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DeVries A.,BIO5 Institute | DeVries A.,Arizona Respiratory Center | Vercelli D.,Arizona Respiratory Center | Vercelli D.,Arizona Center for the Biology of Complex Diseases | Vercelli D.,University of Arizona
Annals of the American Thoracic Society | Year: 2016

Asthma and allergic diseases are among the most prevalent chronic noncommunicable diseases of childhood, but the underlying pathogenetic mechanisms are poorly understood. Because epigenetic mechanisms link gene regulation to environmental cues and developmental trajectories, their contribution to asthma and allergy pathogenesis is under active investigation. DNA methylation signatures associated with concurrent disease and with the development of asthma during childhood asthma have been identified, but their significance is not easily interpretable. On the other hand, the characterization of early epigenetic predictors of asthma points to a potential role of epigenetic mechanisms in regulating the inception of, and the susceptibility to, this disease. Copyright © 2016 by the American Thoracic Society.

Researchers at the University of Arizona have invented a device that for the first time allows neurosurgeons, who use microscopes extensively while operating, to see blood flowing inside vessels and more clearly distinguish cancerous from healthy tissue under the microscope. Called augmented microscopy, the technology gives surgeons a much more detailed picture in real time and helps them stay on course in surgeries where being off two millimeters could cause paralysis, blindness and even death. And surgeons get this better view without having to learn new technical skills or adapt to changes in the operating room. "When we started developing this technology, we thought of it like a Google map of a surgical view, providing layers of pertinent information in real time," said Marek Romanowski, UA associate professor of biomedical engineering. "Our augmented technology provides diagnostic information under the microscope on demand and in color, appearing directly over tissue a surgeon is operating on—as if the tissue was painted to help direct the surgeon's work." The new technology overlays an actual, or bright field, image a surgeon sees under a microscope with an electronically processed image using near-infrared fluorescence. NIR fluorescence is a computer-generated imaging technology in which contrast agents are injected in patients to illuminate vital diagnostic information and help surgeons avoid cutting the wrong vessel or removing healthy tissue. Most neurosurgeons must look up from a surgical microscope, or stereomicroscope, to view fluorescence on a display monitor. If they have a microscope adapted to project fluorescence, it switches back and forth between the real and electronic views, the surgeons' field of vision momentarily fading to black in between. Further, the fluorescence shows only contrast in black and white, not anatomical structures or their spatial relationships. So surgeons must visualize how fluorescence lines up with the anatomical structures they see under the microscope. Doctoral student Jeffrey Watson, left, and associate professor Marek Romanowski assemble parts for the prototype microscopy device.Doctoral student Jeffrey Watson, left, and associate professor Marek Romanowski assemble parts for the prototype microscopy device. The new add-on technology developed at the UA removes such interruptions or guesswork by showing surgeons real and fluorescence images simultaneously and in one location. Romanowski describes the invention with lead author Jeffrey Watson, a biomedical engineering doctoral student in the UA Graduate Interdisciplinary Program in Biomedical Engineering; G. Michael Lemole Jr., MD, chief of the division of neurosurgery in the department of surgery at the UA College of Medicine-Tucson; and UA neurosurgery resident Nikolay Martirosyan, MD, in "Augmented microscopy: Real-time overlay of bright-field and near-infrared fluorescence images," published in the Oct. 2015 edition of the SPIE Journal of Biomedical Optics. "Surgeons need more information than can be provided by stereomicroscopes alone," said Jennifer Barton, a UA professor of biomedical engineering and interim director of the UA BIO5 Institute, who specializes in cancer imaging. "Dr. Romanowski's augmented microscopy technology provides critical functional information that can improve surgical accuracy and efficiency." The new device, a small box fitted inside a surgical microscope, combines electronic circuitry and optical technologies to superimpose the fluorescence image on the real one and send the augmented view up through the microscope's right eyepiece to the surgeon. Lemole, a former flight surgeon in the Air Force Reserve, likens the technology to the head-up display in an airplane cockpit. "If you can place your critical gauges directly in the pilot's line of sight, they don't need to look in a different direction while performing critical maneuvers. It won't change the way they fly the plane, but it gives them more information, without distraction." Perhaps the most valuable application for augmented microscopy is treating brain cancer, said Romanowski, who holds appointments with the University of Arizona Cancer Center and BIO5 Institute. More than 20,000 new cases of primary brain cancer are diagnosed in the United States each year, and each year nearly 16,000 patients die from the disease, Romanowski said. Of the half-million patients who die of any other cancer, up to a third has some form of cancer spreading to the brain. "Brain cancer is especially difficult to remove," he said. "Current surgical microscopes limit how much of the cancer tissue surgeons can see and how precisely they can determine its boundaries." Lemole, a skull base neurosurgeon, routinely operates on brain cancer patients, manipulating vessels the width of a pin to remove malignant tumors. He walks a fine line to remove all of the cancer without removing healthy tissue. "Aggressive resection is associated with the risk of removing normal brain tissue and impairing functions of the patient," he and his co-authors write in the Journal of Biomedical Optics. "On the other hand, incomplete resection of tumor results in its immediate relapse in 90 percent of patients. Intraoperative NIR imaging may aid in resection of these challenging tumors." Augmented microscopy also holds promise for aneurysm, a bulging of an artery caused by weakened arterial walls. Neurosurgeons treat aneurysm by sealing it off from connecting vessels to prevent a rupture. Nearly half the patients with ruptured aneurysms die, Lemole said, and at least half the survivors have major mobility and other problems. Augmented technology could improve aneurysm patients' prognosis, by giving surgeons real-time feedback on every delicate and potentially deadly surgical maneuver they make. "When I pick up and clip a vessel, I like to see the implications of what happens in that very moment," he said. Explore further: New high-tech glasses detect cancer cells during surgery More information: Jeffrey R. Watson et al. Augmented microscopy: real-time overlay of bright-field and near-infrared fluorescence images, Journal of Biomedical Optics (2015). DOI: 10.1117/1.JBO.20.10.106002

