random genetic mutations in more than 8,000 mice and screened them using electroencephalography (EEG) to determine which ones had abnormal sleep as a result of the genetic perturbations. "The barrier in the past has been that it's a very laborious process. To do a genetic screen, you should be prepared to screen thousands of animals before you find something interesting, and most people are just not willing to measure EEGs in thousands of mice," explained Takahashi. But by optimizing the surgical methods, electrodes, and the software to analyze the EEGs in automated fashion, the researchers were able to conduct the first unbiased genetic screen of this magnitude for sleep defects in mice, which they report in this week's issue of Nature. The researchers identified two mutations, which they called Sleepy and Dreamless, and subsequently mapped them to locations in the mouse genome. Sleepy mice, which need approximately one third more sleep than normal mice, carry a mutation in the Sik3 kinase gene. Because the Sik3 kinase can phosphorylate many proteins, it is likely to be involved in many signaling pathways, which makes it trickier to characterize. To investigate why Sleepy mice need more sleep, the researchers examined the circadian clock in Sleepy mice, but they did not find a circadian rhythm disturbance. They tested whether the mice had defects in their arousal system by stimulating them with environmental (e.g., a new cage) and pharmacological (e.g, caffeine and modafinil) stimuli, but found that the mice had normal arousal responses. They concluded that the Sleepy mice had an increased sleep need, but the physiological reasons for that remained unclear. "Sleep need still remains a mystery, but what we hope is that this kinase is maybe the key, the initial key to this big door," said Yanagisawa. Dreamless mice, which have reduced rapid eye movement (REM) sleep, carry a mutation in a sodium channel. Understanding the effects of the dreamless mutation was more straightforward. The mutation increases the conductivity of a leaky sodium channel that was previously known to regulate neuronal excitability. The neuronal populations that terminate REM sleep have too much excitability, said Yanagisawa, which is why the mice have reduced REM sleep. The researchers are optimistic that the screen will yield more mutants with sleep defects to investigate. "We really hope that this is opening up some mysteries ... this is just the beginning," said Yanagisawa.


News Article | January 20, 2016
Site: www.nature.com

Alfred Goodman Gilman discovered heterotrimeric G proteins, which help to usher chemical signals into cells. For this work, which reshaped our understanding of hormone and drug action, he shared the 1994 Nobel Prize in Medicine or Physiology with Martin Rodbell. Gilman's impact on biomedical research and education extended much further. He edited several editions of the definitive textbook The Pharmacological Basis of Therapeutics (or 'Goodman and Gilman'), which has served generations of medical and graduate students. At the University of Texas Southwestern Medical Center in Dallas, he chaired the pharmacology department from 1981 to 2004, became dean in 2004 and provost in 2006. And in 2009, he was appointed first chief scientific officer of the Cancer Prevention and Research Institute of Texas (CPRIT), established to disburse US$3 billion in state funding. He died on 23 December 2015. Gilman was born in July 1941 in New Haven, Connecticut, with, in his words, a “scientific silver spoon” in his mouth. His father was the eminent pharmacologist Alfred Gilman Sr, who wrote the aforementioned textbook with his close colleague Louis Goodman. Gilman Sr, in tribute to his friend, gave his son the middle name Goodman. As a child, young Gilman enjoyed trips to his father's labs at Columbia University and the Albert Einstein School of Medicine in New York. After completing a degree in biochemistry at Yale University in New Haven in 1962, he enrolled in one of the first MD–PhD programmes in the United States, at Case Western Reserve University in Cleveland, Ohio. The programme was run by Nobel laureate Earl Sutherland, the discoverer of cyclic AMP, a key intracellular messenger molecule. Here, Gilman solidified his interest in cell-signalling mechanisms. A postdoc at the US National Institute of Health in Bethesda, Maryland, with Marshall Nirenberg, another Nobel laureate, followed. In 1970, in Nirenberg's lab, Gilman independently developed a sensitive technique for detecting cAMP that was immediately widely adopted. But it was his discovery of a family of G proteins made up of three different subunits (known as heterotrimeric G proteins) as a junior faculty member at the University of Virginia in Charlottesville, that transformed the field of cell signalling. In the 1970s, evidence was mounting that the hormone receptors involved in cell signalling were independent entities in the plasma membrane. Gilman and his postdoc Elliott Ross were investigating this problem using membranes from a lymphoma cell line called cyc−. These cells seemed to lack the key enzyme adenylyl cyclase, which catalyses formation of cAMP. The cells retained the β-adrenergic receptor that binds a class of molecules called catecholamines, which includes the hormone adrenaline. This binding stimulates adenylyl cyclase, increasing the formation of cAMP. To these cyc− membranes, Gilman and Ross added an extract of another cell line, mouse L cells, which retained enzyme activity but lacked the hormone receptors. To their delight, this 'reconstitution' system worked and yielded hormone-sensitive adenylyl cyclase activity. However, control experiments indicated that extracts in which the cyclase activity had been inactivated, by heat for example, still led to a fully reconstituted hormone-sensitive enzyme. Through ingenious experiments they demonstrated that these extracts contained a heat-stable, hitherto unknown regulatory component required for activity of the adenylyl cyclase and that the cyc− cells actually did contain the catalytic unit of adenylyl cyclase (and hence had been misnamed). They called the new component G α — the α subunit of the stimulatory guanine nucleotide regulatory protein (G protein), because it bound guanosine triphosphate (GTP) and conferred both hormone and GTP sensitivity on the adenylyl cyclase. Using their reconstitution assay, they purified the protein and determined its DNA sequence. Their discovery, published in a series of papers in 1977–78 (E. M. Ross and A. G. Gilman J. Biol. Chem. 252, 6966–6969; 1977; and E. M. Ross et al. J. Biol. Chem. 253, 6401–6412; 1978) confirmed Rodbell's prediction of the existence of such an intermediate protein linking a cell's hormone receptors to adenylyl cyclase and intracellular signalling. Over the next 15 years the family of heterotrimeric G proteins grew to about 20, and their roles expanded to include almost all physiological processes in species from yeast to mammals. Gilman's subsequent work with Stephen Sprang focused on the detailed characterization by X-ray crystallography of several of the G proteins and the adenylyl cyclases that they regulate. Gilman's writing and speaking style were distinctive. He was critical not just of others' work but also of his own. His wit could border on the acerbic if he felt that the highest standards were not being met. He was beloved by his trainees, to whom he was fiercely loyal. They remember him as a hands-off mentor who was nonetheless approachable and inspirational, and who instilled in them a sense of rigour and integrity. Gilman displayed this personal integrity when, in 2012, he resigned from the CPRIT over concerns that political and commercial factors were exerting undue influence on its awarding of grants. Al's offbeat sense of humour occasionally made its way into his published work. For instance, he designated the irreversibly activated state of adenylyl cyclase induced by a certain guanine nucleotide as the 'P state' (for priapic) because “the enzyme's rate of catalysis was persistently elevated”. Reflecting on the role of serendipity in scientific discovery, Al once commented: “The trick is to recognize good luck when it happens, embrace it, and then commit whatever it takes to extract its full value.” He certainly extracted the full value of his lucky start as a scientific blueblood.


News Article | November 2, 2016
Site: www.eurekalert.org

The first unbiased genetic screen for sleep defects in mice has yielded two interesting mutants, Sleepy, which sleeps excessively, and Dreamless, which lacks rapid eye movement (REM) sleep. The findings are the first step towards discovering the biochemistry that controls the switch from wakefulness to sleep, the researchers say. Although sleep is essential for life and we spend one third of our lives doing it, the function of sleep, and the physiology that regulates it, remain longstanding mysteries in biology. Researchers have hypothesized that there is a substance that builds up when we are awake, and then has to be discharged or recovered while we are sleeping. "But we still don't understand those processes," said HHMI Investigator Joseph Takahashi at the University of Texas Southwestern Medical Center in Dallas. To address these issues, Takahashi and former HHMI Investigator Masashi Yanagisawa, now Professor in the International Institute for Integrative Sleep Medicine at the University of Tsukuba in Japan, decided to take an unbiased exploratory approach. Instead of beginning with a hypothesis about specific genes that might be involved, the researchers introduced random genetic mutations in more than 8,000 mice and screened them using electroencephalography (EEG) to determine which ones had abnormal sleep as a result of the genetic perturbations. "The barrier in the past has been that it's a very laborious process. To do a genetic screen, you should be prepared to screen thousands of animals before you find something interesting, and most people are just not willing to measure EEGs in thousands of mice," explained Takahashi. But by optimizing the surgical methods, electrodes, and the software to analyze the EEGs in automated fashion, the researchers were able to conduct the first unbiased genetic screen of this magnitude for sleep defects in mice, which they report in this week's issue of Nature. The researchers identified two mutations, which they called Sleepy and Dreamless, and subsequently mapped them to locations in the mouse genome. Sleepy mice, which need approximately one third more sleep than normal mice, carry a mutation in the Sik3 kinase gene. Because the Sik3 kinase can phosphorylate many proteins, it is likely to be involved in many signaling pathways, which makes it trickier to characterize. To investigate why Sleepy mice need more sleep, the researchers examined the circadian clock in Sleepy mice, but they did not find a circadian rhythm disturbance. They tested whether the mice had defects in their arousal system by stimulating them with environmental (e.g., a new cage) and pharmacological (e.g, caffeine and modafinil) stimuli, but found that the mice had normal arousal responses. They concluded that the Sleepy mice had an increased sleep need, but the physiological reasons for that remained unclear. "Sleep need still remains a mystery, but what we hope is that this kinase is maybe the key, the initial key to this big door," said Yanagisawa. Dreamless mice, which have reduced rapid eye movement (REM) sleep, carry a mutation in a sodium channel. Understanding the effects of the dreamless mutation was more straightforward. The mutation increases the conductivity of a leaky sodium channel that was previously known to regulate neuronal excitability. The neuronal populations that terminate REM sleep have too much excitability, said Yanagisawa, which is why the mice have reduced REM sleep. The researchers are optimistic that the screen will yield more mutants with sleep defects to investigate. "We really hope that this is opening up some mysteries ... this is just the beginning," said Yanagisawa.


News Article | November 16, 2016
Site: www.nature.com

Male C57BL/6J mice (CLEA Japan) were treated with ethylnitrosourea (85 mg kg−1, Sigma-Aldrich) by intraperitoneal injection twice at weekly intervals at the age of 8 weeks. At the age of 25–30 weeks, the sperm of the mice was used for in vitro fertilization with eggs of C57BL/6N mice to obtain F offspring. Mice were provided food and water ad libitum, and were maintained on a 12-h light:12-h dark cycle and housed under controlled temperature and humidity conditions. All procedures were approved by the Institutional Animal Care and Use Committee of the University of Tsukuba and the RIKEN BioResource Center, University of Texas Southwestern Medical Center at Dallas. EEG/EMG electrode implantation was performed as described previously35, with isoflurane (3% for induction, 1% for maintenance) used for anaesthesia. Seven days after surgery, the mice were tethered to a counterbalanced arm (Instech Laboratories) that allowed free movement and exerted minimal weight. At the age of 12 weeks, male mice were implanted with EEG/EMG electrodes and then screened for sleep/wakefulness behaviour. Examined parameters were total time spent in wake, NREMS and REMS states, episode duration of wake, NREMS and REMS states, appearance of muscle atonia during REMS, and rebound sleep after 4-h sleep deprivation by shaking the cages. For quantitative parameters, we selected mice whose phenotypes deviated from the average by at least 3 standard deviations. After confirming the reproducibility of the sleep phenotype, the mice were selected for offspring production by natural mating or IVF with wild-type females to examine the heritability of the sleep phenotypes. If at least 30% of the male littermates showed sleep phenotypes similar to their father, we considered the sleep abnormalities to be heritable. Single nucleotide polymorphisms of N mice were determined using a custom TaqMan Genotyping assay (Thermo Fisher). The custom probes were designed based on the polymorphism data between C57BL/6J and C57BL/6N (ref. 20). QTL analysis was performed using J/qtl software (Jackson Laboratory). Whole exomes were captured with SureSelectXT2 Mouse All Exon (Agilent) and processed to a paired end 2 × 100-bp run on the Illumina HiSeq2000 platform at the UTSW McDermott Center Next Generation Sequencing Core. Reads were mapped to the University of California Santa Cruz mm9 genome reference sequence for C57BL/6J using Burrows–Wheeler aligner and quality filtered using SAMtools. Cleaned BAM files were used to realign data and call variants using the Genome Analysis ToolKit to detect heterozygous mutations. The recording room was kept under 12-h light:12-h dark cycles and a constant temperature (24–25 °C). To examine sleep–wake behaviour under baseline conditions, EEG/EMG signals were recorded for two consecutive days from the onset of the light phase. EEG/EMG data were visualized and analysed using a MatLab (MathWorks)-based, custom semi-automated staging program followed by visual inspection. EEG signals were subjected to fast Fourier transform analysis from 1 to 30 Hz with 1-Hz bin using MatLab-based custom software. Epochs containing movement artefacts were included in the state totals but excluded from subsequent spectral analysis. Sleep/wakefulness was staged into wakefulness, NREMS and REMS. Wakefulness was scored based on the presence of low amplitude, fast EEG, and high amplitude, variable EMG. NREMS was characterized by high amplitude, delta (1–4 Hz) frequency EEG and low EMG tonus, whereas REMS was staged based on theta (6–9 Hz)-dominant EEG and EMG atonia. Hourly delta density during NREMS indicates hourly averages of delta density which is the ratio of delta power to total EEG power at each 20-s epoch. For the power spectrum of sleep/wakefulness, the EEG power of each frequency bins was expressed as a percentage of the total EEG power over all frequency bins (1–30 Hz) and sleep/wakefulness states35, 36. For sleep deprivation, mice were sleep deprived for 2, 4 and 6 h from the onset of the light phase by gently touching the cages when they started to recline and lower their heads. Food and water were available. To evaluate the effect of sleep deprivation, the NREMS delta power during the first hour after sleep deprivation was expressed relative to the same zeitgeber time (ZT) of the basal recording or relative to the mean of the basal recording. For caffeine and modafinil injection experiments, mice were fully acclimatized for intraperitoneal injection before sleep recording. After 24-h baseline recording, mice received caffeine (Sigma), modafinil (Sigma) or vehicle (0.5% methyl cellulose (Wako)) intraperitoneally at ZT0, followed by 12-h recording. Injections were delivered once per week, with each injection followed by a 6–8-day washout period, during which mice remained in the recording chamber. To examine the sleep/wakefulness behaviour under constant darkness, after 48-h recording under a 12-h light:12-h dark cycle, EEG/EMG recording continued in constant darkness for 3 days. Mice were housed individually in a cage (width 23 cm, length 33 cm, height 14 cm) containing a wireless running wheel (Med Associate ENV-044). Cages were placed in a light-tight chamber equipped with green LED light (100 lx at the bottom of the cage). The rotation numbers of wheels were obtained with 1-min bin using Wheel manager software (Med Associate). Mice were entrained to 12-h light:12-h dark cycle for 7 days, and then released into constant darkness for 3 weeks. The free running period was calculated with linear regression analysis of activity onset using MatLab-based custom software. Circadian activity amplitude was calculated by fast Fourier transform of activity data, which were processed with Bartlett window using MatLab-based custom software. Relative amplitude was normalized to the mean amplitude of the wild- type group. A rabbit polyclonal antibody against the C-terminal 171 amino acids of mouse SIK3 was generated using custom antibody production service (Pacific Immunology). Tissues were homogenized using a rotor-stator homogenizer (Polytron) in ice-cold lysis buffer (20 mM HEPES, pH 7.5, 100 mM NaCl, 10 mM Na P O , 1.5% Triton X-100,15 mM NaF, 1× PhosSTOP (Roche), 5 mM EDTA, 1× protease inhibitor (Roche)), and then centrifuged at 13,000g at 4 °C. The supernatants were separated by SDS–PAGE and transferred on PVDF membrane. Western blotting was performed according to standard protocols. In situ hybridization was performed as described previously37. In brief, a 0.7–0.8-kb fragment of Nalcn cDNA was inserted into pGEM-T easy (Promega) and used for DIG-labelled probe synthesis. Mice were deeply anaesthetized with sodium pentobarbital and perfused transcardially with PBS followed by 4% paraformaldehyde (PFA). Forty-micrometre-thick brain sections were treated with 0.3% Triton X-100, digested with 1 μg ml−1 proteinase K, treated with 0.75% glycine, and then treated with 0.25% acetic anhydride in 0.1 M triethanolamine. After overnight incubation with a digoxigenin (DIG)-labelled probe at 60 °C, the sections were washed and then incubated with alkaline phosphatase-conjugated anti-DIG Fab fragments (Roche, 11175041910). The reactions were visualized with a 5-bromo-4-chloro-3-indolyl-phosphate/4-nitroblue tetrazolium (BCIP/NBT) substrate solution (Roche). HEK293 cells (RCB1637) and HEK293T cells (RCB2202) were obtained from the RIKEN BRC Cell Bank. Cells were cultured in DMEM (Wako) supplemented with 10% FBS, 1% GlutaMAX (Thermo Fisher Scientific), and penicillin/streptomycin at 37 °C in a humidified atmosphere of 5% CO . Cell lines were regularly tested for mycoplasma contamination using MycoAlert (Lonza). Cell lines were regularly renewed by obtaining cell stocks from the Cell Bank for authentication. We used HEK293 and HEK293T cells because of their reliable growth, high efficiency in transfection and morphology suitable for electrophysiological experiments. For generating Sik3Slp knock-in mice, a genomic fragment containing exon 13 of the Sik3 gene was isolated from C57BL/6 mouse genomic BAC clone from a RP23 mouse genomic BAC library (Advanced GenoTEchs Co). A 1.7-kb fragment of FRT-PGK-gb2-neo-FRT-loxP cassette (Gene Bridges) flanked by two flippase recognition target (FRT) sites was inserted before exon 12. The targeting vector also contains a G-to-A substitution at the fifth nucleotide from the beginning of intron 13. The targeting vector was linearized and electroporated into the C57BL/6N ES cell line RENKA. Correctly targeted clones were injected into eight-cell stage ICR mouse embryos, which were cultured to produce blastocysts and then transferred to pseudopregnant ICR females. Resulting male chimaeric mice were crossed with female C57BL/6N mice to establish the Sik3Slp-neo/+ line. To remove the neomycin resistance gene with the FLP-FRT system, Sik3Slp-neo/+ mice were crossed with Actb-FLP knock-in mice. The custom-designed ZFN mRNAs targeting the exon 13–intron 13 boundary region of the Sik3 gene were obtained from Sigma-Aldrich’s Composers Custom ZFN service. Before the final assembly of the ZFN products, Sigma-Aldrich validated the designed ZFN binding sequences in silico using their bioinformatics tools and in vitro using Nero2A cell lines, ensuring high cutting efficiency and specificity using mismatch-specific endonuclease CelI according to the manufacturer’s instructions. The ZFN mRNAs were injected into single-cell stage C57BL/6J mouse zygotes at the University of Texas Southwestern Transgenic Core facility. The injected eggs were then transferred to pseudopregnant females to generate F founders. In total, 45 out of 96 F mice were found to be modified at the exon 13–intron 13 boundary region of the Sik3 gene. We crossed one F male mouse that had a 2-bp deletion from the last nucleotide of exon 13 with female C57BL/6N mice to obtain F mice of Sik3Slp/+ ZFN. The F mice were used to confirm the skipping of exon13 in Sik3 mRNA, which was purified from the brains and livers. The F male mice were used for sleep/wakefulness behaviour analysis. To produce a Cas9/single-guide RNA (sgRNA) expression vector, oligonucleotide DNAs (5′-CACCGCGAGCGGCCATCGACCCGC-3′ and 5′-AAACGCGGGTCGATGGCCGCTCGC-3′) were annealed and then inserted into pX330 vector (Addgene). The cleavage activity of the pX330-Sik3Ex1 vector was evaluated by the EGxxFP system38. Genomic DNA containing exon 1 of the Sik3 gene was amplified and inserted into pCAG-EGxxFP to produce pCAG-EGxxFP-Sik3Ex1. The pX330-Sik3Ex1 and pCAG-EGxxFP-Sik3Ex1 were transfected into HEK293 cells. As a donor oligonucleotide, a single-stranded 200-nucleotide DNA was synthesized (Integrated DNA Technologies), which contained a Flag-haemagglutinin-coding sequence in the centre and 70-nucleotide arms at the 5′ and 3′ ends. Female C57BL/6J mice or Sik3Slp knock-in mice were injected with pregnant mare serum gonadotropin and human chorionic gonadotropin at a 48-h interval, and mated with male C57BL/6J mice. The fertilized one-cell embryos were collected from the oviducts. Then, 5 ng μl−1 of pX330-Sik3Ex1 vector and 10 ng μl−1 of the donor oligonucleotide were injected into the pronuclei of these one-cell-stage embryos. The injected one-cell embryos were then transferred into pseudopregnant ICR mice. F mice were genotyped for the presence of Flag-coding sequence in exon1 of the Sik3 gene and for the presence of the Sik3Slp mutation. F mice containing Flag–SIK3 were further examined for the presence of the Cas9 transgene and off-target effects. Candidate off-target sites were identified based on a complete match of 16 bp at the 3′ end, including the PAM sequence. F mice were mated with C57BL/6N mice to obtain F offspring. NalcnDrl mice were produced as described above. To produce the sgRNA expression vector, pX330-NalcnEx9, oligonucleotide DNAs (5′-CACCAGCAATAAACACATTCTGAA-3′ and 5′-AAACTTCAGAATGTGTTTATTGCT-3′) were used. Genomic DNA containing exon 9 of the Nalcn gene was amplified and inserted into pCAG-EGxxFP to produce pCAG-EGxxFP-NalcnEx9. As a donor oligonucleotide, a single-stranded 199-nucleotide DNA containing a T-to-A substitution at the centre was synthesized (Integrated DNA Technologies). Nalcn mutant mice of N –N generation were used for sleep/wakefulness analysis. To evaluate Flag-tagged SIK3 protein in brains, we performed peptide mapping of the purified Flag–SIK3 protein. The brains of Flag-Sik3 knock-in mice and Flag-Sik3Slp knock-in mice were quickly dissected after cervical dislocation. Brains were homogenized in detergent-free buffer and then centrifuged (100,000g, 30 min, 4 °C). The supernatant was immunoprecipitated with anti-DDDDK antibody beads (MBL 3325). The eluate was run on a polyacrylamide gel and stained with SilverQuest Silver staining kit (Life technologies). Flag–SIK3 band (150 kDa) was dissected with a fresh blade. The proteins in the bands were reduced with 10 mM dithiothreitol and alkylated with 40 mM iodoacetamide. Each sample was digested with trypsin (4 μg ml−1; Trypsin Gold, Promega) at 37 °C overnight. The extracted peptides were then separated via nano flow LC (Advance LC, Michrom Bioresources) using a C18 column. The LC eluate was coupled to a nano-ionspray source attached to a Orbitrap Velos Pro mass spectrometer (Thermo Fisher Scientific). All MS/MS spectra were searched using Proteome Discoverer 1.3 software (Thermo Fisher Scientific). Peptides were mapped through mouse SIK3 (NP_081774) with 56% coverage. To examine the effect of sleep deprivation on the phosphorylation status of SIK3 protein, five Flag-Sik3 knock-in mice or five Flag-Sik3Slp knock-in mice were ad libitum slept (S) or sleep-deprived (SD) for 4 h by gentle handling immediately after light onset (ZT0–ZT4). Five wild-type mice were used as a negative control. At ZT4, mouse brains were quickly dissected after cervical dislocation, rinsed with cold PBS, and snap frozen in liquid nitrogen. Each half of the brains was lysed in 2 ml of ice-cold lysis buffer (20 mM HEPES, pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 2 mM MgCl , 15 mM NaF, 10 mM Na P O ) freshly supplemented with protease/phosphatase inhibitor cocktail tablets (Roche), and homogenized in a glass tissue homogenizer. After brain homogenate was incubated for 30 min and centrifuged at 13,000g for 20 min at 4 °C, the supernatant was pre-cleared by IgG and Protein G beads for 30 min before immunoprecipitation. Each pre-cleared lysate was added to 50 μl of anti-Flag antibody-conjugated Sepharose beads (Sigma, A2220) and rotated overnight at 4 °C. After washing the beads five times with cold wash buffer (20 mM HEPES, pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 2 mM MgCl , 15 mM NaF, 10 mM Na P O ), 50 μl of elution buffer (2% SDS, 60 mM Tris-HCl, pH 6.8, 50 mM DTT, 10% glycerol) was added and rotated for 10 min at 4 °C. Elution was repeated twice and combined into one eluate and analysed by western blotting. For each group of Flag knock-in mice, the five eluates of were mixed and equally split into two or three samples for mass spectrometric analysis. Thus, a total of six (Flag-Sik3) or nine (Flag-Sik3 and Flag-Sik3Slp) samples were reduced, alkylated, and trypsin digested overnight. After desalting, each sample was labelled with a different Tandem Mass Tag (TMT) reagent (Thermo Fisher Scientific), then all samples were combined into one mixture for HPLC fractionation using a C18 column. A total of 12 fractions were collected, and analysed separately on the Orbitrap-Fusion mass spectrometry platform (Thermo Fisher Scientific) using a reverse-phase liquid chromatography tandem mass spectrometry (LC–MS/MS) method. We performed data analysis to identify peptides and quantified reporter ion relative abundance using Proteome Discoverer 2.1 (Thermo Fisher Scientific). The relative abundance of quantified SIK3 phosphorylation sites was normalized with wild-type negative control and total SIK3 protein abundance. To express wild-type NALCN, we used pTracer-CMV2-ratNALCN-EF1α-EGFP (a gift from D. Ren)39. A single nucleotide substitution was induced to make pTracer-CMV2-ratNALCN(DRL)-EF1α-EGFP using a KOD plus Mutagenesis kit (Toyobo). HEK293T cells were grown to ~50% confluency in 12-well plates. Using Lipofectamine LTX (2 μl) and PLUS (1 μl) reagents (Thermo Fisher Scientific), the cells were cotransfected with 0.3 μg of each plasmid DNA encoding rat NALCN-EGFP (wild type or DRL), mouse UNC-80, and mouse SRC (Y529F) (constitutively active Src) in 12-well plates. UNC-80 and SRC kinase activate NALCN27, 28. In some experiments, the cells were incubated with 10 μM Gd3+ to inhibit NALCN. The cells were dissociated and plated on 18-mm coverslips coated with poly-l-lysine in fresh culture medium before patch-clamp recordings. All patch-clamp recordings from HEK293T cells were performed >72 h after transfection. Recording patch pipettes were pulled from glass capillaries (1B150F-4, World Precision Instruments) using a micropipette puller (P-97, Sutter Instrument) to give a resistance of ~9 MΩ. The series resistance of whole-cell recordings was ~40 MΩ, which was not compensated. Patch pipettes were filled with solution containing 150 mM CsOH, 120 mM methanesulfonic acid, 10 mM NaCl, 10 mM EGTA, 2 mM Mg ATP and 10 mM HEPES (pH 7.4 adjusted with methanesulfonic acid; osmolarity, 290−299 mOsm l−1 adjusted with CsCl). The cells on coverslips were transferred to a recording chamber under a fluorescence upright microscope (Axio Examiner D1, Zeiss) and continuously perfused with the bath solutions containing 150 mM NaCl, 3.5 mM KCl, 10 mM HEPES, 20 mM glucose, 5 mM NaOH, 2 mM MgCl and 1.2 mM CaCl (pH 7.4 adjusted; osmolarity, 300−310 mOsm l−1). The transfected cells were identified by enhance green fluorescent protein (eGFP) fluorescence. Patch-clamp recordings were performed at room temperature (24 °C) using a computer-controlled amplifier (MultiClamp 700B, Molecular Devices). The signals were digitized with A/D converter (Digidata 1440A, Molecular Devices), and acquired with Clampex (Molecular Devices) at a sampling rate of 50 kHz, and low-pass filtered at 5 kHz. At the end of recording, Gd3+ (10 μM) was used to confirm that the whole-cell currents were mediated through NALCN39. Data were analysed using Clampfit (Molecular Devices). The equilibrium potentials were calculated from I–V curves. Mean membrane conductance was estimated from the regression lines fitted to I–V curves from individual cells. Current, membrane conductance and charge transfer were normalized to membrane capacitance. Patch pipettes and recording system were the same as those used in recordings from HEK293 cells. Acute brain slices containing the DpMe were prepared from post-natal day 12–23 Nalcn+/+ or NalcnDrl/+ mice. After the induction of deep anaesthesia with isoflurane, mice were decapitated and the brains were rapidly removed into an ice-cold cutting solution containing 2.5 mM KCl, 1.25 mM NaH PO , 26 mM NaHCO , 25 mM glucose, 185 mM sucrose, 0.5 mM CaCl and 10 mM MgCl (pH 7.4, when bubbled with 95% O and 5% CO ). The brains were cut coronally into 200–250 μm-thick slices with a vibratome (VT-1200S, Leica). The slices were incubated at 37 °C for 1 h in artificial cerebrospinal fluid (aCSF) containing 125 mM NaCl, 2.5 mM KCl, 1.25 mM NaH PO , 26 mM NaHCO , 10 mM glucose, 2 mM CaCl and 1 mM MgCl (pH 7.4, when bubbled with 95% O and 5% CO ) before recordings. Slices were transferred to a recording chamber perfused with aCSF under an upright microscope (Axio Examiner D1, Zeiss). Patch pipettes were filled with solution containing 125 mM K-gluconate, 10 mM KCl, 10 mM HEPES, 0.5 mM EGTA, 8 mM phosphocreatine-Na , 4 mM ATP-Mg and 0.3 mM GTP-Na (pH 7.3 adjusted with KOH; osmolality, 290 mOsm l−1). The DpMe was identified with axon bundles. Recordings were made from cells located in the medial part of the DpMe. Cells showing no action potentials following current injection (>1 nA, > 5 ms) were discarded from analysis. Membrane potentials were recorded for 1–10 min. Sik3 hypomorph and UAS-Sik3, UAS-Sik3(S563A) transgenic flies were gifts from M. Montminy and J. B. Thomas40. elav-GS (GeneSwitch) stocks were from the Bloomington stock centre. Flies were reared at 25 °C under 12-h light:12-h dark cycle in 50–60% relative humidity on a standard fly food consisting of corn meal, yeast, glucose, wheat germ and agar. Sleep analysis was performed as described previously41. In brief, male flies (2–5 days old) were individually housed in glass tubes (length, 65 mm; inside diameter, 3 mm) containing standard fly food at one end and a cotton plug on the other end. Sucrose-agar (1% agar supplemented with 5% sucrose) food was used for the GeneSwitch system assay, instead of standard food. The glass tubes were placed in the Drosophila activity monitor (DAM) (Trikinetics) and the locomotor activity of each fly was recorded as the number of infrared beam crossings in 1-min bin. Sleep was defined as periods of inactivity lasting 5 min or longer. Sleep assay were performed for 3 d under 12-h light:12-h dark cycle conditions and then constant darkness conditions. For 12-h light:12-h dark cyles, zeitgeber time (ZT) was used, and for constant darkness, circadian time (CT), with CT0 as 12 h after lights-off of the last 12-h light:12-h dark conditions, was used to indicate the daily time. For conditional expression analysis, we used the GeneSwitch system42 where expression is induced by a steroid hormone antagonist RU486. Flies are monitored for 3 days in tubes without drug in constant darkness and then transferred to new tubes either with vehicle (0.5% DMSO) alone or with 0.5 mM RU486 and then further monitored under constant darkness conditions. The expression of endogenous or transgenic Sik3 genes was confirmed by RT–PCR using RNA from fly heads. The wild-type strain N and the mutant strain PY1479 kin-29(oy38) X were obtained from the Caenorhabditis Genetics Center (CGC)43. All worms were maintained at 20 °C on nematode growth medium (NGM) agar plates seeded with E. coli HB101. For construction of P ::kin-29, kin-29 cDNA was amplified by RT–PCR and inserted into the plasmid pPD-DEST (a gift from Y. Iino) to generate pDEST-KIN-29. Next, we carried out the LR-recombinase reaction (Gateway System, Life Technologies) between pENTR-P (a gift from Y. Iino) and pDEST-KIN-29 to generate P ::kin-29. P ::kin-29 was injected at 30 ng μl−1 together with the injection marker P ::mcherry (10 ng μl−1) and the empty vector pPD49_26 (60 ng μl−1) into the kin-29(oy38) mutant worms. Quiescence during the L4 to adult lethargus was measured using the microfluidic-chamber based assay44. In brief, polydimethylsiloxane-made microfluidic chambers containing liquid NGM and the E. coli HB101 were loaded with early L4 larvae and sealed with a cover glass plus 2% agarose, and set under the microscope. Images were taken every 2 s for 12 to 20 h at 20 ± 0.5 °C using the microscope M205FA (Leica) equipped with the camera MC120HD (Leica) (pixel size: 1,024 μm × 768 μm) controlled by Leica Application Suite V4.3 or the microscope SZX16 (Olympus) equipped with the camera GR500BCM2 (Shodensha) (pixel size: 1,024 μm × 768 μm) controlled by μManager (UCSF). Subtraction between serial images was carried out using Image J, and worms were regarded as quiescent at a specific time point if the difference from the preceding time point was less than 1% of the total body size. The fraction of quiescence was defined as the number of quiescent time points divided by the total number of time points during a period of 10 min. The onset of lethargus quiescence was defined as the time point after which the fraction of quiescence was higher than 0.05 for at least 20 min, whereas the end point was defined as the time point after which the fraction of quiescence was lower than 0.05 for at least 20 min. Occasionally, brief episodes of quiescence were observed outside of lethargus both in wild-type and mutant worms; these episodes were excluded by setting a threshold of 60 min for the minimum duration of lethargus quiescence. Sample sizes were determined using R software based on averages and standard deviations that were obtained from small scale experiments. No method of randomization was used in any of the experiments. The experimenters were blinded to genotypes and treatment assignment. Statistical analysis was performed using SPSS Statistics 22 (IBM) and R software. All data were tested for Gaussian distribution and variance. Homogeneity of variances was tested with Levene’s test. We used Student’s t-test for pairwise comparisons, one-way ANOVA for multiple comparisons, one-way repeated measure ANOVA for multiple comparisons with multiple data points, and two-way ANOVA for multiple comparisons involving two independent variables. ANOVA analyses were subjected to Tukey’s post-hoc test. When deviation from normality and lack of homogeneity of variances occurred (P < 0.05), Mann–Whitney U test was used for group comparison. P < 0.05 was considered statistically significant. The datasets generated and/or analysed during the current study are available from the corresponding author on reasonable request.


BURLINGTON, Mass.--(BUSINESS WIRE)--ArQule, Inc. (Nasdaq: ARQL) today announced that preclinical data was presented on its proprietary AKT inhibitor, ARQ 092, in an oral presentation by the University of Illinois College of Medicine at the American Society of Hematology (ASH) Annual Meeting. The presentation highlighted preclinical studies of ARQ 092 in sickle cell disease (SCD). ARQ 092 is an orally available, selective pan-AKT inhibitor. Title: Specific inhibition of AKT with ARQ 092, an orally-available selective allosteric AKT inhibitor, attenuates acute vaso-occlusive events in sickle cell disease The presentation can be viewed at https://www.arqule.com/wp-content/uploads/ASH-2016-ARQ-092-in-Sickle-Cell-Disease-.pdf. “SCD is a debilitating lifetime disease with a worldwide presence. A report from the American Society of Hematology shows that approximately 100,000 people suffer from this disease in the U.S.” said Dr. Jaehyung Cho, PhD., of the University of Illinois College of Medicine. “Our research has been focused on identifying novel therapeutic targets to prevent and treat vaso-occlusive diseases. The work done at the University of Illinois, in collaboration with ArQule, shows the importance of the AKT pathway in inducing vaso-occlusive events during SCD and the beneficial effect of ARQ 092 in attenuating adhesive function of neutrophils and platelets in SCD mice. This work warrants further exploration of ARQ 092 in patients with SCD.” “Our AKT program, specifically ARQ 092, continues to progress with the phase 1 trial in Proteus syndrome sponsored by the National Institutes of Health (NIH) and with the recently approved Investigational New Drug (IND) application for expansion into the PROS family of rare over-growth diseases,” said Dr. Brian Schwartz, M.D., Head of Research and Development and Chief Medical Officer at ArQule. “The PI3K/AKT pathway has been implicated in multiple diseases including SCD. We would like to thank the University of Illinois College of Medicine for lending its expertise and helping to establish valuable proof of concept for ARQ 092 in this disease setting.” About the AKT Pathway and ARQ 092 ARQ 092 is an orally bioavailable, selective small molecule inhibitor of the AKT kinases. The AKT pathway when abnormally activated is implicated in multiple oncogenic processes such as cell proliferation and apoptosis. This pathway has emerged as a target of potential therapeutic relevance for compounds that inhibit its activity, which has been linked to a variety of cancers as well as to select non-oncology indications. ARQ 092, the lead compound in ArQule’s AKT program, has completed phase 1a clinical testing and has advanced into phase 1b expansion testing in cohorts of patients with endometrial cancer, lymphomas and tumors harboring either AKT or PI3K mutations. It is also in a phase 1 trial being conducted by the NIH for Proteus syndrome, a rare over-growth disease from the PROS family. Collaborators are exploring, in preclinical testing, other indications for ARQ 092 including sickle cell disease. ArQule is a biopharmaceutical company engaged in the research and development of targeted therapeutics to treat cancers and rare diseases. Our mission is to discover, develop and commercialize novel small molecule drugs in areas of high unmet need that will dramatically extend and improve the lives of our patients. Our clinical-stage pipeline consists of five drug candidates, all of which are in targeted, biomarker-defined patient populations, making ArQule a leader among companies our size in precision medicine. ArQule’s lead product, in phase 3 clinical development, is tivantinib (ARQ 197), an oral, selective inhibitor of the c-MET receptor tyrosine kinase, for second-line treatment of hepatocellular carcinoma in partnership with Daiichi Sankyo in the West and Kyowa Hakko Kirin in Asia. ArQule’s proprietary pipeline includes: ARQ 087, a multi-kinase inhibitor designed to preferentially inhibit the fibroblast growth factor receptor (FGFR) family, in phase 2 for iCCA and in phase 1b for multiple oncology indications; ARQ 092, a selective inhibitor of the AKT serine/threonine kinase, in phase 1 for multiple oncology indications as well as ultra-rare Proteus syndrome, in partnership with the National Institutes of Health (NIH); ARQ 751, a next generation AKT inhibitor, in phase 1 for patients with AKT1 and PI3K mutations; and ARQ 761, a β-lapachone analog being evaluated as a promoter of NQO1-mediated programmed cancer cell necrosis, in phase 1/2 in multiple oncology indications in partnership with the University of Texas Southwestern Medical Center. In addition, we have advanced ARQ 531, an investigational, orally bioavailable, potent and reversible inhibitor of both wild type and C481S-mutant BTK, into toxicology testing and plan to file an Investigational New Drug Application in early 2017. ArQule’s current discovery efforts are focused on the identification and development of novel kinase inhibitors, leveraging the Company’s proprietary library of compounds. You can follow us on Twitter and LinkedIn. This press release contains forward-looking statements regarding certain pre-clinical experiments conducted by the Company’s collaborators and the Company’s clinical trials with ARQ 092. These statements are based on the Company’s current beliefs and expectations, and are subject to risks and uncertainties that could cause actual results to differ materially. Positive information about pre-clinical, and early stage clinical trial, results, including in SCD and Proteus syndrome, does not ensure that later stage or larger scale clinical trials will be successful. For example, ARQ 092 may not demonstrate promising therapeutic effect; in addition, the drug candidate may not demonstrate appropriate safety profiles in current or later stage or larger scale clinical trials as a result of known or as yet unanticipated side effects. The results achieved in later stage trials may not be sufficient to meet applicable regulatory standards or to justify further development. Problems or delays may arise during clinical trials or in the course of developing, testing or manufacturing the compound that could lead the Company or its partners, including the National Institutes of Health, to discontinue development. Even if later stage clinical trials are successful, unexpected concerns may arise from subsequent analysis of data or from additional data. Obstacles may arise or issues may be identified in connection with review of clinical data with regulatory authorities. Regulatory authorities may disagree with the Company’s view of the data or require additional data or information or additional studies. Drug development involves a high degree of risk. Only a small number of research and development programs result in the commercialization of a product. Positive pre-clinical data may not be supported in later stages of development. Furthermore, ArQule may not have the financial or human resources to successfully pursue drug discovery in the future. For more detailed information on the risks and uncertainties associated with the Company’s drug development and other activities, see the Company’s periodic reports filed with the Securities and Exchange Commission. The Company does not undertake any obligation to publicly update any forward-looking statements.


