Type 1 Diabetes: Cellular, Molecular & Clinical ImmunologyChapter 2 - The Pancreatic Beta-CellSuparna A. Sarkar, 3/1/2012 Update of chapter by Kirstine Juhl, John C. Hutton and George S. EisenbarthCell Therapy of Diabetes PowerPoint slide set-Updated 7/06Proprotein Processing and Pancreatic Islet Function PowerPoint slide set-Updated 11/06Stimulus-Secretion Coupling in the Pancreatic Beta-Cell PowerPoint slide set- Updated 7/08Human Fetal Pancreas Development slide setUpdated 03/12IntroductionThe two most common forms of diabetes in man (Type 1A and Type 2) have very different etiologies and different clinical presentation1. Nevertheless, the underlying loss of islet beta cell function has similar consequences in terms of glycemic control and the emergence of long-term complications. type 1 Diabetes (T1D) is a polygenic T-cell dependent autoimmune disease, characterized by the selective destruction of the -cells of the islets of Langerhans1-6and that susceptible individuals have inherent defects in critical immunomodulatory mechanisms7that increase the risk of a pathogenic rather than protective immune response to self6, 8, 9. Type 2 Diabetes is typically linked to dysmetabolism or metabolic syndrome and the presence of insulinresistance, however a large subset of T1D patients routinely exhibits insulin resistance10-13contributing to the metabolic distress in islets.With the rising incidence of T1D and T2D, it is now being argued that both T1D and T2D are essentially disorders of altered insulin resistance set against the backdrop of genetic susceptibility14and the inflammatory process; in T1D brought about by the autoimmune component of the disease process15.In type 1A (autoimmune diabetes) the loss of beta cells is often close to absolute with less than 1% of beta cells remaining in patients with long-term diabetes16-18with prolonged C peptide production19.In most patients with almost no remaining beta cells, essentially all of the islets are devoid of beta cells while islets contain cells expressing glucagon and somatostatin. Such islets are termed pseudoatrophic islets20. Nevertheless, some beta cells remain often as scattered single cells in the parenchyma and ducts.In a small subset of patients, even with long-term type 1A diabetes, significant C-peptide is present and lobules of pancreas remain where all the islets contain beta cells and appear essentially normal in terms of expression of insulin while the rest of the pancreas is devoid of beta cells in islets20-22. The figure below illustrates a section of one such pancreas from the nPOD collection (jdrfnpod.org) where the pancreatic lobule on the right is stained dark and at higher power one can observe that all of the islets in this lobule lack insulin23.In contrast to the lobule on the left, all of the islets contain insulin.The dark staining of the right lobule likely results from pancreatic acinar atrophy that occurs with severe loss of pancreatic insulin.Shrinkage in overall pancreatic mass in patients with type 1 diabetes has long been noted24-26. Analysis of decreased pancreatic volume was recently combined with imaging of iron particle pancreatic accumulation to help distinguish patients with type 1 diabetes from normal controls27-29.In fact,C-peptide secretion in long-standing diabetic patients has now been explained by two different patterns of beta cell survival, which possibly reflect different subsets of type 1 diabetes.In a recent study20associatedPattern A with type 1A diabetes that histologically had lobular retention of islet areas with abnormal beta cells producing the apoptosis inhibitor BIRC5 (survivin) and HLA class I. In pattern B, 100% of all islets contained normal-appearing but quantitatively reduced beta cells without survivin or HLA class I.Baculoviral IAP repeat 5 BiRC530is anapoptosis inhibitor that is produced in the beta cells of fetal human pancreas31, but not in adult islets. It is also found in the beta cells in areas of pancreatitis32. The presence of survivin, in all surviving islet beta cells Pattern A patients, may result from 1) inflammatory changes that did not result in beta cell destruction of a subset of islets or 2) be protected from destruction and further lymphocytic infiltration. An alternative hypothesis extended by the authors is that lobular regions with beta cells of Pattern A pancreas represent areas of beta cell regeneration. Althoughthe sudden onset of type 1A belies the fact that the underlying loss of beta cell mass is the culmination of many years of gradual and progressive loss of beta cells in the face of autoimmune attack which is first evident with the appearance of autoantibodies to islet proteins in the preceding years (see other chapters)33-38. In the NOD mouse the infiltration of the islets with immune and inflammatory cells that initiates the disease first appears in the islets of the pancreatic periphery, affects a subpopulation of islets and is possibly benign or at least kept in check by the presence of regulatory T cells39-42. The invasive insulitis seen in NOD mice closer to disease onset may reflect a change in the balance of destructive and protective responses in favor of the former. The histological changes in man are comparatively mild and may reflect the slower progression of the disease or possibly a different immune process. The islet tends to be viewed as the source of autoantigen that supports or initiates the immune attack and ultimately the victim of the crime.Histopathological