Spiering R.,University Utrecht |
Margry B.,University Utrecht |
Keijzer C.,University Utrecht |
Petzold C.,Foundation University |
And 7 more authors.
Journal of Immunology | Year: 2015
Previous studies in mouse models of autoimmune diabetes and encephalomyelitis have indicated that the selective delivery of self-antigen to the endocytic receptor DEC205 on steady-state dendritic cells (DCs) may represent a suitable approach to induce Ag-specific immune tolerance. In this study, we aimed to examine whether DEC205+ DC targeting of a single immunodominant peptide derived from human cartilage proteoglycan (PG) can promote immune tolerance in PG-induced arthritis (PGIA). Besides disease induction by immunization with whole PG protein with a high degree of antigenic complexity, PGIA substantially differs from previously studied autoimmune models not only in the target tissue of autoimmune destruction but also in the nature of pathogenic immune effector cells. Our results show that DEC205+ DC targeting of the PG peptide 70-84 is sufficient to efficiently protect against PGIA development. Complementary mechanistic studies support a model in which DEC205+ DC targeting leads to insufficient germinal center B cell support by PG-specific follicular helper T cells. Consequently, impaired germinal center formation results in lower Ab titers, severely compromising the development of PGIA. Overall, this study further corroborates the potential of prospective tolerogenic DEC205+ DC vaccination to interfere with autoimmune diseases, such as rheumatoid arthritis. Copyright © 2015 by The American Association of Immunologists, Inc.
News Article | November 7, 2016
Certain proteins in the blood of children can predict incipient type 1 diabetes, even before the first symptoms appear. A team of scientists at the Helmholtz Zentrum München, partners in the German Center for Diabetes Research (DZD), reported these findings in the Diabetologia journal. The work was based on two large studies that are intended to explain the mechanisms behind the development of type 1 diabetes (BABYDIAB and BABYDIET*). The study participants are children who have a first-degree relative with type 1 diabetes and who consequently have an increased risk of developing the disease due to the familial predisposition. This autoimmune process does not develop from one day to the next, however: Often the young patients go through longer asymptomatic preliminary stages that see the formation of the first antibodies against the child's own insulin-producing cells in the pancreas; these are the so-called autoantibodies. Biomarkers that indicate whether and when this is the case and how quickly the clinical symptoms will appear could significantly improve the treatment of patients at-risk. A team of scientists, led by Dr. Stefanie Hauck, head of the Research Unit Protein Science and the Core Facility Proteomics, and Prof. Dr. Anette-G. Ziegler, Director of the Institute of Diabetes Research (IDF) at the Helmholtz Zentrum München, analyzed blood samples from 30 children with autoantibodies who had developed type 1 diabetes either very rapidly or with a very long delay. The researchers compared the data with data on children who displayed neither autoantibodies nor diabetes symptoms. In a second step with samples from another 140 children, the researchers confirmed the protein composition differences that they found in this approach. "Altogether, we were able to identify 41 peptides** from 26 proteins that distinguish children with autoantibodies from those without," reports Dr. Christine von Toerne, a scientist in the Research Unit Protein Science who shared first authorship of the work with Michael Laimighofer, a doctoral candidate in Jan Krumsiek's junior research group at the Institute of Computational Biology. Striking in their evaluations: A large number of these proteins are associated with lipid metabolism. "Two peptides -- from the proteins apolipoprotein M and apolipoprotein C-IV -- were particularly conspicuous and were especially differently expressed in the two groups," von Toerne adds. In autoantibody-positive children, it was furthermore possible to reach a better estimate of the speed of the diabetes development using the peptide concentrations of three proteins (hepatocyte growth factor activator, complement factor H and ceruloplasmin) in combination with the age of the particular child. The researchers are confident that the protein signatures they have discovered will be helpful as biomarkers for future diagnostics. "The progression of type 1 diabetes into a clinical disease takes place over a period of time that varies from individual to individual and that at this time is insufficiently predictable," explains Prof. Ziegler. "The biomarkers that we have identified allow a more precise classification of this presymptomatic stage and they are relatively simple to acquire from blood samples." * The BABYDIAB study, which was established in 1989 as the world's first diabetes prospective birth cohort, is a pioneering study in the field of type 1 diabetes pathogenesis research. More than 1650 children of parents with type 1 diabetes have been observed since their birth, or for a period of 25 years. The objective of the BABYDIAB study is to determine when islet autoantibodies first appear, which genetic factors and environmental factors influence their development, and which characteristics of the autoantibodies are most strongly associated with the development of type 1 diabetes. The participants in the study are reexamined every three years by means of blood samples and questionnaires. The BABYDIET is examining the influence of food containing gluten on the development of type 1 diabetes. Of the 2,441 children included in the two studies, so far 124 have developed a precursor to diabetes. 