News Article | September 8, 2016
Site: www.sciencenews.org

President Barack Obama’s “Cancer Moonshot” now has a scientific flight plan. It calls for better cooperation among researchers and institutions, aggressive pursuit of immunotherapy and making better use of proven cancer prevention strategies. Called the Blue Ribbon Panel Report, the document was approved September 7 by the National Cancer Advisory Board, part of the National Cancer Institute. Five months in the making, the report’s 10 recommendations for research priorities were put together by a 28-member group of cancer experts appointed last April. It’s the most specific direction yet for the moonshot (SN: 4/2/16, p. 20), launched when Obama announced the intention to make the United States “the country that cures cancer once and for all” in his State of the Union address in January. Vice President Joe Biden, whose son Beau died in 2015 from brain cancer at age 46, has been leading the charge. If the vice president formally adopts the recommendations, they could form the foundation for research grants awarded through the National Institutes of Health. But that depends largely on the U.S. Congress providing funding for the moonshot, which has not yet happened. The report doesn’t contain dramatic surprises and doesn’t veer far off course from current cancer research. That’s largely by design. The goal of the panel was to come up with a plan to make “10 years of progress in five years” by hastening those areas with the most promise, which also stand to affect large numbers of patients. “We want it to be a pushy evolution, not a revolution,” says Stan Gerson, director of the Case Comprehensive Cancer Center in Cleveland, who was not on the panel. “There are some things here that are right on the money where we ought to be focused.” The Cancer Moonshot’s Blue Ribbon Panel presented 10 research recommendations to the National Cancer Advisory Board on September 7. Patients would enter genetic information about their cancer into a national database, making it easier to find and participate in clinical trials. A clinical trials network devoted exclusively to immunotherapies would focus on what’s considered one of the most promising advances in cancer treatment. More research is needed to understand drug resistance, a major cause of death from cancer. This system would link many of the nation’s largest data repositories, enabling free, one-stop access for researchers, doctors and patients to share data. Intensify research on the major drivers of childhood cancers More research into the biology of pediatric and adolescent cancers could lead to improved treatments. Guidelines are needed to minimize and manage the often excruciating side effects of treatment. Expand use of proven prevention and early detection strategies Researchers need to identify ways to increase adherence to proven strategies, such as the HPV vaccine, especially in medically underserved populations and those with inherited genetic risks. Researchers should analyze stored tumor tissue from patients who received standard treatment to search for genetic and other factors that predict which people would benefit from standard care versus experimental treatment. Aweb-based catalog that maps the genetic and cellular evolution of tumors would enable researchers to develop predictive models of tumor progression and response to treatment. More public–private sector collaborations are needed to develop new technologies to help doctors deliver more effective therapies to patients. If there is any underlying thread to the report, it is that progress depends on greater cooperation in research and improvements in patient engagement. “There’s not a lot new under the sun in terms of the areas they have targeted. The most important concept here with the vice president’s efforts is that he can serve as an accelerant at a time when we’re at an inflection point in treating cancer,” says Gary Gilliland, president and director of the Fred Hutchinson Cancer Research Center in Seattle. “Within 10 years if we don’t get to a place where we’ve got curative approaches to essentially all cancers then we’ve failed — and shame on us.” For treatment, the report singles out immunotherapy, which harnesses a patient’s own immune system to fight cancer. The strategy is widely regarded as one of the most significant advances in cancer care, even though so far only 10 to 20 percent of patients receiving such treatments show long-term benefit. “When I speak to my patients, I tell them that the greatest risk is disappointment,” says medical oncologist David Gerber of the University of Texas Southwestern Medical Center in Dallas. Nonetheless, he and others remain optimistic about immunotherapy’s potential, and he agrees with recommendations to speed up research. In proposing an immunotherapy clinical trials network, the report states that current treatments “represent only the tip of the iceberg of what is possible.” Since the moonshot began, Biden has crisscrossed the country, touring cancer research centers, holding photo ops and meeting with doctors. In June, he presided over a cancer summit at Howard University in Washington, D.C. One theme Biden has stressed at these events — and was reflected in the panel’s report — is the need for better data sharing. Traditionally, raw scientific data remains the property of the institutions and researchers who conduct studies. But this can impede collaboration — between institutions or across disciplines — and make it harder to find patterns within genetics and biology that might reveal how cancer appears and grows. The new report acknowledges the problem with research silos, stating that “our ability to accelerate progress against cancer demands that researchers, clinicians and patients across the country collaborate in sharing their collective data and knowledge about the disease.” Among the recommendations is the creation of a National Cancer Data Ecosystem, a one-stop, free collection of data that will allow patients to upload and receive data about their specific type of tumor. While Obama heralded the moonshot as a cure for cancer, Gerson, from the Case Comprehensive Cancer Center, would also like to see more scientists take on less flashy issues, finding better ways to save lives in known ways. Among them: “How do we get people to stop smoking?” he says. “We’re not doing it well.” Tyler Jacks, the report cochair and director of the Koch Institute for Integrative Cancer Research at MIT, said during a news conference that the panel recognizes that progress against cancer is about “emphasizing the use of known prevention strategies. It’s not just about treatment.” In discussing prevention, the report notes low rates of adoption for the human papillomavirus vaccine, which protects against the virus that causes cervical and other cancers, and for colorectal cancer, or CRC, screening. “If we understood better the reasons these proven cancer prevention strategies are not being widely used and how we could increase uptake of these strategies,” the report states, “we could reduce deaths due to cervical cancer by 90 percent, CRC by up to 70 percent and lung cancer by as much as 95 percent.” The authors also note that many people carry inherited genetic risks for cancer and don’t know it, and improved screening for genetic predisposition could save lives. What happens to the moonshot after this year depends largely on Congress. The report will eventually be forwarded to the vice president and the moonshot task force he oversees. The panel did not say how much the recommendations would cost. But in his budget request for fiscal year 2017, Obama asked for $680 million for additional funding to pay for the moonshot. The panel members confined themselves to scientific matters but noted that barriers to cancer progress are not just about research. Disparities keep many patients from getting treatments today, much less improvements coming tomorrow. “The one concern I have is that they put off the side policy issues as being out of scope for the Blue Ribbon Panel. I think it’s appropriate, but in the end it’s the policy issues that are going to determine the success of the entire program,” says Gilliland. “How do you get coverage and reimbursement? How do you ensure privacy if you’re sequencing everybody’s genome? How do you address access to clinical trials? Those are some of the very hardest problems, and if we don’t solve them it’s not going to matter how clever we are in the laboratory or how sensitive our techniques are for early detection.”


Motta-Mena L.B.,University of Texas Southwestern Medical Center | Heyd F.,University of Pennsylvania | Lynch K.W.,University of Texas Southwestern Medical Center | Lynch K.W.,University of Pennsylvania
Molecular Cell | Year: 2010

Splicing regulatory proteins often have distinct activities when bound to exons versus introns. However, less clear is whether variables aside from location can influence activity. HnRNP L binds to a motif present in both CD45 variable exons 4 and 5 to affect their coordinate repression. Here, we show that, in contrast to its direct repression of exon 4, hnRNP L represses exon 5 by countering the activity of a neighboring splicing enhancer. In the absence of the enhancer, hnRNP L unexpectedly activates exon inclusion. As the splice sites flanking exon 4 and 5 are distinct, we directly examined the effect of varying splice site strength on the mechanism of hnRNP L function. Remarkably, binding of hnRNP L to an exon represses strong splice sites but enhances weak splice sites. A model in which hnRNP L stabilizes snRNP binding can explain both effects in a manner determined by the inherent snRNP-substrate affinity. © 2010 Elsevier Inc. All rights reserved.


Balcer L.J.,University of Pennsylvania | Frohman E.M.,University of Texas Southwestern Medical Center
Neurology | Year: 2010

Background: Disturbances in visual function are common in patients with multiple sclerosis (MS) and are often accompanied by substantial impairments in daily functioning and quality of life. Lesions associated with these impairments frequently involve the afferent visual pathway. Expert Clinical Opinion: Because these impairments are often not readily apparent on commonly used high-contrast acuity tests, low-contrast charts (e.g., low-contrast Sloan letter charts) have gained validity in the assessment of visual dysfunction in patients with MS. Decrements in low-contrast letter acuity are associated with MS and correlate with increasing disability, MRI abnor-malities, and reduced retinal nerve fiber layer (RNFL) thickness as measured by optical coherence tomography (OCT). These findings suggest that low-contrast letter acuity testing is a potentially useful addition to disability scales such as the Multiple Sclerosis Functional Composite, serving as another surrogate marker for MS disability. Assessment of RNFL thickness by OCT, which is also associated with visual impairment, also may be considered for inclusion in clinical trials eval-uating treatments for MS. Future Directions: The effects of disease-modifying therapies on visual dysfunction in patients with MS have been evaluated only recently. Two phase 3 studies of natalizumab showed that low-contrast letter acuity testing, included as an exploratory outcome, demonstrated treatment effects. Other ongoing studies have incorporated low-contrast acuity and OCT measures of RNFL thickness. The availability and wider use of low-contrast letter acuity tests, in combination with ocular imaging techniques, may improve assessment of treatment efficacy in patients with MS. Copyright © 2010 by AAN Enterprises, Inc. All rights reserved.


Sonnenberg A.,Portland Medical Center P3 | Sonnenberg A.,Oregon Health And Science University | Genta R.M.,Miraca Life science | Genta R.M.,University of Texas Southwestern Medical Center | Genta R.M.,Dallas Medical Center
American Journal of Gastroenterology | Year: 2013

OBJECTIVES:It has been suggested that Helicobacter pylori (H. pylori) constitutes a risk for the development of adenomatous polyps and adenocarcinoma of the colon. Our aim was to study the association between H. pylori-positive gastritis and the occurrence of any colonic neoplasm.METHODS:From a computerized database of surgical pathology reports, we selected 156,000 subjects who underwent colonoscopy and esophago-gastro-duodenoscopy with biopsy results from both procedures.RESULTS:Compared with normal gastric mucosa, H. pylori gastritis occurred more frequently among patients with hyperplastic polyps (OR=1.24, 95% CI: 1.18-1.30), adenomatous polyps (1.52, 1.46-1.57), advanced adenomas (1.80, 1.69-1.92), villous adenomas or adenomas with high-grade dysplasia (1.97, 1.82-2.14), and adenocarcinomas (2.35, 1.98-2.80). Similarly, the strength of the association between H. pylori-positive gastritis and colonic neoplasm increased with size and number of the adenomas. The association between H. pylori gastritis and the occurrence of colonic neoplasm was similar for different locations of the large bowel. Other gastric conditions etiologically associated with H. pylori, such as intestinal metaplasia, adenoma, lymphoma, and adenocarcinoma, were also significantly associated with an increased risk of colonic neoplasm.CONCLUSIONS:Various forms of gastritis related to H. pylori infection confer an increased risk for colonic neoplasm. In the past, when H. pylori infection was more prevalent, its attributable risk to the occurrence of colorectal neoplasm may have been quite substantial. © 2013 by the American College of Gastroenterology.


Popugaeva E.,Saint Petersburg State University | Bezprozvanny I.,University of Texas Southwestern Medical Center
Frontiers in Molecular Neuroscience | Year: 2013

Alzheimer disease (AD) is a major threat of twenty-first century that is responsible for the majority of dementia in the elderly. Development of effective AD-preventing therapies are the top priority tasks for neuroscience research. Amyloid hypothesis of AD is a dominant idea in the field, but so far all amyloid-targeting therapies have failed in clinical trials. In addition to amyloid accumulation, there are consistent reports of abnormal calcium signaling in AD neurons. AD neurons exhibit enhanced intracellular calcium (Ca2+) liberation from the endoplasmic reticulum (ER) and reduced store-operated Ca2+ entry (SOC). These changes occur primarily as a result of ER Ca2+ overload. We argue that normalization of intracellular Ca2+ homeostasis could be a strategy for development of effective disease-modifying therapies. The current review summarizes recent data about changes in ER Ca2+ signaling in AD. Ca2+ channels that are discussed in the current review include: inositol trisphosphate receptors, ryanodine receptors, presenilins as ER Ca2+ leak channels, and neuronal SOC channels. We discuss how function of these channels is altered in AD and how important are resulting Ca2+ signaling changes for AD pathogenesis. © 2013 Popugaeva and Bezprozvanny.


Friedman D.I.,University of Texas Southwestern Medical Center | Liu G.T.,University of Pennsylvania | Digre K.B.,University of Utah
Neurology | Year: 2013

The pseudotumor cerebri syndrome (PTCS) may be primary (idiopathic intracranial hypertension) or arise from an identifiable secondary cause. Characterization of typical neuroimaging abnormalities, clarification of normal opening pressure in children, and features distinguishing the syndrome of intracranial hypertension without papilledema from intracranial hypertension with papilledema have furthered our understanding of this disorder. We propose updated diagnostic criteria for PTCS to incorporate advances and insights into the disorder realized over the past 10 years. © 2013 American Academy of Neurology.


Rohatgi A.,University of Texas Southwestern Medical Center | Khera A.,University of Texas Southwestern Medical Center | Berry J.D.,University of Texas Southwestern Medical Center | Givens E.G.,University of Texas Southwestern Medical Center | And 7 more authors.
New England Journal of Medicine | Year: 2014

Background: It is unclear whether high-density lipoprotein (HDL) cholesterol concentration plays a causal role in atherosclerosis. A more important factor may be HDL cholesterol efflux capacity, the ability of HDL to accept cholesterol from macrophages, which is a key step in reverse cholesterol transport. We investigated the epidemiology of cholesterol efflux capacity and its association with incident atherosclerotic cardiovascular disease outcomes in a large, multiethnic population cohort.Methods: We measured HDL cholesterol level, HDL particle concentration, and cholesterol efflux capacity at baseline in 2924 adults free from cardiovascular disease who were participants in the Dallas Heart Study, a probability-based population sample. The primary end point was atherosclerotic cardiovascular disease, defined as a first nonfatal myocardial infarction, nonfatal stroke, or coronary revascularization or death from cardiovascular causes. The median follow-up period was 9.4 years.Results: In contrast to HDL cholesterol level, which was associated with multiple traditional risk factors and metabolic variables, cholesterol efflux capacity had minimal association with these factors. Baseline HDL cholesterol level was not associated with cardiovascular events in an adjusted analysis (hazard ratio, 1.08; 95% confidence interval [CI], 0.59 to 1.99). In a fully adjusted model that included traditional risk factors, HDL cholesterol level, and HDL particle concentration, there was a 67% reduction in cardiovascular risk in the highest quartile of cholesterol efflux capacity versus the lowest quartile (hazard ratio, 0.33; 95% CI, 0.19 to 0.55). Adding cholesterol efflux capacity to traditional risk factors was associated with improvement in discrimination and reclassification indexes.Conclusions: Cholesterol efflux capacity, a new biomarker that characterizes a key step in reverse cholesterol transport, was inversely associated with the incidence of cardiovascular events in a population-based cohort. Copyright © 2014 Massachusetts Medical Society.


Sonnenberg A.,Oregon Health And Science University | Genta R.M.,Caris Diagnostics | Genta R.M.,University of Texas Southwestern Medical Center
Alimentary Pharmacology and Therapeutics | Year: 2012

Background There is some preliminary evidence to suggest that patients with inflammatory bowel disease (IBD) are less frequently infected with Helicobacter pylori than the general population. Aim To examine whether the prevalence of Helicobacter pylori (H. pylori) is lower among IBD patients compared with non-IBD individuals based on results from surgical pathology. Methods From a database of surgical pathology reports, we recruited a sample of unique patients who underwent a same-day bidirectional gastrointestinal endoscopy with biopsies. Of the total 65 515 patients, 1061 served as cases with IBD and 64 451 as controls without IBD. The histological presence of H. pylori was correlated with the patients' demographic characteristics and histological presence of any oesophageal disease, Crohn's disease (CD), ulcerative colitis (UC) and indeterminate colitis (IND). Results were expressed as odds ratios (OR), using multivariate logistic regression to adjust for the cofounding influence of comorbidities and demographic characteristics. Results The presence of H. pylori was inversely associated with IBD, the adjusted OR and their 95% confidence intervals being 0.48 (0.27-0.79) for CD, 0.59 (0.39-0.84) for UC and 0.43 (0.15-0.95) for IND. In contradistinction, H. pylori-negative gastritis was positively associated with IBD, the adjusted OR being 11.06 (7.98-15.02) for CD, 2.25 (1.31-3.60) for UC and 6.91 (3.50-12.30) for IND. Conclusions Our study confirms an inverse association between H. pylori and IBD and a positive association between the H. pylori-negative gastritis and IBD. These relationships may open new avenues to study the pathogenesis of IBD. © 2012 Blackwell Publishing Ltd.


Yin J.,University of Texas Southwestern Medical Center | Mobarec J.C.,University of Marburg | Kolb P.,University of Marburg | Rosenbaum D.M.,University of Texas Southwestern Medical Center
Nature | Year: 2015

The orexin (also known as hypocretin) G protein-coupled receptors (GPCRs) respond to orexin neuropeptides in the central nervous system to regulate sleep and other behavioural functions in humans. Defects in orexin signalling are responsible for the human diseases of narcolepsy and cataplexy; inhibition of orexin receptors is an effective therapy for insomnia. The human OX 2 receptor (OX 2 R) belongs to the β branch of the rhodopsin family of GPCRs, and can bind to diverse compounds including the native agonist peptides orexin-A and orexin-B and the potent therapeutic inhibitor suvorexant. Here, using lipid-mediated crystallization and protein engineering with a novel fusion chimaera, we solved the structure of the human OX 2 R bound to suvorexant at 2.5 Å resolution. The structure reveals how suvorexant adopts a π-stacked horseshoe-like conformation and binds to the receptor deep in the orthosteric pocket, stabilizing a network of extracellular salt bridges and blocking transmembrane helix motions necessary for activation. Computational docking suggests how other classes of synthetic antagonists may interact with the receptor at a similar position in an analogous π-stacked fashion. Elucidation of the molecular architecture of the human OX 2 R expands our understanding of peptidergic GPCR ligand recognition and will aid further efforts to modulate orexin signalling for therapeutic ends. © 2015, Macmillan Publishers Limited. All rights reserved.


DUARTE, CA--(Marketwired - Feb 16, 2017) - Prolacta Bioscience®, the pioneer in human milk-based neonatal nutritional products, announced today that the first patient has been enrolled in a clinical study to evaluate the effect of a specially fortified exclusive human milk diet (EHMD)1 to improve growth in infants who have undergone surgery for a serious heart defect known as single ventricle cardiac physiology. Prolacta developed a first-of-its-kind, human milk-based fortifier to address the post-surgical nutrition needs of this fragile infant population. Single ventricle cardiac physiology occurs in five out of 100,000 live births,2 and results in a range of clinical problems resulting from the heart having only one adequate ventricle or pumping chamber.2 The condition is usually fatal unless treated by a series of surgeries.3 Babies with single ventricle cardiac physiology often have difficulty breathing, feeding and growing.3,4 "These medically frail newborns are at high risk for growth failure, due to increased caloric demands due to their heart and pulmonary conditions. This presents unique nutritional challenges," said Principal Investigator Cynthia Blanco, M.D., University of Texas Health Science Center. "We believe a densely fortified human milk diet may be a real benefit to these infants with an ultimate goal of optimizing their nutritional status prior to their second surgery." "While not preemies, these babies have a similar need for rich nutrition delivered in small volumes," said Scott Elster, CEO of Prolacta. "This study is the first to explore the benefits of human milk-based fortifiers in a population of fragile infants other than preemies. The medical centers will be using a new formulation that we developed specifically for these babies, and we hope that it will make a difference to their health and healing." The randomized controlled study will evaluate a minimum of 84 infants, starting at age seven days or younger, who require surgery to correct a single ventricle cardiac physiology present at birth, and who were fed an EHMD prior to study enrollment. The study will measure growth rate and clinical outcomes for up to 30 days following surgery in these infants. One group will be fed an EHMD that includes a new formulation of Prolacta's highest caloric density exclusive human milk-based fortifier, while the other group will receive a standard diet that includes some cow milk-based nutrition. The hypothesis is that infants fed the EHMD will have short-term benefits, including improved growth, reduced episodes of feeding intolerance, sepsis and necrotizing enterocolitis (damaged intestinal tissue), reduced stay in the hospital and potentially improved long-term neurodevelopmental outcomes. The study is titled, "A Randomized Controlled Trial to Evaluate Growth Velocity and Clinical Outcomes of Infants with Single Ventricle Physiology Fed an Exclusive Human Milk Diet with Early Nutritional Fortification Following Surgical Repair." It is expected to take place at 12 centers: University of Texas Health Science Center at San Antonio (lead center), San Antonio, Texas; University of Texas Southwestern Medical Center, Dallas, Texas; Texas Children's Hospital, Houston, Texas; Cook Children's Medical Center, Fort Worth, Texas; Children's Hospital of Philadelphia, Philadelphia, Pa.; Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio; Children's Hospital of Orange County, Orange, Calif.; Ann & Robert H. Lurie Children's Hospital of Chicago, Chicago, Ill.; Columbia University Medical Center, New York, N.Y.; Emory University Hospital, Atlanta, Ga.; University of Oklahoma Health Sciences Center, Oklahoma City, Ok.; and UF Health Shands Children's Hospital, Gainesville, Fla. About Prolacta Bioscience Prolacta Bioscience, Inc. is a privately-held life sciences company dedicated to Advancing the Science of Human Milk®. The company pioneered the development of human milk-based neonatal nutritional products to meet the needs of critically ill, premature infants in the NICU. Prolacta leads the industry in the quality and safety of nutritional products made from donor breast milk and operates the first and only pharmaceutical-grade manufacturing facility for the processing of human milk. For more information, please visit www.prolacta.com. 1 An exclusive human milk diet (EHMD) is when 100 percent of the protein, fat and carbohydrates in an infant's intake are derived solely from human milk. 2 Adult Congenital Heart Association. "Single Ventricle Defects and the Fontan." Accessed Nov. 13, 2016. 3 Cadet, J.V., "Children's Hospital of Philadelphia: Treating Single Ventricle Defects." Cardiovascular Business. July 26, 2010. Accessed Nov. 13, 2016. 4 American Heart Association. "Single Ventricle Defects." Updated Oct. 21, 2015. Accessed Nov. 13, 2016.


News Article | December 6, 2016
Site: www.eurekalert.org

A new biopharmaceutical company, Tenaya Therapeutics Inc., will build on discoveries in cardiovascular disease research made at the Gladstone Institutes, concentrating on regenerative medicine and drug discovery for heart failure. The new company combines Gladstone's basic science expertise with the resources and translational know-how of the biotechnology industry. Cardiovascular disease -- including heart attacks and congenital heart defects -- is the world's leading cause of death. Heart failure alone afflicts more than 20 million people around the world. "When heart muscle is damaged, the body is unable to repair the dead or injured cells," explained Deepak Srivastava, MD, director of the Gladstone Institute of Cardiovascular Disease and a co-founder of the new company. "Right now, the only possible cure for heart failure is a heart transplant. We hope that this new venture will bring us closer to a more scalable cure." "There has been limited commercial investment in the cardiac field, despite the remarkable progress made at Gladstone and elsewhere," added Stephen Freedman, PhD, vice president for corporate liaison and ventures at Gladstone. "Tenaya will advance some of the innovative work coming out of Gladstone, with the aim of developing new therapeutics for heart failure." Tenaya is supported by a $50 million Series A financing from The Column Group. David Goeddel, PhD, a managing partner at The Column Group and a pioneer of the biotechnology industry, is the board chair of Tenaya. JJ Kang, PhD, an associate at The Column Group, is a board director and president of the new company. "The key ingredients for a successful company are a great team and great ideas," said Goeddel. "With the stellar group of scientists that we've assembled, we're confident that we will be able to build something transformative from the cutting-edge research of the Gladstone Institutes." Tenaya will leverage Gladstone's pioneering work in cellular reprogramming to search for cures for heart failure. One program will translate cellular reprogramming technology to the clinic to regenerate heart muscle cells in patients with heart failure. Other programs will use cellular models of heart disease created from stem cells to identify potential new drug targets. The new company is the first formed out of BioFulcrum, an entrepreneurial initiative within Gladstone that takes a milestone driven approach to science. BioFulcrum aims to accelerate the discovery of cures by bringing together scientists, multiple non-profit institutions, and industry partners such as Goeddel, who sits on the board of BioFulcrum. "Inspired by the collaborative research at BioFulcrum, we wanted to establish Tenaya to integrate new findings and tools in the cardiac field and advance them towards clinical translation," said Kang. "Our goal is to build a science-focused company that discovers novel therapies for heart failure patients." Scientific co-founders of the company include Srivastava and Gladstone Investigators Benoit Bruneau, PhD, Bruce Conklin, MD, Sheng Ding, PhD, and Saptarsi Haldar, MD, as well as Eric Olson, PhD, from the University of Texas Southwestern Medical Center. Other Gladstone scientists will serve as scientific partners (Katherine Pollard, PhD, Todd McDevitt, PhD, Nevan Krogan, PhD) or founding employees of the company, including Kathy Ivey, PhD, former director of the Gladstone Stem Cell Core and the new director of research operations at Tenaya. "The launch of Tenaya Therapeutics exemplifies Gladstone's fierce dedication to scientific discovery and to putting those discoveries on a path to pioneering new therapeutics," said Gladstone President R. Sanders "Sandy" Williams, MD, who is on the board of Tenaya. "Deepak assembled a group of investigators adept at discovery within Gladstone, and our novel BioFulcrum initiative accelerated their progress. Plus, in Dave Goeddel and JJ Kang from The Column Group, we found the perfect partners for this new enterprise." To ensure our work does the greatest good, the Gladstone Institutes focuses on conditions with profound medical, economic, and social impact--unsolved diseases of the brain, the heart, and the immune system. Affiliated with the University of California, San Francisco, Gladstone is an independent, nonprofit life science research organization that uses visionary science and technology to overcome disease.


News Article | February 27, 2017
Site: www.eurekalert.org

Lifestyle patterns, including physical activity and body mass index (BMI), are associated with a risk of overall heart failure but are more strongly associated with the heart failure subtype HFpEF, according to a study published today in the Journal of the American College of Cardiology. Heart failure is a medical condition defined by the inability of the heart to meet the demands of the body, particularly during exertion. Heart failure with preserved ejection fraction (HFpEF) is a subtype of heart failure that involves the heart and other organs and is characterized a stiff heart muscle that is unable to fill adequately with blood, resulting in fluid backing up into the lungs and body. Heart failure with preserved ejection fraction accounts for up to 50 percent of heart failure cases and is associated with poor outcomes. It has also proven to be resistant to available therapies, leading to prevention being a critical part of controlling the growing burden of this disease. "We consistently found an association between physical activity, BMI and overall heart failure risk," said Jarett D. Berry, MD, associate professor in the department of internal medicine and clinical sciences and director of cardiac rehabilitation at the University of Texas Southwestern Medical Center in Dallas, and the study's senior author. "This was not unexpected, however the impact of these lifestyle factors on heart failure subtypes was quite different." Researchers analyzed data from three cohort studies that included 51,541 participants with 3,180 heart failure events. The three studies used to pool data were the Women's Health Initiative, the Multiethnic Study of Atherosclerosis, and the Cardiovascular Health Study. The current study included all participants from the three cohort studies free from cardiovascular disease at baseline had quantitative measures of physical activity and BMI. In addition, medical experts reviewed all hospitalizations in these study participants to determine whether they were hospitalized for heart failure over the subsequent several years after they enrolled into these studies. Study participants with higher levels of physical activity were most often white, male and had higher annual income and education levels. Those with a higher level of physical activity had a lower prevalence of traditional risk factors, such as hypertension, diabetes, smoking and obesity. Participants with high BMIs were younger, had lower levels of physical activity and had a higher prevalence of cardiovascular risk factors. A total of 3,180 heart failure events were observed from the pooled data; of these events, 39.4 percent were HFpEF, 28.7 percent were heart failure with reduced ejection fraction (HFrEF, the type of heart failure that is associated with a weak heart muscle that doesn't pump well), and 31.9 percent were unclassified. When compared to no physical activity, low levels of physical activity were associated with 6 percent lower risk of heart failure. Researchers found that higher levels of physical activity had even lower risk of heart failure--11 percent lower risk for those who met the guideline-recommended amount of activity and 22 percent lower risk for greater than guideline-recommended physical activity. Overall, the incidence of HFpEF was significantly lower among study participants with higher levels of physical activity. For those achieving physical activity levels above the guideline-recommended amount, they had a 19 percent lower risk of HFpEF. However, there was no association between higher levels of physical activity and risk of HFrEF. Similarly, higher BMI was associated with increased overall heart failure risk. The cumulative incidence of HFpEF was higher among those with higher BMI. Once again, the association between higher BMI and HFrEF was more attenuated compared to HFpEF. Ambarish Pandey, MD, a cardiology fellow at the University of Texas Southwestern Medical Center in Dallas and first author on the study said, "There was a distinct relationship between both physical activity and BMI and the different heart failure subtypes, which may have important clinical and public health implications. These data suggest the importance of modifying lifestyle patterns to help prevent HFpEF in the general population." Limitations include the inability to prove a cause-and-effect relationship due to its observational nature. In an accompanying editorial, Sanjiv J. Shah, MD, professor of medicine and director of the Northwestern HFpEF Program at Northwestern University Feinberg School of Medicine in Chicago, said the researchers "have provided strong evidence that lack of physical activity is associated with incident HFpEF." Shah said there is a "critical need to focus on primary prevention in order to control HFpEF at the population level." The American College of Cardiology is a 52,000-member medical society that is the professional home for the entire cardiovascular care team. The mission of the College is to transform cardiovascular care and to improve heart health. The ACC leads in the formation of health policy, standards and guidelines. The College operates national registries to measure and improve care, offers cardiovascular accreditation to hospitals and institutions, provides professional medical education, disseminates cardiovascular research and bestows credentials upon cardiovascular specialists who meet stringent qualifications. For more, visit acc.org. The Journal of the American College of Cardiology is the most widely read cardiovascular journal in the world and is the top ranked cardiovascular journal for its scientific impact. JACC is the flagship for a family of journals that publish peer-reviewed research on all aspects of cardiovascular disease. JACC: Cardiovascular Interventions, JACC: Cardiovascular Imaging and JACC: Heart Failure also rank among the top ten cardiovascular journals for impact. JACC: Clinical Electrophysiology and JACC: Basic to Translational Science are the newest journals in the JACC family. Learn more at JACC.org.


News Article | February 15, 2017
Site: www.prweb.com

Akonni Biosystems, a molecular diagnostics (MDx) company that develops, manufactures, and intends to market integrated MDx systems, announced today the appointments of key members to its leadership team. Michael Murphy, M.Sc. joined as Vice President, Regulatory Affairs; Sandra Foster, Ph.D. as Director of Quality Assurance and Michael Reinemann, MPH as Director of Business Development. The new additions to Akonni’s leadership team fill critical gaps needed to ensure Akonni’s successful commercialization of its robust product lines. These experienced individuals further strengthen Akonni’s leadership team as the company prepares for its first FDA submission for a pharmacogenomic test on the Akonni TruDiagnosis® system. Michael Murphy is an industry pioneer and thought-leader in the field of Pharmacogenomics, with more than 33 years of scientific and business experience. He is a serial entrepreneur in the personalized medicine space and in 1997 was founder of Intek Labs, the first international Pharmacogenomics company. Following the acquisition of Intek Labs by PPD, Inc., Mr. Murphy was the co-founder, President and CEO of Gentris Corporation. Gentris was acquired by Cancer Genetics Inc. in 2015 while Mr. Murphy served on the Board of Directors at Gentris. In 2007, Gentris spun off its diagnostic group and Mr. Murphy served as the President and CEO of ParagonDx, one of the first companies to win FDA clearance of a Rapid Genotyping Kit for patients taking the anticoagulant, Warfarin. He has also held Executive Vice President management positions with PPGx and Clingenix. Prior to joining Akonni, Mr. Murphy served for 7 years as President of Conatus Consulting, a regulatory consulting practice based in Raleigh, NC. Mr. Murphy is a frequent lecturer and author on Pharmacogenomic topics, and currently sits on the editorial review board of the journal, Pharmacogenomics. He has been responsible for over 15 successful 510(k) submissions, FDA audits and inspections. He brings the expertise in FDA regulations and Quality Management Systems for medical devices that Akonni needs to advance its commercialization and registration efforts. Sandra Foster brings a unique blend of scientific knowledge and quality experience. She began her scientific career as a Medical Technologist (MT, ASCP), working in hospital laboratories. From the clinical lab she transitioned to research before going to graduate school. She earned a Ph.D. in Immunology from the University of Texas Southwestern Medical Center, followed by a post-doctoral fellowship at Duke University. Dr. Foster spent 12 years in clinical-phase biotechnology companies in roles of increasing responsibility from pre-clinical research and product design and development, to directing the manufacture of clinical trial materials and leading Quality Assurance and Regulatory Compliance. She designed ISO 14644-compliant clean room facilities for the manufacture of cellular therapy products for clinical trials, and implemented quality systems to support those activities. She designed, wrote and implemented process validations, operator qualifications, aseptic process simulations, comparability protocols, and authored multiple CMC sections for INDs. Immediately prior to joining Akonni, Dr. Foster owned her own consulting company, Triangle GxP Solutions, LLC as well as worked collaboratively with Mr. Murphy at Conatus. Client projects included translating R&D protocols into cGMP compliant SOPs, implementing quality systems, conducting client staff training, BLA, and pre-approval inspection (PAI) readiness. In her role as Director of Quality Assurance for Akonni, she leads design control efforts for product development, manages Device History Files, and prepares Akonni’s first audit for ISO 13485 Certification and FDA submission. Michael Reinemann has an exceptional track record of developing and implementing strategic, data-driven marketing and sales initiatives for diagnostic products resulting in strong double-digit growth and increased market share. Mr. Reinemann brings a diverse background, with experience in both technical and business roles. While earning his Master’s in Public Health at Columbia University in New York and working at the Mailman School of Public Health’s Center for Infection and Immunity, Mr. Reinemann worked on pioneering research projects in immunotherapy and pathogen discovery, and implemented cutting-edge technologies for highly multiplexed analysis and next-generation sequencing. Prior to joining Akonni in June of 2016, Mr. Reinemann served in various Commercial Operations roles at Qiagen. During his time at Qiagen, Mr. Reinemann led marketing and sales efforts that accelerated the growth of what has become the company’s single biggest revenue-contributing product. As Regional Marketing Manager of North America, Mr. Reinemann’s achievements included year-over-year growth of 55%, and the introduction of innovative co-marketing initiatives with strategic accounts, resulting in customer-specific growth of 75%. As Senior Global Product Manager, Mr. Reinemann managed a $150M product line with an annual growth rate of 25%, leading cross-functional project teams on commercial efforts as well as product development and product launches. His international business experience positions Akonni for success as the company navigates late-stage product development, registration, and commercialization of its technologies. “We are very excited to announce these essential additions to Akonni’s leadership team,” said Charles Daitch, Ph.D., President and CEO of Akonni Biosystems. “Each of these talented individuals bring valuable experience, demonstrated core competencies and dynamic industry insights from successful careers in clinical diagnostics. Our ability to hire people of this caliber speaks to the competitiveness of our product lines and their readiness to move expeditiously through the regulatory process. We are confident that we have the expertise needed to achieve our first FDA clearance and successful commercial launch of our TruDiagnosis platform.” Akonni is aggressively pursuing regulatory clearance of two product platforms – the TruDx®2000 platform and the TruTip® Automated Workstation. TruDx2000 is Akonni’s modular version of the TruDiagnosis® system, consisting of TruArray® three-dimensional (3D) gel-drop microarray diagnostic test devices and the TruDx Imager, complete with custom software for data analysis and reporting; the TruDx2000 can be bundled with or without the TruTip sample prep workstation depending on the needs of each clinical lab. The TruArray microfluidic device incorporates new, proprietary on-chip PCR technology, resulting in a much more user-friendly workflow, multiplexed detection, and a closed-amplicon system that virtually eliminates the risk of PCR contamination. The proprietary 3D gel-drop microarray nano-test-tubes can be tailored to detect genetic, protein or metabolite markers, providing the potential for access to a much broader range of diagnostic information from a single platform. The TruTip Automated Workstation is a small, affordable, fully-automated benchtop instrument. TruTip is a revolutionary technology that simplifies sample preparation by combining the complex protocols of DNA or RNA purification into just a few easy steps. The TruTip Automated Workstation is Akonni’s new nucleic acid purification instrument, which, in addition to blood and saliva, can homogenize and purify difficult samples such as sputum, stool and tissue. For more information visit: http://www.akonni.com About Akonni Biosystems Akonni Biosystems was founded in 2003 and has been issued 17 US and 24 International patents primarily covering sample preparation, microfluidic devices, bioinstrumentation, and integrated systems.    Product development has been supported by a series of government grants and contracts from NIH, CDC, DOE, DOD, NIJ, and NSF. The company significantly advanced the original technology by improving the system’s capabilities from sample preparation to test result. Commercial products in Akonni’s near-term pipeline include rapid sample preparation technologies for nucleic acid extraction and multiplex panel assays for detecting clinically relevant genotypes for pharmacogenomics, human chronic diseases, and genotypes for infectious diseases such as multidrug-resistant tuberculosis (MDR-TB), extensively drug-resistant tuberculosis (XDR-TB), upper respiratory infections, viral encephalitis, and hospital-acquired infections (MRSA).


