Michele Solimena

Technische Universität Dresden, Dresden, Saxony, Germany

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Publications (124)1077.03 Total impact

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    ABSTRACT: Insulin secretion from pancreatic β-cells in response to sudden glucose stimulation, is biphasic. Prolonged secretion in vivo requires synthesis, delivery to the plasma membrane (PM) and exocytosis of insulin secretory granules (SGs). Here, we provide the first agent-based space-resolved model for SG dynamics in pancreatic β-cells. Using recent experimental data, we consider a single β-cell with identical SGs moving on a phenomenologically represented cytoskeleton network. A single exocytotic machinery mediates SG exocytosis on the PM. This novel model reproduces the measured spatial organization of SGs and insulin secretion patterns under different stimulation protocols. It proposes that the insulin potentiation effect and the rising second-phase secretion are mainly due to the increasing number of docking sites on the PM. Furthermore, it shows that for 6 min after glucose stimulation, the "newcomer" SGs are recruited from a region within less than 600 nm from the PM. This article is protected by copyright. All rights reserved.
    Traffic 03/2015; DOI:10.1111/tra.12286 · 4.71 Impact Factor
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    ABSTRACT: Insulin secretion is key for glucose homeostasis. Insulin secretory granules (SGs) exist in different functional pools, with young SGs being more mobile and preferentially secreted. However, the principles governing the mobility of age-distinct SGs remain undefined. Using the time-reporter insulin-SNAP to track age-distinct SGs we now show that their dynamics can be classified into three components: highly dynamic, restricted, and nearly immobile. Young SGs display all three components, whereas old SGs are either restricted or nearly immobile. Both glucose stimulation and F-actin depolymerization recruit a fraction of nearly immobile young, but not old, SGs for highly dynamic, microtubule-dependent transport. Moreover, F-actin marks multigranular bodies/lysosomes containing aged SGs. These data demonstrate that SGs lose their responsiveness to glucose stimulation and competence for microtubule-mediated transport over time while changing their relationship with F-actin.
    Proceedings of the National Academy of Sciences 02/2015; DOI:10.1073/pnas.1409542112 · 9.81 Impact Factor
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    ABSTRACT: The type-1 diabetes autoantigen ICA512/IA-2/RPTPN is a receptor protein tyrosine phosphatase of the insulin secretory granules, which regulates the size of granule stores, possibly via cleavage/signaling of its cytosolic tail. The role of its extracellular region, instead, remains unknown. Structural studies indicated that β2- or β4-strands in the mature ectodomain (ME ICA512) form dimers in vitro. Here we show that ME ICA512 prompts proICA512 dimerization in the endoplasmic reticulum. Perturbation of ME ICA512 β2-strand N-glycosylation upon S508A replacement allows for proICA512 dimerization, O-glycosylation, targeting to granules and conversion, which are instead precluded upon G553D replacement in the ME ICA512 β4-strand. S508A/G553D or N506A/G553D double mutants dimerize, but remain in the endoplasmic reticulum. Removal of the N-terminal fragment (ICA512-NTF) preceding ME ICA512 allows instead an ICA512-ΔNTF G553D mutant to exit the endoplasmic reticulum and ICA512-ΔNTF is constitutively delivered to the cell surface. The signal for SG sorting is located within the NTF RESP18-homology domain (RESP18-HD), whereas soluble NTF is retained in the endoplasmic reticulum. Hence, we propose that the ME ICA512 β2-strand fosters proICA512 dimerization until NTF prevents N506 glycosylation. Removal of this constraint allows for proICA512 β4-strand induced dimerization, exit from the endoplasmic reticulum, O-glycosylation and RESP18-HD-mediated targeting to granules. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Molecular and Cellular Biology 01/2015; DOI:10.1128/MCB.00994-14 · 5.04 Impact Factor
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    ABSTRACT: Lateral compositional and physicochemical heterogeneity is a ubiquitous feature of cellular membranes on various length scales, from molecular assemblies to micrometric domains. Segregated lipid domains of increased local order, referred to as rafts, are believed to be prominent features in eukaryotic plasma membranes; however, their exact nature (i.e. size, lifetime, composition, homogeneity) in live cells remains difficult to define. Here we present evidence that both synthetic and natural plasma membranes assume a wide range of lipid packing states with varying levels of molecular order. These states may be adapted and specifically tuned by cells during active cellular processes, as we show for stimulated insulin secretion. Most importantly, these states regulate both the partitioning of molecules between coexisting domains and the bioactivity of their constituent molecules, which we demonstrate for the ligand binding activity of the glycosphingolipid receptor GM1. These results confirm the complexity and flexibility of lipid-mediated membrane organization and reveal mechanisms by which this flexibility could be functionalized by cells.