82 of these meanwhile display a clinical disease (as of November 2014). ** Peptides are molecules that, like proteins, are constructed from amino acids. However, they are smaller and to some extent result as fragments during protein breakdown. The transition is therefore relatively fluid. The study was financed by the Juvenile Diabetes Research Foundation (JDRF), which has headquarters in the USA. The number of new cases of type 1 diabetes each year continues to rise. New immunotherapeutic approaches aim at stopping this development. A precise assessment of the individual stage of disease development is an important criterion for the targeted use of new treatments. The described study shows that children already display proteomic changes in the blood during the presymptomatic stage. This information allows a better assessment of the time until clinical manifestation of the disease. Recently scientists in the Protein Science Research Unit were also able to identify biomarkers for the precursor to type 2 diabetes: https:/ Von Toerne, C. & Laimighofer, M. et al. (2016): Peptide serum markers in islet autoantibody-positive children. Diabetologia, doi: 10.1007/s00125-016-4150-x http://link. The presence of certain proteins in blood samples can predict incipient type 1 diabetes. The researchers identify these in their measurements using so-called peptide peaks (see selection in red). Source: Helmholtz Zentrum München The Helmholtz Zentrum München, the German Research Center for Environmental Health, pursues the goal of developing personalized medical approaches for the prevention and therapy of major common diseases such as diabetes and lung diseases. To achieve this, it investigates the interaction of genetics, environmental factors and lifestyle. The Helmholtz Zentrum München is headquartered in Neuherberg in the north of Munich and has about 2,300 staff members. It is a member of the Helmholtz Association, a community of 18 scientific-technical and medical-biological research centers with a total of about 37,000 staff members. http://www. The independent Research Unit Protein Science (PROT) investigates the composition of protein complexes and their integration into cellular processes and protein networks. One focus is the analysis of the interaction of genetic variance and environmental factors in neurodegenerative and metabolic diseases. The aim of this research is to identify biological systems and disease-associated disorders on a systemic level, thus contributing to a molecular understanding of diseases. http://www. The Core Facility Proteomics is an instrumental analysis platform at the Helmholtz Zentrum München. It provides interested research groups with access to comprehensive proteome analyses conducted with highly sensitive mass spectrometers. The portfolio ranges from technical and scientific consultation during project design and sample preparation to the development of optimized analysis methods to actual sample measurement and data evaluation. The Institute of Diabetes Research (IDF) focuses on the pathogenesis and prevention of type 1 diabetes and type 2 diabetes and the long-term effects of gestational diabetes. A major project is the development of an insulin vaccination against type 1 diabetes. The IDF conducts long-term studies to examine the link between genes, environmental factors and the immune system for the pathogenesis of type 1 diabetes. Findings of the BABYDIAB study, which was established in 1989 as the world's first prospective birth cohort study, identified risk genes and antibody profiles. These permit predictions to be made about the pathogenesis and onset of type 1 diabetes and will lead to changes in the classification and the time of diagnosis. The IDF is part of the Helmholtz Diabetes Center (HDC). http://www. The German Center for Diabetes Research (DZD) is a national association that brings together experts in the field of diabetes research and combines basic research, translational research, epidemiology and clinical applications. The aim is to develop novel strategies for personalized prevention and treatment of diabetes. Members are Helmholtz Zentrum München - German Research Center for Environmental Health, the German Diabetes Center in Düsseldorf, the German Institute of Human Nutrition in Potsdam-Rehbrücke, the Paul Langerhans Institute Dresden of the Helmholtz Zentrum München at the University Medical Center Carl Gustav Carus of the TU Dresden and the Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Zentrum München at the Eberhard-Karls-University of Tuebingen together with associated partners at the Universities in Heidelberg, Cologne, Leipzig, Lübeck and Munich. https:/
News Article | February 16, 2017