News Article | November 11, 2016
Site: www.newsmaker.com.au

According to study results from Minneapolis Medical Research Foundation, in 2012, over 280,000 patients were living with bone metastasis in the U.S. with breast and prostate cancers being the most common primary tumor types. Metastasis is the unique hallmark of cancer cells which differentiates them from the benign tumors, which refers to the spread of cancer from its primary site to other body parts. Bone is the third most common site for the spread of cancer and invasion of such cancerous cells to bone gives rise to the deadly condition of bone metastasis. Bone metastasis is the third most common condition in metastatic cancers and occurs in 60-65% of patients with metastatic cancers. The spine is the most common site of bone metastasis. Other common sites include ribs, skull, pelvis (hip bone), femur (leg bone) and humerus (arm bone). Bone metastatic cancers are rarely able to be cured, and treatment of bone metastasis involves medications to shrink, stop or slow down the growth of the tumor. Treatment options available for bone metastasis are symptomatic treatments which are meant for pain control, treatment, and prevention of fractures, improvement in functional disability, etc. Randomized trials on metastatic patients by UBM Medica, LLC showed that 45-75% patients develop functional disability every 3-4 months, 10-15% patients develop hypercalcemia and 10-20% patients develop long bone fractures. Treatment of bone metastasis is the interdisciplinary field involving pain management, orthopedics, and other medical specialties. Primary factors driving the growth of bone metastasis therapeutics market are the global increase in cancer prevalence and delayed diagnosis of cancer in low-income countries. According to University of Texas Southwestern Medical Center Dallas, over 600,000 of bone metastasis are diagnosed in the U.S. every year. Rapid innovation in the field of personalized medicine and identification of new therapeutic targets for bone metastasis presents a huge opportunity to manufacturers of targeted therapy agents. A large number of treatment methods are still under investigation. However, heterogeneous nature of cancer and high development cost of neoplastic agents are the factors limiting the growth of global bone metastasis therapeutics market. Increasing investment by multinational companies in cancer research is expected to drive the growth of global bone metastasis therapeutics market during the forecast period. Based on treatment type, the global market for bone metastasis therapeutics has been classified as drug treatment, Tumor ablation therapy, and surgery. Drug treatment segment is foreseen to lead the market owing to easy availability of drugs and better reimbursement policies. Targeted therapeutic agents sub-segment of drug treatment segment is expected grow rapidly due to increasing acceptance of biological therapy owing to lesser side effects. By end user, the global bone metastasis therapeutics market has been segmented into hospitals, Clinics, cancer rehabilitation center and ambulatory surgical centers. Hospital end user segment is anticipated to contribute the maximum share among end users owing to the requirement of advanced healthcare infrastructure for management of bone metastasis symptoms. According to a study conducted by Amgen Inc., average hospitalization stay of cancer bone metastasis patients is 5.9 to 11.6 days, and almost 59% of the inpatient cost is attributed to skeletal-related events (SREs) such as radiation to the bone, spinal cord compression, pathologic fracture, etc. By regional presence, bone metastasis therapeutics market is segmented into five key regions viz.  North America, Latin America, Europe, Asia-Pacific, and the Middle East & Africa. North America will continue to dominate the global bone metastasis therapeutics market for due to high prevalence of malignant neoplasm. Europe is expected to hold second largest market share in global bone metastasis therapeutics market. Request TOC (desk of content material), Figures and Tables of the report: http://www.persistencemarketresearch.com/toc/11731 Some of the major players in global bone metastasis therapeutics market include F. Hoffmann-La Roche Ltd., Bayer AG, Merck & Co., Pfizer Inc. Novartis AG, Amgen Inc., Pharmalucence, Inc., Fresenius Kabi AG, Omega Laboratories Ltd., Eli Lilly and Company, and others. The majority of the key players are involved in development of new methods for treatment of cancer bone metastasis in collaboration with cancer research institutes such as M. D. Anderson Cancer Center, Memorial Sloan-Kettering Cancer Center, etc.


COLUMBIA, Md., Oct. 26, 2016 (GLOBE NEWSWIRE) -- Osiris Therapeutics, Inc. (NASDAQ:OSIR), a leading regenerative medicine company focused on developing and marketing products for wound care, orthopaedics, and sports medicine, announced today that a new peer-reviewed manuscript “Innovative Treatment of Chronic Diabetic Foot Ulcer in a Controlled Randomized Clinical Trial Produces Fewer Adverse Events, Faster Wound Closure, and Lower Costs” has been published in the Journal of Clinical Diabetes and Practice and is available online.   A cost comparative model was applied to patient data from the Grafix multicenter, controlled, randomized, blinded, clinical trial for chronic diabetic foot ulcers (DFUs) (Lavery et al., Int Wound J, 2014, 11: 554-560) by investigators at the Division of Health Care Policy and Research at the University of Colorado, Anschutz Medical Campus. This group evaluated the cost effectiveness and cost benefits of Grafix compared to Good Wound Care (GWC). Data demonstrated that the use of Grafix for the treatment of chronic DFUs is cost effective in comparison to GWC. The estimated savings during the trial based on associated adverse events and serious adverse events were approximately $14,000/patient. There were no Grafix-related reportable adverse or serious adverse events during the trial. When closed and non-closed wounds were compared, the estimated cost savings for closed wounds were also approximately $14,000/patient. Overall, lower costs were associated with patients managed with Grafix, and these patients experienced fewer adverse events, fewer serious adverse events, and fewer hospitalizations due to faster wound closure. “A DFU is a common complication of diabetes that is costly to treat. Poor or ineffective management of such wounds will lead to the use of intensive services, thus further increasing cost. Cost effectiveness should be an integral part for assessments of all wound care treatment modalities,” says Dr. Eugene J. Nuccio, an Assistant Professor within the Division of Health Care Policy and Research at the University of Colorado, Anschutz Medical Campus, the Principal Investigator of the Grafix cost comparative analysis. “We were very pleased to see significant reduction of wound related infections in the Grafix DFU trial,” said Dr. Lawrence Lavery, D.P.M., M.P.H., a Principal Investigator and Professor of Plastic Surgery, University of Texas Southwestern Medical Center. “The outcome of this study is critical evidence for every wound care provider and payer to consider. This study shows that adding Grafix in addition to GWC prevents adverse events and reduces the overall cost of care.” About Grafix Grafix is a cryopreserved placental membrane comprised of an extracellular matrix (ECM) rich in collagen, growth factors and viable cells native to the tissue. Grafix is processed using Osiris' proprietary BioSmart™ technology; it is flexible and conforming and designed for application directly to hard-to-treat acute and chronic wounds, including but not limited to diabetic foot ulcers, venous leg ulcers and thermal burns. About Osiris Therapeutics Osiris Therapeutics, Inc., based in Columbia, Maryland, is a world leader in researching, developing and marketing regenerative medicine products that improve health and lives of patients and lower overall healthcare costs. Having developed the world’s first approved stem cell drug, the company continues to advance its research and development in biotechnology by focusing on innovation in regenerative medicine – including bioengineering, stem cell research and viable tissue based products. Osiris has achieved commercial success with products in orthopaedics, sports medicine and wound care, including BIO4 ™, Cartiform®,  Grafix®, TruSkin ™ and Stravix™. Osiris, Grafix, Cartiform, TruSkin and Stravix are registered trademarks of Osiris Therapeutics, Inc., and BIO4 is a trademark of Howmedica Osteonics Corp. More information can be found on the company's website, www.Osiris.com. Forward-Looking Statements This press release contains forward-looking statements. Forward-looking statements include statements about our expectations, beliefs, plans, objectives, intentions, assumptions and other statements that are not historical facts. Words or phrases such as "anticipate," "believe," "continue," "ongoing," "estimate," "expect," "intend," "may," "plan," "potential," "predict," "project" or similar words or phrases, or the negatives of those words or phrases, may identify forward-looking statements, but the absence of these words does not necessarily mean that a statement is not forward-looking. Examples of forward-looking statements may include, without limitation, statements regarding the anticipated efficiencies, advantages, or cost-savings of products or services, as individual results may vary depending on the circumstances, means of use, costs of goods or services, and other factors. Forward-looking statements are subject to known and unknown risks and uncertainties and are based on potentially inaccurate assumptions that could cause actual results to differ materially from those expected or implied by the forward-looking statements. Accordingly, you should not unduly rely on these forward-looking statements. We undertake no obligation to publicly revise any forward-looking statement to reflect circumstances or events after the date of this press release or to reflect the occurrence of unanticipated events.


Gatica D.,University of Michigan | Chiong M.,University of Chile | Lavandero S.,University of Chile | Lavandero S.,University of Texas Southwestern Medical Center | Klionsky D.J.,University of Michigan
Circulation Research | Year: 2015

Autophagy is a catabolic recycling pathway triggered by various intra-or extracellular stimuli that is conserved from yeast to mammals. During autophagy, diverse cytosolic constituents are enveloped by double-membrane vesicles, autophagosomes, which later fuse with lysosomes or the vacuole to degrade their cargo. Dysregulation in autophagy is associated with a diverse range of pathologies including cardiovascular disease, the leading cause of death in the world. As such, there is great interest in identifying novel mechanisms that govern the cardiovascular response to disease-related stress. First described in failing hearts, autophagy within the cardiovascular system has been characterized widely in cardiomyocytes, cardiac fibroblasts, endothelial cells, and vascular smooth muscle cells. In all cases, a window of optimal autophagic activity seems to be critical to the maintenance of cardiovascular homeostasis and function; excessive or insufficient levels of autophagic flux can each contribute to heart disease pathogenesis. Here, we review the molecular mechanisms that govern autophagosome formation and analyze the link between autophagy and cardiovascular disease. © 2015 American Heart Association, Inc.


Unger R.H.,University of Texas Southwestern Medical Center | Unger R.H.,Veterans Affairs Medical Center | Scherer P.E.,University of Texas Southwestern Medical Center
Trends in Endocrinology and Metabolism | Year: 2010

Once considered divine retribution for sins, comorbidities of obesity (metabolic syndrome) are today attributed to obesity-induced metabolic defects. Here, we propose that obesity and hyperleptinemia protect lipid-intolerant nonadipose organs against lipotoxic lipid spillover during sustained caloric surplus. Metabolic syndrome is ascribed to lipotoxicity caused by age-related resistance to antilipotoxic protection by leptin. © 2010.


White P.,University of Texas Southwestern Medical Center | Bachega T.S.S.,University of Sao Paulo
Seminars in Reproductive Medicine | Year: 2012

The most frequent form of congenital adrenal hyperplasia (CAH) is steroid 21-hydroxylase deficiency, accounting for more than 90% of cases. Affected patients cannot synthesize cortisol efficiently. Thus the adrenal cortex is stimulated by corticotropin (ACTH) and overproduces cortisol precursors. Some precursors are diverted to sex hormone biosynthesis, causing signs of androgen excess including ambiguous genitalia in newborn females and rapid postnatal growth in both sexes. In the most severe "salt wastingo" form of CAH (~75% of severe or "classico" cases), concomitant aldosterone deficiency may lead to salt wasting with consequent failure to thrive, hypovolemia, and shock. Newborn screening minimizes delays in diagnosis, especially in males, and reduces morbidity and mortality from adrenal crises. CAH is a recessive disorder caused by mutations in the CYP21 (CYP21A2) gene, most of which arise from recombination between CYP21 and a nearby pseudogene, CYP21P (CYP21A1P). Phenotype is generally correlated with genotype. Classic CAH patients require chronic glucocorticoid treatment at the lowest dose that adequately suppresses adrenal androgens and maintains normal growth and weight gain, and most require mineralocorticoid (fludrocortisone). Transition of care of older patients to adult physicians should be planned in advance as a structured, ongoing process. Copyright © 2012 by Thieme Medical Publishers, Inc., 333 Seventh Avenue, New York, NY 10001, USA.


Vaishnava S.,University of Texas Southwestern Medical Center | Yamamoto M.,University of Texas Southwestern Medical Center | Severson K.M.,University of Texas Southwestern Medical Center | Ruhn K.A.,University of Texas Southwestern Medical Center | And 5 more authors.
Science | Year: 2011

The mammalian intestine is home to ~100 trillion bacteria that performimportant metabolic functions for their hosts. The proximity of vast numbers of bacteria to host intestinal tissues raises the question of how symbiotic host-bacterial relationships are maintained without eliciting potentially harmful immune responses. Here, we show that RegIIIg, a secreted antibacterial lectin, is essential for maintaining a ~50-micrometer zone that physically separates the microbiota from the small intestinal epithelial surface. Loss of host-bacterial segregation in RegIIIγ-/- mice was coupled to increased bacterial colonization of the intestinal epithelial surface and enhanced activation of intestinal adaptive immune responses by the microbiota. Together, our findings reveal that Reglllγ is a fundamental immune mechanism that promotes host-bacterial mutualism by regulating the spatial relationships between microbiota and host.


Huang H.,University of Texas Southwestern Medical Center | Fan X.,Dalian University of Technology | Williamson D.E.,University of Texas Health Science Center at San Antonio | Rao U.,Southwestern Medical Center
Neuropsychopharmacology | Year: 2011

Alterations in white matter integrity of several cortical and subcortical circuits have been reported in relation to unipolar major depressive disorder. It is not clear whether these white matter changes precede the onset of illness. In all, 13 adolescent volunteers with no personal or family history of a psychiatric disorder (controls) and 18 adolescent volunteers with no personal history of a psychiatric illness including depression, but who were at high risk for developing unipolar depression by virtue of parental depression (high-risk youth), underwent diffusion tensor imaging studies. An automated tract-based spatial statistics method, a whole-brain voxel-by-voxel analysis, was used to analyze the scans. Population average diffusion parameter values were also calculated for each tract. Adolescents at high risk for unipolar depression had lower fractional anisotropy (FA) values in the left cingulum, splenium of the corpus callosum, superior longitudinal fasciculi, uncinate, and inferior fronto-occipital fasciculi than did controls. Altered white matter integrity in healthy adolescents at familial risk for unipolar depression suggests that it might serve as a vulnerability marker for the illness. © 2011 American College of Neuropsychopharmacology. All rights reserved.


Girardi A.C.C.,University of Sao Paulo | di Sole F.,University of Texas Southwestern Medical Center
American Journal of Physiology - Cell Physiology | Year: 2012

The Na+/H+ exchanger-3 (NHE3) belongs to the mammalian NHE protein family and catalyzes the electro-neutral exchange of extracellular sodium for intracellular proton across cellular membranes. Its transport function is of essential importance for the maintenance of the body's salt and water homeostasis as well as acid-base balance. Indeed, NHE3 activity is finely regulated by a variety of stimuli, both acutely and chronically, and its transport function is fundamental for a multiplicity of severe and world-wide infection-pathological conditions. This review aims to provide a concise overview of NHE3 physiology and discusses the role of NHE3 in clinical conditions of prominent importance, specifically in hypertension, diabetic nephropathy, heart failure, acute kidney injury, and diarrhea. Study of NHE3 function in models of these diseases has contributed to the deciphering of mechanisms that control the delicate ion balance disrupted in these disorders. The majority of the findings indicate that NHE3 transport function is activated before the onset of hypertension and inhibited thereafter; NHE3 transport function is also upregulated in diabetic nephropathy and heart failure, while it is reported to be downregulated in acute kidney injury and in diarrhea. The molecular mechanisms activated during these pathological conditions to regulate NHE3 transport function are examined with the aim of linking NHE3 dysfunction to the analyzed clinical disorders. © 2012 the American Physiological Society.


Chow T.T.,University of Texas Southwestern Medical Center | Zhao Y.,University of Texas Southwestern Medical Center | Zhao Y.,Sun Yat Sen University | Mak S.S.,University of Texas Southwestern Medical Center | And 2 more authors.
Genes and Development | Year: 2012

Telomere overhangs are essential for telomere end protection and telomerase extension, but how telomere overhangs are generated is unknown. Leading daughter strands synthesized by conventional semiconservation DNA replication are initially blunt, while lagging daughter strands are shorter by at least the size of the final RNA primer, which is thought to be located at extreme chromosome ends. We developed a variety of new approaches to define the steps in the processing of these overhangs. We show that the final lagging RNA primer is not terminal but is randomly positioned ~70-100 nucleotides from the ends and is not removed for more than an hour. This identifies an important intrinsic step in replicative aging. Telomeric termini are processed in two distinct phases. During the early phase, which occupies 1-2 h following replication of the duplex telomeric DNA, several steps occur on both leading and lagging daughters. Leading telomere processing remains incomplete until late S/G2, when the C-terminal nucleotide is specified-referred to as the late phase. These observations suggest the presence of previously unsuspected complexes and signaling events required for the replication of the ends of human chromosomes. © 2012 by Cold Spring Harbor Laboratory Press.


Sarode R.,University of Texas Southwestern Medical Center | Milling T.J.,University of Texas at Austin | Refaai M.A.,University of Rochester | Mangione A.,CSL Behring LLC | And 3 more authors.
Circulation | Year: 2013

BACKGROUND - : Patients experiencing major bleeding while taking vitamin K antagonists require rapid vitamin K antagonist reversal. We performed a prospective clinical trial to compare nonactivated 4-factor prothrombin complex concentrate (4F-PCC) with plasma for urgent vitamin K antagonist reversal. METHODS AND RESULTS - : In this phase IIIb, multicenter, open-label, noninferiority trial, nonsurgical patients were randomized to 4F-PCC (containing coagulation factors II, VII, IX, and X and proteins C and S) or plasma. Primary analyses examined whether 4F-PCC was noninferior to plasma for the coprimary end points of 24-hour hemostatic efficacy from start of infusion and international normalized ratio correction (≤1.3) at 0.5 hour after end of infusion. The intention-to-treat efficacy population comprised 202 patients (4F-PCC, n=98; plasma, n=104). Median (range) baseline international normalized ratio was 3.90 (1.8-20.0) for the 4F-PCC group and 3.60 (1.9-38.9) for the plasma group. Effective hemostasis was achieved in 72.4% of patients receiving 4F-PCC versus 65.4% receiving plasma, demonstrating noninferiority (difference, 7.1% [95% confidence interval, -5.8 to 19.9]). Rapid international normalized ratio reduction was achieved in 62.2% of patients receiving 4F-PCC versus 9.6% receiving plasma, demonstrating 4F-PCC superiority (difference, 52.6% [95% confidence interval, 39.4 to 65.9]). Assessed coagulation factors were higher in the 4F-PCC group than in the plasma group from 0.5 to 3 hours after infusion start (P<0.02). The safety profile (adverse events, serious adverse events, thromboembolic events, and deaths) was similar between groups; 66 of 103 (4F-PCC group) and 71 of 109 (plasma group) patients experienced ≥1 adverse event. CONCLUSIONS - : 4F-PCC is an effective alternative to plasma for urgent reversal of vitamin K antagonist therapy in major bleeding events, as demonstrated by clinical assessments of bleeding and laboratory measurements of international normalized ratio and factor levels.


Mauvais-Jarvis F.,Northwestern University | Clegg D.J.,University of Texas Southwestern Medical Center | Hevener A.L.,University of California at Los Angeles
Endocrine Reviews | Year: 2013

Estrogens play a fundamental role in the physiology of the reproductive, cardiovascular, skeletal, and central nervous systems. In this report, we review the literature in both rodents and humans on the role of estrogens and their receptors in the control of energy homeostasis and glucose metabolism in health and metabolic diseases. Estrogen actions in hypothalamic nuclei differentially control food intake, energy expenditure, and white adipose tissue distribution. Estrogen actions in skeletal muscle, liver, adipose tissue, and immune cells are involved in insulin sensitivity as well as prevention of lipid accumulation and inflammation. Estrogen actions in pancreatic islet β-cells also regulate insulin secretion, nutrient homeostasis, and survival. Estrogen deficiency promotes metabolic dysfunction predisposing to obesity, the metabolic syndrome, and type 2 diabetes. We also discuss the effect of selective estrogen receptor modulators on metabolic disorders. © 2013 by The Endocrine Society.


Ye S.,Zhejiang University | Li Y.,CAS Shanghai Institute of Materia Medica | Jiang Y.,University of Texas Southwestern Medical Center
Nature Structural and Molecular Biology | Year: 2010

K+ channels are highly selective for K+ over Na +. Here we present several crystal structures of the MthK K + channel pore at up to 1.45-Åresolution. The MthK selectivity filter maintains a conductive conformation even in the absence of K+, allowing the channel to conduct Na+. The high-resolution structures, along with single-channel recordings, allow for an accurate analysis of how K+ competes with Na+ in a conductive selectivity filter. At high K+ concentrations, two K+ ions equivalently occupy the four sites in the selectivity filter, whereas at low K+/high Na+ concentrations, a single K+ ion remains bound in the selectivity filter, preferably at site 1 or site 3. This single K+ binding at low concentration effectively blocks the permeation of Na +, providing a structural basis for the anomalous mole-fraction effect, a key property of multi-ion pores. © 2010 Nature America, Inc. All rights reserved.


Doyle J.M.,University of Texas Southwestern Medical Center | Gao J.,Tsinghua University | Wang J.,Tsinghua University | Yang M.,Tsinghua University | Potts P.R.,University of Texas Southwestern Medical Center
Molecular Cell | Year: 2010

The melanoma antigen (MAGE) family consists of more than 60 genes, many of which are cancer-testis antigens that are highly expressed in cancer and play a critical role in tumorigenesis. However, the biochemical and cellular functions of this enigmatic family of proteins have remained elusive. Here, we identify really interesting new gene (RING) domain proteins as binding partners for MAGE family proteins. Multiple MAGE family proteins bind E3 RING ubiquitin ligases with specificity. The crystal structure of one of these MAGE-RING complexes, MAGE-G1-NSE1, reveals structural insights into MAGE family proteins and their interaction with E3 RING ubiquitin ligases. Biochemical and cellular assays demonstrate that MAGE proteins enhance the ubiquitin ligase activity of RING domain proteins. For example, MAGE-C2-TRIM28 is shown to target p53 for degradation in a proteasome-dependent manner, consistent with its tumorigenic functions. These findings define a biochemical and cellular function for the MAGE protein family. © 2010 Elsevier Inc.


Zong H.,University of Virginia | Parada L.F.,University of Texas Southwestern Medical Center | Baker S.J.,St Jude Childrens Research Hospital
Cold Spring Harbor Perspectives in Biology | Year: 2015

Malignant glioma remains incurable despite tremendous advancementinbasic research and clinical practice. The identification of the cell(s) of origin should provide deep insights into leverage points for one to halt disease progression. Here we summarize recent studies that support the notion that neural stem cell (NSC), astrocyte, and oligodendrocyte precursor cell (OPC) can all serveasthe celloforigin.Wealso layout important considerationsontechnical rigor for further exploring this subject. Finally, weshare perspectivesonhow one could apply the knowledge of cell of origin to develop effective treatment methods. Although it will be a difficult battle, victory should be within reach as along as we continue to assimilate new information and facilitate the collaboration among basic scientists, translational researchers, and clinicians. © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.


Hah N.,Cornell University | Danko C.G.,Cornell University | Core L.,Cornell University | Waterfall J.J.,Cornell University | And 4 more authors.
Cell | Year: 2011

We report the immediate effects of estrogen signaling on the transcriptome of breast cancer cells using global run-on and sequencing (GRO-seq). The data were analyzed using a new bioinformatic approach that allowed us to identify transcripts directly from the GRO-seq data. We found that estrogen signaling directly regulates a strikingly large fraction of the transcriptome in a rapid, robust, and unexpectedly transient manner. In addition to protein-coding genes, estrogen regulates the distribution and activity of all three RNA polymerases and virtually every class of noncoding RNA that has been described to date. We also identified a large number of previously undetected estrogen-regulated intergenic transcripts, many of which are found proximal to estrogen receptor binding sites. Collectively, our results provide the most comprehensive measurement of the primary and immediate estrogen effects to date and a resource for understanding rapid signal-dependent transcription in other systems. © 2011 Elsevier Inc.


Nakamura N.,Kyoto Sangyo University | Wei J.-H.,University of Texas Southwestern Medical Center | Seemann J.,University of Texas Southwestern Medical Center
Current Opinion in Cell Biology | Year: 2012

The Golgi apparatus is essential for post-translational modifications and sorting of proteins in the secretory pathway. In addition, it further performs a broad range of specialized functions. This functional diversity is achieved by combining basic morphological modules of cisternae into higher ordered structures. Linking cisternae into stacks that are further connected through tubules into a continuous Golgi ribbon greatly increases its efficiency and expands its repertoire of functions. During cell division, the different modules of the Golgi are inherited by different mechanisms to maintain its functional and morphological composition. © 2012 Elsevier Ltd.


Hah N.,Cornell University | Hah N.,Salk Institute for Biological Studies | Murakami S.,University of Texas Southwestern Medical Center | Nagari A.,University of Texas Southwestern Medical Center | And 3 more authors.
Genome Research | Year: 2013

We have integrated and analyzed a large number of data sets from a variety of genomic assays using a novel computational pipeline to provide a global view of estrogen receptor 1 (ESR1; a.k.a. ERa) enhancers in MCF-7 human breast cancer cells. Using this approach, we have defined a class of primary transcripts (eRNAs) that are transcribed uni- or bidirectionally from estrogen receptor binding sites (ERBSs) with an average transcription unit length of ~3-5 kb. The majority are up-regulated by short treatments with estradiol (i.e., 10, 25, or 40 min) with kinetics that precede or match the induction of the target genes. The production of eRNAs at ERBSs is strongly correlated with the enrichment of a number of genomic features that are associated with enhancers (e.g., H3K4me1, H3K27ac, EP300/CREBBP, RNApolymerase II, open chromatin architecture), as well as enhancer looping to target gene promoters. In the absence of eRNA production, strong enrichment of these features is not observed, even though ESR1 binding is evident. We find that flavopiridol, a CDK9 inhibitor that blocks transcription elongation, inhibits eRNA production but does not affect other molecular indicators of enhancer activity, suggesting that eRNA production occurs after the assembly of active enhancers. Finally, we show that an enhancer transcription "signature" based on GRO-seq data can be used for de novo enhancer prediction across cell types. Together, our studies shed new light on the activity of ESR1 at its enhancer sites and provide new insights about enhancer function. © 2013, Published by Cold Spring Harbor Laboratory Press.


BOSTON, Feb. 27, 2017 (GLOBE NEWSWIRE) -- Exonics Therapeutics, Inc., a newly formed biotechnology company focused on developing gene editing technologies like CRISPR/Cas9 to permanently correct a majority of mutations causing Duchenne muscular dystrophy and other neuromuscular diseases, today announced a commitment of $5 million in seed financing from CureDuchenne Ventures, LLC, a subsidiary of the nonprofit CureDuchenne. The initial seed funding will allow Exonics to advance the preclinical research of its scientific founder and chief science advisor Eric Olson, PhD. Dr. Olson’s laboratory has demonstrated the ability to use adeno-associated virus (AAV) to deliver a payload based on CRISPR/Cas9 technology that can identify and correct exon mutations that prevent the production of dystrophin, a protein that helps stabilize and protect muscle fibers. Dystrophin is the key protein missing in boys with Duchenne, and published preclinical data suggest that this approach has the potential to permanently treat up to 80 percent of children suffering from the disease. Additional preclinical data is expected to be published in March 2017. “This represents the next generation of potential Duchenne muscular dystrophy therapies. By leveraging the revolutionary CRISPR/Cas9 method to permanently correct errors in the DNA sequence, it is our hope that we can develop a one-time therapy that provides lifelong benefit to Duchenne patients,” said Dr. Olson, who also serves as Professor and Chairman of the Department of Molecular Biology at the University of Texas Southwestern Medical Center (UTSW), from which Exonics’ technology is licensed. Duchenne is a rare X-linked genetic progressive muscle disease affecting nearly 15,000 boys in the U.S. and more than 300,000 boys worldwide. There is no cure for Duchenne. Children with the disease start missing development milestones at age 3 and often lose their ability to walk by age 12. All Duchenne patients will suffer from reduced mobility and independence, and ultimately respiratory or cardiac failure results in a reduced life expectancy in the mid-20s. “We are building on a scientific program of exceptional quality from Dr. Olson’s laboratory with the support of our funder CureDuchenne Ventures, an organization with deep understanding of this disease, therapeutic landscape and large unmet need. This funding underscores their recognition of the potential of Exonics’ gene editing method to develop a novel therapy for Duchenne,” said Cristina Csimma, PharmD, MHP, Executive Chair of the Board of Directors of Exonics, former President and Chief Executive Officer of Cydan, and long-time drug development leader. “Exonics is extremely well-positioned to advance our preclinical program as we follow a rigorous approach to enable translation to clinical trials.” “We look forward to working closely with the Duchenne community as we aggressively advance gene editing technology to address the significant unmet need for a curative therapy that would dramatically improve the lives of patients with Duchenne and their families,” said Jak Knowles, MD, President and Interim Chief Executive Officer of Exonics. “We are honored to advance the groundbreaking work of Dr. Olson’s laboratory and are eager to translate this approach into an important therapy for the Duchenne community.” Dr. Knowles will continue to serve the Duchenne community as Managing Director of CureDuchenne Ventures, and VP of Medical and Scientific Affairs at CureDuchenne. “We are delighted to support Dr. Olson and the Exonics team as they advance novel gene editing technology toward a potential cure for Duchenne. CureDuchenne Ventures’ unique model enables us to share our significant scientific resources, expertise and deep connections with the Duchenne community to accelerate scientific breakthroughs, such as Exonics’ program,” said Debra Miller, President of CureDuchenne Ventures and Founder and Chief Executive Officer of CureDuchenne.                                                                                       About Exonics Therapeutics Exonics Therapeutics is advancing gene editing technologies like CRISPR/Cas9 to permanently correct the majority of Duchenne muscular dystrophy mutations. Preclinical data suggest Exonics’ novel gene editing approach has the potential to permanently treat up to 80 percent of children who suffer from Duchenne. Exonics’ technology is licensed from the University of Texas Southwestern Medical Center and is based on the research of its scientific founder and chief science advisor, Eric Olson, PhD. The Company’s corporate office is located in Boston, Mass., and research activities are being conducted in Dallas, Tex. For more information, please visit www.exonicstx.com. CureDuchenne Ventures LLC collaborates with pharmaceutical and biotechnology companies to facilitate the development of drugs to treat Duchenne muscular dystrophy. CureDuchenne Ventures LLC was formed by CureDuchenne, a national nonprofit that has funded nine research projects that have advanced to human clinical trials.


News Article | February 27, 2017
Site: www.technologyreview.com

Knowles says the $5 million investment in Exonics is the largest ever by CureDuchenne in a single company, reflecting its belief that gene editing could go far beyond existing drugs for Duchenne. “CRISPR is not some repurposed drug doing the bare minimum,” says Knowles. “It’s something with high impact.” Exonics will advance research underway at the University of Texas Southwestern Medical Center, where scientist Eric Olson and colleagues have cured mice of muscular dystrophy using CRISPR, stirring intense hopes among patients. MIT Technology Review profiled Olson’s efforts last year. Olson says his next step is to treat larger animals, such as dogs or monkeys. “If that is as positive as we think it will be, we would move to humans,” says Olson, who is a cofounder of the company and owns a stake in it. Exonics did not provide a timeline for when a human study could begin. Olson says he had the chance to raise money from traditional investors or to partner with a large biotech company, but decided a company backed by a patient group would move the treatment along faster. “I get e-mails from mothers every day,” says Olson. “I think this enables me to move forward in the most effective way.” Officially, CRISPR drug technology is dominated by three public biotechnology companies—Editas Medicine, Intellia Therapeutics, and CRISPR Therapeutics—all based in Cambridge, Massachusetts, and which have raised more than $1 billion among them. Two, Editas and CRISPR Therapeutics, list muscular dystrophy among the diseases they are interested in, but it’s not a top priority. The companies are primarily developing treatments for blindness, blood disorders, liver disease, and cancer. Knowles says CureDuchenne feared the muscle disease was not receiving enough attention by the bigger biotechs. “We want to move fast and we don’t have the conflict of priorities a larger company does,” says Knowles. More patients groups could soon hatch their own break-away CRISPR plans. That’s because the technique is versatile enough that it could help with scores of ultra-rare inherited diseases, many of which are now untreatable. The gene that goes wrong in Duchenne muscular dystrophy, called dystrophin, was discovered 30 years ago. Boys who lack a working copy of dystrophin become paralyzed when their muscles waste away and usually die of heart failure before they turn 25. Olson and others have already shown CRISPR technology can repair the dystrophin gene in mice. But gene-editing ingredients have never been directly injected into a living person, which is Exonics’s goal. Patients would receive an injection of trillions of viruses, each harboring the instructions to edit the DNA of the dystrophin gene in their muscle cells. If enough muscle cells get corrected—perhaps 15 percent—the progression of the disease could be halted, Olson thinks. Because many different mutations in the dystrophin gene can lead to muscular dystrophy, initially a CRISPR treatment wouldn’t fix all of them. Olson says the treatment he’s working on targets part of the gene known as “exon 51” and, if it works, would help about 13 percent of boys with the disease. Drug development experts caution that CRISPR studies may not work out as planned. “There is a concern in the Duchenne community that patients have become overly excited that CRISPR will let them get up and walk,” says Susan J. Ward, executive director of the Collaborative Trajectory Analysis Project, which helps drugmakers develop better ways of testing new drugs. In fact, delays lasting years, or decades, are common when scientists attempt to craft treatments from a new technology, only to encounter unexpected roadblocks or safety issues. “I think CRISPR is truly exciting,” says Ward. “But it’s not a slam dunk yet.”