    PLoS ONE 01/2015; 10(4):e0123930. DOI:10.1371/journal.pone.0123930 · 3.53 Impact Factor
  • Hormone and Metabolic Research 12/2014; 47(01). DOI:10.1055/s-0034-1394453 · 2.04 Impact Factor
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    ABSTRACT: Studies on the cellular function of the pancreas are typically performed in vitro on its isolated functional units, the endocrine islets of Langerhans and the exocrine acini. However, these approaches are hampered by preparation-induced changes of cell physiology and the lack of an intact surrounding. We present here a detailed protocol for the preparation of pancreas tissue slices. This procedure is less damaging to the tissue and faster than alternative approaches, and it enables the in situ study of pancreatic endocrine and exocrine cell physiology in a conserved environment. Pancreas tissue slices facilitate the investigation of cellular mechanisms underlying the function, pathology and interaction of the endocrine and exocrine components of the pancreas. We provide examples for several experimental applications of pancreas tissue slices to study various aspects of pancreas cell biology. Furthermore, we describe the preparation of human and porcine pancreas tissue slices for the validation and translation of research findings obtained in the mouse model. Preparation of pancreas tissue slices according to the protocol described here takes less than 45 min from tissue preparation to receipt of the first slices.
    Nature Protocols 12/2014; 9(12):2809-22. DOI:10.1038/nprot.2014.195 · 7.78 Impact Factor
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    ABSTRACT: Phogrin/IA-2β and ICA512/IA-2 are two paralogs receptor-type protein-tyrosine phosphatases (RPTP) that localize in secretory granules of various neuroendocrine cells. In pancreatic islet β-cells, they participate in the regulation of insulin secretion, ensuring proper granulogenesis, and β-cell proliferation. The role of their cytoplasmic tail has been partially unveiled, while that of their luminal region remains unclear. To advance the understanding of its structure-function relationship, the X-ray structure of the mature ectodomain of phogrin (ME phogrin) at pH 7.4 and 4.6 has been solved at 1.95- and 2.01-Å resolution, respectively. Similarly to the ME of ICA512, ME phogrin adopts a ferredoxin-like fold: a sheet of four antiparallel β-strands packed against two α-helices. Sequence conservation among vertebrates, plants and insects suggests that the structural similarity extends to all the receptor family. Crystallized ME phogrin is monomeric, in agreement with solution studies but in striking contrast with the behavior of homodimeric ME ICA512. The structural details that may cause the quaternary structure differences are analyzed. The results provide a basis for building models of the overall orientation and oligomerization state of the receptor in biological membranes.
    Journal of Structural and Functional Genomics 11/2014; 16(1). DOI:10.1007/s10969-014-9191-0
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    ABSTRACT: Glucose and GLP-1 stimulate not only insulin secretion, but also the post-transcriptional induction of insulin granule biogenesis. This process involves the nucleocytoplasmic translocation of the RNA binding protein PTBP1. Binding of PTBP1 to the 3’-UTRs of mRNAs for insulin and other cargoes of beta cell granules increases their stability. Here we show that glucose enhances also the binding of PTBP1 to the 5’-UTRs of these transcripts, which display IRES activity, and their translation exclusively in a cap-independent fashion. Accordingly, glucose-induced biosynthesis of granule cargoes was unaffected by pharmacological, genetic or Coxsackievirus-mediated inhibition of cap-dependent translation. Infection with Coxsackieviruses, which also depend on PTBP1 for their own cap-independent translation, reduced instead granule stores and insulin release. These findings provide insight into the mechanism for glucose induction of insulin granule production and on how Coxsackieviruses, which have been implicated in the pathogenesis of type 1 diabetes, can foster beta cell failure.