examination of pancreata from diabetic organ donors procured from nPOD was examined with the goal to provide a foundation for the informed selection of potential therapeutic targets within the chemokine/receptor family43. CCL5, CCL8, CCL22, CXCL9, CXCL10 and CX3CL1 were the major chemokines transcribed and translated by human islet cells in response toin vitroinflammatory stimuli. CXCL10 was identified as the dominant chemokine expressedin vivoin the islet environment of prediabetic animals and T1D patients, while CCL5, CCL8, CXCL9 and CX3CL1 proteins were present at lower levels in the islets of both species. Importantly, additional expression of the same chemokines in human acinar tissues emphasized an underappreciated involvement of the exocrine pancreas in the natural course of T1D that will require consideration for further T1D pathogenesis and immune intervention studies.Undoubtedly, much more needs to be learned about the reaction of the islet to cytokine mediators of the immune response and about how the beta cell manages to survive so long or replenish its population from progenitor cells in the pancreas. Since the mechanism of autoimmune destruction by effector cells may be mediated by CD4+ cells, and thus indirect, there is also the question of whether the beta cell is uniquely susceptible to oxygen and nitrogen free radicals or cytokine mediators of cell death which may account for the fact that other islet cells exposed to same molecules survive while the beta cell dies.The focus of the following review is to discuss the wealth of information regarding the physiological and pathophysiological responses of the islet to nutrient secretagogues and pharmacological agents and to emphasize how the beta cell differs from its neighbors and from other endocrine tissues and how it may participate in its own demise in type 1 diabetes. The review also illustrates the challenges faced by investigators wishing to genetically engineer non-cells for cellular therapy of type 1 diabetes or wanting to introduce specific genes into the beta cell population to afford it greater protection from autoimmune attack.

Development of the Human PancreasSimilar to the mouse pancreas, the human pancreas develops from two endodermal diverticula, the dorsal and ventral44, which fuses around 56 days post coitum of development45. The pancreas comprises of 3 important cell lineages: Endocrine, acinar and ductal (which together make up the exocrine pancreas). The morphogenesis of the endocrine tissue, however, is unlikely to be equivalent given the differences in gestation (260 vs 20 days) and the larger relative volume of the human pancreas46.Human fetal pancreases obtained at gestational ages 923weeks were processed in parallel for immunohistochemistry and gene expression profiling by Affymetrix microarray47. At 911weeks, the pancreas was made up principally of mesenchymal tissue interspersed with PDX1 positive branched epithelial structures containing scattered hormone-negative neurogenin3-positive endocrine cells. Protoacinar structures marked by carboxy esterase lipase (CEL) expression were noted by 1519weeks, along with clusters of endocrine cells producing either glucagon or insulin. By 2023weeks, vascularized islet-like structures appeared. Analysis of Ki67 immunoreactivity showed that the replicative rate of endocrine cells was low and suggested that the endocrine expansion was derived from hormone-negative precursors. Insulin, glucagon, somatostatin, ghrelin and pancreatic polypeptide transcripts were present at 910weeks as confirmed by quantitative PCR and increased progressively, commensurate with the expansion of endocrine cell volume. The human equivalent of a mouse endocrine secondary transition was not evident, neither in terms of morphology nor in dramatic changes in endocrine-specific transcriptional regulators. By contrast, exocrine genes showed a marked transition at around 11weeks, associated with a greater than six-fold increase in exocrine gene transcripts.The terminal differentiation of human endocrine tissue into late gestation and the presence of NEUROG3 are in contrast with findings in the mouse, where neurog3 is transiently expressed from e12.5e15.5. This indicates that the human fetal pancreas could provide an abundant islet precursor cell population that could be expanded ex vivo for therapeutic transplantation for the treatment of brittle and unstable type 1 diabetes. The ductal cells also develop from the PDX1 expressing primordial pancreatic epithelium and expresses Cytokeratin 19 (CK19), cystic fibrosis transmembrane receptor, DBA lectin, Carbonic anhydrase 248. (For images of the human fetal pancreas development,please refer to power-point slides).Physiology of the islet of Langerhans

The endocrine pancreas is arranged in clusters of secretory cells the Islets of Langerhans scattered throughout the exocrine glandular tissue (Fig. 1)49, 50. In man, the pancreas contains around one million islets that comprise 1-2% of the total mass of the gland. The islets are separated from the exocrine tissue by a capsule made up of connective tissue fibers and by glial like cells and human islets vary in size from less than 50 up to several thousand cells. Four different endocrine cell types are contained in the islets; beta cells, which produce insulin and constitute 60-80% of the endocrine cell mass, glucagon secreting a-cells (10-20%), somatostatin producing d-cells (~5%) and pancreatic polypeptide secreting PP-cells (