The global proliferation of overweight and obese people and people with type 2 diabetes is often associated with the consumption of saturated fats. Scientists at the German Diabetes Center (Deutsches Diabetes-Zentrum, DDZ) and the Helmholtz Center in Munich (HMGU) have found that even the one-off consumption of a greater amount of palm oil reduces the body's sensitivity to insulin and causes increased fat deposits as well as changes in the energy metabolism of the liver. The results of the study provide information on the earliest changes in the metabolism of the liver that in the long term lead to fatty liver disease in overweight persons as well as in those with type 2 diabetes. In the current issue of the "Journal of Clinical Investigation", DZD researchers working at the German Diabetes Center, in conjunction with the Helmholtz Center in Munich and colleagues from Portugal, published a scientific investigation conducted on healthy, slim men, who were given at random a flavored palm oil drink or a glass of clear water in a control experiment. The palm oil drink contained a similar amount of saturated fat as two cheeseburgers with bacon and a large portion of French fries or two salami pizzas. The scientists showed that this single high-fat meal sufficed to reduce the insulin action, e.g. cause insulin resistance and increase the fat content of the liver. In addition, changes in the energy balance of the liver were proven. The observed metabolic changes were similar to changes observed in persons with type 2 diabetes or non-alcoholic fatty liver disease (NAFLD). NAFLD is the most common liver disease in the industrial nations and associated with obesity, the so-called "metabolic syndrome," and is associated with an increased risk in developing type 2 diabetes. Furthermore, NAFLD in advanced stages can result in severe liver damage. "The surprise was that a single dosage of palm oil has such a rapid and direct impact on the liver of a healthy person and that the amount of fat administered already triggered insulin resistance", explained Prof. Dr. Michael Roden, scientist, Managing Director and Chairman at the DDZ and the German Center for Diabetes Research (Deutsches Zentrum für Diabetesforschung, DZD). "A special feature of our study is that we monitored the liver metabolism of people with a predominantly non-invasive technology, e.g. by magnetic resonance spectroscopy. This allows us to track the storage of sugar and fat as well as the energy metabolism of the mitochondria (power plants of the cell)." Thanks to the new methods of investigation, the scientists were able to verify that the intake of palm oil affects the metabolic activity of muscles, liver and fatty tissue. The induced insulin resistance leads to an increased new formation of sugar in the liver with a concomitant decreased sugar absorption in the skeletal muscles - a mechanism that makes the glucose level rise in persons afflicted with type 2 diabetes and its pre-stages. In addition, the insulin resistance of the fatty tissue causes an increased release of fats into the blood stream, which in turn continues to foster the insulin resistance. The increased availability of fat leads to an increased workload for the mitochondria, which can in the long term overtax these cellular power plants and contribute to the emergence of a liver disease. The team of Prof. Roden suspects that healthy people, depending on genetic predisposition, can easily manage this direct impact of fatty food on the metabolism. The long-term consequences for regular eaters of such high-fat meals can be far more problematic, however. This paper is promoted by the Federal Ministry of Health, the Ministry for Innovation, Science and Research of the state of North Rhine-Westphalia, the Federal Ministry for Education and Research (Deutsches Zentrum für Diabetesforschung e.V.), as well as the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG), the German Diabetes Society (DDG) and the Schmutzler Foundation. Elisa Álvarez Hernández, Sabine Kahl, Anett Seelig, Paul Begovatz, Martin Irmler, Yuliya Kupriyanova, Bettina Nowotny, Peter Nowotny, Christian Herder, Cristina Barosa, Filipa Carvalho, Jan Rozman, Susanne Neschen, John G. Jones, Johannes Beckers, Martin Hrab? de Angelis and Michael Roden, Acute dietary fat intake initiates alterations in energy metabolism and insulin resistance, J Clin Invest. 2017, January 23, 2017. doi:10.1172/JCI89444. The German Diabetes Center (DDZ) is the German reference center for diabetes. The goal is to contribute to the prevention, early detection, diagnosis and treatment of diabetes mellitus. At the same time, the research center aims at improving the epidemiological data situation in Germany. DDZ is in charge of the multi-center German Diabetes Study. It is the point of contact for all players in the health sector. In addition, it prepares scientific information on diabetes mellitus and makes it available to the public. DDZ is part of "Wissenschaftsgemeinschaft Gottfried Wilhelm Leibniz" (WGL) and is a partner of the German Center for Diabetes Research (DZD e.V.). The German Center for Diabetes Research (DZD) is a national association that brings together experts in the field of diabetes research and combines basic research, translational research, epidemiology and clinical applications. The aim is to develop novel strategies for personalized prevention