News Article | November 25, 2015
Site: www.nature.com

All mice were maintained on a C57BL/6 background, including ScfGFP (ref. 19), Scffl/+ (ref. 19), Cxcl12DsRed (ref. 18), Cxcl12fl/+ (ref. 18), R26tdTomato (ref. 26), Vav1-cre (ref. 24), Leprcre (ref. 27), Tcf21cre/ER (ref. 21) and α-catulinGFP. To induce Cre/ER activity in Tcf21cre/ER mice, 4–6-week-old mice were administered 2 mg tamoxifen (Sigma) daily by oral gavage for 12 consecutive days. For induction of EMH, mice were injected at day 0 with a single dose of 4 mg cyclophosphamide followed by daily injections of 5 μg G-CSF for 4–21 days. Both male and female mice were used. All mice were housed in the Animal Resource Center at the University of Texas Southwestern Medical Center (UTSW). All procedures were approved by the UTSW Institutional Animal Care and Use Committee. Bone marrow cells were isolated by flushing the femur or tibia with Ca2+- and Mg2+-free HBSS with 2% heat-inactivated bovine serum using a 3 ml syringe fitted with a 25-gauge needle. Spleen cells were obtained by crushing the spleen between two frosted slides. The cells were dissociated to a single-cell suspension by gently passing through the needle several times and then filtering through a 40-μm nylon mesh. Blood was collected by cardiac puncture, and white blood cells were isolated by ficoll centrifugation according to the manufacturer’s instructions (GE Healthcare). The following antibodies were used to isolate HSCs: anti-CD150 (TC15-12F12.2), anti-CD48 (HM48-1), anti-Sca-1 (E13-161.7), anti-c-kit (2B8) and the following antibodies against lineage markers (anti-Ter119, anti-B220 (6B2), anti-Gr-1 (8C5), anti-CD2 (RM2-5), anti-CD3 (17A2), anti-CD5 (53-7.3) and anti-CD8 (53-6.7)). Haematopoietic progenitors were identified by flow cytometry using the following antibodies: anti-Sca-1 (E13-161.7), anti-c-Kit (2B8) and the following antibodies against lineage markers (anti-Ter119, anti-B220 (6B2), anti-Gr-1 (8C5), anti-CD2 (RM2-5), anti-CD3 (17A2), anti-CD5 (53-7.3) and anti-CD8 (53-6.7)), anti-CD34 (RAM34), anti-CD135 (Flt3) (A2F10), anti-CD16/32 (FcγR) (93), anti-CD127 (IL7Rα) (A7R34), anti-CD24 (M1/69), anti-CD43 (1B11), anti-B220 (6B2), anti-IgM (II/41), anti-CD3 (17A2), anti-Gr-1 (8C5), anti-Mac-1 (M1/70), anti-CD41 (MWReg30), anti-CD71 (C2) and anti-Ter119. 4′,6-Diamidino-2-phenylindole (DAPI) was used to exclude dead cells. Antibodies were obtained from eBioscience or BD Bioscience. To isolate bone marrow stromal cells the marrow was gently flushed out of the bone marrow cavity with a 3-ml syringe fitted with a 23-guage needle and then transferred into 1 ml pre-warmed bone marrow digestion solution (200 U ml−1 DNase I (Sigma), 250 μg ml−1 LiberaseDL (Roche) in HBSS plus Ca2+ and Mg2+) and incubated at 37 °C for 30 min with gentle shaking. To isolate splenic stromal cells, the spleen capsule was cut into ~1 mm3 fragments using scissors and then digested as described earlier in spleen digestion solution (200 U ml−1 DNase I, 250 μg ml−1 LiberaseDL, 1 mg ml−1 collagenase, type 4 (Roche) and 500 μg ml−1 collagenase D (Roche) in HBSS plus Ca2+ and Mg2+). After a brief vortex, the spleen fragments were allowed to sediment for ~3 min and the supernatant was transferred to another tube on ice. The sedimented (undigested) spleen fragments were subjected to a second round of digestion. The two fractions of digested cells were pooled and filtered through a 100-μm nylon mesh. Anti-PDGFR-α (APA5), anti-PDGFR-β (APB5), anti-LepR (R&D), anti-CD45 (30F-11) and anti-Ter119 antibodies were used to isolate stromal cells. For analysis of endothelial cells, mice were injected intravenously into the retro-orbital venous sinus with 10 μg Alexa-Fluor-660-conjugated anti-VE-cadherin antibody (BV13) 10 min before being killed. Samples were analysed using a FACSAria or FACSCanto II flow cytometer (BD Biosciences). To assess BrdU incorporation into spleen cells after EMH induction, mice were intraperitoneally injected with a single dose of BrdU (2 mg BrdU per mouse) then maintained on 0.5 mg BrdU per ml drinking water for 7 days. Endothelial cells were labelled by intravenous injection of an anti-VE-cadherin antibody (eBioscience). Enzymatically dissociated spleen cells were stained with antibodies against surface markers and the target cell populations were sorted then resorted to ensure purity. The sorted cells were then fixed, and stained with an anti-BrdU antibody using the BrdU APC Flow Kit (BD Biosciences) according to the manufacturer’s instructions. Adult recipient mice were irradiated using an XRAD 320 X-ray irradiator (Precision X-Ray) with two doses of 540 rad (total 1,080 rad) delivered at least 2 h apart. Cells were injected into the retro-orbital venous sinus of anaesthetized mice. Sorted doses of splenocytes from donor mice with EMH were transplanted along with 3 × 105 recipient bone marrow cells. Recipient mice were bled every 4 weeks to assess the level of donor-derived blood cells, including myeloid, B and T cells for at least 16 weeks. Blood was subjected to ammonium chloride/potassium red cell lysis before antibody staining. Antibodies including anti-CD45.2 (104), anti-CD45.1 (A20), anti-Gr1 (8C5), anti-Mac-1 (M1/70), anti-B220 (6B2) and anti-CD3 (KT31.1) were used for flow cytometric analysis. For bone marrow sections, freshly dissected bones were fixed in 4% paraformaldehyde overnight followed by 3 days of decalcification in 10% EDTA dissolved in PBS. Bones were sectioned using the CryoJane tape-transfer system (Instrumedics). For spleen sections, freshly dissected spleens were fixed in 4% paraformaldehyde for 1 h followed by 1 day incubation in 10% sucrose in PBS. Frozen spleens were sectioned with a cryostat (Leica). For whole mount imaging, spleens were sectioned into ~2 mm pieces. Spleen sections were blocked in PBS with 10% horse serum for 1 h and then stained overnight with chicken-anti-GFP (Aves) and/or rabbit-anti-laminin (Abcam) antibodies. Donkey-anti-chicken Alexa Fluor 488 and/or donkey-anti-rabbit Alexa Fluor 647 were used as secondary antibodies (Invitrogen). Specimens were mounted with anti-fade prolong gold (Invitrogen) and images were acquired with either a Zeiss LSM780 confocal microscope or a Leica SP8 confocal microscope equipped with a resonant scanner. Three-dimensional images were achieved using Bitplane Imaris v.7.7.1 software. Spleens were harvested and fixed for 4 h in 4% PFA at 4 °C. Since the spleen capsule is highly autofluorescent, spleens were sectioned perpendicular to the long axis into 300-μm-thick sections using a Leica VT100S vibrotome. These 300-μm sections were fixed for an additional 2 h in 4% PFA and blocked overnight in staining solution (10% dimethylsulfoxide (DMSO), 0.5% IgePal630 (Sigma) and 5% donkey serum (Jackson Immunoresearch) in PBS). All staining steps were performed in staining solution on a rotator at room temperature. Spleen sections were stained for 3 days in primary antibodies, washed overnight in several changes of PBS then stained for 3 days in secondary antibodies. The stained sections were dehydrated in a methanol dehydration series then incubated for 3 h in 100% methanol with several changes. The methanol was then exchanged with benzyl alcohol:benzyl benzoate 1:2 mix (BABB clearing28). The tissues were incubated in BABB for 3 h to overnight with several exchanges of fresh BABB. Spleen sections were mounted in BABB between two coverslips and sealed with silicone (Premium waterproof silicone II clear; General Electric). We found it necessary to clean the BABB of peroxides (which can accumulate as a result of exposure to air and light) by adding 10 g of activated aluminium oxide (Sigma) to 40 ml of BABB and rotating for at least 1 h, then centrifuging at 2,000 g for 10 min to remove the suspended aluminium oxide particles. Images were acquired using a Zeiss LSM780 confocal microscope with a Zeiss LD LCI Plan-Apo ×25/0.8 multi-immersion objective lens, which has a 570 μm working distance. Images were taken at 512 × 512 pixel resolution with 2 μm Z-steps, pinhole for the internal detector at 47.7 μm. Random spots were inserted into images by generating randomized X, Y, and Z coordinates using the random integer generator at http:// www.random.org. After mouse anaesthesia by ketamine/xylazine, a ventral midline incision was made and the peritoneum was breached. The splenic blood vessels were ligated with an absorbable suture (4-0 vicryl). The splenic vessels were cut distal to the suture and the spleen was removed. The vessels were cauterized and the abdomen was sutured with non-absorbable sutures (3-0 Tevdek III). Buprenorphine was administered every 12 h for 3 days to minimize postoperative pain and mice were maintained with ampicillin-containing water to avoid infection. Complete blood counts were measured one month after the survival surgery. EMH was induced by repeated bleeding over a 2-week period according to a published protocol2. Briefly, 4–6 month-old mice were bled via the tail vein five times, every 3 days, removing approximately 250 μl of blood each time, then the mice were killed for analysis 2 days after the last bleed. Approximately 30,000 CD45−Ter119−VE-cadherin+ splenic endothelial cells were flow cytometrically sorted into 50 μl of 66% trichoracetic acid (TCA) in water. Extracts were incubated on ice for at least 15 min and centrifuged at 16,100 g at 4 °C for 10 min. Precipitates were washed in acetone twice and the dried pellets were solubilized in 9 M urea, 2% Triton X-100, and 1% dithiothreitol (DTT). Samples were separated on 4–12% Bis-Tris polyacrylamide gels (Invitrogen) and transferred to PVDF membrane (Millipore). The blots were incubated with primary antibodies overnight at 4 °C and then with secondary antibodies. Blots were developed with the SuperSignal West Femtochemiluminescence kit (Thermo Scientific). Primary antibodies used: rabbit-anti-SCF (Abcam, 1:1,000) and mouse-anti-actin (Santa Cruz, clone AC-15, 1:20,000). Cells were sorted directly into Trizol (Life Technologies). Total RNA was extracted according to the manufacturer’s instructions. Total RNA was reverse transcribed using SuperScript III Reverse Transcriptase (Life Technologies). Quantitative real-time PCR was performed using SYBR green on a LightCycler 480 (Roche). β-Actin was used to normalize the RNA content of samples. Primers used in this study were Scf: 5′-GCCAGAAACTAGATCCTTTACTCCTGA-3′ and 5′-CATAAATGGTTTTGTGACACTGACTCTG-3′; β-actin: 5′-GCTCTTTTCCAGCCTTCCTT-3′ and 5′-CTTCTGCATCCTGTCAGCAA-3′. Three independent samples of 5,000 spleen Scf-GFP+VE-cadherin− spleen stromal cells and two independent samples of 5,000 unfractionated spleen cells were flow cytometrically sorted into Trizol. Total RNA was extracted, amplified, and sense strand cDNA was generated using the Ovation Pico WTA System V2 (NuGEN) according to the manufacturer’s instructions. cDNA was fragmented and biotinylated using the Encore Biotin Module (NuGEN) according to the manufacturer’s instructions. Labelled cDNA was hybridized to Affymetrix Mouse Gene ST 1.0 chips according to the manufacturer’s instructions. Expression values for all probes were normalized and determined using the robust multi-array average (RMA) method29. Panels in all figures represented multiple independent experiments performed on different days with different mice. Sample sizes were not based on power calculations. No randomization or blinding was performed. No animals were excluded from analysis. Variation is always indicated using standard deviation. For analysis of the statistical significance of differences between two groups we generally performed two-tailed Student’s t-tests. For analysis of the statistical significance of differences among more than two groups, we performed repeated measures one-way analysis of variance (ANOVA) tests with Greenhouse–Geisser correction (variances between groups were not equal) and Tukey’s multiple comparison tests with individual variances computed for each comparison. To assess the statistical significance of differences in fetal mass between paired control and mutant mice (Fig. 5j and Extended Data Fig. 8v), we performed a two-way ANOVA.


October 28, 2016 - How should plastic surgeons choose the best implant type and size for women undergoing breast augmentation surgery? Implant size selection systems based on breast tissue measurements may provide better outcomes, suggests a research review in the November issue of Plastic and Reconstructive Surgery®, the official medical journal of the American Society of Plastic Surgeons (ASPS). Tissue-based planning systems--using clinical guidelines to determine the optimal breast implant dimensions for individual patients--appear superior to approaches relying more on the patient's or surgeon's preference, according to the study by Drs. William P. Adams, Jr., of University of Texas Southwestern Medical Center, Dallas, and Daniel McKee of McMaster University, Hamilton, Ont., Canada. But further studies will be needed to clarify how breast implant size selection systems affect the outcomes of breast augmentation. The researchers performed a "data-driven review" of methods used by plastic surgeons to select the appropriate implant size for breast augmentation surgery. Breast augmentation is the most popular cosmetic plastic surgery procedure in the United States, with nearly 280,000 procedures performed in 2015, according to ASPS statistics. The review identified 33 articles on implant sizing systems. Studies evaluating TBP sizing systems were of higher quality than those in the other two categories. "The top ten studies based on methodological quality all used patients' breast dimensions before selecting final implant dimensions, and this should now be considered standard of practice based on our analysis," Drs. Adams and McKee write. The TBP studies reported low rates of repeat surgery, compared to industry standards and accepted research values. The researchers emphasize some major limitations of the available evidence on implant sizing systems. Just four out of 33 studies reported clinical outcomes that could be compared to any standard, while none of the studies compared two or more sizing systems. Overall, 60 percent of studies scored zero on the quality rating scale used--including some popular sizing systems that were "not grounded on any published data or evidence." The topic of implant selection can be an emotional one, with tension between the plastic surgeon's roles as "Artist" versus "Engineer." The researchers note that some TBP systems with the highest quality of evidence take a "middle-of-the-road" approach--based on measurements, but also considering the patient's aesthetic desires. Based on their data, Drs. Adams and McKee are evaluating a new "implant-specific" TBP system designed to guide the surgeon to a selection of manufactured implant styles and models. "Going forward," they write, "new published systems should [use] rigorous quantitative methods so that comparisons can be made in terms of patient outcomes." Plastic and Reconstructive Surgery® is published by Wolters Kluwer. Click here to read "Matching the Implant to the Breast: A Systematic Review of Implant Size Selection Systems for Breast Augmentation." Article: "Matching the Implant to the Breast: A Systematic Review of Implant Size Selection Systems for Breast Augmentation" (doi: 10.1097/PRS.0000000000002623) For more than 70 years, Plastic and Reconstructive Surgery® (http://www. ) has been the one consistently excellent reference for every specialist who uses plastic surgery techniques or works in conjunction with a plastic surgeon. The official journal of the American Society of Plastic Surgeons, Plastic and Reconstructive Surgery® brings subscribers up-to-the-minute reports on the latest techniques and follow-up for all areas of plastic and reconstructive surgery, including breast reconstruction, experimental studies, maxillofacial reconstruction, hand and microsurgery, burn repair, and cosmetic surgery, as well as news on medico-legal issues The American Society of Plastic Surgeons is the largest organization of board-certified plastic surgeons in the world. Representing more than 7,000 physician members, the Society is recognized as a leading authority and information source on cosmetic and reconstructive plastic surgery. ASPS comprises more than 94 percent of all board-certified plastic surgeons in the United States. Founded in 1931, the Society represents physicians certified by The American Board of Plastic Surgery or The Royal College of Physicians and Surgeons of Canada. Wolters Kluwer is a global leader in professional information services. Professionals in the areas of legal, business, tax, accounting, finance, audit, risk, compliance and healthcare rely on Wolters Kluwer's market leading information-enabled tools and software solutions to manage their business efficiently, deliver results to their clients, and succeed in an ever more dynamic world. Wolters Kluwer reported 2015 annual revenues of €4.2 billion. The group serves customers in over 180 countries, and employs over 19,000 people worldwide. The company is headquartered in Alphen aan den Rijn, the Netherlands. Wolters Kluwer shares are listed on Euronext Amsterdam (WKL) and are included in the AEX and Euronext 100 indices. Wolters Kluwer has a sponsored Level 1 American Depositary Receipt program. The ADRs are traded on the over-the-counter market in the U.S. (WTKWY). Wolters Kluwer Health is a leading global provider of information and point of care solutions for the healthcare industry. For more information about our products and organization, visit http://www. , follow @WKHealth or @Wolters_Kluwer on Twitter, like us on Facebook, follow us on LinkedIn, or follow WoltersKluwerComms on YouTube.


BOSTON, Feb. 27, 2017 (GLOBE NEWSWIRE) -- Exonics Therapeutics, Inc., a newly formed biotechnology company focused on developing gene editing technologies like CRISPR/Cas9 to permanently correct a majority of mutations causing Duchenne muscular dystrophy and other neuromuscular diseases, today announced a commitment of $5 million in seed financing from CureDuchenne Ventures, LLC, a subsidiary of the nonprofit CureDuchenne. The initial seed funding will allow Exonics to advance the preclinical research of its scientific founder and chief science advisor Eric Olson, PhD. Dr. Olson’s laboratory has demonstrated the ability to use adeno-associated virus (AAV) to deliver a payload based on CRISPR/Cas9 technology that can identify and correct exon mutations that prevent the production of dystrophin, a protein that helps stabilize and protect muscle fibers. Dystrophin is the key protein missing in boys with Duchenne, and published preclinical data suggest that this approach has the potential to permanently treat up to 80 percent of children suffering from the disease. Additional preclinical data is expected to be published in March 2017. “This represents the next generation of potential Duchenne muscular dystrophy therapies. By leveraging the revolutionary CRISPR/Cas9 method to permanently correct errors in the DNA sequence, it is our hope that we can develop a one-time therapy that provides lifelong benefit to Duchenne patients,” said Dr. Olson, who also serves as Professor and Chairman of the Department of Molecular Biology at the University of Texas Southwestern Medical Center (UTSW), from which Exonics’ technology is licensed. Duchenne is a rare X-linked genetic progressive muscle disease affecting nearly 15,000 boys in the U.S. and more than 300,000 boys worldwide. There is no cure for Duchenne. Children with the disease start missing development milestones at age 3 and often lose their ability to walk by age 12. All Duchenne patients will suffer from reduced mobility and independence, and ultimately respiratory or cardiac failure results in a reduced life expectancy in the mid-20s. “We are building on a scientific program of exceptional quality from Dr. Olson’s laboratory with the support of our funder CureDuchenne Ventures, an organization with deep understanding of this disease, therapeutic landscape and large unmet need. This funding underscores their recognition of the potential of Exonics’ gene editing method to develop a novel therapy for Duchenne,” said Cristina Csimma, PharmD, MHP, Executive Chair of the Board of Directors of Exonics, former President and Chief Executive Officer of Cydan, and long-time drug development leader. “Exonics is extremely well-positioned to advance our preclinical program as we follow a rigorous approach to enable translation to clinical trials.” “We look forward to working closely with the Duchenne community as we aggressively advance gene editing technology to address the significant unmet need for a curative therapy that would dramatically improve the lives of patients with Duchenne and their families,” said Jak Knowles, MD, President and Interim Chief Executive Officer of Exonics. “We are honored to advance the groundbreaking work of Dr. Olson’s laboratory and are eager to translate this approach into an important therapy for the Duchenne community.” Dr. Knowles will continue to serve the Duchenne community as Managing Director of CureDuchenne Ventures, and VP of Medical and Scientific Affairs at CureDuchenne. “We are delighted to support Dr. Olson and the Exonics team as they advance novel gene editing technology toward a potential cure for Duchenne. CureDuchenne Ventures’ unique model enables us to share our significant scientific resources, expertise and deep connections with the Duchenne community to accelerate scientific breakthroughs, such as Exonics’ program,” said Debra Miller, President of CureDuchenne Ventures and Founder and Chief Executive Officer of CureDuchenne.                                                                                       About Exonics Therapeutics Exonics Therapeutics is advancing gene editing technologies like CRISPR/Cas9 to permanently correct the majority of Duchenne muscular dystrophy mutations. Preclinical data suggest Exonics’ novel gene editing approach has the potential to permanently treat up to 80 percent of children who suffer from Duchenne. Exonics’ technology is licensed from the University of Texas Southwestern Medical Center and is based on the research of its scientific founder and chief science advisor, Eric Olson, PhD. The Company’s corporate office is located in Boston, Mass., and research activities are being conducted in Dallas, Tex. For more information, please visit www.exonicstx.com. CureDuchenne Ventures LLC collaborates with pharmaceutical and biotechnology companies to facilitate the development of drugs to treat Duchenne muscular dystrophy. CureDuchenne Ventures LLC was formed by CureDuchenne, a national nonprofit that has funded nine research projects that have advanced to human clinical trials.


News Article | January 13, 2016
Site: www.nature.com

Three teams of researchers have used CRISPR–Cas9 gene editing to treat mice that have the most common and severe form of muscular dystrophy. Duchenne muscular dystrophy is a fatal disease caused by mutations that disable the gene encoding dystrophin, an important muscle protein. Teams led by Charles Gersbach of Duke University in Durham, North Carolina; Amy Wagers of Harvard University in Cambridge, Massachusetts; and Eric Olson of the University of Texas Southwestern Medical Center in Dallas used the CRISPR–Cas9 gene-editing technique to repair the dystrophin gene in mice that have such mutations. The three teams used viruses to shuttle the components of the CRISPR–Cas9 system into the muscle cells of infant and adult mice. Treated mice made functional dystrophin and showed improvements in cardiac and skeletal muscle function.


BOSTON, Feb. 27, 2017 (GLOBE NEWSWIRE) -- Exonics Therapeutics, Inc., a newly formed biotechnology company focused on developing gene editing technologies like CRISPR/Cas9 to permanently correct a majority of mutations causing Duchenne muscular dystrophy and other neuromuscular diseases, today announced a commitment of $5 million in seed financing from CureDuchenne Ventures, LLC, a subsidiary of the nonprofit CureDuchenne. The initial seed funding will allow Exonics to advance the preclinical research of its scientific founder and chief science advisor Eric Olson, PhD. Dr. Olson’s laboratory has demonstrated the ability to use adeno-associated virus (AAV) to deliver a payload based on CRISPR/Cas9 technology that can identify and correct exon mutations that prevent the production of dystrophin, a protein that helps stabilize and protect muscle fibers. Dystrophin is the key protein missing in boys with Duchenne, and published preclinical data suggest that this approach has the potential to permanently treat up to 80 percent of children suffering from the disease. Additional preclinical data is expected to be published in March 2017. “This represents the next generation of potential Duchenne muscular dystrophy therapies. By leveraging the revolutionary CRISPR/Cas9 method to permanently correct errors in the DNA sequence, it is our hope that we can develop a one-time therapy that provides lifelong benefit to Duchenne patients,” said Dr. Olson, who also serves as Professor and Chairman of the Department of Molecular Biology at the University of Texas Southwestern Medical Center (UTSW), from which Exonics’ technology is licensed. Duchenne is a rare X-linked genetic progressive muscle disease affecting nearly 15,000 boys in the U.S. and more than 300,000 boys worldwide. There is no cure for Duchenne. Children with the disease start missing development milestones at age 3 and often lose their ability to walk by age 12. All Duchenne patients will suffer from reduced mobility and independence, and ultimately respiratory or cardiac failure results in a reduced life expectancy in the mid-20s. “We are building on a scientific program of exceptional quality from Dr. Olson’s laboratory with the support of our funder CureDuchenne Ventures, an organization with deep understanding of this disease, therapeutic landscape and large unmet need. This funding underscores their recognition of the potential of Exonics’ gene editing method to develop a novel therapy for Duchenne,” said Cristina Csimma, PharmD, MHP, Executive Chair of the Board of Directors of Exonics, former President and Chief Executive Officer of Cydan, and long-time drug development leader. “Exonics is extremely well-positioned to advance our preclinical program as we follow a rigorous approach to enable translation to clinical trials.” “We look forward to working closely with the Duchenne community as we aggressively advance gene editing technology to address the significant unmet need for a curative therapy that would dramatically improve the lives of patients with Duchenne and their families,” said Jak Knowles, MD, President and Interim Chief Executive Officer of Exonics. “We are honored to advance the groundbreaking work of Dr. Olson’s laboratory and are eager to translate this approach into an important therapy for the Duchenne community.” Dr. Knowles will continue to serve the Duchenne community as Managing Director of CureDuchenne Ventures, and VP of Medical and Scientific Affairs at CureDuchenne. “We are delighted to support Dr. Olson and the Exonics team as they advance novel gene editing technology toward a potential cure for Duchenne. CureDuchenne Ventures’ unique model enables us to share our significant scientific resources, expertise and deep connections with the Duchenne community to accelerate scientific breakthroughs, such as Exonics’ program,” said Debra Miller, President of CureDuchenne Ventures and Founder and Chief Executive Officer of CureDuchenne.                                                                                       About Exonics Therapeutics Exonics Therapeutics is advancing gene editing technologies like CRISPR/Cas9 to permanently correct the majority of Duchenne muscular dystrophy mutations. Preclinical data suggest Exonics’ novel gene editing approach has the potential to permanently treat up to 80 percent of children who suffer from Duchenne. Exonics’ technology is licensed from the University of Texas Southwestern Medical Center and is based on the research of its scientific founder and chief science advisor, Eric Olson, PhD. The Company’s corporate office is located in Boston, Mass., and research activities are being conducted in Dallas, Tex. For more information, please visit www.exonicstx.com. CureDuchenne Ventures LLC collaborates with pharmaceutical and biotechnology companies to facilitate the development of drugs to treat Duchenne muscular dystrophy. CureDuchenne Ventures LLC was formed by CureDuchenne, a national nonprofit that has funded nine research projects that have advanced to human clinical trials.


News Article | October 7, 2016
Site: news.yahoo.com

The number of PSA tests ordered in the U.S. to screen men for prostate cancer hasn't changed in recent years, despite new guidelines that say men shouldn't get this test, a new study finds. Researchers analyzed information from more than 275,000 men who visited the University of Texas Southwestern Medical Center between 2010 and 2015. The researchers were looking to see whether the number of PSA tests ordered changed after 2012, the year when the U.S. Preventive Services Task Force (an expert panel that advises the federal government) recommended that men not undergo routine screening for prostate cancer with the PSA test, no matter their age. The test, which is a blood test, was not reliable enough at detecting prostate cancer in men who had it, and also gave too many false positive results in men who didn't have it, the task force had concluded. In the study, the researchers examined patients' electronic medical records and found that there were more than 63,000 PSA tests ordered during the study period. They found that the number of tests ordered yearly was similar before and after the announcement of the new guidelines in 2012. Some previous studies have suggested that the 2012 guidelines caused a big change in prostate cancer screening, but these studies were based on surveys, which might not be as accurate as electronic medical records, the researchers said. [5 Myths About the Male Body] "We used actual, real-world data," study co-author Dr. Yair Lotan, a professor of urology at UT Southwestern, said in a statement. The PSA test, which stands for prostate-specific antigen test, looks for levels of a protein called PSA in the blood. Abnormally high levels of PSA can mean that a man has prostate cancer, but not always. The task force recommended against using the PSA test because the panel said that the harms of the test do not outweigh its potential benefits. Potential harms of the test include false-positives, which can lead to anxiety and unnecessary follow-up tests, and "overtreatment," which occurs when doctors treat a cancer that would not have caused problems in a patient's lifetime. This can happen with prostate cancer because, in many cases, the cancer doesn't grow, or it grows so slowly that it will never cause problems, the task force said. Side effects of prostate cancer treatment include erectile dysfunction, urinary incontinence and problems with bowel control. Not all organizations recommended against PSA testing. The American Cancer Society recommends that men have a discussion with their doctor about whether to start PSA screening at age 50 if they are at average risk for prostate cancer, and at age 40 to 45 if they have a family history of prostate cancer. Most cases of prostate cancer occur after age 60. Just this week, actor Ben Stiller revealed that he was diagnosed with prostate cancer 2 years ago at  age 48, after getting a PSA test. He first started getting the PSA test at age 46 at the recommendation of his doctor, even though he did not have a family history of prostate cancer, Stiller said. The new study was published online Sept. 22 in the journal Cancer.


BOSTON, Feb. 27, 2017 (GLOBE NEWSWIRE) -- Exonics Therapeutics, Inc., a newly formed biotechnology company focused on developing gene editing technologies like CRISPR/Cas9 to permanently correct a majority of mutations causing Duchenne muscular dystrophy and other neuromuscular diseases, today announced a commitment of $5 million in seed financing from CureDuchenne Ventures, LLC, a subsidiary of the nonprofit CureDuchenne. The initial seed funding will allow Exonics to advance the preclinical research of its scientific founder and chief science advisor Eric Olson, PhD. Dr. Olson’s laboratory has demonstrated the ability to use adeno-associated virus (AAV) to deliver a payload based on CRISPR/Cas9 technology that can identify and correct exon mutations that prevent the production of dystrophin, a protein that helps stabilize and protect muscle fibers. Dystrophin is the key protein missing in boys with Duchenne, and published preclinical data suggest that this approach has the potential to permanently treat up to 80 percent of children suffering from the disease. Additional preclinical data is expected to be published in March 2017. “This represents the next generation of potential Duchenne muscular dystrophy therapies. By leveraging the revolutionary CRISPR/Cas9 method to permanently correct errors in the DNA sequence, it is our hope that we can develop a one-time therapy that provides lifelong benefit to Duchenne patients,” said Dr. Olson, who also serves as Professor and Chairman of the Department of Molecular Biology at the University of Texas Southwestern Medical Center (UTSW), from which Exonics’ technology is licensed. Duchenne is a rare X-linked genetic progressive muscle disease affecting nearly 15,000 boys in the U.S. and more than 300,000 boys worldwide. There is no cure for Duchenne. Children with the disease start missing development milestones at age 3 and often lose their ability to walk by age 12. All Duchenne patients will suffer from reduced mobility and independence, and ultimately respiratory or cardiac failure results in a reduced life expectancy in the mid-20s. “We are building on a scientific program of exceptional quality from Dr. Olson’s laboratory with the support of our funder CureDuchenne Ventures, an organization with deep understanding of this disease, therapeutic landscape and large unmet need. This funding underscores their recognition of the potential of Exonics’ gene editing method to develop a novel therapy for Duchenne,” said Cristina Csimma, PharmD, MHP, Executive Chair of the Board of Directors of Exonics, former President and Chief Executive Officer of Cydan, and long-time drug development leader. “Exonics is extremely well-positioned to advance our preclinical program as we follow a rigorous approach to enable translation to clinical trials.” “We look forward to working closely with the Duchenne community as we aggressively advance gene editing technology to address the significant unmet need for a curative therapy that would dramatically improve the lives of patients with Duchenne and their families,” said Jak Knowles, MD, President and Interim Chief Executive Officer of Exonics. “We are honored to advance the groundbreaking work of Dr. Olson’s laboratory and are eager to translate this approach into an important therapy for the Duchenne community.” Dr. Knowles will continue to serve the Duchenne community as Managing Director of CureDuchenne Ventures, and VP of Medical and Scientific Affairs at CureDuchenne. “We are delighted to support Dr. Olson and the Exonics team as they advance novel gene editing technology toward a potential cure for Duchenne. CureDuchenne Ventures’ unique model enables us to share our significant scientific resources, expertise and deep connections with the Duchenne community to accelerate scientific breakthroughs, such as Exonics’ program,” said Debra Miller, President of CureDuchenne Ventures and Founder and Chief Executive Officer of CureDuchenne.                                                                                       About Exonics Therapeutics Exonics Therapeutics is advancing gene editing technologies like CRISPR/Cas9 to permanently correct the majority of Duchenne muscular dystrophy mutations. Preclinical data suggest Exonics’ novel gene editing approach has the potential to permanently treat up to 80 percent of children who suffer from Duchenne. Exonics’ technology is licensed from the University of Texas Southwestern Medical Center and is based on the research of its scientific founder and chief science advisor, Eric Olson, PhD. The Company’s corporate office is located in Boston, Mass., and research activities are being conducted in Dallas, Tex. For more information, please visit www.exonicstx.com. CureDuchenne Ventures LLC collaborates with pharmaceutical and biotechnology companies to facilitate the development of drugs to treat Duchenne muscular dystrophy. CureDuchenne Ventures LLC was formed by CureDuchenne, a national nonprofit that has funded nine research projects that have advanced to human clinical trials.


NEWPORT BEACH, Calif.--(BUSINESS WIRE)--CureDuchenne Ventures, a subsidiary of the nonprofit CureDuchenne that funds research to find a cure for Duchenne muscular dystrophy, announced today that they have committed $5 million in seed financing in Exonics Therapeutics, a new biotechnology company focused on utilizing gene editing technologies like CRISPR/Cas9 to advance the development of a treatment for Duchenne muscular dystrophy. Exonics’ CRISPR/Cas9 technology is a potential one-time treatment that would make a permanent correction of the mutation that causes Duchenne. Exonics Therapeutics will use proceeds from the seed financing to advance the preclinical research of its scientific founder and chief science advisor Eric Olson, PhD, to a clinic-ready therapy. Dr. Olson’s laboratory has demonstrated the ability to use adeno-associated virus (AAV) to deliver a payload based on CRISPR/Cas9 technology that can identify and correct exon mutations that prevent the production of dystrophin, a protein that helps stabilize and protect muscle fibers. Dystrophin is the key protein missing in boys with Duchenne, and published preclinical data suggest that this approach has the potential to permanently treat up to 80 percent of children suffering from Duchenne. Watch this video for more information about the research being conducted in the Olson lab, on which Exonics’ technology is based. “The Duchenne community needs novel and diverse approaches to treat and cure this devastating disease,” said Debra Miller, Founder and CEO of CureDuchenne. “Exonics’ CRISPR/Cas9 technology has the potential to dramatically improve the lives of those who live with Duchenne. We are hopeful that a one-time treatment with gene editing therapy could provide a lifelong benefit to Duchenne patients.” “We are delighted to provide seed funding for Exonics and over the past year, have worked closely with Dr. Olson and his team, to create a company which is well positioned to advance promising gene editing treatments for Duchenne,” said Dr. Jak Knowles, President and Interim Chief Executive Officer of Exonics, and Managing Director of CureDuchenne Ventures. “This technology could be a breakthrough for the Duchenne community because of the ability to use a patient’s own DNA to fix the mutated gene.” “The creation of Exonics, in conjunction with CureDuchenne Ventures, will allow us to work toward a one-time therapy that would make a big impact on the lives of Duchenne patients,” said Dr. Olson, who also serves as Professor and Chairman of the Department of Molecular Biology at the University of Texas Southwestern Medical Center, from which Exonics’ technology is licensed. “CureDuchenne has strong relationships with the Duchenne community including patients, families, physicians and scientists in academia and industry. These relationships, CureDuchenne’s deep knowledge of Duchenne and all the therapeutic approaches for the disease; and their strong leadership team make them a great partner to help accelerate drug development.” To donate to the CRISPR/Cas9 gene editing research, click here. Exonics Therapeutics is advancing gene editing technologies like CRISPR/Cas9 to permanently correct the majority of Duchenne muscular dystrophy mutations. Preclinical data suggest Exonics’ novel gene editing approach has the potential to permanently treat up to 80 percent of children who suffer from Duchenne. Exonics’ technology is licensed from the University of Texas Southwestern Medical Center (UTSW) and is based on the research of its scientific founder and chief science advisor, Eric Olson, PhD. The Company’s corporate office is located in Boston, Mass., and research activities are being conducted in Dallas, Tex. For more information, please visit www.exonicstx.com. CureDuchenne Ventures LLC collaborates with pharmaceutical and biotechnology companies to facilitate the development of drugs to treat Duchenne muscular dystrophy. CureDuchenne Ventures LLC was formed by CureDuchenne, a national nonprofit that has funded nine research projects that have advanced to human clinical trials. CureDuchenne has leveraged more than $1.3 billion in follow-on investment from pharmaceutical companies and biotech to fund research. CureDuchenne has been working to treat the whole disease with a multi-pronged approach to find treatments for the many effects that Duchenne has on the body. CureDuchenne was founded in 2003 with a focus on saving the lives of those with Duchenne muscular dystrophy, a disease that affects more than 300,000 boys worldwide. With support from CureDuchenne, nine research projects have advanced to human clinical trials. CureDuchenne also is the innovator bringing physical therapy and standard of care to local communities around the country through CureDuchenne Cares. For more information, please visit CureDuchenne.org and follow us on Facebook, Twitter, Instagram and YouTube.


Joset A.,University of Zürich | Dodd D.A.,University of Texas Southwestern Medical Center | Halegoua S.,State University of New York at Stony Brook | Schwab M.E.,University of Zürich
Journal of Cell Biology | Year: 2010

Nogo-A is one of the most potent myelin-associated inhibitors for axonal growth, regeneration, and plasticity in the adult central nervous system. The Nogo-A - specific fragment NogoΔ20 induces growth cone collapse, and inhibits neurite outgrowth and cell spreading by activating RhoA. Here, we show that NogoΔ20 is internalized into neuronal cells by a Pincher- and rac-dependent, but clathrin- and dynamin-independent, mechanism. Pincher-mediated macroendocytosis results in the formation of NogoΔ20-containing signalosomes that direct RhoA activation and growth cone collapse. In compartmentalized chamber cultures, NogoΔ20 is endocytosed into neurites and retrogradely transported to the cell bodies of dorsal root ganglion neurons, triggering RhoA activation en route and decreasing phosphorylated cAMP response element binding levels in cell bodies. Thus, Pincher-dependent macroendocytosis leads to the formation of Nogo-A signaling endosomes, which act both within growth cones and after retrograde transport in the cell body to negatively regulate the neuronal growth program. © 2010 Joset et al.


Unger R.H.,University of Texas Southwestern Medical Center | Unger R.H.,Veterans Affairs North Texas Health Care System | Orci L.,University of Geneva
Proceedings of the National Academy of Sciences of the United States of America | Year: 2010

New results have brought to light the importance of the regulation of glucagon by β-cells in the development of diabetes. In this perspective, we examine the normal paracrinology of α- and β-cells in nondiabetic pancreatic islets. We propose a Sherringtonian model of coordinated reciprocal secretory responses of these juxtaposed cells that secrete glucagon and insulin, hormones with opposing actions on the liver. As insulin is a powerful inhibitor of glucagon, we propose that within-islet inhibition of α-cells by β-cells creates an insulin-to-glucagon ratio that maintains glycemic stability even in extremes of glucose influx or efflux. By contrast, in type 1 diabetes mellitus, α-cells lack constant action of high insulin levels from juxtaposed β-cells. Replacement with exogenous insulin does not approach paracrine levels of secreted insulin except with high doses that "overinsulinize" the peripheral insulin targets, thereby promoting glycemic volatility. Based on the stable normoglycemia of mice with type 1 diabetes during suppression of glucagon with leptin, we conclude that, in the absence of paracrine regulation of α-cells, tonic inhibition of α-cells improves the dysregulated glucose homeostasis. These results have considerable medical implications, as they suggest new approaches to normalize the extreme volatility of glycemia in diabetic patients.


