Organizer: M. Slak Rupnik (Vienna, Austria); Co-chair: O. Pechanova (Bratislava, Slovakia)
|β cell diversity is required for normal islet function
1 University of Birmingham, Institute of Metabolism and Systems Research (IMSR), Birmingham, United Kingdom
2 Birmingham Health Partners, Centre for Endocrinology, Diabetes and Metabolism, Birmingham, United Kingdom
3 Imperial College London, Section of Cell Biology and Functional Genomics, Department of Medicine, London, United Kingdom
4 Harvard University, Department of Stem Cell and Regenerative Biology, Harvard Stem Cell Institute, Cambridge, United States
Abstract text :
Aim: Insulin-secreting β cells are a heterogeneous islet population whose acitivity is guided by a subgroup of "pacemakers" termed hubs. Since normal islet function may depend on maintaining this diversity, we looked into the effects of overexpression-induced β cell maturity on gene expression, signalling and insulin secretion.
Materials and methods: Maturity in adult mouse islets was forced using an adenoviral polycistronic construct for Ngn3, MafA, Pdx1 and mCherry (Ad3-NPM). Non-transduced islets (CT) served as controls. Gene expression was assessed by QRT-PCR. Pdx1 and insulin were detected by immunohistochemistry. Ca , ATP and cAMP dynamics in live islets were assessed by Nipkow spinning disk microscopy. Glucose-stimulated (GSIS) and incretin-stimulated insulin secretion (ISIS) were HTRF-measured.
Results: Ad3-NPM increased the expression of Pdx1 and MafA, without changing Ngn3. Pdx1 overexpression was predominantly localized to the immature beta cells, reducing cellular heterogeneity. There was a marked decrease in the magnitude of Ca responses to glucose (ΔF=0.81 vs 0.44 AU, CT vs Ad3-NPM; P<0.01), along with a reduction in β cell-β cell coordination and hub number (12.6 vs 5.6 % hubs, CT vs Ad3-NPM; P<0.05). Ad3-NPM islets showed an increase in the ATP/ADP ratio and a marked reduction of the cAMP. Basal insulin secretion was increased post-overexpression, with impaired GSIS and ISIS (7.5-fold vs 5-fold post-glucose and 98-fold vs 50-fold after exendin-4 stimulation; CT vs Ad3-NPM).
Summary: Disrupting β cell heterogeneity through forced overexpression weakens islet function and lowers insulin secretion. This suggests that cell diversity plays a critical role in islet performance.
|Induction of pancreatic beta-cell neogenesis
1 INSERM U1091, , Nice, France
Abstract text :
The recent discovery that genetically-modified pancreatic alpha-cells can regenerate and convert into beta-like cells in vivo holds great promise for diabetes research. However, to eventually translate these findings to human, it is crucial to discover compounds with similar activities. Herein, we report the identification of GABA as an inducer of alpha-to-beta-like cell conversion in vivo. This conversion induces alpha-cell replacement mechanisms through the mobilization of duct-lining precursor cells that adopt an alpha-cell identity prior to being converted into beta-like cells, solely upon sustained GABA exposure. Importantly, these neo-generated beta-like cells are functional and can repeatedly reverse chemically-induced diabetes in vivo. Similarly, the treatment of transplanted human islets with GABA results in a loss of alpha-cells and a concomitant increase in beta-like cell counts, suggestive of alpha-to-beta-like cell conversion processes also in humans. This newly discovered GABA-induced alpha-cell-mediated beta-like cell neogenesis could therefore represent an unprecedented hope towards improved therapies for diabetes.