    08/2014; 3(5). DOI:10.1016/j.molmet.2014.05.002
  • Experimental and Clinical Endocrinology & Diabetes 03/2014; 122(03). DOI:10.1055/s-0034-1372025 · 1.76 Impact Factor
  • Experimental and Clinical Endocrinology & Diabetes 03/2014; 122(03). DOI:10.1055/s-0034-1371979 · 1.76 Impact Factor
  • Experimental and Clinical Endocrinology & Diabetes 03/2014; 122(03). DOI:10.1055/s-0034-1371987 · 1.76 Impact Factor
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    ABSTRACT: IntroductionPancreatic islet transplantation is currently restricted to patients with critical metabolic lability due to long-term need for immunosuppression and a persistent shortage of donor organs [1–3]. To overcome these obstacles we have developed a strategy for islet macroencapsulation that provides sufficient immune-isolation whereas regulated islet graft function is maintained [4–8].Case Report and MethodsA 63 year old patient with type 1 diabetes and severe metabolic lability was transplanted with isolated islets (2,000 islets/kgBW) encapsulated in an oxygenated chamber system composed of immune-isolating alginate and polymembrane covers. Via a small abdominal incision, a pre-peritoneal pocket for the chamber was dissected, connected oxygen ports were implanted subcutaneously. No immunosuppressive therapy was applied.ResultsThe procedure was surgically straightforward and without complications. We could demonstrate persistent graft function by detection of endogenous insulin and c-peptide secretion proving islet viability and function. This observation was accompanied by persistent lowering in HbA1c despite reduction in insulin requirement.For oxygenation of the non-vascularized and therefore immune-shielded islet graft, the chamber-integrated gas reservoir was replenished daily via the implanted ports without complications.Conclusion This encapsulation strategy was for the first time applied to allogeneic human islet transplantation in man. We demonstrated a persistent graft function with regulated insulin secretion without any immunosuppressive therapy. This novel concept may allow for future widespread application for cell-based therapies.References[1] 2007 update on allogeneic islet transplantation from the Collaborative Islet Transplant Registry (CITR). Cell Transplant 2009; 18: 753–767.[2] Ludwig, B., Ludwig, S., Steffen, A., Saeger, H.D., Bornstein, S.R. Islet versus pancreas transplantation in type 1 diabetes: competitive or complementary? Curr Diab Rep 2010; 10: 506–511.[3] Mccall, M., James Shapiro, A.M. Update on islet transplantation. Cold Spring Harb Perspect Med 2012; 2: a007823.[4] Barkai, U., Weir G.C., Colton C.K. et al. Enhanced Oxygen Supply Improves Islet Viability in a New Bioartificial Pancreas. Cell Transplant 2013; 22(8): 1463–1476[5] Ludwig, B., Rotem A., Schmid J. et al. Improvement of islet function in a bioartificial pancreas by enhanced oxygen supply and growth hormone releasing hormone agonist. Proc Natl Acad Sci U S A 2012; 109: 5022–5027.[6] Ludwig, B., Zimmermann B., Steffen A. et al. A novel device for islet transplantation providing immune protection and oxygen supply. Horm Metab Res 2010; 42: 918–922.[7] Neufeld, T., Ludwig B., Barkai U. et al. The efficacy of an immunoisolating membrane system for islet xenotransplantation in minipigs. PLoS One 2013; 8: e70150.[8] Ludwig, B., Reichel A, Steffen A et al. Transplantation of human islets without immunosuppression. PNAS 2013; 110: 19054–19058.