Wang H.,China National Institute of Biological Sciences | Sun L.,China National Institute of Biological Sciences | Su L.,University of Texas Southwestern Medical Center | Rizo J.,University of Texas Southwestern Medical Center | And 4 more authors.
Molecular Cell | Year: 2014

Programmed necrotic cell death induced by the tumor necrosis factor alpha (TNF-α) family of cytokines is dependent on a kinase cascade consisting of receptor-interacting kinases RIP1 and RIP3. How these kinase activities cause cells to die by necrosis is not known. The mixed lineage kinase domain-like protein MLKL is a functional RIP3 substrate that binds to RIP3 through its kinase-like domain but lacks kinase activity of its own. RIP3 phosphorylates MLKL at the T357 and S358 sites. Reported here is the development of a monoclonal antibody that specifically recognizes phosphorylated MLKL in cells dying of this pathway and in human liver biopsy samples from patients suffering from drug-induced liver injury. The phosphorylated MLKL forms an oligomer that binds to phosphatidylinositol lipids and cardiolipin. This property allows MLKL to move from the cytosol to the plasma and intracellular membranes, where it directly disrupts membrane integrity, resulting in necrotic death. © 2014 Elsevier Inc.


Burd C.E.,University of North Carolina at Chapel Hill | Sorrentino J.A.,University of North Carolina at Chapel Hill | Clark K.S.,University of North Carolina at Chapel Hill | Darr D.B.,University of North Carolina at Chapel Hill | And 6 more authors.
Cell | Year: 2013

Monitoring cancer and aging in vivo remains experimentally challenging. Here, we describe a luciferase knockin mouse (p16LUC), which faithfully reports expression of p16INK4a, a tumor suppressor and aging biomarker. Lifelong assessment of luminescence in p16+/LUC mice revealed an exponential increase with aging, which was highly variable in a cohort of contemporaneously housed, syngeneic mice. Expression of p16 INK4a with aging did not predict cancer development, suggesting that the accumulation of senescent cells is not a principal determinant of cancer-related death. In 14 of 14 tested tumor models, expression of p16 LUC was focally activated by early neoplastic events, enabling visualization of tumors with sensitivity exceeding other imaging modalities. Activation of p16INK4a was noted in the emerging neoplasm and surrounding stromal cells. This work suggests that p16INK4a activation is a characteristic of all emerging cancers, making the p16 LUC allele a sensitive, unbiased reporter of neoplastic transformation. © 2013 Elsevier Inc.


Wu S.-Y.,University of Texas Southwestern Medical Center | Lee A.-Y.,University of Texas Southwestern Medical Center | Lee A.-Y.,Daewoong Bio Co. | Lai H.-T.,University of Texas Southwestern Medical Center | And 2 more authors.
Molecular Cell | Year: 2013

Bromodomain-containing protein 4 (Brd4) is an epigenetic reader and transcriptional regulator recently identified as a cancer therapeutic target for acute myeloid leukemia, multiple myeloma, and Burkitt's lymphoma. Although chromatin targeting is a crucial function of Brd4, there is little understanding of how bromodomains that bind acetylated histones are regulated, nor how the gene-specific activity of Brd4 is determined. Via interaction screen and domain mapping, we identified p53 as a functional partner of Brd4. Interestingly, Brd4 association with p53 is modulated by casein kinase II (CK2)-mediated phosphorylation of a conserved acidic region in Brd4 that selectively contacts either a juxtaposed bromodomain or an adjacent basic region to dictate the ability of Brd4 binding to chromatin and also the recruitment of p53 to regulated promoters. The unmasking of bromodomains and activator recruitment, concurrently triggered by the CK2 phospho switch, provide an intriguing mechanism for gene-specific targeting by a universal epigenetic reader. © 2013 Elsevier Inc.


Hedayati S.S.,Veterans Affairs North Texas Health Care System | Hedayati S.S.,University of Texas Southwestern Medical Center | Yalamanchili V.,University of Texas Southwestern Medical Center | Finkelstein F.O.,Yale University
Kidney International | Year: 2012

Depression is a common, under-recognized, and under-treated problem that is independently associated with increased morbidity and mortality in CKD patients. However, only a minority of CKD patients with depression are treated with antidepressant medications or nonpharmacologic therapy. Reasons for low treatment rates include a lack of properly controlled trials that support or refute efficacy and safety of various treatment regimens in CKD patients. The aim of this manuscript is to provide a comprehensive review of studies exploring depression treatment options in CKD. Observational studies as well as small trials suggest that certain serotonin-selective reuptake inhibitors may be safe to use in patients with advanced CKD and ESRD. These studies were limited by small sample sizes, lack of placebo control, and lack of formal assessment for depression diagnosis. Nonpharmacologic treatments were explored in selected ESRD samples. The most promising data were reported for frequent hemodialysis and cognitive behavioral therapy. Alternative proposed therapies include exercise training regimens, treatment of anxiety, and music therapy. Given the association of depression with cardiovascular events and mortality, and the excessive rates of cardiovascular death in CKD, it becomes imperative to not only investigate whether treatment of depression is efficacious, but also whether it would result in a reduction in morbidity and mortality in this patient population. © 2012 International Society of Nephrology.


Coppari R.,University of Texas Southwestern Medical Center | Coppari R.,University of Geneva | Coppari R.,University of California at Irvine | Bjorbaek C.,Beth Israel Deaconess Medical Center
Nature Reviews Drug Discovery | Year: 2012

Since the discovery of leptin in 1994, we now have a better understanding of the cellular and molecular mechanisms underlying its biological effects. In addition to its established anti-obesity effects, leptin exerts antidiabetic actions that are independent of its regulation of body weight and food intake. In particular, leptin can correct diabetes in animal models of type 1 and type 2 diabetes. In addition, long-term leptin replacement therapy improves glycaemic control, insulin sensitivity and plasma triglycerides in patients with severe insulin resistance due to lipodystrophy. These results have spurred enthusiasm for the use of leptin therapy to treat diabetes. Here, we review the current understanding of the glucoregulatory functions of leptin, emphasizing its central mechanisms of action and lessons learned from clinical studies, and discuss possible therapeutic applications of leptin in the treatment of type 1 and type 2 diabetes. © 2012 Macmillan Publishers Limited. All rights reserved.


Fujikawa T.,University of Texas Southwestern Medical Center | Coppari R.,University of Geneva
Aging | Year: 2014

Diabetes afflicts hundreds of millions worldwide. People affected by type 1 diabetes mellitus (T1DM; the insulindeficient form of diabetes) or type 2 diabetes mellitus (T2DM; the insulin-resistant form of diabetes) have significantly reduced life expectancy compared to normal individuals. This is due in part to the fact that (despite improvements) current anti-diabetic approaches are suboptimal. Indeed, severe morbidities (e.g.: cardiovascular disease, hypertension) are still too often associated with diabetes. Recent preclinical results indicate that different types of hypothalamic neurons are endowed with the ability to mediate the hyperglycemia-lowering action of the adipocyte-derived hormone leptin in an insulin-dependent and insulin-independent fashion. These results may pave the way for better anti-diabetic approaches and therefore positively impact on life expectancy of diabetic subjects. © Fujikawa and Coppari.


Jain N.,University of Texas Southwestern Medical Center | Reilly R.F.,Veterans Affairs North Texas Health Care System
Nature Reviews Nephrology | Year: 2014

Traditional strategies for management of patients with chronic kidney disease (CKD) have not resulted in any change in the growing prevalence of CKD worldwide. A historic belief that eating healthily might ameliorate kidney disease still holds credibility in the 21 st century. Dietary sodium restriction to <2.3 g daily, a diet rich in fruits and vegetables and increased water consumption corresponding to a urine output of 3-4 l daily might slow the progression of early CKD, polycystic kidney disease or recurrent kidney stones. Current evidence suggests that a reduction in dietary net acid load could be beneficial in patients with CKD, but the supremacy of any particular diet has yet to be established. More trials of dietary interventions are needed, especially in diabetic nephropathy, before evidence-based recommendations can be made. In the meantime, nephrologists should discuss healthy dietary habits with their patients and provide individualized care aimed at maximizing the potential benefits of dietary intervention, reducing the incidence of CKD and delaying its progression to end-stage renal disease. Keeping in mind the lack of data on hard outcomes, dietary recommendations should take into account barriers to adherence and be tailored to different cultures, ethnicities and geographical locations. © 2014 Macmillan Publishers Limited. All rights reserved.


Berger J.R.,University of Texas Southwestern Medical Center | Susan Hedayati S.,University of Texas Southwestern Medical Center | Susan Hedayati S.,Veterans Affairs North Texas Health Care System
Clinical Journal of the American Society of Nephrology | Year: 2012

ESRD has become an important problem for elderly patients. The segment of the ESRD population age 65 years or older has grown considerably, and this growth is expected to accelerate in coming years. Nephrologists caring for the elderly with advanced kidney disease will encounter patients with comorbid conditions common in younger patients, as well as physical, psychological, and social challenges that occur with increased frequency in the aging population. These challenging factors must be addressed to help inform decisions regarding the option to initiate dialysis, the choice of dialysismodality,whether to pursue kidney transplantation, and end-of-life care. This article will highlight some common problems encountered by elderly patients with ESRD and review data on the clinical outcomes of elderly patients treatedwith differentmodalities of dialysis, outcomes of kidney transplantation in the elderly, and nondialytic management of CKD stage 5. © 2012 by the American Society of Nephrology.


Al Mutair A.N.,King Saud bin Abdulaziz University for Health Sciences | Nasrat G.H.,King Saud bin Abdulaziz University for Health Sciences | Russell D.W.,University of Texas Southwestern Medical Center
Journal of Clinical Endocrinology and Metabolism | Year: 2012

Context: Inherited forms of vitamin D deficiency are rare causes of rickets and to date have been traced to mutations in three genes, VDR, encoding the 1α,25-dihydroxyvitamin D receptor, CYP27B1, encoding the vitamin D 1α-hydroxylase, and CYP2R1, encoding a microsomal vitamin D 25-hydroxylase. Results: Multiple mutations have been identified in VDR and CYP27B1 in patients with rickets, and thus, the roles of these two genes in vitamin D metabolism are unassailable. The case is less clear for CYP2R1, in which only a single mutation, L99P in exon 2 of the gene, has been identified in Nigerian families, and because multiple enzymes with vitamin D 25-hydroxylase activity have been identified. Here we report molecular genetic studies on two siblings from a Saudi family who presented with classic symptoms of vitamin D deficiency. The affected offspring inherited two different CYP2R1 mutations (367+1, G→A; 768, iT), which are predicted to specify null alleles. Conclusion: We conclude that CYP2R1 is a major vitaminD25-hydroxylase that plays a fundamental role in activation of this essential vitamin. Copyright © 2012 by The Endocrine Society.


Frank A.,University of Texas Southwestern Medical Center | Brown L.M.,North Carolina A&T State University | Clegg D.J.,University of Texas Southwestern Medical Center
Frontiers in Neuroendocrinology | Year: 2014

Estrogens regulate key features of metabolism, including food intake, body weight, energy expenditure, insulin sensitivity, leptin sensitivity, and body fat distribution. There are two 'classical' estrogen receptors (ERs): estrogen receptor alpha (ERS1) and estrogen receptor beta (ERS2). Human and murine data indicate ERS1 contributes to metabolic regulation more so than ESR2. For example, there are human inactivating mutations of ERS1 which recapitulate aspects of the metabolic syndrome in both men and women. Much of our understanding of the metabolic roles of ERS1 was initially uncovered in estrogen receptor α-null mice (ERS1-/-); these mice display aspects of the metabolic syndrome, including increased body weight, increased visceral fat deposition and dysregulated glucose intolerance. Recent data further implicate ERS1 in specific tissues and neuronal populations as being critical for regulating food intake, energy expenditure, body fat distribution and adipose tissue function. This review will focus predominantly on the role of hypothalamic ERs and their critical role in regulating all aspects of energy homeostasis and metabolism. © 2014 Elsevier Inc.


Kim T.-K.,University of Texas Southwestern Medical Center | Hemberg M.,Boston Childrens Hospital | Gray J.M.,Harvard University
Cold Spring Harbor Perspectives in Biology | Year: 2015

Recent studies have revealed that active enhancers are transcribed, producing a class of noncoding RNAs called enhancer RNAs (eRNAs). eRNAs are distinct from long noncoding RNAs (lncRNAs), but these two species of noncoding RNAs may share a similar role in the activation of mRNA transcription. Emerging studies, showing that eRNAs function in controlling mRNA transcription, challenge the idea that enhancers are merely sites of transcription factor assembly. Instead, communicationbetween promoters andenhancers canbebidirectional with promoters required to activate enhancer transcription. Reciprocally, eRNAs may then facilitate enhancer–promoter interaction or activate promoter-driven transcription. © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.


Inzucchi S.E.,Yale University | Lipska K.J.,Yale University | Mayo H.,University of Texas Southwestern Medical Center | Bailey C.J.,Aston University | McGuire D.K.,University of Texas Southwestern Medical Center
JAMA - Journal of the American Medical Association | Year: 2014

IMPORTANCE: Metformin is widely viewed as the best initial pharmacological option to lower glucose concentrations in patients with type 2 diabetes mellitus. However, the drug is contraindicated in many individuals with impaired kidney function because of concerns of lactic acidosis.OBJECTIVE: To assess the risk of lactic acidosis associated with metformin use in individuals with impaired kidney function.EVIDENCE ACQUISITION: In July 2014, we searched the MEDLINE and Cochrane databases for English-language articles pertaining tometformin, kidney disease, and lactic acidosis in humans between 1950 and June 2014.We excluded reviews, letters, editorials, case reports, small case series, and manuscripts that did not directly pertain to the topic area or that met other exclusion criteria. Of an original 818 articles, 65 were included in this review, including pharmacokinetic/metabolic studies, large case series, retrospective studies, meta-analyses, and a clinical trial.RESULTS: Although metformin is renally cleared, drug levels generally remain within the therapeutic range and lactate concentrations are not substantially increased when used in patients with mild to moderate chronic kidney disease (estimated glomerular filtration rates, 30-60 mL/min per 1.73m2). The overall incidence of lactic acidosis in metformin users varies across studies from approximately 3 per 100 000 person-years to 10 per 100 000 person-years and is generally indistinguishable from the background rate in the overall population with diabetes. Data suggesting an increased risk of lactic acidosis in metformin-treated patients with chronic kidney disease are limited, and no randomized controlled trials have been conducted to test the safety ofmetformin in patients with significantly impaired kidney function. Population-based studies demonstrate that metformin may be prescribed counter to prevailing guidelines suggesting a renal risk in up to 1 in 4 patients with type 2 diabetes mellitus-use which, in most reports, has not been associated with increased rates of lactic acidosis. Observational studies suggest a potential benefit from metformin on macrovascular outcomes, even in patients with prevalent renal contraindications for its use.CONCLUSIONS AND RELEVANCE: Available evidence supports cautious expansion of metformin use in patients with mild to moderate chronic kidney disease, as defined by estimated glomerular filtration rate, with appropriate dosage reductions and careful follow-up of kidney function.


Antonarakis S.E.,University of Geneva | Chakravarti A.,Johns Hopkins University | Cohen J.C.,University of Texas Southwestern Medical Center | Hardy J.,University College London
Nature Reviews Genetics | Year: 2010

For the past century, Mendelian and multifactorial traits have existed at opposite ends of the disease spectrum in humans. Furthermore, the recent emphasis on genome-wide association studies for uncovering variants that underlie common diseases has risked deepening the divide ĝ€" or has it? Four experienced human geneticists express their views on the changing landscape of human disease studies and the impact of new technologies and study designs on the age-old aim of connecting a genomic variant with its phenotypic consequences. © 2010 Macmillan Publishers Limited. All rights reserved.


Kumbhani D.J.,University of Texas Southwestern Medical Center | Bavry A.A.,University of Florida | Desai M.Y.,Cleveland Clinic | Bangalore S.,New York University | Bhatt D.L.,Harvard University
Journal of the American College of Cardiology | Year: 2013

Objectives This meta-analysis was designed to update data on clinical outcomes with aspiration thrombectomy or mechanical thrombectomy before primary percutaneous coronary intervention (PCI) compared with conventional primary PCI alone. Background The clinical efficacy of thrombectomy in acute myocardial infarction (AMI) remains uncertain. Methods Clinical trials that randomized AMI patients to aspiration (18 trials, n = 3,936) or mechanical thrombectomy (7 trials, n = 1,598) before PCI compared with conventional PCI alone were included. Results The weighted mean duration of clinical follow-up was 6 months. Aspiration thrombectomy vs. conventional primary PCI (18 trials, n=3,936): Major adverse cardiac events (MACE) (risk ratio [RR]: 0.76; 95% confidence interval [CI]: 0.63 to 0.92; p = 0.006) and all-cause mortality (RR: 0.71; 95% CI: 0.51 to 0.99; p = 0.049) were significantly reduced with aspiration thrombectomy. Beneficial trends were noted for recurrent MI (p = 0.11) and target vessel revascularization (p = 0.06). Final infarct size (p = 0.64) and ejection fraction (p = 0.32) at 1 month were similar. ST-segment resolution (STR) at 60 min (RR: 1.31; 95% CI: 1.16 to 1.48; p < 0.0001) and Thrombolysis In Myocardial Infarction blush grade (TBG) 3 post-procedure (RR: 1.37; 95% CI: 1.19 to 1.59; p < 0.0001) were both improved with aspiration thrombectomy. Mechanical thrombectomy vs. conventional primary PCI (7 trials, n = 1,598): there was no difference between the mechanical thrombectomy and conventional primary PCI arms in the incidence of MACE (RR: 1.10; 95% CI: 0.59 to 2.05; p = 0.77), mortality (p = 0.57), recurrent MI (p = 0.32), target vessel revascularization (p = 0.19), or final infarct size (p = 0.47). A benefit in STR at 60 min (RR: 1.25; 95% CI: 1.06 to 1.47; p = 0.007), but not TBG 3 (RR: 1.09; 95% CI: 0.86 to 1.38; p = 0.48) was noted. Conclusions Thrombectomy during AMI by manual catheter aspiration, but not mechanically, is beneficial in reducing MACE, including mortality, at 6 to 12 months compared with conventional primary PCI alone. © 2013 by the American College of Cardiology Foundation Published by Elsevier Inc.


Grant
Agency: Department of Health and Human Services | Branch: | Program: STTR | Phase: Phase I | Award Amount: 150.00K | Year: 2013

DESCRIPTION (provided by applicant): Agave BioSystems and Professor Makoto Kuro-o at the University of Texas Southwestern Medical Center are proposing a collaborative effort to screen for novel small molecules acting as agonists or antagonists of the Klotho and Klotho- dependent endocrine Fibroblast Growth Factors. The expected outcome of this Phase I effort will be the validation of a high-throughput screening methodology and the identification of confirmed hits which modulate the activity of endocrine Fibroblast Growth Factors FGF21 and FGF23. The effect of each hit will be confirmed in FGF- specific cell-based assays verifying changes in known activities of the Fibroblast Growth Factors. The small scale proof of concept screening campaign of the Phase Iwill be followed by a larger campaign in the Phase II do identify series of novel agonists and antagonists offering to uncover a broader set of structures and mechanisms of action for this novel effectors of endocrine Fibroblast Growth Factors. Selected hits will undergo early hit-to-lead optimization and be evaluated for cytotoxicity prior t small animal testing. Rodent disease models will be treated and the expected molecular, cellular and physiological changes of endocrine FGF activity modulation will be verified. This translational research project for therapeutic target validation will lead to the development of potential new drugs against chronic kidney disease, diabetes, obesity and cancer. PUBLIC HEALTH RELEVANCE PUBLIC HEALTH RELEVANCE:Agave BioSystems and Professor Makoto Kuro-o at the University of Texas Southwestern Medical Center are proposing a collaborative effort to screen for novel small molecules acting as antagonists of the Klotho-dependent endocrine Fibroblast Growth Factor FGF23 and as agonists of the Klotho-dependent FGF21. The identification of such compounds will be used for therapeutic target validation and potentially lead to the development of novel drugs against chronic kidney disease, diabetes, obesity and aging.


News Article | March 1, 2017
Site: www.eurekalert.org

Leesburg, VA, March 1, 2017-- FDG PET/CT is a valuable imaging tool for treatment assessment of patients with lung cancer, though systematic evidence for its comparative effectiveness with conventional imaging, such as chest CT, is still evolving. Authors of the study titled "The Value of FDG PET/CT in Treatment Response Assessment, Follow-Up, and Surveillance of Lung Cancer" published their findings in the February 2017 issue of the American Journal of Roentgenology. The study is available on the ARRS website accessible here. In this review, the authors summarized the existing evidence in the literature concerning use of PET/CT for both assessing the efficacy of treatment response and performing posttreatment follow-up of lung cancer. "FDG PET/CT is most useful when there is clinical suspicion or other evidence for disease recurrence or metastases," said study coauthor Rathan M. Subramaniam, of the department of radiology, University of Texas Southwestern Medical Center, Dallas. "Using FDG PET/CT for routine surveillance without any clinical suspicion should be discouraged until its value for patient survival outcomes is fully established." The National Comprehensive Cancer Network (NCCN) recommends the use of FDG PET/CT for appropriately staging lung cancer and avoiding futile thoracotomies. It also recommends the imaging for accurate radiation therapy (RT) planning for both non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC). However, NCCN does not recommend routine use of FDG PET/CT for treatment response evaluation and follow-up in lung cancer. According to the study authors, FDG PET/CT is usually recommended to be performed 12 weeks after completion of concurrent chemoradiation therapy to minimize radiation-related inflammatory uptake leading to false-positive studies. In some cases, radiation-related therapy changes, especially with stereotactic body radiotherapy, can last for many months. In these circumstances, a follow-up FDG PET/CT in 3 months is suggested to ensure resolution of therapy-related FDG uptake. FDG PET/CT can be performed 4 weeks after completion of chemotherapy or surgery (without concurrent radiation), because the therapy-related inflammatory uptake is less and subsides within a shorter time. The sequencing, cost analysis, and comparative effectiveness of FDG PET/CT and conventional imaging modalities in the follow-up setting need to be investigated. Lung cancer remains the leading cause of cancer-related mortality worldwide, accounting for about 1.6 million deaths yearly. Despite significant advances in both diagnostic and therapeutic approaches, the overall 5-year survival rate for lung cancer is 17.4%. However, this is largely dependent on the stage of diagnosis, with a survival rate of 54.8% for localized and 4.2% for disease that has metastasized in other parts of the body. Lung cancer has historically been divided into two main types: NSCLC (85% of cases) and SCLC (10-15% of cases). A multidisciplinary approach including the use of advanced imaging techniques for early accurate staging of disease and delivery of treatment is needed to avoid futile treatments and improve overall survival, which, in turn, influence the patient's quality of life. Surgery is usually the primary treatment modality for localized disease in patients with lung cancer. Even after curative surgery, patients remain at risk for the development of recurrence or a secondary pulmonary malignancy. Posttreatment follow-up usually consists of a combination of physical examination, laboratory tests, and imaging. No single modality is simultaneously sensitive, specific, and cost effective; thus, a combined approach is needed for the detection of tumor recurrence. PET is a noninvasive imaging modality that is promising in the evaluation of lung cancer and is currently used clinically for initial antitumor treatment strategy (for staging, treatment planning, and delivery of radiation treatment) and for subsequent antitumor treatment strategy (treatment response assessment and detection of recurrence in follow-up). Founded in 1900, ARRS is the first and oldest radiology society in the United States, and is an international forum for progress in radiology. The Society's mission is to improve health through a community committed to advancing knowledge and skills in radiology. ARRS achieves its mission through an annual scientific and educational meeting, publication of the American Journal of Roentgenology (AJR) and InPractice magazine, topical symposia and webinars, and print and online educational materials. ARRS is located in Leesburg, VA.


MAPLE GROVE, Minn.--(BUSINESS WIRE)--NxThera, Inc., a medical device company pioneering the application of its convective radiofrequency water vapor thermal therapy platform technology to treat endourological conditions, today announced the in press publication of the 2-year data from the Rezūm II pivotal study in The Journal of Urology. The Rezūm II clinical study enrolled 197 patients in this randomized, controlled trial across 15 centers in the United States and documents the durable symptom reduction and preserved sexual function in patients who underwent the treatment with Rezūm. Of the 61 patients in the original control arm, 53 elected and qualified for crossover active treatment. The Rezūm System convectively and uniformly disperses the thermal energy created with RF current through the tissue interstices to disrupt the tissue cell membranes to effect tissue cell death and necrosis. The body’s immune system then simply resorbs the dead tissue cells, thereby removing the obstructive prostate tissue to successfully treat Lower Urinary Tract Symptoms (LUTS) secondary to Benign Prostatic Hyperplasia (BPH). “One of the primary factors in considering BPH therapies for our patients is durability,” said Dr. Claus Roehrborn, co-principal investigator and Professor and Chair, Division of Urology at University of Texas Southwestern Medical Center. “This important data demonstrates that BPH treatments with the Rezūm System provides clinically significant symptom relief out to 2 years while preserving sexual function. Given its strong safety profile and sustained outcomes, Rezūm warrants consideration as a first-line therapeutic alternative to medical therapy for the treatment of BPH.” “We are excited about the sustained clinical and sexual health benefits out to two years for both clinicians and their patients,” said Bob Paulson, President & CEO of NxThera. “The sustained and compelling clinical results from our Rezūm II pivotal study demonstrate a minimally invasive treatment option that can be performed in a variety of clinical settings, including as an in-office procedure. We are committed to provide urologists with world-class clinical training and patient education to support them in their efforts to improve the lives of men suffering from BPH.” About NxThera NxThera pioneered its convective radiofrequency water vapor thermal therapy platform technology to treat a variety of endourological conditions beginning with BPH. The Company is furthering the application of the technology platform through advanced research and development targeting the treatment of prostate cancer and kidney cancer. NxThera’s FDA cleared Rezūm System to treat BPH uses radiofrequency energy to create sterile water vapor, or steam, to convectively deliver targeted, precise thermal energy treatments in a simple procedure with minimal discomfort, and provide improvements in LUTS symptoms, urine flow and quality of life. Founded in 2008, NxThera is located in Maple Grove, Minnesota.


Fleischmann R.,University of Texas Southwestern Medical Center | Fleischmann R.,Metroplex Clinical Research Center
Current Opinion in Rheumatology | Year: 2012

Purpose of Review: Since the introduction of biologic therapies into the treatment paradigm of rheumatoid arthritis (RA), there has been hope that oral small molecule immune modulators would be developed that would have a risk:benefit profile at least similar to biologic therapies, be more convenient for the patient and, hopefully, be less expensive. This article reviews the progress made in the development of these compounds over the past year. Recent Finding: Additional information has become available in the past year on five oral compounds including kinase inhibitors (tofacitinib, fostamatinib, VX-509), an S1P lyase inhibitor (LX 3305) and a chemokine receptor-1 antagonist (CCX354-C). Efficacy has been shown in phase III with tofacitinib and in phase II with fostamatinib and VX-509; safety was the primary endpoint of the trials of CCX354-C and LX3305. Regarding side effects, liver test elevation and neutropenia occurred with tofacitinib, VX-509 and fostamatinib; lipid elevation with tofacitinib and VX-509; creatinine elevation and anemia with tofacitinib, and hypertension and diarrhea with fostamatinib. Summary: Compounds that inhibit tyrosine kinase pathways involved in cellular signalling have been shown to be effective in the treatment of RA with a reasonable risk:benefit ratio. It is too early to tell about inhibitors of other pathways. © 2012 Wolters Kluwer Health | Lippincott Williams & Wilkins.


Unger R.H.,University of Texas Southwestern Medical Center | Unger R.H.,VA North Texas Health Care System | Roth M.G.,University of Texas Southwestern Medical Center
Cell Metabolism | Year: 2015

A variety of leptin actions require a re-examination of classic concepts of metabolic diseases. Here we present evidence for two physiologic pathways: a pathway that protects nonadipose tissues from overaccumulation of potentially toxic lipids and unrecognized paracrine interactions between α and β cells revealed by leptin's ability to suppress diabetic hyperglucagonemia. These observations strongly point to new therapeutic possibilities for both type 1 and type 2 diabetes. © 2015 Elsevier Inc.


Lowrey P.L.,Rider University | Takahashi J.S.,University of Texas Southwestern Medical Center
Advances in Genetics | Year: 2011

The mammalian circadian system is a complex hierarchical temporal network which is organized around an ensemble of uniquely coupled cells comprising the principal circadian pacemaker in the suprachiasmatic nucleus of the hypothalamus. This central pacemaker is entrained each day by the environmental light/dark cycle and transmits synchronizing cues to cell-autonomous oscillators in tissues throughout the body. Within cells of the central pacemaker and the peripheral tissues, the underlying molecular mechanism by which oscillations in gene expression occur involves interconnected feedback loops of transcription and translation. Over the past 10 years, we have learned much regarding the genetics of this system, including how it is particularly resilient when challenged by single-gene mutations, how accessory transcriptional loops enhance the robustness of oscillations, how epigenetic mechanisms contribute to the control of circadian gene expression, and how, from coupled neuronal networks, emergent clock properties arise. Here, we will explore the genetics of the mammalian circadian system from cell-autonomous molecular oscillations, to interactions among central and peripheral oscillators and ultimately, to the daily rhythms of behavior observed in the animal. © 2011 Elsevier Inc.


Pankotai T.,French National Center for Scientific Research | Bonhomme C.,French National Center for Scientific Research | Chen D.,University of Texas Southwestern Medical Center | Soutoglou E.,French National Center for Scientific Research
Nature Structural and Molecular Biology | Year: 2012

DNA double-strand break (DSB) repair interferes with ongoing cellular processes, including replication and transcription. Although the process of replication stalling upon collision of replication forks with damaged DNA has been extensively studied, the fate of elongating RNA polymerase II (RNAPII) that encounters a DSB is not well understood. We show that the occurrence of a single DSB at a human RNAPII-transcribed gene leads to inhibition of transcription elongation and reinitiation. Upon inhibition of DNA protein kinase (DNAPK), RNAPII bypasses the break and continues transcription elongation, suggesting that it is not the break per se that inhibits the processivity of RNAPII, but the activity of DNAPK. We also show that the mechanism of DNAPK-mediated transcription inhibition involves the proteasome-dependent pathway. The results point to the pivotal role of DNAPK activity in the eviction of RNAPII from DNA upon encountering a DNA lesion. © 2012 Nature America, Inc. All rights reserved.


Sohn J.-W.,University of Texas Southwestern Medical Center | Harris L.E.,University of Bristol | Berglund E.D.,University of Texas Southwestern Medical Center | Liu T.,University of Texas Southwestern Medical Center | And 5 more authors.
Cell | Year: 2013

Melanocortin 4 receptors (MC4Rs) in the central nervous system are key regulators of energy and glucose homeostasis. Notably, obese patients with MC4R mutations are hyperinsulinemic and resistant to obesity-induced hypertension. Although these effects are probably dependent upon the activity of the autonomic nervous system, the cellular effects of MC4Rs on parasympathetic and sympathetic neurons remain undefined. Here, we show that MC4R agonists inhibit parasympathetic preganglionic neurons in the brainstem. In contrast, MC4R agonists activate sympathetic preganglionic neurons in the spinal cord. Deletion of MC4Rs in cholinergic neurons resulted in elevated levels of insulin. Furthermore, re-expression of MC4Rs specifically in cholinergic neurons (including sympathetic preganglionic neurons) restores obesity-associated hypertension in MC4R null mice. These findings provide a cellular correlate of the autonomic side effects associated with MC4R agonists and demonstrate a role for MC4Rs expressed in cholinergic neurons in the regulation of insulin levels and in the development of obesity-induced hypertension. © 2013 Elsevier Inc.


North C.S.,VA North Texas Health Care System | North C.S.,University of Texas Southwestern Medical Center | Pfefferbaum B.,The University of Oklahoma Health Sciences Center
JAMA - Journal of the American Medical Association | Year: 2013

IMPORTANCE: Exposure to a disaster is common, and one-third or more of individuals severely exposed may develop posttraumatic stress disorder or other disorders. A systematic approach to the delivery of timely and appropriate disaster mental health servicesmay facilitate their integration into the emergency medical response. OBJECTIVE: To review and summarize the evidence for how best to identify individuals in need of disaster mental health services and triage them to appropriate care. EVIDENCE REVIEW: Search of the peer-reviewed English-language literature on disaster mental health response in PsycINFO, PubMed, Cochrane Database of Systematic Reviews, Academic Search Complete, and Google Scholar (inception to September 2012) and PILOTS (inception to February 2013), using a combination of subject headings and text words (Disasters, Natural Disasters, Mental Health, Mental Health Programs, Public Health Services, Mental Disorders, Mental Health Services, Community Mental Health Services, Emergency Services Psychiatric, Emotional Trauma, Triage, and Response). FINDINGS: Unlike physical injuries, adverse mental health outcomes of disasters may not be apparent, and therefore a systematic approach to case identification and triage to appropriate interventions is required. Symptomatic individuals in postdisaster settings may experience new-onset disaster-related psychiatric disorders, exacerbations of preexisting psychopathology, and/or psychological distress. Descriptive disaster mental health studies have found that many (11%-38%) distressed individuals presenting for evaluation at shelters and family assistance centers have stress-related and adjustment disorders; bereavement, major depression, and substance use disorders were also observed, and up to 40% of distressed individuals had preexisting disorders. Individuals with more intense reactions to disaster stress were more likely to accept referral to mental health services than those with less intense reactions. Evidence-based treatments are available for patients with active psychiatric disorders, but psychosocial interventions such as psychological first aid, psychological debriefing, crisis counseling, and psychoeducation for individuals with distress have not been sufficiently evaluated to establish their benefit or harm in disaster settings. CONCLUSION AND RELEVANCE: In postdisaster settings, a systematic framework of case identification, triage, and mental health interventions should be integrated into emergency medicine and trauma care responses.


Wang Z.,University of Texas Southwestern Medical Center | Jiang H.,University of Texas Southwestern Medical Center | Jiang H.,China National Institute of Biological Sciences | Chen S.,China National Institute of Biological Sciences | And 3 more authors.
Cell | Year: 2012

The programmed necrosis induced by TNF-α requires the activities of the receptor-interacting serine-threonine kinases RIP1 and RIP3 and their interaction with the mixed lineage kinase domain-like protein MLKL. We report the identification of RIP1- and RIP3-containing protein complexes that form specifically in response to necrosis induction. One component of these complexes is the mitochondrial protein phosphatase PGAM5, which presents as two splice variants, PGAM5L (long form) and PGAM5S (short form). Knockdown of either form attenuated necrosis induced by TNF-α as well as reactive oxygen species (ROS) and calcium ionophore, whereas knockdown of RIP3 and MLKL blocked only TNF-α-mediated necrosis. Upon necrosis induction, PGAM5S recruited the mitochondrial fission factor Drp1 and activated its GTPase activity by dephosphorylating the serine 637 site of Drp1. Drp1 activation caused mitochondrial fragmentation, an early and obligatory step for necrosis execution. These data defined PGAM5 as the convergent point for multiple necrosis pathways. © 2012 Elsevier Inc.


Marin-Valencia I.,University of Texas Southwestern Medical Center | Guerrini R.,University of Florence | Gleeson J.G.,Howard Hughes Medical Institute
Epilepsia | Year: 2014

Focal cortical dysplasias (FCDs) constitute a prevalent cause of intractable epilepsy in children, and is one of the leading conditions requiring epilepsy surgery. Despite recent advances in the cellular and molecular biology of these conditions, the pathogenetic mechanisms of FCDs remain largely unknown. The purpose if this work is to review the molecular underpinnings of FCDs and to highlight potential therapeutic targets. A systematic review of the literature regarding the histologic, molecular, and electrophysiologic aspects of FCDs was conducted. Disruption of the mammalian target of rapamycin (mTOR) signaling comprises a common pathway underlying the structural and electrical disturbances of some FCDs. Other mechanisms such as viral infections, prematurity, head trauma, and brain tumors are also posited. mTOR inhibitors (i.e., rapamycin) have shown positive results on seizure management in animal models and in a small cohort of patients with FCD. Encouraging progress has been achieved on the molecular and electrophysiologic basis of constitutive cells in the dysplastic tissue. Despite the promising results of mTOR inhibitors, large-scale randomized trials are in need to evaluate their efficacy and side effects, along with additional mechanistic studies for the development of novel, molecular-based diagnostic and therapeutic approaches. A PowerPoint slide summarizing this article is available for download in the Supporting Information section here. © Wiley Periodicals, Inc. © 2014 International League Against Epilepsy.


Choudhury D.,University of Texas Southwestern Medical Center | Levi M.,University of Colorado at Denver
Nature Reviews Nephrology | Year: 2011

The aging process affects all organs, including the kidneys. As part of this process, progressive scarring and a measurable decline in renal function occur in most people over time. The improved understanding of the processes that can lead to and/or hasten scarring and loss of renal function over time parallels advances in our understanding of the aging process. Clinical factors, including hypertension, diabetes mellitus, obesity, abnormal lipid levels and vitamin D deficiency, have been associated with increasing renal sclerosis with age. In addition, tissue factors such as angiotensin II, advanced glycation end products, oxidative stress and Klotho are associated with renal aging. These associations and possible interventions, including the control of blood pressure, blood sugar, weight, diet and calorie restriction might make renal aging more preventable than inevitable. © 2011 Macmillan Publishers Limited. All rights reserved.