|The patterns of synchronicity and functional connectivity in islets of Langerhans
1 University of Maribor, Institute of Physiology, Maribor, Slovenia
Abstract text :
Classical morphological and physiological approaches, such as gross inspection, light and electron microscopy, blood analyses and secretion assays, in concert with classical descriptive and inferential statistics, have taught us a lot about the structure and function of islets of Langerhans, revealing assemblies of coupled heterogenous beta cells producing oscillations in metabolism, membrane potential, cytosolic calcium, and secretion of insulin that are probably crucial for oscillations in plasma insulin and normal glucose tolerance. Recently, high resolution in situ morphological and functional live cell imaging experiments, together with graph theoretical analyses and mathematical modelling, have revealed that heterogeneity, coupling, and oscillations are probably inseparably linked and necessary to ensure efficient synchronization, robustness, and adaptability of beta cell assemblies in the long run. These insights opened doors to modern network science in islet physiology and are forcing us to rethink what we know about the structure and function of islets of Langerhans and possibly other endocrine tissues, and are beginning to change our understanding of diseases, such as diabetes mellitus.
|Heterogeneity on all levels: insight into pancreatic islet function with modeling
1 University of Padova, Information Engineering, Padova, Italy
Abstract text :
Hormone secretion from pancreatic islets results from complex regulatory mechanisms operating on multiple biological scales. On each of these levels, there is pronounced heterogeneity between the functional units. At the lowest subcellular level, the members of ion channel populations are exposed to uneven biophysical conditions, and secretory granules show heterogeneity in, e.g., positioning with respect to calcium sources and local protein abundance. At higher levels, there are cell-to-cell and islet-to-islet differences in e.g. glucose sensitivity. Mathematical modeling has been used to investigate how the pancreas exploits such heterogeneity to create particular secretion patterns, while – at the same time – taming heterogeneous behavior to provide robust responses to changing glucose levels. Our recent modeling has suggested that cell-to-cell variation in combination with electrical coupling within the islets can lead to so-called functional small-world behavior as a result of wave propagation. Concerning the subcellular levels, I will show how to exploit heterogeneity in granule properties to quantify how e.g. local protein abundance controls exocytosis with advanced statistical methods. I will discuss how neglecting such subcellular heterogeneity may lead to misleading results concerning exocytosis or ion channel function. Finally, multi-scale modeling can be used to link the physiological levels to provide insight into how pancreatic secretion and cell function is influenced by subcellular and molecular events.
|Investigating SNAP-25b (Synaptosomal-associated protein 25) function in mouse islet physiology beyond its classical role in membrane fusion
1 Karolinska Institutet, The Rolf Luft Research Center for Diabetes and Endocrinology, Department of Molecular Medicine and Surgery, Stockholm, Sweden
2 Institute of Physiology, Faculty of Medicine, University of Maribor, , Maribor, Slovenia
3 Department of Physics, Faculty of Natural Sciences and Mathematics, University of Maribor, , Maribor, Slovenia
4 Center for Physiology and Pharmacology, Medical University of Vienna, , Vienna, Austria
Abstract text :
SNAP-25 is a core protein of the SNARE complex mediating stimulus-dependent release of insulin from pancreatic β cells. The protein exists as two alternative spliced isoforms, SNAP-25a and SNAP-25b, differing in 9 out of 206 amino acids but their specific roles in β cells remain unclear.
We explored the effect of SNAP-25b-deficiency on glucose-stimulated insulin release in islets and found increased secretion both in vivo and in vitro. However, slow photo-release of caged Ca2 in β cells within pancreatic slices showed no significant differences in Ca2 -sensitivity, amplitude or rate of exocytosis between SNAP-25b-deficient and wild-type littermates. Therefore, we next investigated if SNAP-25b-deficiency affected Ca2 handling in glucose-stimulated β cells using intracellular Ca2 -imaging and found premature activation and delayed termination of [Ca2 ]i elevations. These finding were accompanied by less synchronized Ca2 -oscillations and hence more segregated functional networks between individual β cells. Islet gross morphology and architecture were maintained in mutant mice, although sex specific compensatory changes were observed.
In summary, our observations suggest that SNAP-25b in pancreatic β cells, except for participating in the core SNARE complex, plays an important role in regulating insulin secretion by affecting Ca2 dynamics.
Keywords: exocytosis, β cell, Ca2