    Xenotransplantation 03/2014; 21(2). DOI:10.1111/xen.12083_10 · 1.78 Impact Factor
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    Maria Grazia Magro, Michele Solimena
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    ABSTRACT: β-cells of the pancreatic islets are highly specialized and high-throughput units for the production of insulin, the key hormone for maintenance of glucose homeostasis. Elevation of extracellular glucose and/or GLP-1 levels triggers a rapid upregulation of insulin biosynthesis through the activation of post-transcriptional mechanisms. RNA-binding proteins are emerging as key factors in the regulation of these mechanisms as well as in other aspects of β-cell function and glucose homeostasis at large, and thus may be implicated in the pathogenesis of diabetes. Here we review current research in the field, with a major emphasis on RNA-binding proteins that control biosynthesis of insulin and other components of the insulin secretory granules by modulating the stability and translation of their mRNAs.
    11/2013; 2(4):348-355. DOI:10.1016/j.molmet.2013.09.003
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    ABSTRACT: Transplantation of pancreatic islets is emerging as a successful treatment for type-1 diabetes. Its current stringent restriction to patients with critical metabolic lability is justified by the long-term need for immunosuppression and a persistent shortage of donor organs. We developed an oxygenated chamber system composed of immune-isolating alginate and polymembrane covers that allows for survival and function of islets without immunosuppression. A patient with type-1 diabetes received a transplanted chamber and was followed for 10 mo. Persistent graft function in this chamber system was demonstrated, with regulated insulin secretion and preservation of islet morphology and function without any immunosuppressive therapy. This approach may allow for future widespread application of cell-based therapies.
    Proceedings of the National Academy of Sciences 10/2013; 110(47). DOI:10.1073/pnas.1317561110 · 9.81 Impact Factor
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    ABSTRACT: Obese adipose tissue (AT) inflammation contributes critically to development of insulin resistance. The complement anaphylatoxin C5a receptor (C5aR) has been implicated in inflammatory processes and as regulator of macrophage activation and polarization. However, the role of C5aR in obesity and AT inflammation has not been addressed. We engaged the model of diet-induced obesity and found that expression of C5aR was significantly upregulated in the obese AT, compared with lean AT. In addition, C5a was present in obese AT in the proximity of macrophage-rich crownlike structures. C5aR-sufficient and -deficient mice were fed a high-fat diet (HFD) or a normal diet (ND). C5aR deficiency was associated with increased AT weight upon ND feeding in males, but not in females, and with increased adipocyte size upon ND and HFD conditions in males. However, obese C5aR(-/-) mice displayed improved systemic and AT insulin sensitivity. Improved AT insulin sensitivity in C5aR(-/-) mice was associated with reduced accumulation of total and proinflammatory M1 macrophages in the obese AT, increased expression of IL-10, and decreased AT fibrosis. In contrast, no difference in β cell mass was observed owing to C5aR deficiency under an HFD. These results suggest that C5aR contributes to macrophage accumulation and M1 polarization in the obese AT and thereby to AT dysfunction and development of AT insulin resistance.
    The Journal of Immunology 09/2013; 191(8). DOI:10.4049/jimmunol.1300038 · 5.36 Impact Factor
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    ABSTRACT: Insulin is stored within the secretory granules of pancreatic beta cells and impairment of its release is the hallmark of type 2 diabetes. Preferential exocytosis of newly synthesized insulin suggests that granule aging is a key factor influencing insulin secretion. Here we illustrate a technology that enables the study of granule aging in insulinoma cells and beta cells of knock-in mice through the conditional and unequivocal labeling of insulin fused to the SNAP tag. This approach, which overcomes the limits encountered with previous strategies based on radiolabeling or fluorescence timer proteins, allowed us to formally demonstrate the preferential release of newly-synthesized insulin and reveal that the motility of cortical granules significantly changes over time. Exploitation of this approach may enable the identification of molecular signatures associated with granule aging and unravel possible alterations of granule turnover in diabetic beta cells. Furthermore, the method is of general interest for the study of membrane traffic and aging.