Cutrell J.,University of Texas Southwestern Medical Center | Bedimo R.,VA North Texas Health Care System | Bedimo R.,University of Texas Southwestern Medical Center
Current HIV/AIDS Reports | Year: 2013

In the highly active antiretroviral therapy (HAART) era, the incidence of non-AIDS-defining cancers (NADC) has increased and contributes to a growing proportion of mortality in the aging HIV-infected population. The underlying pathogenic mechanisms of increased cancer risk are incompletely understood. Potential contributors include oncogenic effects of the HIV virus, immunosuppression, chronic inflammation and immune activation, exposure to HAART, higher rates of oncogenic viral coinfections and traditional cancer risk factors. HIV-infected patients often present with NADC at younger ages with more aggressive or advanced stage disease. However, when standard cancer therapy is given, treatment outcomes appear similar to the non-HIV population. These facts highlight the importance of clinicians' maintaining a high index of suspicion, performing age-appropriate screening, and optimizing cancer therapy. Development of novel strategies for screening, prevention, and treatment of NADC will be required to reverse these epidemiologic trends and improve the survival of HIV-infected patients. © 2013 Springer Science+Business Media New York (outside the USA).


Kieseier B.C.,Heinrich Heine University Düsseldorf | Stuve O.,University of Texas Southwestern Medical Center
Nature Reviews Neurology | Year: 2011

Interferon β 2 and glatiramer acetate have been available for the treatment of multiple sclerosis (MS) for over a decade and their efficacy and safety are well established. These agents have detectable effects on the immune system, but have not been associated with a breakdown of immune surveillance. Novel MS therapies have been approved, or are awaiting approval, that differ from established immunotherapies with regard to their mechanisms of action, modes of administration, adverse-effect profiles and, possibly, the clinical and paraclinical benefits that they may provide for patients. Neurologists will soon be required to make complex treatment decisions with their patients on the basis of very limited clinical data and evidence. Under these circumstances, optimal assessment of risks and benefits will be challenging. In this Review, the anticipated benefits of novel therapies, including reduction in disease activity, possible prevention of disability, and improvement in quality of life, are outlined. In addition, the current acceptance of potential risks-including serious or even life-threatening adverse effects, the likelihood of which may rise with increased cumulative exposure to a particular agent-by patients with MS will be reviewed. © 2011 Macmillan Publishers Limited. All rights reserved.


Ramesh A.,University of Texas Southwestern Medical Center | Winkler W.C.,University of Maryland University College
Biochimica et Biophysica Acta - Gene Regulatory Mechanisms | Year: 2014

Catalysis in the biological context was largely thought to be a protein-based phenomenon until the discovery of RNA catalysts called ribozymes. These discoveries demonstrated that many RNA molecules exhibit remarkable structural and functional versatility. By virtue of these features, naturally occurring ribozymes have been found to be involved in catalyzing reactions for fundamentally important cellular processes such as translation and RNA processing. Another class of RNAs called riboswitches directly binds ligands to control downstream gene expression. Most riboswitches regulate downstream gene expression by controlling premature transcription termination or by affecting the efficiency of translation initiation. However, one riboswitch class couples ligand-sensing to ribozyme activity. Specifically, the glmS riboswitch is a nucleolytic ribozyme, whose self-cleavage activity is triggered by the binding of GlcN6P. The products of this self-cleavage reaction are then targeted by cellular RNases for rapid degradation, thereby reducing glmS expression under conditions of sufficient GlcN6P. Since the discovery of the glmS ribozyme, other metabolite-binding ribozymes have been identified. Together, these discoveries have expanded the general understanding of noncoding RNAs and provided insights that will assist future development of synthetic riboswitch-ribozymes. A very broad overview of natural and synthetic ribozymes is presented herein with an emphasis on the structure and function of the glmS ribozyme as a paradigm for metabolite-binding ribozymes that control gene expression. This article is part of a Special Issue entitled: Riboswitches. © 2014 Elsevier B.V.


Cai X.,University of Texas Southwestern Medical Center | Chiu Y.-H.,Beth Israel Deaconess Medical Center | Chen Z.J.,University of Texas Southwestern Medical Center
Molecular Cell | Year: 2014

The innate immune system deploys a variety of sensors to detect signs of infection. Nucleic acids represent amajor class of pathogen signatures that can trigger robust immune responses. The presence of DNA inthecytoplasm of mammalian cells is a danger signal that activates innate immune responses; however, how cytosolic DNA triggers these responses remained unclear until recently. In this review, we focus onthe mechanism of DNA sensing by the newly discovered cGAS-cGAMP-STING pathway and highlight recent progress in dissecting the invivo functions of this pathway in immune defense as well as autoimmunity. © 2014 Elsevier Inc.


Souza R.F.,VA North Texas Health Care System | Souza R.F.,University of Texas Southwestern Medical Center
Biochemical Society Transactions | Year: 2010

The precise mechanisms whereby gastro-oesophageal reflux disease causes reflux oesophagitis and Barrett's oesophagus are not clear, even though these diseases have been known to be linked for many years. Recent studies indicate a role for the reflux-induced inflammatory response of oesophageal squamous epithelial cells and the immune cells in the pathogenesis of reflux oesophagitis. Although reflux oesophagitis commonly heals with oesophageal squamous cell regeneration, in some individuals the oesophagus heals through the process of metaplasia, a condition termed Barrett's oesophagus. Recent studies indicate that individual differences in the reflux-mediated response of oesophageal squamous epithelial cells in the type of immune response and/or in signalling pathways that regulate cell proliferation or cell phenotype may determine whether the oesophagus heals with the regeneration of squamous cells or through Barrett's metaplasia. ©The Authors.


Patent
University of Maryland University College and University of Texas Southwestern Medical Center | Date: 2014-12-17

Provided are compositions and methods related to heme transporters and high-throughput methods of identifying agents that can modulate heme transporters. An approach for identifying a modulator of a eukaryotic heme transporter involves adding a toxic heme analog and at least one test agent to a culture of cells, wherein the cells express a recombinant heterologous eukaryotic heme transporter. The cells are incubated with the toxic heme analog and the test agent for a period of time. A change in toxic effect of the toxic heme analog relative to a control is indicative that the test agent is a modulator of the eukaryotic heme transporter. Heme transporter agonists and antagonists can be identified. Also provided is a cell culture comprising a plurality of cells which express a recombinant heterologous eukaryotic heme transporter. The plurality of cells are divided into a plurality of reaction chambers, each of which may contain a test agent, and may further contain a toxic heme analog.


News Article | September 6, 2016
Site: www.biosciencetechnology.com

In rare cases -- for instance, among siblings in two families from Pakistan and Oman described in a new study -- children have been born with an unnamed neurological disorder. Now researchers have not only identified the genetic mutations involved, but also replicated them in lab cultures and mouse models to produce an initial understanding of how the mutations cause the disease. The study in the Proceedings of the National Academy of Sciences highlights both new medical and scientific opportunities, said Dr. Eric Morrow, associate professor of biology and of psychiatry and human behavior in the Warren Alpert Medical School of Brown University. "This is a clear, new neurogenetic disorder due to mutations in GPT2," said Morrow, who also sees patients at Bradley Hospital and is affiliated with the Brown Institute for Brain Science. "In addition to the relevance this has to the diagnosis of developmental disorders, and potentially therapeutics, it is also a window into how the brain develops and how the brain functions." The paper reveals specific findings of basic neurodevelopment, Morrow said. The gene at issue, GPT2, is expressed in the nucleus of cells, but the enzyme it generates appears vital to metabolic pathways in the mitochondria, organelles which provide energy and biosynthetic building blocks to cells. The consequences of the mutations appear to be in leaving developing brains without biosynthetic abilities to grow properly, and to deficits in metabolites that could help prevent degeneration. Moreover, the G in the gene name stands for glutamate, an important neurotransmitter that governs how brain cells, or neurons, connect and interact. "To find a glutamate metabolizing enzyme that is associated with a brain disease is an opportunity to understand how that neurotransmitter might work or be modulated," Morrow said. Morrow is co-corresponding author along with collaborators David Housman at Massachusetts Institute of Technology and Ganeshwaran Mochida at Boston Children's Hospital. The study's co-lead authors are Brown investigator Qing Ouyang and Boston Children's Hospital researcher Tojo Nakayama. The second author, Ozan Baytas, is a Ph.D. candidate in Brown's neuroscience graduate training program. The team, which also includes collaborators in Pakistan and Oman, began the investigation more than five years ago when they were studying two families in those countries with children whose symptoms included below-normal postnatal brain growth, intellectual disability and progressively worsening motor problems. The children, 14 in all across the two large families, typically were able to walk by age 3, yet a majority lost that ability later as motor control diminished in their legs, as a condition called spastic paraplegia emerged. Spastic paraplegia is generally considered to involve a neurodegenerative cause, Morrow said. The team traced a genetic mutation to chromosome 16, and as next-generation sequencing technology became available, they were able to find two specific mutations in GPT2. In the interim, a few other research groups had also linked GPT2 mutations to neurological disease in other families, including a family in the U.S. Taken together, Morrow said, the studies provide strong evidence that the gene is relevant to neurological disease. With gene mutations identified, Morrow and his collaborators went much further to learn how the mutations could cause the disease. To do that the team created models in which the mutations were induced in human cells and also in mice. Like the children with GPT2 mutations, developing mice with the mutations also showed reduced neural and brain growth. In these lab models the researchers were able to study in detail the different biochemistry at play with and without the mutations. In the human cells they saw that mutations led to reduced enzyme activity. They also determined that the protein locates in the mitochondria. Mutant mice engineered with a GPT2 enzyme deficiency showed abnormal brain metabolism. For example, some of the differences undermined a process called the TCA cycle, which is important for producing energy and generating building blocks for cells. While these metabolic pathways have been well studied in rapidly dividing cells such as in cancer, Morrow said, they have not been as thoroughly studied in differentiating neurons growing extensions and connections during early childhood. To do that, Morrow and colleagues teamed up with experts in cancer metabolism, Ralph DeBerardinis at the University of Texas Southwestern Medical Center and Shawn Davidson and David Housman at MIT, to develop experiments pertinent to GPT2 and brain. When the researchers looked at the neurons of developing mice, they found that ones with the GPT2 mutations produced fewer synapses, the connections between neurons that make up brain circuits. The researchers conducted large-scale profiling of metabolites in the brains of the mutant mice. They found in the GPT2 mutant mice abnormal metabolite levels related to amino acid metabolism, TCA and pathways required for protecting neuron health. The deficiencies in these neuroprotective metabolites, Morrow said, might explain why the disease appears to have a degenerative course. Morrow's team at Brown is now developing new hypotheses and testing them in the mouse model they developed. Not only are they seeking to refine and deepen their understanding of how those metabolic pathways regulate brain disease and function, but also, they are eager to test potential ways to rescue development and prevent disease progression. "I believe there is hope that if these children were identified early as having this genetic condition, there may be an intervention that could prevent the progression," Morrow said. The researchers are also interested to learn more about how these mitochondrial metabolic pathways play key roles in the developing brain and how these pathways may contribute to brain health. Several public and private organizations helped to support the research.


News Article | December 14, 2016
Site: www.sciencenews.org

Rashes are the temporary tattoos of childhood. The prickly, red bumps can blossom across the skin for a host of reasons: an ear infection, a virus or even an allergic reaction to a penicillin antibiotic. What’s hard to tell, though, is whether the penicillin or the illness itself triggers the rash. To be safe, doctors label some children as allergic to penicillin, but a skin test to verify the diagnosis rarely happens. “These kids march into adulthood with a penicillin allergy label that’s never really addressed,” says Allison Ramsey, an allergist at Rochester Regional Health in New York. About 10 percent of U.S. adults and children believe they have a penicillin allergy, the most commonly reported drug allergy. But 90 percent of people who think they’re allergic to penicillin actually aren’t, according to a 2010 report in Annals of Allergy, Asthma & Immunology. There is a “massive problem with the overreporting of penicillin allergy,” Ramsey says. When researchers from the University of Texas Southwestern Medical Center in Dallas recently skin tested 228 “penicillin allergic” patients, almost 98 percent of the patients turned out not to be allergic. The team reported the findings November 12 in San Francisco at the annual meeting of the American College of Allergy, Asthma & Immunology. In reality, Ramsey says, people either never had the allergy or they got over it with time. To avoid the chance of triggering a severe allergic reaction, doctors often give people who are considered allergic to penicillin a broad-spectrum, second-line antibiotic. Compared with penicillin, these drugs are often more expensive, less effective against certain bacteria and come with more side effects. On a troubling societal level, using the more general antibiotics may encourage the spread of antibiotic resistance (SN: 10/4/14, p. 22). Overdiagnosis of penicillin allergy is not benign, Ramsey says. Surveying 276 physicians, physician assistants, nurse practitioners and pharmacists at two Rochester Regional Health hospitals, Ramsey and colleagues found very low levels of allergy testing. More than 85 percent of respondents reported that they never consulted with an allergist or immunologist for antibiotic allergies or skin tests, or they did so only once a year. More than 40 percent didn’t know that a penicillin allergy can resolve over time. Ramsey presented the results November 14 at the allergy meeting. Taking the time to confirm or rule out a penicillin allergy can cut down on the use of second-line antibiotics. In the Dallas study, after penicillin allergy testing, the use of vancomycin, a powerful, last-resort antibiotic, decreased by 34 percent and use of the costly aztreonam dropped by 68 percent. “Those are big numbers,” says Ramsey. It’s important that people know that childhood penicillin allergies can be revisited, she adds. “It’s not a lifetime label.”


Curthoys N.P.,Colorado State University | Moe O.W.,University of Texas Southwestern Medical Center
Clinical Journal of the American Society of Nephrology | Year: 2014

The human kidneys produce approximately 160–170 L of ultrafiltrate per day. The proximal tubule contributes to fluid, electrolyte, and nutrient homeostasis by reabsorbing approximately 60%–70% of the water and NaCl, a greater proportion of the NaHCO3, and nearly all of the nutrients in the ultrafiltrate. The proximal tubule is also the site of active solute secretion, hormone production, and many of the metabolic functions of the kidney. This review discusses the transport of NaCl, NaHCO3, glucose, amino acids, and two clinically important anions, citrate and phosphate. NaCl and the accompanying water are reabsorbed in an isotonic fashion. The energy that drives this process is generated largely by the basolateral Na+/K+-ATPase, which creates an inward negative membrane potential and Na+-gradient. Various Na+-dependent countertransporters and cotransporters use the energy of this gradient to promote the uptake of HCO3 - and various solutes, respectively. A Na+-dependent cotransporter mediates the movement of HCO3 - across the basolateral membrane, whereas various Na+- independent passive transporters accomplish the export of various other solutes. To illustrate its homeostatic feat, the proximal tubule alters its metabolism and transport properties in response to metabolic acidosis. The uptake and catabolism of glutamine and citrate are increased during acidosis, whereas the recovery of phosphate from the ultrafiltrate is decreased. The increased catabolism of glutamine results in increased ammoniagenesis and gluconeogenesis. Excretion of the resulting ammonium ions facilitates the excretion of acid, whereas the combined pathways accomplish the net production of HCO3 - ions that are added to the plasma to partially restore acid-base balance. © 2014 by the American Society of Nephrology.


Cantey J.B.,University of Texas Southwestern Medical Center | Milstone A.M.,Johns Hopkins University
Clinics in Perinatology | Year: 2015

Bloodstream infections in the neonatal intensive care unit (NICU) are associated with many adverse outcomes in infants, including increased length of stay and cost, poor neurodevelopmental outcomes, and death. Attention to the insertion and maintenance of central lines, along with careful review of when the catheters can be safely discontinued, can minimize central-line-associated bloodstream infections rates. Good antibiotic stewardship can further decrease the incidence of bloodstream infections, minimize the emergence of drug-resistant organisms or Candida as pathogens in the NICU, and safeguard the use of currently available antibiotics for future infants. © 2015 Elsevier Inc.


Mealy M.A.,Johns Hopkins University | Wingerchuk D.M.,Mayo Medical School | Greenberg B.M.,University of Texas Southwestern Medical Center | Levy M.,Johns Hopkins University
Archives of Neurology | Year: 2012

Background: Rare diseases require integrated multicenter clinical networks to facilitate clinical research. Neuromyelitis optica (NMO) and NMO spectrum disorders (NMOSDs) are uncommon neuroinflammatory syndromes that are distinct from multiple sclerosis and associated with NMO-IgG, a serologic antibody against aquaporin 4. Objective: To develop a national multicenter NMO clinical consortium and report initial demographic, clinical, and radiographic features of a cohort of patients with NMO/NMOSD in the United States. Design: Review of medical records from patients undergoing evaluation during a 5-year period. We used uniform diagnostic criteria and clinical, laboratory, and neuroimaging definitions to describe the cohort. Setting: Three academic medical centers. Patients: One hundred eighty-seven patients with NMO/NMOSD. Results: Of the 187 patients included in the analysis, 86 had NMO-IgG-seropositive NMO; 40, NMO-IgG-seronegative NMO; and 61, NMO-IgG-seropositive NMOSD. Altogether, 29.4% of our patients were initially misdiagnosed with multiple sclerosis. The average age at onset of NMO/NMOSD was 41.1 years with a strong female predilection, similar to other autoimmune disorders. Nonwhite patients constituted 52.4% of the cohort. The hallmark of NMO and NMOSD is recurrent longitudinally extensive transverse myelitis, but patients with NMO tend to initially present with optic neuritis. Conclusions: A national multicenter consortium to study NMO/NMOSD is feasible and facilitates accurate clinical diagnosis. This network establishes a foundation for determining disease prevalence, translational research, and clinical trials. ©2012 American Medical Association. All rights reserved.


Kourrich S.,University of Texas Southwestern Medical Center | Calu D.J.,U.S. National Institute on Drug Abuse | Bonci A.,U.S. National Institute on Drug Abuse | Bonci A.,Johns Hopkins University
Nature Reviews Neuroscience | Year: 2015

Exposure to drugs of abuse, such as cocaine, leads to plastic changes in the activity of brain circuits, and a prevailing view is that these changes play a part in drug addiction. Notably, there has been intense focus on drug-induced changes in synaptic excitability and much less attention on intrinsic excitability factors (that is, excitability factors that are remote from the synapse). Accumulating evidence now suggests that intrinsic factors such as K + channels are not only altered by cocaine but may also contribute to the shaping of the addiction phenotype. © 2015 Macmillan Publishers Limited. All rights reserved.


Kanatous S.B.,Colorado State University | Mammen P.P.A.,University of Texas Southwestern Medical Center
Journal of Experimental Biology | Year: 2010

Myoglobin is a well-characterized, cytoplasmic hemoprotein that is expressed primarily in cardiomyocytes and oxidative skeletal muscle fibers. However, recent studies also suggest low-level myoglobin expression in various non-muscle tissues. Prior studies incorporating molecular, pharmacological, physiological and transgenic technologies have demonstrated that myoglobin is an essential oxygen-storage hemoprotein capable of facilitating oxygen transport and modulating nitric oxide homeostasis within cardiac and skeletal myocytes. Concomitant with these studies, scientific investigations into the transcriptional regulation of myoglobin expression have been undertaken. These studies have indicated that activation of key transcription factors (MEF2, NFAT and Sp1) and co-activators (PGC-1α) by locomotor activity, differential intracellular calcium fluxes and low intracellular oxygen tension collectively regulate myoglobin expression. Future studies focused on tissue-specific transcriptional regulatory pathways and post-translational modifications governing myoglobin expression will need to be undertaken. Finally, further studies investigating the modulation of myoglobin expression under various myopathic processes may identify myoglobin as a novel therapeutic target for the treatment of various cardiac and skeletal myopathies.


Dutchak P.A.,University of Texas Southwestern Medical Center | Katafuchi T.,University of Texas Southwestern Medical Center | Bookout A.L.,University of Texas Southwestern Medical Center | Choi J.H.,Dana-Farber Cancer Institute | And 4 more authors.
Cell | Year: 2012

Fibroblast growth factor-21 (FGF21) is a circulating hepatokine that beneficially affects carbohydrate and lipid metabolism. Here, we report that FGF21 is also an inducible, fed-state autocrine factor in adipose tissue that functions in a feed-forward loop to regulate the activity of peroxisome proliferator-activated receptor γ (PPARγ), a master transcriptional regulator of adipogenesis. FGF21 knockout (KO) mice display defects in PPARγ signaling including decreased body fat and attenuation of PPARγ-dependent gene expression. Moreover, FGF21-KO mice are refractory to both the beneficial insulin-sensitizing effects and the detrimental weight gain and edema side effects of the PPARγ agonist rosiglitazone. This loss of function in FGF21-KO mice is coincident with a marked increase in the sumoylation of PPARγ, which reduces its transcriptional activity. Adding back FGF21 prevents sumoylation and restores PPARγ activity. Collectively, these results reveal FGF21 as a key mediator of the physiologic and pharmacologic actions of PPARγ. © 2012 Elsevier Inc.


Chen Y.,CAS Kunming Institute of Zoology | Jiang J.,University of Texas Southwestern Medical Center
Cell Research | Year: 2013

Hedgehog (Hh) signaling plays pivotal roles in embryonic development and adult tissue homeostasis, and its deregulation leads to numerous human disorders including cancer. Binding of Hh to Patched (Ptc), a twelve-transmembrane protein, alleviates its inhibition of Smoothened (Smo), a seven-transmembrane protein related to G-protein-coupled receptors (GPCRs), leading to Smo phosphorylation and activation. Smo acts through intracellular signaling complexes to convert the latent transcription factor Cubitus interruptus (Ci)/Gli from a truncated repressor to a full-length activator, leading to derepression/activation of Hh target genes. Increasing evidence suggests that phosphorylation participates in almost every step in the signal relay from Smo to Ci/Gli, and that differential phosphorylation of several key pathway components may be crucial for translating the Hh morphogen gradient into graded pathway activities. In this review, we focus on the multifaceted roles that phosphorylation plays in Hh signal transduction, and discuss the conservation and difference between Drosophila and mammalian Hh signaling mechanisms. © 2013 IBCB, SIBS, CAS All rights reserved.


Reynolds K.A.,University of Texas Southwestern Medical Center | McLaughlin R.N.,University of Texas Southwestern Medical Center | McLaughlin R.N.,Fred Hutchinson Cancer Research Center | Ranganathan R.,University of Texas Southwestern Medical Center
Cell | Year: 2011

Recent work indicates a general architecture for proteins in which sparse networks of physically contiguous and coevolving amino acids underlie basic aspects of structure and function. These networks, termed sectors, are spatially organized such that active sites are linked to many surface sites distributed throughout the structure. Using the metabolic enzyme dihydrofolate reductase as a model system, we show that: (1) the sector is strongly correlated to a network of residues undergoing millisecond conformational fluctuations associated with enzyme catalysis, and (2) sector-connected surface sites are statistically preferred locations for the emergence of allosteric control in vivo. Thus, sectors represent an evolutionarily conserved "wiring" mechanism that can enable perturbations at specific surface positions to rapidly initiate conformational control over protein function. These findings suggest that sectors enable the evolution of intermolecular communication and regulation. © 2011 Elsevier Inc.


Pelosof L.C.,University of Texas Southwestern Medical Center | Pelosof L.C.,Johns Hopkins University | Gerber D.E.,University of Texas Southwestern Medical Center
Mayo Clinic Proceedings | Year: 2010

Recent medical advances have improved the understanding, diagnosis, and treatment of paraneoplastic syndromes. These disorders arise from tumor secretion of hormones, peptides, or cytokines or from immune cross-reactivity between malignant and normal tissues. Paraneoplastic syndromes may affect diverse organ systems, most notably the endocrine, neurologic, dermatologic, rheumatologic, and hematologic systems. The most commonly associated malignancies include small cell lung cancer, breast cancer, gynecologic tumors, and hematologic malignancies. In some instances, the timely diagnosis of these conditions may lead to detection of an otherwise clinically occult tumor at an early and highly treatable stage. Because paraneoplastic syndromes often cause considerable morbidity, effective treatment can improve patient quality of life, enhance the delivery of cancer therapy, and prolong survival. Treatments include addressing the underlying malignancy, immunosuppression (for neurologic, dermatologic, and rheumatologic paraneoplastic syndromes), and correction of electrolyte and hormonal derangements (for endocrine paraneoplastic syndromes). This review focuses on the diagnosis and treatment of paraneoplastic syndromes, with emphasis on those most frequently encountered clinically. Initial literature searches for this review were conducted using PubMed and the keyword paraneoplastic in conjunction with keywords such as malignancy, SIADH, and limbic encephalitis, depending on the particular topic. Date limitations typically were not used, but preference was given to recent articles when possible. © 2010 Mayo Foundation for Medical Education and Research.


Pfeiffer B.E.,Johns Hopkins University | Pfeiffer B.E.,University of Texas Southwestern Medical Center | Foster D.J.,Johns Hopkins University
Science | Year: 2015

Neuronal circuits produce self-sustaining sequences of activity patterns, but the precise mechanisms remain unknown. Here we provide evidence for autoassociative dynamics in sequence generation. During sharp-wave ripple (SWR) events, hippocampal neurons express sequenced reactivations, which we show are composed of discrete attractors. Each attractor corresponds to a single location, the representation of which sharpens over the course of several milliseconds, as the reactivation focuses at that location. Subsequently, the reactivation transitions rapidly to a spatially discontiguous location. This alternation between sharpening and transition occurs repeatedly within individual SWRs and is locked to the slow-gamma (25 to 50 hertz) rhythm. These findings support theoretical notions of neural network function and reveal a fundamental discretization in the retrieval of memory in the hippocampus, together with a function for gamma oscillations in the control of attractor dynamics. Copyright 2015 by the American Association for the Advancement of Science; all rights reserved.


Patent
Johns Hopkins University and University of Texas Southwestern Medical Center | Date: 2014-08-07

The present invention is directed to an all-fiber-optic scanning endomicroscope capable of high-resolution second harmonic generation (SHG) imaging of biological tissues. The endomicroscope has an overall 2.0 mm diameter and consists of a single customized double-clad fiber, a compact rapid two-dimensional beam scanner, and a miniature compound objective lens for excitation beam delivery, scanning, focusing, and efficient SHG signal collection. Endomicroscopic SHG images of murine cervical tissue sections at different stages of normal pregnancy reveal progressive, quantifiable changes in cervical collagen morphology with resolution similar to that of bench-top SHG microscopy. A device according to an embodiment of the present invention can also be used to assess other pathologies, such as cancer. fibrosis, and inflammation. The present invention allows for diagnosis, monitoring of the effect of therapeutics, and surgical or interventional guidance. The present invention enables visualization of histology in-vivo, in the patient, and is label-free.


News Article | November 10, 2015
Site: www.biosciencetechnology.com

Five scientists, working in the area of life sciences, each received a $3 million award on Sunday night as winners of the Breakthrough Prize. The awards ceremony, which included a live performance by musical star Pharrell Williams and celebrity presenters including Russell Crowe and Hilary Swank, doled out a combined total of $21.9 million to scientists in life sciences, fundamental physics, and mathematics and one high school student. According to the New York Times, founders of the Breakthrough Prize are Google’s Sergey Brin; Facebook’s Mark Zuckerberg and his wife Priscilla Chan; 23andme’s Anne Wojcicki; Alibab’s Jack Ma and his wife Cathy Zhang; and Internet entrepreneur Yuri Milner, and his wife, Julia Milner. Here are the five Breakthrough Prize laureates for life sciences and their research, according to a released statement: Edward S. Boyden, Massachusetts Institute of Technology Karl Deisseroth, Stanford University and researcher at Howard Hughes Medical Institute: Prizes went to both professors for their development and implementation of optogenetics, which is a technique that uses light to control the electrical activity of neurons that have been genetically modified to express light-activated ion channels and pumps. John Hardy, University College London Hardy was awarded for his discovery of gene mutations in the Amyloid Precursor Protein gene (APP) that cause early onset Alzheimer’s disease. He linked the development of Alzheimer’s to accumulation of APP-derived beta-amyloid peptide to the development of Alzheimer’s, and the NYT reported Hardy’s work was the first that shed light on what might start the crippling damage of brain cells in Alzheimer’s disease. This opened the way to new avenues in the search for a cure. Helen Hobbs, University of Texas Southwestern Medical Center and investigator at Howard Hughes Medical Institute Hobbs was awarded for her discovery of human genetic variants, specifically a mutation in a gene PCSK9 that alter levels of cholesterol. In her research she found that the less PCSK9 made, the lower LDL levels were and more protected against cardiovascular disease.  Hobbs work inspired new strategies for preventing cardiovascular and liver disease, and lead to the creation of cholesterol-lowering drugs that imitate a PCSK9 mutation, according to the NYT. Svante Pääbo, Max Planck Institute for Evolutionary Anthropology Pääbo received a prize for his pioneering work sequencing ancient DNA and genomes that helped reveal the origins of modern humans and shed light on relationships with extinct relatives such as Neanderthals.  The NYT reported that Pääbo and his colleagues discovered that modern humans interbred with Neanderthals at least twice before their extinction 40,000 years ago.


News Article | October 29, 2016
Site: www.sciencedaily.com

How should plastic surgeons choose the best implant type and size for women undergoing breast augmentation surgery? Implant size selection systems based on breast tissue measurements may provide better outcomes, suggests a research review in the November issue of Plastic and Reconstructive Surgery®, the official medical journal of the American Society of Plastic Surgeons (ASPS). Tissue-based planning systems -- using clinical guidelines to determine the optimal breast implant dimensions for individual patients -- appear superior to approaches relying more on the patient's or surgeon's preference, according to the study by Drs. William P. Adams, Jr., of University of Texas Southwestern Medical Center, Dallas, and Daniel McKee of McMaster University, Hamilton, Ont., Canada. But further studies will be needed to clarify how breast implant size selection systems affect the outcomes of breast augmentation. 'Tissue-Based Planning' Seems Best -- But Evidence Is Limited The researchers performed a "data-driven review" of methods used by plastic surgeons to select the appropriate implant size for breast augmentation surgery. Breast augmentation is the most popular cosmetic plastic surgery procedure in the United States, with nearly 280,000 procedures performed in 2015, according to ASPS statistics. Implant size selection systems were divided into three groups: • No breast measurements. Implants are chosen based solely on the patient's or surgeon's preference. • Dimensional analysis systems. Implants are chosen in order to establish a desired result, with measurements performed to determine the implant needed to achieve that result. • Tissue-based planning (TBP). Breast tissue measurements are used to set "clear and narrow boundaries" for implant selection based on clinical guidelines, with limited to no flexibility. The review identified 33 articles on implant sizing systems. Studies evaluating TBP sizing systems were of higher quality than those in the other two categories. "The top ten studies based on methodological quality all used patients' breast dimensions before selecting final implant dimensions, and this should now be considered standard of practice based on our analysis," Drs. Adams and McKee write. The TBP studies reported low rates of repeat surgery, compared to industry standards and accepted research values. The researchers emphasize some major limitations of the available evidence on implant sizing systems. Just four out of 33 studies reported clinical outcomes that could be compared to any standard, while none of the studies compared two or more sizing systems. Overall, 60 percent of studies scored zero on the quality rating scale used -- including some popular sizing systems that were "not grounded on any published data or evidence." The topic of implant selection can be an emotional one, with tension between the plastic surgeon's roles as "Artist" versus "Engineer." The researchers note that some TBP systems with the highest quality of evidence take a "middle-of-the-road" approach -- based on measurements, but also considering the patient's aesthetic desires. Based on their data, Drs. Adams and McKee are evaluating a new "implant-specific" TBP system designed to guide the surgeon to a selection of manufactured implant styles and models. "Going forward," they write, "new published systems should [use] rigorous quantitative methods so that comparisons can be made in terms of patient outcomes."


Nomura T.,Karolinska Institutet | Goritz C.,Karolinska Institutet | Catchpole T.,University of Texas Southwestern Medical Center | Henkemeyer M.,University of Texas Southwestern Medical Center | Frisen J.,Karolinska Institutet
Cell Stem Cell | Year: 2010

Stem cells remain in specialized niches over the lifespan of the organism in many organs to ensure tissue homeostasis and enable regeneration. How the niche is maintained is not understood, but is probably as important as intrinsic stem cell self-renewal capacity for tissue integrity. We here demonstrate a high degree of phenotypic plasticity of the two main niche cell types, ependymal cells and astrocytes, in the neurogenic lateral ventricle walls in the adult mouse brain. In response to a lesion, astrocytes give rise to ependymal cells and ependymal cells give rise to niche astrocytes. We identify EphB2 forward signaling as a key pathway regulating niche cell plasticity. EphB2 acts downstream of Notch and is required for the maintenance of ependymal cell characteristics, thereby inhibiting the transition from ependymal cell to astrocyte. Our results show that niche cell identity is actively maintained and that niche cells retain a high level of plasticity. © 2010 Elsevier Inc.


Pieper A.A.,University of Iowa | McKnight S.L.,University of Texas Southwestern Medical Center | Ready J.M.,University of Texas Southwestern Medical Center
Chemical Society Reviews | Year: 2014

A novel neuroprotective small molecule was discovered using a target-agnostic in vivo screen in living mice. This aminopropyl carbazole, named P7C3, is orally bioavailable, crosses the blood-brain barrier, and is non-toxic at doses several fold higher than the efficacious dose. The potency and drug-like properties of P7C3 were optimized through a medicinal chemistry campaign, providing analogues for detailed examination. Improved versions, such as (-)-P7C3-S243 and P7C3-A20, displayed neuroprotective properties in rodent models of Parkinson's disease, amyotrophic lateral sclerosis, traumatic brain injury and age-related cognitive decline. Derivatives appended with immobilizing moieties may reveal the protein targets of the P7C3 class of neuroprotective compounds. Our results indicate that unbiased, in vivo screens might provide starting points for the development of treatments for neurodegenerative diseases as well as tools to study the biology underlying these disorders. This journal is © the Partner Organisations 2014.


Wang H.,National Health Research Institute | Falck J.R.,University of Texas Southwestern Medical Center | Hall T.M.T.,National Health Research Institute | Shears S.B.,National Health Research Institute
Nature Chemical Biology | Year: 2012

Inositol pyrophosphates (such as IP7 and IP8) are multifunctional signaling molecules that regulate diverse cellular activities. Inositol pyrophosphates have 'high-energy' phosphoanhydride bonds, so their enzymatic synthesis requires that a substantial energy barrier to the transition state be overcome. Additionally, inositol pyrophosphate kinases can show stringent ligand specificity, despite the need to accommodate the steric bulk and intense electronegativity of nature's most concentrated three-dimensional array of phosphate groups. Here we examine how these catalytic challenges are met by describing the structure and reaction cycle of an inositol pyrophosphate kinase at the atomic level. We obtained crystal structures of the kinase domain of human PPIP5K2 complexed with nucleotide cofactors and either substrates, product or a MgF 3 - transition-state mimic. We describe the enzyme's conformational dynamics, its unprecedented topological presentation of nucleotide and inositol phosphate, and the charge balance that facilitates partly associative in-line phosphoryl transfer. © 2011 Nature America, Inc. All rights reserved.


Mendoza J.L.,University of Texas Southwestern Medical Center | Schmidt A.,University of Texas Southwestern Medical Center | Li Q.,University of Texas Southwestern Medical Center | Nuvaga E.,University of Texas Southwestern Medical Center | And 5 more authors.
Cell | Year: 2012

Misfolding of ΔF508 cystic fibrosis (CF) transmembrane conductance regulator (CFTR) underlies pathology in most CF patients. F508 resides in the first nucleotide-binding domain (NBD1) of CFTR near a predicted interface with the fourth intracellular loop (ICL4). Efforts to identify small molecules that restore function by correcting the folding defect have revealed an apparent efficacy ceiling. To understand the mechanistic basis of this obstacle, positions statistically coupled to 508, in evolved sequences, were identified and assessed for their impact on both NBD1 and CFTR folding. The results indicate that both NBD1 folding and interaction with ICL4 are altered by the ΔF508 mutation and that correction of either individual process is only partially effective. By contrast, combination of mutations that counteract both defects restores ΔF508 maturation and function to wild-type levels. These results provide a mechanistic rationale for the limited efficacy of extant corrector compounds and suggest approaches for identifying compounds that correct both defective steps. © 2012 Elsevier Inc.


Katz E.S.,Harvard University | Mitchell R.B.,University of Texas Southwestern Medical Center | D'Ambrosio C.M.,Tufts Medical Center and Tufts Medical School
American Journal of Respiratory and Critical Care Medicine | Year: 2012

Obstructive sleep apnea in infants has a distinctive pathophysiology, natural history, and treatment compared with that of older children and adults. Infants have both anatomical and physiological predispositions toward airway obstruction andgas exchange abnormalities; including a superiorly placed larynx, increased chestwall compliance, ventilation-perfusion mismatching, and ventilatory control instability. Congenital abnormalities of the airway, such as laryngomalacia, hemangiomas, pyriform aperture stenosis, choanal atresia, and laryngeal webs, may also have adverse effects on airway patency. Additional exacerbating factors predisposing infants toward airway collapse include neck flexion, airway secretions, gastroesophageal reflux, and sleep deprivation. Obstructive sleep apnea in infants has been associatedwith failure to thrive, behavioral deficits, and sudden infant death. The proper interpretation of infant polysomnography requires an understanding of normative data related to gestation and postconceptual age for apnea, arousal, and oxygenation. Direct visualization of the upper airway is an important diagnostic modality in infants with obstructive apnea. Treatment options for infant obstructive sleep apnea are predicated on the underlying etiology, including supraglottoplasty for severe laryngomalacia, mandibular distraction formicrognathia, tonsillectomy and/or adenoidectomy, choanal atresia repair, and/or treatment of gastroesophageal reflux. Copyright © 2012 by the American Thoracic Society.


