Rodent pancreatic islet biology in health and disease

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During 2009, we further developed multicolor flow cytometry and quantitative gene expression systems to characterize endocrine cell number and function in both healthy mice and mice developing or with recent autoimmune diabetes. Specific observations: 1. Using beta cell DNA content analysis (validated using standard BrdU incorporation techniques), we found increased (at least 5-6 fold) beta cell proliferation at diabetes onset compared to beta cells from healthy mice. The observed beta cell proliferation at diabetes onset is consistent with results from severe insulin-resistance mouse models which typically develop marked beta cell hyperplasia over time. We tested whether the beta cell proliferation in autoimmune diabetes is caused by hyperglycemia or the islet inflammation, with data clearly suggesting the former. For instance, proliferating cell frequency is markedly lower in prediabetic mice compared to recently diabetic animals, even though prediabetic mouse islets are severely infiltrated and substantial islet injury has occurred. A manuscript reporting these observations was published this year. 2. Using flow cytometry and fluorescence immunohistochemistry, we have determined the frequency of mouse pancreatic beta cells relative to alpha- or delta-cells during diabetes development. In addition to the expected beta cell depletion, we observed an equally severe loss of alpha and delta cells. This other-than-beta-cell depletion from islets was much more prevalent in larger islets compared with small, sub-islet-sized endocrine cell clusters. Also, during autoimmune beta cell destruction, islet alpha- and delta-cell proliferation rates did not increase, unlike beta cells. Our data suggest that the distribution of endocrine cells between larger islets and small endocrine cell clusters changes as autoimmune diabetes develops, and may play a role in the paradoxical hormonal responses observed in diabetic subjects. For instance, while patients with diabetes typically display higher than normal circulating glucagon levels, the normal glucagon response to hypoglycemia is typically blunted. A manuscript reporting these observations was published this year. 3. We have developed a technique that can quickly and with some precision measure total pancreatic endocrine cell numbers. Using current techniques, a number of agents have been reported to increase "islet cell mass", but such assessments are typically drawn using laborious and imprecise immunohistological techniques. With the flow cytometric systems we are developing, we can track both the decline in beta cell numbers involved in the processes leading to autoimmune diabetes, and the increase in beta cell numbers believed to occur in insulin resistant states or following treatment with certain islet growth factors (e.g. glucagon-like peptide 1 receptor agonists). Further, we have identified techniques for measuring the gene transcripts within individual islet cell subsets, and a manuscript reporting these observations is not "in press."