and treatment of diabetes. Members are Helmholtz Zentrum München - German Research Center for Environmental Health, the German Diabetes Center in Düsseldorf, the German Institute of Human Nutrition in Potsdam-Rehbrücke, the Paul Langerhans Institute Dresden of the Helmholtz Zentrum München at the University Medical Center Carl Gustav Carus of the TU Dresden and the Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Zentrum München at the Eberhard-Karls-University of Tuebingen together with associated partners at the Universities in Heidelberg, Cologne, Leipzig, Lübeck and Munich. The Helmholtz Zentrum München, the German Research Center for Environmental Health, pursues the goal of developing personalized medical approaches for the prevention and therapy of major common diseases such as diabetes and lung diseases. To achieve this, it investigates the interaction of genetics, environmental factors and lifestyle. The Helmholtz Zentrum München is headquartered in Neuherberg in the north of Munich and has about 2,300 staff members. It is a member of the Helmholtz Association, a community of 18 scientific-technical and medical-biological research centers with a total of about 37,000 staff members
News Article | December 22, 2016
The extra pounds you gain during the holidays will not only show up on your hips but will also affect your DNA. This is the result of a large-scale international study coordinated by Helmholtz Zentrum München, a partner in the German Center for Diabetes Research, which has now been published in 'Nature'. The study shows that a high BMI leads to epigenetic changes at nearly 200 loci of the genome - with effects on gene expression. While our genes do not change in the course of life, our lifestyle can directly influence their surroundings. Scientists speak here of the epigenome (Greek epi: over, outside of, around), which refers to everything that happens on or around the genes. Up to now there has not been much research on how the epigenome is altered as a result of being overweight. "This issue is particularly relevant because an estimated one and a half billion people throughout the world are overweight," said first author Dr. Simone Wahl of the Research Unit Molecular Epidemiology (AME) at Helmholtz Zentrum München, "especially considering that being overweight can have adverse consequences and lead to diabetes and diseases of the cardiovascular and metabolic systems." For this reason, the international research team led by Dr. Christian Gieger and Dr. Harald Grallert of the AME (as well as Jaspal Kooner and John Chambers of Imperial College London) examined possible correlations between body mass index (BMI) and epigenetic changes.* Using state-of-the-art technology, the team carried out the world's largest study so far on the subject. The scientists examined the blood samples of over 10,000 women and men from Europe. A large proportion of these were inhabitants of London of Indian ancestry, who according to the authors are at high risk for obesity and metabolic diseases. In a first step with 5,387 samples **, the research team identified 207 gene loci that were epigenetically altered dependent on the BMI. They then tested these candidate loci in blood samples of an additional 4,874 subjects and were able to confirm 187 of these***. Further studies and long-term observations also indicated that the changes were predominantly a consequence of being overweight - not the cause. "In particular, significant changes were found in the expression of genes responsible for lipid metabolism and substrate transport, but inflammation-related gene loci were also affected," said group leader Harald Grallert. From the data, the team was also able to identify epigenetic markers that could predict the risk of type 2 diabetes. "Our results allow new insights into which signaling pathways are influenced by obesity", said Christian Gieger, head of the AME. "We hope that this will lead to new strategies for predicting and possibly preventing type 2 diabetes and other consequences of being overweight." Next, within the framework of translational research in the German Center for Diabetes Research, the researchers want to investigate in detail how the epigenetic changes affect the expression of the underlying genes. * Specifically, the team investigated the methylation patterns, i.e. the presence or absence of methyl groups on the DNA. By means of high-throughput measurements, these methylation patterns can now be investigated relatively quickly and on a large scale. ** Among others from the Augsburg KORA study, the London LOLIPOP study and a part of the EPICOR study population from Italy *** Some of these have also been confirmed in adipose tissue, indicating that changes in gene regulation in disease-relevant tissues are also visible in the blood. Helmholtz Zentrum München has extensive expertise in the field of genetic and epigenetic causal research on metabolic diseases: As recently as July 2016, the researchers were involved in the world's largest genetic study on type 2 diabetes, which was likewise published in the renowned journal Nature. Link to press release: https:/ Furthermore, already in March 2016 scientists of Helmholtz München showed that diet-induced obesity and diabetes can be passed on epigenetically to the offspring