    Diabetes 08/2013; 62(11). DOI:10.2337/db12-1819 · 8.47 Impact Factor
  • Diabetologie und Stoffwechsel 04/2013; 8(S 01). DOI:10.1055/s-0033-1341765 · 0.31 Impact Factor
  • Diabetologie und Stoffwechsel 04/2013; 8(S 01). DOI:10.1055/s-0033-1341713 · 0.31 Impact Factor
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    ABSTRACT: AIMS/HYPOTHESIS: Immunosuppressive drugs used in human islet transplantation interfere with the balance between beta cell renewal and death, and thus may contribute to progressive graft dysfunction. We analysed the influence of immunosuppressants on the proliferation of transplanted alpha and beta cells after syngeneic islet transplantation in streptozotocin-induced diabetic mice. METHODS: C57BL/6 diabetic mice were transplanted with syngeneic islets in the liver and simultaneously abdominally implanted with a mini-osmotic pump delivering BrdU alone or together with an immunosuppressant (tacrolimus, sirolimus, everolimus or mycophenolate mofetil [MMF]). Glycaemic control was assessed for 4 weeks. The area and proliferation of transplanted alpha and beta cells were subsequently quantified. RESULTS: After 4 weeks, glycaemia was significantly higher in treated mice than in controls. Insulinaemia was significantly lower in mice treated with everolimus, tacrolimus and sirolimus. MMF was the only immunosuppressant that did not significantly reduce beta cell area or proliferation, albeit its levels were in a lower range than those used in clinical settings. CONCLUSIONS/INTERPRETATION: After transplantation in diabetic mice, syngeneic beta cells have a strong capacity for self-renewal. In contrast to other immunosuppressants, MMF neither impaired beta cell proliferation nor adversely affected the fractional beta cell area. Although human beta cells are less prone to proliferate compared with rodent beta cells, the use of MMF may improve the long-term outcome of islet transplantation.
    Diabetologia 03/2013; DOI:10.1007/s00125-013-2895-z · 6.88 Impact Factor
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    ABSTRACT: Laser microdissection (LMD) is a technique that allows the recovery of selected cells and tissues from minute amounts of parenchyma (1,2). The dissected cells can be used for a variety of investigations, such as transcriptomic or proteomic studies, DNA assessment or chromosomal analysis (2,3). An especially challenging application of LMD is transcriptome analysis, which, due to the lability of RNA (4), 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 (5). 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 (6). 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.
    Journal of Visualized Experiments 01/2013; DOI:10.3791/50231

Publication Stats

6k Citations
1,077.03 Total Impact Points

Institutions

  • 2003–2015
    • Technische Universität Dresden
      • • Molecular Diabetology
      • • Faculty of Medicine Carl Gustav Carus
      • • Center for Internal Medicine
      Dresden, Saxony, Germany
  • 2014
    • German Diabetes Center
      Düsseldorf, North Rhine-Westphalia, Germany
  • 2013
    • Dresden International University
      Dresden, Saxony, Germany
  • 2009–2013
    • Max Planck Institute of Molecular Cell Biology and Genetics
      Dresden, Saxony, Germany
  • 2010
    • Carl Gustav Carus-Institut
      Pforzheim, Baden-Württemberg, Germany
  • 2005
    • Johns Hopkins Medicine
      • Department of Neurology
      Baltimore, MD, United States
  • 1990–2001
    • Yale-New Haven Hospital
      • Department of Pathology
      New Haven, Connecticut, United States
  • 1994–1998
    • Howard Hughes Medical Institute
      Ashburn, Virginia, United States
    • University of Miami Miller School of Medicine
      • Diabetes Research Institute (DRI)
      Miami, Florida, United States
  • 1993
    • Yale University
      • Department of Cell Biology
      New Haven, Connecticut, United States
  • 1988
    • National Research Council
      Roma, Latium, Italy