Fulgham P.F.,Texas Health Presbyterian Dallas | Assimos D.G.,Wake forest University | Pearle M.S.,University of Texas Southwestern Medical Center | Preminger G.M.,Duke University
Journal of Urology | Year: 2013

Purpose: This technology assessment addresses the optimal use of imaging in the evaluation and treatment of patients with suspected or documented ureteral stones. Materials and Methods: A comprehensive literature search addressing 4 guiding questions was performed for full text in English articles published between January 1990 and July 2011. The search focused on major subtopics associated with the imaging of ureteral calculi, and included specific imaging modalities used in the diagnosis and management of ureteral calculous disease such as unenhanced (noncontrast) computerized tomography, conventional radiography, ultrasound, excretory urography, magnetic resonance imaging and nuclear medicine studies. Protocols (in the form of decision tree algorithms) were developed based on this literature review and in some instances on panel opinion. The 4 questions addressed were 1) What imaging study should be performed for suspected ureteral calculous disease? 2) What information should be obtained? 3) After diagnosis of a ureteral calculus, what followup imaging should be used? 4) After treatment of a ureteral calculus, what followup imaging studies should be obtained? Results: Based on these protocols, noncontrast computerized tomography is recommended to establish the diagnosis in most cases, with a low energy protocol advocated if body habitus is favorable. Conventional radiography and ultrasound are endorsed for monitoring the passage of most radiopaque stones as well as for most patients undergoing stone removal. Other studies may be indicated based on imaging findings, and patient, stone and clinical factors. Conclusions: The protocols generated assist the clinician in establishing the diagnosis of ureteral calculous disease, monitoring stone passage and following patients after treatment. The protocols take into account not only clinical effectiveness but also cost-effectiveness and risk/harm associated with the various imaging modalities. © 2013 American Urological Association Education and Research, Inc.


Gonzalez-Lima F.,University of Texas at Austin | Barksdale B.R.,University of Texas at Austin | Rojas J.C.,University of Texas Southwestern Medical Center
Biochemical Pharmacology | Year: 2014

This paper focuses on brain mitochondrial respiration as a therapeutic target for neuroprotection and cognitive enhancement. We propose that improving brain mitochondrial respiration is an important future direction in research and treatment of Alzheimer's disease (AD) and other conditions associated with cognitive impairment and neurodegeneration. The central thesis is that supporting and improving brain mitochondrial respiration constitutes a promising neurotherapeutic principle, with potential applications in AD as well as in a wide variety of neuropsychological conditions. We propose three different interventional approaches to improve brain mitochondrial respiration based on (a) pharmacology, (b) photobiomodulation and (c) nutrition interventions, and provide detailed examples for each type of intervention. First, low-dose USP methylene blue is described as a pharmacological intervention that can successfully increase mitochondrial respiration and result in memory enhancement and neuroprotection. Second, transcranial low-level light/laser therapy with near-infrared light is used to illustrate a photobiomodulation intervention with similar neurometabolic mechanisms of action as low-dose methylene blue. Finally, a nutrition intervention to improve mitochondrial respiration is proposed by increasing ketone bodies in the diet. The evidence discussed for each intervention supports a fundamental neurotherapeutic strategy based on improving oxidative energy metabolism while at the same time reducing the pro-oxidant tendencies of the nervous system. Targeting brain mitochondrial respiration with these three types of interventions is proposed as part of a holistic neurotherapeutic approach to improve brain energy metabolism and antioxidant defenses. This strategy represents a promising new bioenergetics direction for treatment of AD and other neuropsychological disorders featuring cognitive impairment and neurodegeneration. © 2013 Elsevier Inc. All rights reserved.


Neunert C.,University of Texas Southwestern Medical Center | Lim W.,McMaster University | Crowther M.,Worcestershire Royal Hospital | Cohen A.,Children's Hospital of Philadelphia | And 2 more authors.
Blood | Year: 2011

Immune thrombocytopenia (ITP) is commonly encountered in clinical practice. In 1996 the American Society of Hematology published a landmark guidance paper designed to assist clinicians in the management of this disorder. Since 1996 there have been numerous advances in the management of both adult and pediatric ITP. These changes mandated an update in the guidelines. This guideline uses a rigorous, evidence-based approach to the location, interpretation, and presentation of the available evidence. We have endeavored to identify, abstract, and present all available methodologically rigorous data informing the treatment of ITP. We provide evidence-based treatment recommendations using the GRADE system in those areas in which such evidence exists. We do not provide evidence in those areas in which evidence is lacking, or is of lower quality - interested readers are referred to a number of recent, consensus-based recommendations for expert opinion in these clinical areas. Our review identified the need for additional studies in many key areas of the therapy of ITP such as comparative studies of "front-line" therapy for ITP, the management of serious bleeding in patients with ITP, and studies that will provide guidance about which therapy should be used as salvage therapy for patients after failure of a first-line intervention. © 2011 by The American Society of Hematology.


Taniguchi M.,University of Texas Southwestern Medical Center | Carreira M.B.,University of Texas Southwestern Medical Center | Smith L.N.,University of Texas Southwestern Medical Center | Zirlin B.C.,University of Texas Southwestern Medical Center | And 2 more authors.
Neuron | Year: 2012

Chromatin remodeling by histone deacetylases (HDACs) is a key mechanism regulating behavioral adaptations to cocaine use. We report here that cocaine and cyclic adenosine monophosphate (cAMP) signaling induce the transient nuclear accumulation of HDAC5 in rodent striatum. We show that cAMP-stimulated nuclear import of HDAC5 requires a signaling mechanism that involves transient, protein phosphatase 2A (PP2A)-dependent dephosphorylation of a Cdk5 site (S279) found within the HDAC5 nuclear localization sequence. Dephosphorylation of HDAC5 increases its nuclear accumulation, by accelerating its nuclear import rate and reducing its nuclear export rate. Importantly, we show that dephosphorylation of HDAC5 S279 in the nucleus accumbens suppresses the development, but not expression, of cocaine reward behavior invivo. Together, our findings reveal a molecular mechanism by which cocaine regulates HDAC5 function to antagonize the rewarding impact of cocaine, likely by putting a brake on drug-stimulated gene expression that supports drug-induced behavioral changes. Taniguchi etal. discover that cocaine and cAMP signaling regulate the transient nuclear accumulation of the histone deacetylase HDAC5. Nuclear HDAC5 limits the development of cocaine reward-associated behaviors, likely by providing a braking mechanism for drug-induced gene expression programs. © 2012 Elsevier Inc.


Brown L.M.,University of North Carolina at Greensboro | Clegg D.J.,University of Texas Southwestern Medical Center
Journal of Steroid Biochemistry and Molecular Biology | Year: 2010

In recent years, obesity and its associated health disorders and costs have increased. Accumulation of adipose tissue, or fat, in the intra-abdominal adipose depot is associated with an increased risk of developing cardiovascular problems, type-2 diabetes mellitus, certain cancers, and other disorders like the metabolic syndrome. Males and females differ in terms of how and where their body fat is stored, in their hormonal secretions, and in their neural responses to signals regulating weight and body fat distribution. Men and post-menopausal women accumulate more fat in their intra-abdominal depots than pre-menopausal women, resulting in a greater risk of developing complications associated with obesity. The goal of this review is to discuss the current literature on sexual dimorphisms in body weight regulation, adipose tissue accrual and deposition. © 2009 Elsevier Ltd.


Holmes D.L.,University of Texas Southwestern Medical Center | Lancaster A.K.,Whitehead Institute For Biomedical Research | Lindquist S.,Whitehead Institute For Biomedical Research | Lindquist S.,Massachusetts Institute of Technology | And 2 more authors.
Cell | Year: 2013

Prion proteins undergo self-sustaining conformational conversions that heritably alter their activities. Many of these proteins operate at pivotal positions in determining how genotype is translated into phenotype. But the breadth of prion influences on biology and their evolutionary significance are just beginning to be explored. We report that a prion formed by the Mot3 transcription factor, [MOT3+], governs the acquisition of facultative multicellularity in the budding yeast Saccharomyces cerevisiae. The traits governed by [MOT3+] involved both gains and losses of Mot3 regulatory activity. [MOT3+]-dependent expression of FLO11, a major determinant of cell-cell adhesion, produced diverse lineage-specific multicellular phenotypes in response to nutrient deprivation. The prions themselves were induced by ethanol and eliminated by hypoxia - conditions that occur sequentially in the natural respiro-fermentative cycles of yeast populations. These data demonstrate that prions can act as environmentally responsive molecular determinants of multicellularity and contribute to the natural morphological diversity of budding yeast. PaperFlick: © 2013 Elsevier Inc.


Perello M.,Multidisc Institute Of Cell Biology Argentine Research Council And Scientific Research Commission | Zigman J.M.,University of Texas Southwestern Medical Center
Biological Psychiatry | Year: 2012

The peptide hormone ghrelin acts in the central nervous system as a potent orexigenic signal. Not only is ghrelin recognized as playing an important role in feeding circuits traditionally thought of as affecting body weight homeostasis, but also an accumulating number of scientific studies have identified ghrelin as being a key regulator of reward-based, hedonic eating behaviors. In the current article, we review ghrelin's orexigenic actions, the evidence linking ghrelin to food reward behavior, potential mechanisms by which ghrelin mediates reward-based eating behavior, and those studies suggesting an obligatory role for ghrelin in the changed eating behaviors induced by stress. © 2012 Society of Biological Psychiatry.


Sharifi N.,Cleveland Clinic | Sharifi N.,University of Texas Southwestern Medical Center
Molecular Endocrinology | Year: 2013

The decades-old terminology of androgen independence has been replaced in recent years with castration-resistant prostate cancer. Biological and clinical evidence have together conspired to support the use of this revised terminology by demonstrating that in the vast majority of cases tumors are neither truly depleted of androgens, nor are they free of the requirement for androgens to sustain growth and progression. Abiraterone acetate, an androgen synthesis inhibitor, and enzalutamide, a potent androgen receptor antagonist, both exploit the continued requirement for androgens. A central question, given the therapeutic gains enabled by further suppression of the androgen axis with these newer agents, is whether there may be additional clinical benefit gained by moving the goal posts of androgen suppression even further. The answer lies in part with the mechanisms utilized by tumors that enable resistance to these therapies. The aims of this review were to give a broad outline of steroidogenesis in prostate cancer and to highlight recent developments in understanding resistance to hormonal therapies. © 2013 by The Endocrine Society.


Carr C.M.,Texas A&M University | Rizo J.,University of Texas Southwestern Medical Center
Current Opinion in Cell Biology | Year: 2010

Sec1/Munc18 (SM) proteins bind to and function with soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) at each vesicle fusion site in the cell. The purpose for these interactions is becoming clearer, as what had been interpreted as functional divergence between SM proteins acting at different vesicle trafficking steps, or in specialized cells, is giving way to more recent evidence for common functions among all SM proteins. What is emerging is a picture of SM proteins acting not merely as SNARE regulators, but also as central components of the membrane fusion apparatus. The available data suggest sequential models that describe how the soluble SM protein might first regulate SNARE complex assembly and then cooperate with SNAREs to stimulate membrane fusion. © 2010 Elsevier Ltd.


Yanagisawa H.,University of Texas Southwestern Medical Center | Davis E.C.,McGill University
International Journal of Biochemistry and Cell Biology | Year: 2010

Evolution of elastic fibers is associated with establishment of the closed circulation system. Primary roles of elastic fibers are to provide elasticity and recoiling to tissues and organs and to maintain the structural integrity against mechanical strain over a lifetime. Elastic fibers are comprised of an insoluble elastin core and surrounding mantle of microfibrils. Elastic fibers are formed in a regulated, stepwise manner, which includes the formation of a microfibrillar scaffold, deposition and integration of tropoelastin monomers into the scaffold, and cross-linking of the monomers to form an insoluble, functional polymer. In recent years, an increasing number of glycoproteins have been identified and shown to be located on or surrounding elastic fibers. Among them, the short fibulins-3, -4 and -5 particularly drew attention because of their potent elastogenic activity. Fibulins-3, -4 and -5 are characterized by tandem repeats of calcium binding EGF-like motifs and a C-terminal fibulin module, which is conserved throughout fibulin family members. Initial biochemical characterization and gene expression studies predicted that fibulins might be involved in structural support and/or matrix-cell interactions. Recent analyses of short fibulin knockout mice have revealed their critical roles in elastic fiber development in vivo. We review recent findings on the elastogenic functions of short fibulins and discuss the molecular mechanism underlying their activity in vitro and in vivo. © 2010 Elsevier Ltd.


Gilbreath J.J.,Uniformed Services University of the Health Sciences | Cody W.L.,University of Texas Southwestern Medical Center | Merrell D.S.,Uniformed Services University of the Health Sciences | Hendrixson D.R.,University of Texas Southwestern Medical Center
Microbiology and Molecular Biology Reviews | Year: 2011

Microbial evolution and subsequent species diversification enable bacterial organisms to perform common biological processes by a variety of means. The epsilonproteobacteria are a diverse class of prokaryotes that thrive in diverse habitats. Many of these environmental niches are labeled as extreme, whereas other niches include various sites within human, animal, and insect hosts. Some epsilonproteobacteria, such as Campylobacter jejuni and Helicobacter pylori, are common pathogens of humans that inhabit specific regions of the gastrointestinal tract. As such, the biological processes of pathogenic Campylobacter and Helicobacter spp. are often modeled after those of common enteric pathogens such as Salmonella spp. and Escherichia coli. While many exquisite biological mechanisms involving biochemical processes, genetic regulatory pathways, and pathogenesis of disease have been elucidated from studies of Salmonella spp. and E. coli, these paradigms often do not apply to the same processes in the epsilonproteobacteria. Instead, these bacteria often display extensive variation in common biological mechanisms relative to those of other prototypical bacteria. In this review, five biological processes of commonly studied model bacterial species are compared to those of the epsilonproteobacteria C. jejuni and H. pylori. Distinct differences in the processes of flagellar biosynthesis, DNA uptake and recombination, iron homeostasis, interaction with epithelial cells, and protein glycosylation are highlighted. Collectively, these studies support a broader view of the vast repertoire of biological mechanisms employed by bacteria and suggest that future studies of the epsilonproteobacteria will continue to provide novel and interesting information regarding prokaryotic cellular biology. Copyright © 2011, American Society for Microbiology. All Rights Reserved.


Troutman T.D.,University of Texas Southwestern Medical Center | Bazan J.F.,NeuroScience Inc. | Pasare C.,University of Texas Southwestern Medical Center
Cell Cycle | Year: 2012

TLRs are a family of pattern recognition receptors that recognize conserved molecular structures/products from a wide variety of microbes.1,2Following recognition of ligands, TLRs recruit signaling adapters to initiate a pro-inflammatory signaling cascade culminating in the activation of several transcription factor families. Additionally, TLR signals lead to activation of PI3K, affecting many aspects of the cellular response, including cell survival, proliferation and regulation of the pro-inflammatory response.3-10 The recent discovery of BCAP as a TLR signaling adaptor, crucial for linking TLRs to PI3K activation, allows new questions of the importance of PI3K activation downstream of TLRs. Here, we summarize the current understanding of signaling pathways activated by TLRs and provide our perspective on TLR mediated activation of PI3K and its impact on regulating cellular processes. © 2012 Landes Bioscience.


Zhu C.,University of Texas Southwestern Medical Center | Li G.,University of Texas Southwestern Medical Center | Ess D.H.,Brigham Young University | Falck J.R.,University of Texas Southwestern Medical Center | Kurti L.,University of Texas Southwestern Medical Center
Journal of the American Chemical Society | Year: 2012

Herein, we disclose the first metal-free synthesis of primary aromatic amines from arylboronic acids, a reaction that has eluded synthetic chemists for decades. This remarkable transformation affords structurally diverse primary arylamines in good chemical yields, including a variety of halogenated primary anilines that often cannot be prepared via transition-metal-catalyzed amination. The reaction is operationally simple, requires only a slight excess of aminating agent, proceeds under neutral or basic conditions, and, importantly, can be scaled up to provide multigram quantities of primary anilines. Density functional calculations reveal that the most likely mechanism involves a facile 1,2-aryl migration and that the presence of an ortho nitro group in the aminating agent plays a critical role in lowering the free energy barrier of the 1,2-aryl migration step. © 2012 American Chemical Society.


Wang J.,University of Texas Southwestern Medical Center | Wang J.,Lady Davis Institute for Medical Research | Pantopoulos K.,University of Texas Southwestern Medical Center | Pantopoulos K.,Lady Davis Institute for Medical Research | Pantopoulos K.,McGill University
Biochemical Journal | Year: 2011

Iron is an essential but potentially hazardous biometal. Mammalian cells require sufficient amounts of iron to satisfy metabolic needs or to accomplish specialized functions. Iron is delivered to tissues by circulating transferrin, a transporter that captures iron released into the plasma mainly from intestinal enterocytes or reticuloendothelial macrophages. The binding of iron-laden transferrin to the cell-surface transferrin receptor 1 results in endocytosis and uptake of the metal cargo. Internalized iron is transported to mitochondria for the synthesis of haem or iron-sulfur clusters, which are integral parts of several metalloproteins, and excess iron is stored and detoxified in cytosolic ferritin. Iron metabolism is controlled at different levels and by diverse mechanisms. The present review summarizes basic concepts of iron transport, use and storage and focuses on the IRE (iron-responsive element)/IRP (iron-regulatory protein) system, a well known post-transcriptional regulatory circuit that not only maintains iron homoeostasis in various cell types, but also contributes to systemic iron balance. © 2010 The Author(s).


Singal A.G.,University of Texas Southwestern Medical Center | Singal A.G.,Southwestern University | Pillai A.,Emory University | Tiro J.,Southwestern University | Tiro J.,University of Texas Southwestern Medical Center
PLoS Medicine | Year: 2014

Background:Surveillance for hepatocellular carcinoma (HCC) has level I evidence among patients with hepatitis B but only level II evidence in patients with cirrhosis. This lack of randomized data has spurred questions regarding the utility of HCC surveillance in this patient population; however, lack of randomized data does not equate to a lack of data supporting the efficacy of surveillance. The aim of our study was to determine the effect of HCC surveillance on early stage tumor detection, receipt of curative therapy, and overall survival in patients with cirrhosis.Methods and Findings:We performed a systematic literature review using Medline from January 1990 through January 2014 and a search of national meeting abstracts from 2009-2012. Two investigators identified studies that reported rates of early stage tumor detection, curative treatment receipt, or survival, stratified by HCC surveillance status, among patients with cirrhosis. Both investigators independently extracted data on patient populations, study methods, and results using standardized forms. Pooled odds ratios, according to HCC surveillance status, were calculated for each outcome using the DerSimonian and Laird method for a random effects model.We identified 47 studies with 15,158 patients, of whom 6,284 (41.4%) had HCC detected by surveillance. HCC surveillance was associated with improved early stage detection (odds ratio [OR] 2.08, 95% CI 1.80-2.37) and curative treatment rates (OR 2.24, 95% CI 1.99-2.52). HCC surveillance was associated with significantly prolonged survival (OR 1.90, 95% CI 1.67-2.17), which remained significant in the subset of studies adjusting for lead-time bias. Limitations of current data included many studies having insufficient duration of follow-up to assess survival and the majority not adjusting for liver function or lead-time bias.Conclusions:HCC surveillance is associated with significant improvements in early tumor detection, receipt of curative therapy, and overall survival in patients with cirrhosis.Please see later in the article for the Editors' Summary. © 2014 Singal et al.


Casey B.,University of Texas Southwestern Medical Center | De Veciana M.,Eastern Virginia Medical School
American Journal of Obstetrics and Gynecology | Year: 2014

The adverse impact of overt hypothyroidism that complicates pregnancy outcomes is well-established and not debated. For more than a decade, however, endocrinologists and obstetricians have been debating whether screening for subclinical thyroid disorders during pregnancy should be routine or should continue to be based on symptoms and risk factors. Several observational studies have suggested that offspring of women with asymptomatic thyroid dysfunction were at increased risk for impaired neurodevelopment. Other studies have suggested that pregnant women with subclinical thyroid disease, particularly those identified with an elevated thyroid-stimulating hormone (TSH) level may be at increased risk for pregnancy complications such as fetal death, preterm birth, or placental abruption. These data have prompted both obstetric and endocrinologic professional societies to draft recommendations regarding screening for thyroid disease during pregnancy, some of which are not entirely based on available evidence. The prevalence of overt thyroid disease is estimated to be 1-2 per 1000 pregnancies and historically has not been considered high enough to justify routine screening. Lower TSH thresholds (>2.5 mU/L) for the diagnosis of hypothyroidism have been promoted, and women with subclinical thyroid dysfunction commonly are included in estimates of thyroid disease during pregnancy, both of which exaggerate prevalence rates. The most compelling recent evidence on this issue has come from the Controlled Antenatal Thyroid Screening trial. After almost 22,000 pregnant women were screened for either isolated high TSH or isolated low free thyroxine level, 390 children of treated women with either diagnosis were compared with 404 children of similar women who were not treated during pregnancy. Treatment had no effect on mean offspring IQ at age 3 years or the number of children with an IQ <85. Authors of this landmark study concluded that antenatal screening and maternal treatment for women with subclinical thyroid dysfunction did not result in improved cognitive function. An ongoing intervention trial conducted by the Eunice Kennedy-Shriver National Institute of Child Health and Human Development's Maternal-Fetal Medicine Units Network will provide further clarity to this important question. In the interim, the debating authors have concluded, after careful review of the currently published literature, that routine screening for subclinical thyroid dysfunction during pregnancy is not currently warranted at this time. © 2014 Elsevier Inc. All rights reserved.


Goldstein J.L.,NorthShore University Health System | Cryer B.,University of Texas Southwestern Medical Center
Drug, Healthcare and Patient Safety | Year: 2015

Nonsteroidal anti-inflammatory drugs (NSAIDs) are effective anti-inflammatory and analgesic agents and are among the most commonly used classes of medications worldwide. However, their use has been associated with potentially serious dose-dependent gastrointestinal (GI) complications such as upper GI bleeding. GI complications resulting from NSAID use are among the most common drug side effects in the United States, due to the widespread use of NSAIDs. The risk of upper GI complications can occur even with short-term NSAID use, and the rate of events is linear over time with continued use. Although gastroprotective therapies are available, they are underused, and patient and physician awareness and recognition of some of the factors influencing the development of NSAID-related upper GI complications are limited. Herein, we present a case report of a patient experiencing a gastric ulcer following NSAID use and examine some of the risk factors and potential strategies for prevention of upper GI mucosal injuries and associated bleeding following NSAID use. These risk factors include advanced age, previous history of GI injury, and concurrent use of medications such as anticoagulants, aspirin, corticosteroids, and selective serotonin reuptake inhibitors. Strategies for prevention of GI injuries include anti-secretory agents, gastroprotective agents, alternative NSAID formulations, and nonpharmacologic therapies. Greater awareness of the risk factors and potential therapies for GI complications resulting from NSAID use could help improve outcomes for patients requiring NSAID treatment. © 2015 Goldstein and Cryer.


Gao H.,University of Texas Southwestern Medical Center | Xu Q.-L.,University of Texas Southwestern Medical Center | Yousufuddin M.,University of Texas at Arlington | Ess D.H.,Brigham Young University | Kurti L.,University of Texas Southwestern Medical Center
Angewandte Chemie - International Edition | Year: 2014

We disclose an efficient and operationally simple protocol for the preparation of fused N-heterocycles starting from readily available 2-nitrobiaryls and PhMgBr under mild conditions. More than two dozen N-heterocycles, including two bioactive natural products, have been synthesized using this method. A stepwise electrophilic aromatic cyclization mechanism was proposed by DFT calculations. Controlled fusion: A transition-metal-free, low-temperature, and regioselective intramolecular amination of aromatic C(sp2)-H bonds provides fused N-heterocycles. This reaction is operationally simple and scalable (1-10 mmol) and the scope of substrates is wide (see scheme). Density functional calculations indicate that a stepwise electrophilic aromatic cyclization mechanism may be operative. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


McMurray J.J.V.,University of Glasgow | Packer M.,University of Texas Southwestern Medical Center | Desai A.S.,Brigham and Women's Hospital | Gong J.,Novartis | And 8 more authors.
New England Journal of Medicine | Year: 2014

Background We compared the angiotensin receptor-neprilysin inhibitor LCZ696 with enalapril in patients who had heart failure with a reduced ejection fraction. In previous studies, enalapril improved survival in such patients.Methods: In this double-blind trial, we randomly assigned 8442 patients with class II, III, or IV heart failure and an ejection fraction of 40% or less to receive either LCZ696 (at a dose of 200 mg twice daily) or enalapril (at a dose of 10 mg twice daily), in addition to recommended therapy. The primary outcome was a composite of death from cardiovascular causes or hospitalization for heart failure, but the trial was designed to detect a difference in the rates of death from cardiovascular causes.Results: The trial was stopped early, according to prespecified rules, after a median followup of 27 months, because the boundary for an overwhelming benefit with LCZ696 had been crossed. At the time of study closure, the primary outcome had occurred in 914 patients (21.8%) in the LCZ696 group and 1117 patients (26.5%) in the enalapril group (hazard ratio in the LCZ696 group, 0.80; 95% confidence interval [CI], 0.73 to 0.87; P< 0.001). A total of 711 patients (17.0%) receiving LCZ696 and 835 patients (19.8%) receiving enalapril died (hazard ratio for death from any cause, 0.84; 95% CI, 0.76 to 0.93; P< 0.001); of these patients, 558 (13.3%) and 693 (16.5%), respectively, died from cardiovascular causes (hazard ratio, 0.80; 95% CI, 0.71 to 0.89; P< 0.001). As compared with enalapril, LCZ696 also reduced the risk of hospitalization for heart failure by 21% (P< 0.001) and decreased the symptoms and physical limitations of heart failure (P = 0.001). The LCZ696 group had higher proportions of patients with hypotension and nonserious angioedema but lower proportions with renal impairment, hyperkalemia, and cough than the enalapril group.Conclusions: LCZ696 was superior to enalapril in reducing the risks of death and of hospitalization for heart failure. Copyright © 2014 Massachusetts Medical Society.


News Article | November 28, 2016
Site: www.latimes.com

Without sustaining a single concussion, a North Carolina high school football team showed worrisome brain changes after a single season of play, a new study has shown. A detailed effort to capture the on-field experiences of 24 high school football players showed that, at the end of a single season of play, teammates whose heads sustained the most frequent contact with other moving bodies had the most pronounced changes in several measures of brain health. The new research findings, presented Monday at a meeting of the Radiological Society of North America, are a key first step in discovering how the developing brain responds to the repeated blows that come with youth football. Researchers underscored that they don’t know yet whether the young brains they studied might recover completely in the off-season, or whether their expected developmental trajectory might be knocked off-course by the impacts. Only more long-term research on a broader swath of players will tell, said the new study’s authors, who come from the University of Texas Southwestern Medical Center. Those studies are already underway. Across the country, neuroscientists and sports-injury experts have lashed together their efforts to conduct brain-imaging research on participants in youth contact sports. In the course of the high school football season just completed, the resulting consortium conducted studies that will supplement the University of Texas Southwestern Medical Center findings. "It's important to understand the potential changes occurring in the brain related to youth contact sports," said Elizabeth Moody Davenport, a postdoctoral researcher at University of Texas Southwestern Medical Center in Dallas who led the latest study. "We know that some professional football players suffer from a serious condition called chronic traumatic encephalopathy, or CTE,” said Davenport. “We are attempting to find out when and how that process starts, so that we can keep sports a healthy activity for millions of children and adolescents," she added. In the study presented Monday, researchers focused on 24 members of a single high school football squad, wiring up each player’s helmet with six accelerometers. During practices as well as play, those sensors gathered detailed data on the location, strength of impact and direction of any blows to a player’s head. Called the Head Impact Telemetry System, or HITS, this suite of sensors allowed researchers to record exactly how many and what types of blows to the head a given player had over the course of a season, as well as how forceful they were. Data from the helmets were uploaded to a computer for analysis. The study’s 24 subjects were 17 years old on average. None had ever had a diagnosed concussion, and no concussions were seen in the course of the team’s season. When they have detailed before-and-after brain imaging studies, researchers can use the HITS results to infer links between certain kinds of brain impacts and observed brain changes. Before the season began as well as after it ended, the North Carolina players underwent brain-imaging scans that measured the density of key brain structures and analyzed certain patterns of electrical activity generated by their working brain cells. In two different kinds of specialized MRI scans, scientists sought to measure the integrity of the brain’s “white matter” — the bundles of insulated wiring that carry electrical signals both among cells and between groups of cells on either side of the brain’s two hemispheres. Earlier research has suggested that blows to the head that cause the brain to move side-to-side in its skeletal casing — such as those sustained by boxers — can twist and shear these bundles of connective tissue and disrupt their ability to conduct electrical signals efficiently. A second type of scan, a magnetoencephalography (MEG) scan, recorded and analyzed the magnetic fields produced by brain waves. Among the many patterns they might listen for, researchers were attuned to Delta waves, a form of slow-wave activity described by Davenport as “a type of distress signal" emitted by the injured brain. The scans showed that the condition of the brain’s white matter was most affected in players whose helmets had recorded the most linear acceleration — the kind of direct frontal hit a receiver might get when returning a kick, or that a lineman might sustain repeatedly when lunging forward on every play. Changes on the MEG scan were most often seen in players whose helmets showed they had sustained the most rotational impact. In rotational impact, the brain twists within the skull, usually in response to a sideways impact. Boxers, who receive punches to the side of the head, often show the effects of rotational impacts. In football, the brain of a quarterback or ball-runner who is tackled laterally will likely withstand strong rotational forces. Davenport said such results “demonstrate that you need both” brain imaging that measures changes in the brain’s structure and imaging that detects changes in its workings. Different kinds of brain changes are linked to “very different” kinds of impacts, she noted. And as researchers begin to recognize whether long-term behavioral effects — say, depression, memory loss or movement disturbances — come with certain brain changes, they can begin to protect players accordingly. As important, given the North Carolina team’s concussion-free season, is that it might be important to have both types of brain imaging to discern what effects repeated sub-concussive blows to the head might have, Davenport and her colleagues said. Mounting evidence already suggests that sustaining multiple concussions puts a person at higher risk of later depression, memory loss and other neuropsychiatric problems. But researchers suspect that even when they don’t result in a concussion diagnosis, repeated blows to the head may be harmful as well. More, and more sensitive, tests will help reveal whether that is so. Follow me on Twitter @LATMelissaHealy and "like" Los Angeles Times Science & Health on Facebook.


News Article | October 5, 2016
Site: www.nature.com

Molecular biologist Yoshinori Ohsumi has won the 2016 Nobel Prize in Physiology or Medicine for his work in the field of autophagy: the processes by which the cell digests and recycles its own components. The 71-year-old Ohsumi, who is currently a professor at the Tokyo Institute of Technology in Yokohama, was recognized for his experiments in the 1990s, when he used baker's yeast (Saccharomyces cerevisiae) to identify genes that control how cells destroy their own contents. The same kinds of mechanism operate in human cells — and are sometimes involved in genetic disease. "He's a very humble yeast geneticist who basically transformed the field," says Sharon Tooze, a cell biologist at the Francis Crick Institute in London. "He was interested in this weird pathway that turns out to be a vitally important pathway in medicine." The word 'autophagy' — from the Greek for 'self-eating' — was coined in 1963 by the Belgian biochemist Christian de Duve, who saw how cells broke down their parts inside a waste-processing sac that he called a lysosome. Biologists now understand that this process is fundamentally important to living cells. "Without autophagy our cells won't survive," says Juleen Zierath, a physiologist at the Karolinska Institute in Stockholm who was on the selection committee for the medicine Nobel. When cells are starved, they can consume their own proteins for fuel. The same degradation process can be used to eliminate damaged proteins and organelles — effectively, to renew cells and clear out debris — or to ward off invading bacteria and viruses. Ohsumi began studying yeast as a postdoc, turning to yeast DNA replication as a side project when his main research stalled, says Tooze. When Ohsumi first started studying autophagy in 1988, “it was kind of a sleepy backwater of a research topic,” says biochemist Michael Hall of the University of Basel in Switzerland. “It was basically considered the garbage-disposal system of the cell — just bulk, non-specific degradation of junk.” In an interview given to the Tokyo Institute of Technology's website in December 2012, Ohsumi said that all his research findings began with a love of the microscope. "You can answer the most basic and important questions about the nature of life through yeasts," he added. Ohsumi would go on to develop the first yeast genetics screen to identify genes involved in the autophagy pathway. But it was a few years before biologists recognized the importance of the process in physiology and disease. Interest in the field skyrocketed when, in 1999, Beth Levine (now at the University of Texas Southwestern Medical Center in Dallas) and her colleagues reported that a mammalian autophagy gene could suppress tumour growth. That finding launched widespread efforts to learn more about the role of autophagy in cancer. Disruptions in autophagy have also been linked to Parkinson's disease, type 2 diabetes and other disorders — and research is ongoing to develop drugs that can affect the process. Researchers’ understanding of the complex role of autophagy in cancer has become more detailed: the process seems to inhibit tumours in the early stages of their growth, but can also fuel cancer once it has spread, says Hall. Ohsumi, who will collect 8 million Swedish kronor (US$940,000) for the Nobel prize, also won the ¥50-million (US$626,000) Kyoto Prize in basic sciences in 2012 for his autophagy work. Others have made key contributions to the field, and were considered to be contenders for a share of a Nobel. Biochemist Michael Thumm of the University Medical Center Göttingen in Germany, for example, also discovered autophagy genes, as did cell biologist Daniel Klionsky of the University of Michigan in Ann Arbor. “If they’re going to give it to just one, Ohsumi’s the one,” says Hall. “But it also would have been good to include other people.” In Japan, the prize had been widely anticipated for the past few years, with journalists showing up regularly to ask Ohsumi for interviews, says Hitoshi Nakatogawa, a biologist at the Tokyo Institute of Technology who has worked with Ohsumi for a decade. When colleagues heard — around two hours before the official announcement — of Ohsumi’s win, they gathered together to celebrate in the victor’s lab. “We talked about how great it was that he won it alone,” he says. “Ohsumi never overlooks anything even in the most banal kind of experiment,” Nakatogawa adds. “He doesn't care about whether it will lead to something useful, whether a breakthrough can be expected, whether it will lead to more funding. He just follows his curiosity.”


Bezprozvanny I.,University of Texas Southwestern Medical Center | Bezprozvanny I.,Saint Petersburg State Polytechnic University
Science Signaling | Year: 2013

Mutations in presenilins result in familial Alzheimer's disease (FAD). Presenilins encode a catalytic subunit of the γ-secretase complex, and FAD mutations in presenil-ins alter γ-secretase activity. Many FAD mutations in presenilins also affect intracel-lular calcium signaling. To explain these results, it was proposed that presenilins also function as endoplasmic reticulum (ER) calcium leak channels and that this function is disrupted by FAD mutations. Although this hypothesis has been controversial, new research supports the calcium leak channel hypothesis. One group reported the presence of putative ion-conduction pore in the high-resolution crystal structure of bacterial presenilin homolog PSH1. Another group identifi ed an essential role for presenilins in mediating ER calcium leak in a cell-based screen for calcium homeostasis modulators. These results should enable the fi eld to move forward and to focus on exploring connections between FAD mutations in presenilins, changes in γ-secretase and ER Ca2+ leak functions, and development of the disease. Copyright 2008 American Association for the Advancement of Science. All Rights Reserved.


Taurog J.D.,University of Texas Southwestern Medical Center | Chhabra A.,University of Texas Southwestern Medical Center | Colbert R.A.,U.S. National Institutes of Health
New England Journal of Medicine | Year: 2016

Chronic back pain is common worldwide and is cared for by a variety of providers, but specific, satisfactory treatment is often lacking. Ankylosing spondylitis, an inflammatory disorder that in its extreme form can lead to the bony fusion of vertebral joints, is an uncommon but well-established cause of chronic back pain. During the past decade, ankylosing spondylitis has come to be considered as a subset of the broader and more prevalent diagnostic entity referred to as axial spondyloarthritis. The estimated prevalence of axial spondyloarthritis in the United States is 0.9 to 1.4% of the adult population, similar to that of rheumatoid arthritis.1 Axial spondyloarthritis is generally diagnosed and treated by rheumatologists, and there is specific treatment for it. However, prolonged delay in reaching the diagnosis is common and is usually the result of the failure of recognition by nonrheumatologists.2 This review is intended to enhance awareness and understanding of axial spondyloarthritis and ankylosing spondylitis - and the relationship between the two - in order to facilitate prompt and accurate diagnosis and proper treatment. Recent advances in our understanding and treatment of these conditions are discussed. Copyright © 2016 Massachusetts Medical Society.


Reuben A.,Medical University of South Carolina | Koch D.G.,Medical University of South Carolina | Lee W.M.,University of Texas Southwestern Medical Center
Hepatology | Year: 2010

Acute liver failure (ALF) due to drug-induced liver injury (DILI), though uncommon, is a concern for both clinicians and patients. The Acute Liver Failure Study Group has prospectively collected cases of all forms of acute liver failure since 1998. We describe here cases of idiosyncratic DILI ALF enrolled during a 10.5-year period. Data were collected prospectively, using detailed case report forms, from 1198 subjects enrolled at 23 sites in the United States, all of which had transplant services. A total of 133 (11.1%) ALF subjects were deemed by expert opinion to have DILI; 81.1% were considered highly likely, 15.0% probable, and 3.8% possible. Subjects were mostly women (70.7%) and there was overrepresentation of minorities for unclear reasons. Over 60 individual agents were implicated, the most common were antimicrobials (46%). Transplant-free (3-week) survival was poor (27.1%), but with highly successful transplantation in 42.1%, overall survival was 66.2%. Transplant-free survival in DILI ALF is determined by the degree of liver dysfunction, specifically baseline levels of bilirubin, prothrombin time/international normalized ratio, and Model for End-Stage Liver Disease scores. Conclusion: DILI is an uncommon cause of ALF that evolves slowly, affects a disproportionate number of women and minorities, and shows infrequent spontaneous recovery, but transplantation affords excellent survival. Copyright © 2010 American Association for the Study of Liver Diseases.