via both oocytes and sperm. Link to related press release: https:/ Wahl, S. et al. (2016): Epigenome-wide association study of body mass index, and the adverse outcomes of adiposity. Nature, doi:10.1038/nature20784 http://www. The Helmholtz Zentrum München, the German Research Center for Environmental Health, pursues the goal of developing personalized medical approaches for the prevention and therapy of major common diseases such as diabetes and lung diseases. To achieve this, it investigates the interaction of genetics, environmental factors and lifestyle. The Helmholtz Zentrum München is headquartered in Neuherberg in the north of Munich and has about 2,300 staff members. It is a member of the Helmholtz Association, a community of 18 scientific-technical and medical-biological research centers with a total of about 37,000 staff members. http://www. The Research Unit of Molecular Epidemiology (AME) analyses population-based cohorts and case studies for specific diseases, using genomics, epigenomics, transcriptomics, proteomics, metabolomics and functional analyses. The aim of this research unit is to decipher the molecular mechanisms of complex diseases like type 2 diabetes or obesity. The unit administers the biological specimen repository of the Department of Epidemiology and stores the samples for national and international projects. http://www. The Institute of Epidemiology II (EPI II) focuses on the assessment of environmental and lifestyle risk factors which jointly affect major chronic diseases such as diabetes, heart disease and mental health. Research builds on the unique resources of the KORA cohort, the KORA myocardial infarction registry, and the KORA aerosol measurement station. Aging-related phenotypes have been added to the KORA research portfolio within the frame of the Research Consortium KORA-Age. The institute's contributions are specifically relevant for the population as modifiable personal risk factors are being researched that could be influenced by the individual or by improving legislation for the protection of public health. http://www. Research at the Institute of Genetic Epidemiology (IGE) focuses on planning, realization and analysis of projects regarding the identification of genetic factors responsible for complex traits. This involves application, further development and implementation of a variety of statistical methods to address specific aspects such as rare genetic variants, mitochondrial DNA, gene-gene and gene-environment interactions, family studies, and the handling of population structures. The elucidation of disease-relevant genetic factors as well as their inclusion into models of disease risk provides the basis of individualized approaches to treatment or prevention. http://www. The Institute of Human Genetics (IHG) at the Helmholtz Zentrum München and the Technical University of Munich: The Institute is concerned with identifying genes associated with disease and characterizing their functions. The main aim of the research projects is to develop disease-related genetic variation in humans and mice as well as to develop chromosome analysis techniques and new methods for dealing with specific issues in the sphere of pre- and post-natal diagnostics and tumor cytogenetics. http://www. The German Center for Diabetes Research (DZD) is a national association that brings together experts in the field of diabetes research and combines basic research, translational research, epidemiology and clinical applications. The aim is to develop novel strategies for personalized prevention and treatment of diabetes. Members are Helmholtz Zentrum München - German Research Center for Environmental Health, the German Diabetes Center in Düsseldorf, the German Institute of Human Nutrition in Potsdam-Rehbrücke, the Paul Langerhans Institute Dresden of the Helmholtz Zentrum München at the University Medical Center Carl Gustav Carus of the TU Dresden and the Institute for Diabetes Research and Metabolic Diseases of the Helmholtz Zentrum München at the Eberhard-Karls-University of Tuebingen together with associated partners at the Universities in Heidelberg, Cologne, Leipzig, Lübeck and Munich. http://www.
Phieler J.,TU Dresden |
Garcia-Martin R.,TU Dresden |
Lambris J.D.,University of Pennsylvania |
Chavakis T.,TU Dresden |
Chavakis T.,Paul Langerhans Institute Dresden
Seminars in Immunology | Year: 2013
Emerging evidence points to a close crosstalk between metabolic organs and innate immunity in the course of metabolic disorders. In particular, cellular and humoral factors of innate immunity are thought to contribute to metabolic dysregulation of the adipose tissue or the liver, as well as to dysfunction of the pancreas; all these conditions are linked to the development of insulin resistance and diabetes mellitus. A central component of innate immunity is the complement system. Interestingly, the classical view of complement as a major system of host defense that copes with infections is changing to that of a multi-functional player in tissue homeostasis, degeneration, and regeneration. In the present review, we will discuss the link between complement and metabolic organs, focusing on the pancreas, adipose tissue, and liver and the diverse effects of complement system on metabolic disorders. © 2013 Elsevier Ltd.