Yu Z.,Kent State University | Gaerig V.,University of Arizona | Cui Y.,Kent State University | Kang H.,University of Arizona | And 6 more authors.
Journal of the American Chemical Society | Year: 2012

The discovery of G-quadruplexes and other DNA secondary elements has increased the structural diversity of DNA well beyond the ubiquitous double helix. However, it remains to be determined whether tertiary interactions can take place in a DNA complex that contains more than one secondary structure. Using a new data analysis strategy that exploits the hysteresis region between the mechanical unfolding and refolding traces obtained by a laser-tweezers instrument, we now provide the first convincing kinetic and thermodynamic evidence that a higher order interaction takes place between a hairpin and a G-quadruplex in a single-stranded DNA fragment that is found in the promoter region of human telomerase. During the hierarchical unfolding or refolding of the DNA complex, a 15-nucleotide hairpin serves as a common species among three intermediates. Moreover, either a mutant that prevents this hairpin formation or the addition of a DNA fragment complementary to the hairpin destroys the cooperative kinetic events by removing the tertiary interaction mediated by the hairpin. The coexistence of the sequential and the cooperative refolding events provides direct evidence for a unifying kinetic partition mechanism previously observed only in large proteins and complex RNA structures. Not only does this result rationalize the current controversial observations for the long-range interaction in complex single-stranded DNA structures, but also this unexpected complexity in a promoter element provides additional justification for the biological function of these structures in cells. © 2012 American Chemical Society.

Cui Y.,Kent State University | Koirala D.,Kent State University | Kang H.,University of Arizona | Dhakal S.,Kent State University | And 5 more authors.
Nucleic Acids Research | Year: 2014

Minute difference in free energy change of unfolding among structures in an oligonucleotide sequence can lead to a complex population equilibrium, which is rather challenging for ensemble techniques to decipher. Herein, we introduce a new method, molecular population dynamics (MPD), to describe the intricate equilibrium among non-B deoxyribonucleic acid (DNA) structures. Using mechanical unfolding in laser tweezers, we identified six DNA species in a cytosine (C)-rich bcl-2 promoter sequence. Population patterns of these species with and without a small molecule (IMC-76 or IMC-48) or the transcription factor hnRNP LL are compared to reveal the MPD of different species. With a pattern recognition algorithm, we found that IMC-48 and hnRNP LL share 80% similarity in stabilizing i-motifs with 60 s incubation. In contrast, IMC-76 demonstrates an opposite behavior, preferring flexible DNA hairpins. With 120-180 s incubation, IMC-48 and hnRNP LL destabilize i-motifs, which has been previously proposed to activate bcl-2 transcriptions. These results provide strong support, from the population equilibrium perspective, that small molecules and hnRNP LL can modulate bcl-2 transcription through interaction with i-motifs. The excellent agreement with biochemical results firmly validates the MPD analyses, which, we expect, can be widely applicable to investigate complex equilibrium of biomacromolecules. © 2014 The Author(s) 2014.

Melemedjian O.K.,University of Arizona | Tillu D.V.,University of Arizona | Moy J.K.,University of Arizona | Asiedu M.N.,University of Arizona | And 9 more authors.
Molecular Pain | Year: 2014

Transcriptional regulation of genes by cyclic AMP response element binding protein (CREB) is essential for the maintenance of long-term memory. Moreover, retrograde axonal trafficking of CREB in response to nerve growth factor (NGF) is critical for the survival of developing primary sensory neurons. We have previously demonstrated that hindpaw injection of interleukin-6 (IL-6) induces mechanical hypersensitivity and hyperalgesic priming that is prevented by the local injection of protein synthesis inhibitors. However, proteins that are locally synthesized that might lead to this effect have not been identified. We hypothesized that retrograde axonal trafficking of nascently synthesized CREB might link local, activity-dependent translation to nociceptive plasticity. To test this hypothesis, we determined if IL-6 enhances the expression of CREB and if it subsequently undergoes retrograde axonal transport. IL-6 treatment of sensory neurons in vitro caused an increase in CREB protein and in vivo treatment evoked an increase in CREB in the sciatic nerve consistent with retrograde transport. Importantly, co-injection of IL-6 with the methionine analogue azido-homoalanine (AHA), to assess nascently synthesized proteins, revealed an increase in CREB containing AHA in the sciatic nerve 2 hrs post injection, indicating retrograde transport of nascently synthesized CREB. Behaviorally, blockade of retrograde transport by disruption of microtubules or inhibition of dynein or intrathecal injection of cAMP response element (CRE) consensus sequence DNA oligonucleotides, which act as decoys for CREB DNA binding, prevented the development of IL-6-induced mechanical hypersensitivity and hyperalgesic priming. Consistent with previous studies in inflammatory models, intraplantar IL-6 enhanced the expression of BDNF in dorsal root ganglion (DRG). This effect was blocked by inhibition of retrograde axonal transport and by intrathecal CRE oligonucleotides. Collectively, these findings point to a novel mechanism of axonal translation and retrograde trafficking linking locally-generated signals to long-term nociceptive sensitization. © 2014 Melemedjian et al.

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