Mayorga C.A.,University of Texas Southwestern Medical Center | Rockey D.C.,Medical University of South Carolina
Clinical Gastroenterology and Hepatology | Year: 2013

Background & Aims: Recent reductions in mortality after acute upper gastrointestinal hemorrhage among patients with cirrhosis have been attributed to early and aggressive use of guideline-recommended pharmacologic agents, antibiotics, and endoscopic therapy. Studies have shown, however, that adherence to recommended guidelines is low. We investigated whether use of a standardized electronic order set would improve adherence to treatment and timeliness of delivery. Methods: We performed a prospective observational study, implementing an electronic order set for 123 patients with known or suspected cirrhosis who presented with symptoms/signs of upper gastrointestinal hemorrhage at Parkland Memorial Hospital (in Dallas, TX) from July 2011 through June 2012. The order set included standard nursing orders, laboratory tests, medications, orders for consultative services, and a brief evidence-based review of the benefits of octreotide and antibiotics in patients with cirrhosis. Primary outcomes included overall adherence to the administration of octreotide and antibiotics and the performance of upper endoscopy, as well as time to these interventions. Results: Administration of antibiotics increased in patients for whom the order set was used (100% vs 89% for whom it was not used; P= .01); the use of the order set significantly reduced the time to administration of antibiotics (3 h 28 min vs 10 h 4 min; P < .001). The time to administration of octreotide also significantly was reduced for patients for whom the order set was used (2 h 16 min vs 6 h 21 min; P < .002). Although all patients underwent endoscopy, there was no significant difference in the time to procedure between patients for whom the order set was used and not used (17 h 54 min vs 18 h 5 min; P= .95). Conclusions: The use of a standardized electronic order set improved not only overall adherence, but also the timeliness of administration of recommended therapies for patients with known or suspected cirrhosis presenting with upper gastrointestinal hemorrhage. © 2013 AGA Institute.


Egorova P.,Saint Petersburg State Polytechnic University | Popugaeva E.,Saint Petersburg State Polytechnic University | Bezprozvanny I.,University of Texas Southwestern Medical Center
Seminars in Cell and Developmental Biology | Year: 2015

Neurodegenerative disorders, such as spinocerebellar ataxias (SCAs) and Alzheimer's disease (AD) represent a huge scientific and medical question, but the molecular mechanisms of these diseases are still not clear. There is increasing evidence that neuronal calcium signaling is abnormal in many neurodegenerative disorders. Abnormal neuronal calcium release from the endoplasmic reticulum may result in disturbances of cell homeostasis, synaptic dysfunction, and eventual cell death. Neuronal loss is observed in most cases of neurodegenerative diseases. Recent experimental evidence supporting the role of neuronal calcium signaling in the pathogenesis of SCAs and AD is discussed in this review. © 2015 Elsevier Ltd.


Fernando H.C.,Boston Medical Center | Timmerman R.,University of Texas Southwestern Medical Center
Journal of Thoracic and Cardiovascular Surgery | Year: 2012

During the past decade, tremendous interest has arisen in the use of nonoperative therapies for patients with non-small cell lung cancer. Of these therapies, stereotactic body radiotherapy has become established as an effective modality for treating peripheral cancer in medically inoperable patients. Toxicity is low, and the treatment is effective, with excellent local control rates. Several investigators have suggested that stereotactic body radiotherapy could be effective for high-risk operable patients (usually treated with sublobar resection) and even perhaps for standard-risk operable patients (usually treated with lobectomy); however, this is less accepted. A direct comparison of stereotactic body radiotherapy and sublobar resection is difficult for a number of reasons. These include different definitions of recurrence, different populations of patients in these studies (with those undergoing stereotactic body radiotherapy tending to be the medically inoperable group), and different methods of classifying morbidity in the surgical and radiation oncology studies. Imaging follow-up has also not been standardized among the studies. Thus, a randomized study is necessary and timely. Investigators from the American College of Surgeons Oncology Group and the Radiation Therapy and Oncology Group have collaborated to develop a phase III randomized study comparing stereotactic body radiotherapy and sublobar resection (with or without brachytherapy) for high-risk operable patients with non-small cell lung cancer. This study (American College of Surgeons Oncology Group Z4099/Radiation Therapy Oncology Group 1021) has recently opened for accrual. It is hoped that this will help to better define the role of these therapies for patients with non-small cell lung cancer. Copyright © 2012 by The American Association for Thoracic Surgery.


Monteggia L.M.,University of Texas Southwestern Medical Center | Zarate C.,U.S. National Institutes of Health
Current Opinion in Neurobiology | Year: 2015

In the past decade the emergence of glutamate N-methyl- d-aspartate (NMDA) receptor blockers such as ketamine as fast-acting antidepressants fostered a major conceptual advance by demonstrating the possibility of a rapid antidepressant response. This discovery brings unique mechanistic insight into antidepressant action, as there is a substantial amount of basic knowledge on glutamatergic neurotransmission and how blockade of NMDA receptors may elicit plasticity. The combination of this basic knowledge base and the growing clinical findings will facilitate the development of novel fast acting antidepressants. © 2014 Elsevier Ltd.


Sephton C.F.,Laval University | Yu G.,University of Texas Southwestern Medical Center
Cellular and Molecular Life Sciences | Year: 2015

The loss of synapses is a central event in neurodegenerative diseases. Synaptic proteins are often associated with disease neuropathology, but their role in synaptic loss is not fully understood. Of the many processes involved in sustaining the integrity of synapses, local protein translation can directly impact synaptic formation, communication, and maintenance. RNA-binding proteins and their association with RNA granules serve to regulate mRNA transportation and translation at synapses and in turn regulate the synapse. Genetic mutations in RNA-binding proteins FUS and TDP-43 have been linked with causing neurodegenerative diseases: amyotrophic lateral sclerosis and frontotemporal dementia. The observation that mutations in FUS and TDP-43 coincide with changes in RNA granules provides evidence that dysfunction of RNA metabolism may underlie the mechanism of synaptic loss in these diseases. However, we do not know how mutations in RNA-binding proteins would affect RNA granule dynamics and local translation, or if these alterations would cause neurodegeneration. Further investigation into this area will lead to important insights into how disruption of RNA metabolism and local translation at synapses can cause neurodegenerative diseases. © 2015 The Author(s).


Uchida A.,University of Texas Southwestern Medical Center | Zigman J.M.,University of Texas Southwestern Medical Center | Perello M.,CONICET
Frontiers in Neuroscience | Year: 2013

Ghrelin is an octanoylated peptide hormone, produced by endocrine cells of the stomach, which acts in the brain to increase food intake and body weight. Our understanding of the mechanisms underlying ghrelin's effects on eating behaviors has been greatly improved by the generation and study of several genetically manipulated mouse models. These models include mice overexpressing ghrelin and also mice with genetic deletion of ghrelin, the ghrelin receptor [the growth hormone secretagogue receptor (GHSR)] or the enzyme that post-translationally modifies ghrelin [ghrelin O-acyltransferase (GOAT)]. In addition, a GHSR-null mouse model in which GHSR transcription is globally blocked but can be cell-specifically reactivated in a Cre recombinase-mediated fashion has been generated. Here, we summarize findings obtained with these genetically manipulated mice, with the aim to highlight the significance of the ghrelin system in the regulation of both homeostatic and hedonic eating, including that occurring in the setting of chronic psychosocial stress. © 2013 Uchida, Zigman and Perelló.


Becker P.M.,University of Texas Southwestern Medical Center | Sharon D.,Tulane University
Journal of Clinical Psychiatry | Year: 2014

Objective: Restless legs syndrome (RLS), also known as Willis-Ekbom disease, is a sensorimotor disorder that can result in considerable sleep disruption. This narrative review provides an overview of RLS diagnosis and reports epidemiologic evidence for an association between RLS and mood disorders. Possible links between RLS, sleep disturbances, and mood disorders are considered, and theoretical pathophysiologic pathways are discussed. Finally, pharmacologic therapies for RLS are summarized. Data Sources: A PubMed search was performed using the search term restless legs syndrome in combination with affective/anxiety, antidepressants, anxiety/anxiety disorder, attention deficit hyperactivity disorder, depression/depressive disorder, mood/mood disorder, neuropsychiatric, panic/panic disorder, psychiatric disorder, and psychosis. English-language articles published between January 1993 and May 2013 were retrieved. Additional studies were identified from the reference lists of relevant publications. Study Selection: 173 publications were retrieved. Articles related to the association between idiopathic RLS and depression, anxiety, and mood disorders were reviewed. In total, 32 epidemiologic studies were identified. These studies were reviewed in detail and ranked according to quality. Data Extraction: Data were extracted on the basis of relevance to the topic. Epidemiologic studies were assessed using 3 parameters: methodology, data quality, and generalizability of the results. Each factor was scored from 1 (high quality) to 4 (low quality), giving a total score of between 3 and 12 for each study. Results and Conclusions: RLS and mood disorders are frequently comorbid. Recognition and appropriate treatment of comorbid RLS are particularly important in patients with psychiatric disorders, as RLS is a common medical reason for insomnia, and antidepressant use may exacerbate sensory symptoms. © Copyright 2014 Physicians Postgraduate Press, Inc.


Janis J.E.,Ohio State University | Harrison B.,University of Texas Southwestern Medical Center
Plastic and Reconstructive Surgery | Year: 2014

LEARNING OBJECTIVES: After studying this article, the participant should be able to: 1. Describe the basic physiologic events in normal wound healing. 2. Understand the differences in healing among skin, bone, cartilage, and tendon. 3. Identify factors that may compromise or delay wound healing. 4. Describe methods for optimal closure of a wound. SUMMARY: Understanding the physiology and pathophysiology of normal wound healing and potential impediments to its end will allow the plastic surgeon to maximize postoperative outcomes and, in some instances, avoid unnecessary surgical interventions. Continuous advancements in our understanding of this process require frequent reviews of available data to permit reliable, evidence-based recommendations for clinical application. This is the first of a two-part article summarizing the science and clinical recommendations necessary for successful wound healing. Copyright © 2014 by the American Society of Plastic Surgeons.


Mishra R.K.,U.S. National Institutes of Health | Chakraborty P.,University of Texas Southwestern Medical Center | Arnaoutov A.,U.S. National Institutes of Health | Fontoura B.M.A.,University of Texas Southwestern Medical Center | Dasso M.,U.S. National Institutes of Health
Nature Cell Biology | Year: 2010

The metazoan nuclear pore complex (NPC) disassembles during mitosis, and many of its constituents distribute onto spindles and kinetochores, including the Nup107-160 sub-complex. We have found that Nup107-160 interacts with the γ-tubulin ring complex (γ-TuRC), an essential and conserved microtubule nucleator, and recruits γ-TuRC to unattached kinetochores. The unattached kinetochores nucleate microtubules in a manner that is regulated by Ran GTPase; such microtubules contribute to the formation of kinetochore fibres (k-fibres), microtubule bundles connecting kinetochores to spindle poles. Our data indicate that Nup107-160 and γ-TuRC act cooperatively to promote spindle assembly through microtubule nucleation at kinetochores: HeLa cells lacking Nup107-160 or γ-TuRC were profoundly deficient in kinetochore-associated microtubule nucleation. Moreover, co-precipitated Nup107-160-γ-TuRC complexes nucleated microtubule formation in assays using purified tubulin. Although Ran did not regulate microtubule nucleation by γ-TuRC alone, Nup107-160-γ-TuRC complexes required Ran-GTP for microtubule nucleation. Collectively, our observations show that Nup107-160 promotes spindle assembly through Ran-GTP-regulated nucleation of microtubules by γ-TuRC at kinetochores, and reveal a relationship between nucleoporins and the microtubule cytoskeleton. © 2010 Macmillan Publishers Limited. All rights reserved.


Kraus W.L.,University of Texas Southwestern Medical Center | Hottiger M.O.,University of Zürich
Molecular Aspects of Medicine | Year: 2013

Poly(ADP-ribose) polymerase-1 (PARP-1), also referred to as ADP-ribosyltransferase Diphtheria toxin-like 1 (ARTD1), is an abundant nuclear protein that plays key roles in a variety of nuclear processes, including the regulation of transcription. PARP-1 possesses an intrinsic enzymatic activity that catalyzes the transfer of ADP-ribose (ADPR) units from nicotinamide adenine dinucleotide (NAD+) onto target gene regulatory proteins, thereby modulating their activities. Although great strides have been made in the past decade in deciphering the seemingly opposing and varied roles of PARP-1 in gene regulation, many puzzles remain. In this review, we discuss the current state of understanding in this area, especially how PARP-1 interfaces with various components of gene regulatory pathways (e.g., the basal transcription machinery, DNA-binding transcription factors, coregulators, chromatin remodeling, histone modifications, and DNA methylation). In addition, we discuss some gene-specific, cell type-specific, and cell state-specific effects of PARP-1 on gene regulation, which might contribute to its biological functions. Finally, we review some of the recent progress targeting PARPs using chemical inhibitors, some of which may alter PARP-1-dependent gene regulatory programs to promote therapeutic outcomes. © 2013 Elsevier Ltd. All rights reserved.


Janis J.,University of Texas Southwestern Medical Center | Harrison B.,Ohio State University
Plastic and Reconstructive Surgery | Year: 2014

Learning objectives: After studying this article, the participant should be able to: 1. Identify effective methods for irrigation and débridement of wounds. 2. Describe methods for optimal closure of a wound. 3. Develop an algorithm for postoperative scar management. 4. Appreciate adjunctive clinical tools that may facilitate improvements in wound healing. Summary: Treatment of all wounds requires adequate wound bed preparation, beginning with irrigation and débridement. Complicated or chronic wounds may also require treatment adjuncts or specialized wound healing products. An extensive body of research and development has introduced novel wound healing therapies and scar management options. In this second of a two-part continuing medical education series on wound healing, the reader is offered an update on current wound healing technologies and recommendations for obtaining optimal outcomes. Copyright © 2014 by the American Society of Plastic Surgeons.


Livingston E.H.,University of Texas Southwestern Medical Center | Livingston E.H.,University of Texas at Arlington
American Journal of Surgery | Year: 2010

Background: Estimates of the procedure incidence for bariatric surgery have been derived primarily from surveys of bariatric surgeons or from inpatient data sources. New population-representative databases of outpatient surgery are available that enable accurate estimations of bariatric surgery case volumes. Methods: The 2006 National Hospital Discharge Survey, National Inpatient Sample, and National Survey of Ambulatory Surgery were assessed for bariatric surgery procedures. Data were compared with inpatient data from 1993 to 2007. Procedure costs were estimated. Results: The incidence of bariatric surgery has plateaued at approximately 113,000 cases per year. Open gastric bypass now constitutes only 3% of all cases but costs $4,800 less than laparoscopic procedures. Laparoscopic gastric banding is performed in 37% of all bariatric surgery cases and costs the same as laparoscopic gastric bypass to perform. Complication rates have fallen from 10.5% in 1993 to 7.6% of all cases in 2006. Bariatric surgery costs the health economy at least $1.5 billion annually. Conclusions: Despite predictions of continued growth of bariatric surgery, it appears that the annual incidence for these operations has remained stable since 2003. Most operations are performed laparoscopically, but open gastric bypass is substantially less costly than laparoscopic operations. Despite its simplicity, laparoscopic gastric banding costs the same as gastric bypass. There is no cost savings associated with ambulatory bariatric surgery. © 2010 Elsevier Inc.


Zhang Y.,University of Texas at Arlington | Zhang Y.,University of Texas Southwestern Medical Center | Yang J.,Pennsylvania State University
Journal of Materials Chemistry B | Year: 2013

The combination of biodegradable polymer and fluorescent imaging has resulted in an important area of polymeric biomaterials: biodegradable fluorescent polymers. Researchers have made significant efforts in developing versatile fluorescent biomaterials due to their promising applications in biological/biomedical labeling, tracking, monitoring, imaging, and diagnostics, especially in drug delivery, tissue engineering, and cancer imaging. Biodegradable fluorescent polymers can function not only as implant biomaterials but also as imaging probes. Currently, there are two major classes of biodegradable polymers, which are used as fluorescent materials. The first class is the combination of non-fluorescent biodegradable polymers and fluorescent agents such as organic dyes and quantum dots. Another class of polymers shows intrinsic photoluminescence as polymers by themselves carrying integral fluorescent chemical structures in or pendent to their polymer backbone, such as Green Fluorescent protein (GFP), and the recently developed biodegradable photoluminescent polymer (BPLP). Thus there is no need to conjugate or encapsulate additional fluorescent materials for the latter. In the present review, we will review the fluorescent biodegradable polymers with emphases on material fluorescence mechanism, design criteria for fluorescence, and their cutting-edge applications in biomedical engineering. We expect that this review will provide an insightful discussion on the fluorescent biomaterial design and lead to innovations for the next generation of fluorescent biomaterials and fluorescence-based biomedical technology. © 2013 The Royal Society of Chemistry.


Gandhi N.S.,University of Hawaii at Hilo | Tekade R.K.,University of Hawaii at Hilo | Tekade R.K.,University of Texas Southwestern Medical Center | Chougule M.B.,University of Hawaii at Hilo | Chougule M.B.,University of Hawaii at Manoa
Journal of Controlled Release | Year: 2014

Chemotherapeutic agents have certain limitations when it comes to treating cancer, the most important being severe side effects along with multidrug resistance developed against them. Tumor cells exhibit drug resistance due to activation of various cellular level processes viz. activation of drug efflux pumps, anti-apoptotic defense mechanisms, etc. Currently, RNA interference (RNAi) based therapeutic approaches are under vibrant scrutinization to seek cancer cure. Especially small interfering RNA (siRNA) and micro RNA (miRNA), are able to knock down the carcinogenic genes by targeting the mRNA expression, which underlies the uniqueness of this therapeutic approach. Recent research focus in the regime of cancer therapy involves the engagement of targeted delivery of siRNA/miRNA in combinations with other therapeutic agents (such as gene, DNA or chemotherapeutic drug) for targeting permeability glycoprotein (P-gp), multidrug resistant protein 1 (MRP-1), B-cell lymphoma (BCL-2) and other targets that are mainly responsible for resistance in cancer therapy. RNAi-chemotherapeutic drug combinations have also been found to be effective against different molecular targets as well and can increase the sensitization of cancer cells to therapy several folds. However, due to stability issues associated with siRNA/miRNA suitable protective carrier is needed and nanotechnology based approaches have been widely explored to overcome these drawbacks. Furthermore, it has been univocally advocated that the co-delivery of siRNA/miRNA with other chemodrugs significantly enhances their capability to overcome cancer resistance compared to naked counterparts. The objective of this article is to review recent nanocarrier based approaches adopted for the delivery of siRNA/miRNA combinations with other anticancer agents (siRNA/miRNA/pDNA/chemodrugs) to treat cancer. © 2014, Elsevier B.V. All rights reserved.


Wang J.,University of Texas Southwestern Medical Center | Hou T.,Soochow University of China
Journal of Chemical Information and Modeling | Year: 2010

Drug likeness analysis is widely used in modem drug design, However, most drug likeness filters, represented by Lipinski's "Rule of 5", are based on drugs' simple structural features and some physiochemical properties. In this study, we conducted thorough structural analyses for two drug datasets. The first dataset, ADDS, is composed of 1240 FDA-approved drugs, and the second drug dataset, EDDS, is a nonredundant collection of FDA-approved drugs and experimental drugs in different phases of clinical trials from several drug databases (6932 entries). For each molecule, all possible fragments were enumerated using a brutal force approach. Three kinds of building blocks, namely, the drug scaffold, ring system, and the small fragment, were identified and ranked according to the frequencies of their occurrence in drug molecules. The major finding is that most top fragments are essentially common for both drug datasets; the top 50 fragments cover 52.6% and 48.6% drugs for ADDS and EDDS, respectively. The identified building blocks were further ranked according to their relative hit rates in the drug datasets and in a screening dataset, which is a nonredundant collection of screening compounds from many resources. In comparison with the previous reports in the field, we have identified many more high-quality building blocks. The results obtained in this study could provide useful hints to medicinal chemists in designing drug-like compounds as well as prioritizing screening libraries to filter out those molecules lack of functional building blocks. © 2010 American Chemical Society.


Lakoski S.G.,University of Vermont | Kozlitina J.,University of Texas Southwestern Medical Center
Medicine and Science in Sports and Exercise | Year: 2014

PURPOSE: This study aimed to examine ethnic differences in objectively measured physical activity (PA) and the relationship between PA and metabolic risk factors. METHODS: The analysis included 2566 participants of the Dallas Heart Study (53% non-Hispanic black, 32% non-Hispanic white, and 15% Hispanic) who wore an accelerometer for an average of 7 d. PA was assessed as mean activity counts and time spent in moderate and vigorous activity. Outcomes included body mass index (BMI), waist circumference, systolic and diastolic blood pressure, heart rate, fasting glucose, homeostatic model assessment of insulin resistance, and plasma lipid and lipoprotein levels. RESULTS: A higher proportion of Hispanics than either whites or blacks obtained the recommended ≥ 150 min·wk of moderate PA (24%, 14%, and 10%, respectively, P < 0.0001). White males were more likely to engage in vigorous activity than other sex-ethnic groups (P < 0.05). Time in moderate-to-vigorous activity was inversely related to BMI, waist circumference, homeostatic model assessment of insulin resistance, heart rate, and positively associated with high-density lipoprotein cholesterol levels (P < 0.0001) in the combined cohort, and the relationship was similar in all ethnic groups (P interaction > 0.05). A significant inverse association between PA and triglycerides was observed in whites (P = 7.2 × 10). Vigorous activity was associated with greater differences in risk factors than moderate activity (for example, β = -0.30 vs β = -0.02 for BMI). Bouts lasting =10 min were associated with metabolic risk factors independent of <10-min bouts in the overall sample, with similar trends observed within subgroups. CONCLUSIONS: Hispanics had higher levels of moderate activity than whites or blacks, whereas white men had higher levels of vigorous PA than other sex-ethnic groups. The relationship between PA and several metabolic risk factors was similar across ethnicities. Vigorous PA was associated with greater benefits than moderate PA. © 2014 by the American College of Sports Medicine.


Lefebvre K.M.,Widener University | Lavery L.A.,University of Texas Southwestern Medical Center
Clinical Orthopaedics and Related Research | Year: 2011

Background: As a result of the impact of health disparities on the healthcare system such as their influence on arenas significant to healthcare distribution, including cost, quality, and access, identification and resolution of health disparities is a primary national agenda item. Resolution of disparities in amputation is an area of opportunity that warrants further consideration. Questions/purposes: The purposes of our review are to highlight current data on disparities in amputation in minorities and to consider future goals related to an elimination of this disparity. Methods: Studies on disparities in amputation were accessed using the following databases: PubMed, Cinahl, OVID/Medline, Embase, and Cochrane databases. In each database, a search of title/abstract was performed for the search terms "disparities and amputation," "race and amputation," and "diabetes and amputation." Each search was limited by human and English language. Where are we now? A disparity exists in both frequency and level of amputation in minorities both in the presence and absence of a diagnosis of diabetes. Where do we need to go? A need exists for future research involving a more deliberate examination of the use of preventive screening for patients at high risk for amputation across medical settings. How do we get there? Research in this area would benefit from funding, large-scale data collection, and physician exposure to education on high-risk patients and preventive screening opportunities. © 2011 The Association of Bone and Joint Surgeons®.


Zhu C.,Soochow University of China | Falck J.R.,University of Texas Southwestern Medical Center
Advanced Synthesis and Catalysis | Year: 2014

Arylboronic acids and their derivatives have been widely exploited as important synthetic precursors in organic synthesis, materials science, and pharmaceutical development. In addition to numerous applications in transition metal-mediated cross-coupling reactions, transition metal-free transformations involving arylboronic acids and derivatives have recently received a surge of attention for converting the C-B bond to C-C, C-N, C-O, and many other C-X bonds. Consequently, a wide range of useful compounds, e.g., phenols, anilines, nitroarenes, and haloarenes, has been readily synthesized. Amongst these efforts, many versatile reagents have been developed and a lot of practical approaches demonstrated. The research in this promising field is summarized in the current review and organized on the basis of the type of bonds being formed. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Dong X.,University of Texas Southwestern Medical Center | Feng P.,University of Texas Southwestern Medical Center | Feng P.,University of Southern California
PLoS Pathogens | Year: 2011

Upon viral infection, mitochondrial antiviral signaling (MAVS) protein serves as a key adaptor to promote cytokine production. We report here that murine gamma herpesvirus 68 (γHV68), a model virus for oncogenic human gamma herpesviruses, subverts cytokine production via the MAVS adaptor. During early infection, γHV68 hijacks MAVS and IKKβ to induce the site-specific phosphorylation of RelA, a crucial subunit of the transcriptionally active NFκB dimer, which primes RelA for the proteasome-mediated degradation. As such, γHV68 efficiently abrogated NFκB activation and cytokine gene expression. Conversely, uncoupling RelA degradation from γHV68 infection promoted NFκB activation and elevated cytokine production. Loss of MAVS increased cytokine production and immune cell infiltration in the lungs of γHV68-infected mice. Moreover, exogenous expression of the phosphorylation- and degradation-resistant RelA variant restored γHV68-induced cytokine production. Our findings uncover an intricate strategy whereby signaling via the upstream MAVS adaptor is intercepted by a pathogen to nullify the immediate downstream effector, RelA, of the innate immune pathway. © 2011 Dong, Feng.


Diamond M.S.,University of Washington | Schoggins J.W.,University of Texas Southwestern Medical Center
Cell Host and Microbe | Year: 2013

In this issue of Cell Host & Microbe, Varble et al. (2013) engineer a library of RNA viruses to express small interfering RNAs and couple this with the power of virus evolution and selection to screen for host genes that when silenced resulted in greater viral infection in vivo. © 2013 Elsevier Inc.


Wang J.,University of Texas Southwestern Medical Center | Hou T.,Soochow University of China
Journal of Chemical Theory and Computation | Year: 2011

Molecular mechanical force field (FF) methods are useful in studying condensed phase properties. They are complementary to experiments and can often go beyond experiments in atomic details. Even if a FF is specific for studying structures, dynamics, and functions of biomolecules, it is still important for the FF to accurately reproduce the experimental liquid properties of small molecules that represent the chemical moieties of biomolecules. Otherwise, the force field may not describe the structures and energies of macromolecules in aqueous solutions properly. In this work, we have carried out a systematic study to evaluate the General AMBER Force Field (GAFF) in studying densities and heats of vaporization for a large set of organic molecules that covers the most common chemical functional groups. The latest techniques, such as the particle mesh Ewald (PME) for calculating electrostatic energies and Langevin dynamics for scaling temperatures, have been applied in the molecular dynamics (MD) simulations. For density, the average percent error (APE) of 71 organic compounds is 4.43% when compared to the experimental values. More encouragingly, the APE drops to 3.43% after the exclusion of two outliers and four other compounds for which the experimental densities have been measured with pressures higher than 1.0 atm. For the heat of vaporization, several protocols have been investigated, and the best one, P4/ntt0, achieves an average unsigned error (AUE) and a root-mean-square error (RMSE) of 0.93 and 1.20 kcal/mol, respectively. How to reduce the prediction errors through proper van der Waals (vdW) parametrization has been discussed. An encouraging finding in vdW parametrization is that both densities and heats of vaporization approach their "ideal" values in a synchronous fashion when vdW parameters are tuned. The following hydration free energy calculation using thermodynamic integration further justifies the vdW refinement. We conclude that simple vdW parametrization can significantly reduce the prediction errors. We believe that GAFF can greatly improve its performance in predicting liquid properties of organic molecules after a systematic vdW parametrization, which will be reported in a separate paper. © 2011 American Chemical Society.


Livingston E.H.,University of Texas Southwestern Medical Center | Burchell I.,University of Texas at Arlington
Archives of Surgery | Year: 2010

Objective: To determine the effect on travel distance for Medicare patients before and after Centers for Medicare & Medicaid Services required that bariatric procedures be performed at Centers of Excellence (COEs). Design: We calculated the distance traveled to our medical center for the 2 years prior (2004-2005) and 2 years after (2006-2007) COE status was required by Medicare. We also compared the proportion of bariatric cases done in large hospitals with those for esophageal and pancreatic resections, procedures whose effects regionalization would have on patient access have been modeled. Setting: University of Texas Southwestern Medical Center, a high-volume tertiary referral center for bariatric surgery. Patients: Patients undergoing bariatric procedures. Main Outcome Measure: Travel distances. Results: Depending on insurance status, before COEs were required, patients traveled a median of 16 to 25 miles to undergo bariatric operations at University of Texas Southwestern. After COEs were required, the median distance Medicare patients were required to travel increased 76% to 44 miles. Conclusions: Center of Excellence requirements have increased the travel distance required for Medicare patients. Prior research has shown that outcomes at COEs are no different than those at non-COEs suggesting that the reduced access to care resulting from requiring COE status is not beneficial. ©2010 American Medical Association. All rights reserved.


LeComte M.D.,University of Vermont | Shimada I.S.,University of Vermont | Shimada I.S.,University of Texas Southwestern Medical Center | Sherwin C.,University of Vermont | Spees J.L.,University of Vermont
Proceedings of the National Academy of Sciences of the United States of America | Year: 2015

Defining the signaling network that controls reactive astrogliosis may provide novel treatment targets for patients with diverse CNS injuries and pathologies. We report that the radial glial cell antigen RC2 identifies the majority of proliferating glial fibrillary acidic protein-positive (GFAP+) reactive astrocytes after stroke. These cells highly expressed endothelin receptor type B (ETBR) and Jagged1, a Notch1 receptor ligand. To study signaling in adult reactive astrocytes, we developed a model based on reactive astrocyte-derived neural stem cells isolated from GFAP-CreER-Notch1 conditional knockout (cKO) mice. By loss- and gain-of-function studies and promoter activity assays, we found that Jagged1/Notch1 signaling increased ETBR expression indirectly by raising the level of phosphorylated signal transducer and activator of transcription 3 (STAT3), a previously unidentified EDNRB transcriptional activator. Similar to inducible transgenic GFAP-CreER-Notch1-cKO mice, GFAP-CreER-ETBR-cKO mice exhibited a defect in reactive astrocyte proliferation after cerebral ischemia. Our results indicate that the Notch1-STAT3-ETBR axis connects a signaling network that promotes reactive astrocyte proliferation after brain injury. © 2015, National Academy of Sciences. All rights reserved.


Garg R.,Harvard University | Chen W.,Medco Health Solutions | Pendergrass M.,Medco Health Solutions | Pendergrass M.,University of Texas Southwestern Medical Center
Diabetes Care | Year: 2010

OBJECTIVE- Cases of acute pancreatitis have been reported in association with exenatide, sitagliptin, and type 2 diabetes without use of these medications. It remains unknown whether exenatide or sitagliptin increase the risk of acute pancreatitis. RESEARCH DESIGN AND METHODS- A retrospective cohort study of a large medical and pharmacy claims database was performed. Data for 786,656 patients were analyzed. Cox proportional hazard models were built to compare the risk of acute pancreatitis between diabetic and nondiabetic subjects and between exenatide, sitagliptin, and control diabetes medication use. RESULTS- Incidence of acute pancreatitis in the nondiabetic control group, diabetic control group, exenatide group, and sitagliptin group was 1.9, 5.6, 5.7, and 5.6 cases per 1,000 patient years, respectively. The risk of acute pancreatitis was significantly higher in the combined diabetic groups than in the nondiabetic control group (adjusted hazard ratio 2.1 [95% CI 1.7-2.5]). Risk of acute pancreatitis was similar in the exenatide versus diabetic control group (0.9 [0.6 -1.5]) and sitagliptin versus diabetic control group (1.0 [0.7-1.3]). CONCLUSIONS- Our study demonstrated increased incidence of acute pancreatitis in diabetic versus nondiabetic patients but did not find an association between the use of exenatide or sitagliptin and acute pancreatitis. The limitations of this observational claims-based analysis cannot exclude the possibility of an increased risk. © 2010 by the American Diabetes Association.


Malki S.,Carnegie Institution for Science | vanderHeijden G.,Carnegie Institution for Science | O'Donnell K.A.,University of Texas Southwestern Medical Center | Martin S.L.,Aurora University | Bortvin A.,Carnegie Institution for Science
Developmental Cell | Year: 2014

Fetal oocyte attrition (FOA) is a conserved but poorly understood process of elimination of more than two-thirds of meiotic prophase I (MPI) oocytes before birth. We now implicate retrotransposons LINE-1 (L1), activated during epigenetic reprogramming of the embryonic germline, in FOA in mice. We show that wild-type fetal oocytes possess differential nuclear levels of L1ORF1p, an L1-encoded protein essential for L1 ribonucleoprotein particle (L1RNP) formation and L1 retrotransposition. We demonstrate that experimental elevation of L1 expression correlates with increased MPI defects, FOA, oocyte aneuploidy, and embryonic lethality. Conversely, reverse transcriptase (RT) inhibitor AZT has a profound effect on the FOA dynamics and meiotic recombination, and it implicates an RT-dependent trigger in oocyte elimination in early MPI. We propose that FOA serves to select oocytes with limited L1 activity that are therefore best suited for the next generation. © 2014 Elsevier Inc.


Buhr E.D.,University of Washington | Takahashi J.S.,University of Texas Southwestern Medical Center
Handbook of Experimental Pharmacology | Year: 2013

Mammals synchronize their circadian activity primarily to the cycles of light and darkness in the environment. This is achieved by ocular photoreception relaying signals to the suprachiasmatic nucleus (SCN) in the hypothalamus. Signals from the SCN cause the synchronization of independent circadian clocks throughout the body to appropriate phases. Signals that can entrain these peripheral clocks include humoral signals, metabolic factors, and body temperature. At the level of individual tissues, thousands of genes are brought to unique phases through the actions of a local transcription/translation-based feedback oscillator and systemic cues. In this molecular clock, the proteins CLOCK and BMAL1 cause the transcription of genes which ultimately feedback and inhibit CLOCK and BMAL1 transcriptional activity. Finally, there are also other molecular circadian oscillators which can act independently of the transcription-based clock in all species which have been tested. © 2013 Springer-Verlag Berlin Heidelberg.


Pietanza M.C.,Sloan Kettering Cancer Center | Byers L.A.,University of Houston | Minna J.D.,University of Texas Southwestern Medical Center | Rudin C.M.,Sloan Kettering Cancer Center
Clinical Cancer Research | Year: 2015

Small cell lung cancer (SCLC) is an aggressive neuroendocrine malignancy with a unique natural history characterized by a short doubling time, high growth fraction, and early development of widespread metastases. Although a chemotherapy- and radiationsensitive disease, SCLC typically recurs rapidly after primary treatment, with only 6% of patients surviving 5 years from diagnosis. This disease has been notable for the absence of major improvements in its treatment: Nearly four decades after the introduction of a platinum etoposide doublet, therapeutic options have remained virtually unchanged, with correspondingly little improvement in survival rates. Here, we summarize specific barriers and challenges inherent to SCLC research and care that have limited progress in novel therapeutic development to date. We discuss recent progress in basic and translational research, especially in the development of mouse models, which will provide insights into the patterns of metastasis and resistance in SCLC. Opportunities in clinical research aimed at exploiting SCLC biology are reviewed, with an emphasis on ongoing trials. SCLC has been described as a recalcitrant cancer, for which there is an urgent need for accelerated progress. The NCI convened a panel of laboratory and clinical investigators interested in SCLC with a goal of defining consensus recommendations to accelerate progress in the treatment of SCLC, which we summarize here. © 2015 American Association for Cancer Research.


Phillips M.A.,University of Texas Southwestern Medical Center | Rathod P.K.,University of Washington
Infectious Disorders - Drug Targets | Year: 2010

Malaria remains a globally prevalent infectious disease that leads to significant morbidity and mortality. While there are a number of drugs approved for its treatment, drug resistance has compromised most of them, making the development of new drugs for the treatment and prevention of malaria essential. The completion of the Plasmodium falciparum genome and a growing understanding of parasite biology are fueling the search for novel drug targets. Despite this, few targets have been chemically validated in vivo. The pyrimidine biosynthetic pathway illustrates one of the best examples of successful identification of anti-malarial drug targets. This review focuses on recent studies to exploit the fourth enzyme in the de novo pyrimidine biosynthetic pathway of P. falciparum, dihydroorotate dehydrogenase (PfDHODH), as a new target for drug discovery. Several chemical scaffolds have been identified by high throughput screening as potent inhibitors of PfDHODH and these show strong selectivity for the malarial enzyme over that from the human host. Potent activity against parasites in whole cell models with good correlation between activity on the enzyme and the parasite have also been observed for a number of the identified series. Lead optimization of a triazolopyrimidinebased series has identified an analog with prolonged plasma exposure, that is orally bioavailable, and which shows good efficacy against the in vivo mouse model of the disease. These data provide strong evidence that PfDHODH is a validated target for the identification of new antimalarial chemotherapy. The challenge remains to identify compounds with the necessary combination of potency and metabolic stability to allow identification of a clinical candidate. © 2010 Bentham Science Publishers Ltd.


Tuite J.J.,International Security and Risk Management Consultant | Haley R.W.,University of Texas Southwestern Medical Center
Neuroepidemiology | Year: 2013

Background: Coalition bombings on the night of 18-19 January 1991, early in the Gulf War, targeted the Iraqi chemical weapons infrastructure. On 19 January 1991, nerve agent alarms sounded within Coalition positions hundreds of kilometers to the south, and the trace presence of sarin vapor was identified by multiple technologies. Considering only surface dispersion of