Borg D.J.,TU Dresden |
Borg D.J.,Paul Langerhans Institute Dresden |
Weigelt M.,TU Dresden |
Weigelt M.,Paul Langerhans Institute Dresden |
And 11 more authors.
Diabetologia | Year: 2014
Aims/hypothesis: Islet transplantation is used therapeutically in a minority of patients with type 1 diabetes. Successful outcomes are hampered by early islet beta cell loss. The adjuvant co-transplantation of mesenchymal stromal cells (MSCs) has the promise to improve islet transplant outcome. Methods: We used a syngeneic marginal islet mass transplantation model in a mouse model of diabetes. Mice received islets or islets plus 250,000 MSCs. Kidney subcapsule, intra-hepatic and intra-ocular islet transplantation sites were used. Apoptosis, vascularisation, beta cell proliferation, MSC differentiation and laminin levels were determined by immunohistochemical analysis and image quantification post-transplant. Results: Glucose homeostasis after the transplantation of syngeneic islets was improved by the co-transplantation of MSCs together with islets under the kidney capsule (p = 0.01) and by intravenous infusion of MSCs after intra-hepatic islet transplantation (p = 0.05). MSC co-transplantation resulted in reduced islet apoptosis, with reduced numbers of islet cells positive for cleaved caspase 3 being observed 14 days post-transplant. In kidney subcapsule, but not in intra-ocular islet transplant models, we observed increased re-vascularisation rates, but not increased blood vessel density in and around islets co-transplanted with MSCs compared with islets that were transplanted alone. Co-transplantation of MSCs did not increase beta cell proliferation, extracellular matrix protein laminin production or alpha cell numbers, and there was negligible MSC transdifferentiation into beta cells. Conclusions/ interpretation: Co-transplantation of MSCs may lead to improved islet function and survival in the early post-transplantation period in humans receiving islet transplantation. © 2013 Springer-Verlag Berlin Heidelberg.
Boguslawska D.M.,University of Zielona Gora |
Machnicka B.,University of Zielona Gora |
Hryniewicz-Jankowska A.,Wrocław University |
Czogalla A.,Wrocław University |
Czogalla A.,Paul Langerhans Institute Dresden
Cellular and Molecular Biology Letters | Year: 2014
The spectrin-based membrane skeleton is crucial for the mechanical stability and resilience of erythrocytes. It mainly contributes to membrane integrity, protein organization and trafficking. Two transmembrane protein macro-complexes that are linked together by spectrin tetramers play a crucial role in attaching the membrane skeleton to the cell membrane, but they are not exclusive. Considerable experimental data have shown that direct interactions between spectrin and membrane lipids are important for cell membrane cohesion. Spectrin is a multidomain, multifunctional protein with several distinctive structural regions, including lipid-binding sites within CH tandem domains, a PH domain, and triple helical segments, which are excellent examples of ligand specificity hidden in a regular repetitive structure, as recently shown for the ankyrin-sensitive lipid-binding domain of beta spectrin. In this review, we summarize the state of knowledge about interactions between spectrin and membrane lipids. © 2013 Versita Warsaw and Springer-Verlag Wien.
Samikannu B.,Justus Liebig University |
Chen C.,Paul Langerhans Institute Dresden |
Lingwal N.,Justus Liebig University |
Padmasekar M.,Justus Liebig University |
And 2 more authors.
PLoS ONE | Year: 2013
Substitution of pancreatic islets is a potential therapy to treat diabetes and it depends on reconstitution of islet's capillary network. In this study, we addressed the question whether stabilization of Glucagon-Like-Peptide-1 (GLP-1) by inhibiting Dipeptidyl Peptidase-IV (DPP-IV) increases β-cell mass by modulating vascularization. Mouse or porcine donor islets were implanted under kidney capsule of diabetic mice treated with DPP-IV inhibitor sitagliptin. Grafts were analyzed for insulin production, β-cell proliferation and vascularization. In addition, the effect of sitagliptin on sprouting and Vascular Endothelial Growth Factor (VEGF)-A expression was examined ex vivo. The cAMP response element-binding (CREB) and VEGF-A/Vascular Endothelial Growth Factor Receptor (VEGFR)-2 signaling pathway leading to islet vascularization was explored. Sitagliptin increased mean insulin content of islet grafts and area of insulin-positive tissue as well as β-cell proliferation. Interestingly, sitagliptin treatment also markedly increased endothelial cell proliferation, microvessel density and blood flow. Finally, GLP-1 (7-36) stimulated sprouting and VEGF expression, which was significantly enhanced by sitagliptin- mediated inhibition of DPP-IV. Our in vivo data demonstrate that sitagliptin treatment phosphorylated CREB and induced islet vascularization through VEGF-A/VEGFR-2 signaling pathway. This study paves a new pathway for improvement of islet transplantation in treating diabetes mellitus. © 2013 Samikannu et al.
Sturm D.,Paul Langerhans Institute Dresden
Journal of visualized experiments : JoVE | Year: 2013
Laser microdissection (LMD) is a technique that allows the recovery of selected cells and tissues from minute amounts of parenchyma. The dissected cells can be used for a variety of investigations, such as transcriptomic or proteomic studies, DNA assessment or chromosomal analysis. An especially challenging application of LMD is transcriptome analysis, which, due to the lability of RNA, can be particularly prominent when cells are dissected from tissues that are rich of RNases, such as the pancreas. A microdissection protocol that enables fast identification and collection of target cells is essential in this setting in order to shorten the tissue handling time and, consequently, to ensure RNA preservation. Here we describe a protocol for acquiring human pancreatic beta cells from surgical specimens to be used for transcriptomic studies. Small pieces of pancreas of about 0.5-1 cm(3) were cut from the healthy appearing margins of resected pancreas specimens, embedded in Tissue-Tek O.C.T. Compound, immediately frozen in chilled 2-Methylbutane, and stored at -80 °C until sectioning. Forty serial sections of 10 μm thickness were cut on a cryostat under a -20 °C setting, transferred individually to glass slides, dried inside the cryostat for 1-2 min, and stored at -80 °C. Immediately before the laser microdissection procedure, sections were fixed in ice cold, freshly prepared 70% ethanol for 30 sec, washed by 5-6 dips in ice cold DEPC-treated water, and dehydrated by two one-minute incubations in ice cold 100% ethanol followed by xylene (which is used for tissue dehydration) for 4 min; tissue sections were then air-dried afterwards for 3-5 min. Importantly, all steps, except the incubation in xylene, were performed using ice-cold reagents - a modification over a previously described protocol. utilization of ice cold reagents resulted in a pronounced increase of the intrinsic autofluorescence of beta cells, and facilitated their recognition. For microdissection, four sections were dehydrated each time: two were placed into a foil-wrapped 50 ml tube, to protect the tissue from moisture and bleaching; the remaining two were immediately microdissected. This procedure was performed using a PALM MicroBeam instrument (Zeiss) employing the Auto Laser Pressure Catapulting (AutoLPC) mode. The completion of beta cell/islet dissection from four cryosections required no longer than 40-60 min. Cells were collected into one AdhesiveCap and lysed with 10 μl lysis buffer. Each single RNA specimen for transcriptomic analysis was obtained by combining 10 cell microdissected samples, followed by RNA extraction using the Pico Pure RNA Isolation Kit (Arcturus). This protocol improves the intrinsic autofluorescence of human beta cells, thus facilitating their rapid and accurate recognition and collection. Further improvement of this procedure could enable the dissection of phenotypically different beta cells, with possible implications for better understanding the changes associated with type 2 diabetes.
Bluher S.,University of Leipzig |
Schwarz P.,TU Dresden |
Schwarz P.,Paul Langerhans Institute Dresden
Metabolism: Clinical and Experimental | Year: 2014
Up to 30% of obese people do not display the "typical" metabolic obesity-associated complications. For this group of patients, the term "metabolically healthy obese (MHO)" has been established during the past years and has been the focus of research activities. The development and severity of insulin resistance as well as (subclinical) inflammations seems to play a key role in distinguishing metabolically healthy from metabolically non-healthy individuals. However, an internationally consistent and accepted classification that might also include inflammatory markers as well as features of non-alcoholic fatty liver disease is missing to date, and available data - in terms of prevalence, definition and severity - are heterogeneous, both during childhood/adolescence and during adulthood. In addition, the impact of MHO on future morbidity and mortality compared to obese, metabolically non-healthy as well as normal weight, metabolically healthy individuals is absolutely not clear to date and even conflicting. This review summarizes salient literature related to that topic and provides insight into our current understanding of MHO, covering all age spans from childhood to adulthood. © 2014 Elsevier Inc.