Linking human islet structural heterogeneity to beta cell state

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Project Summary/Abstract The detailed structure of the beta cell niche, and that of the islet in general, remains poorly understood; this is particularly the case for human islets. Islet structure appears heterogeneous across the pancreas, and whether conserved structural features exist among islets is unknown. A detailed understanding of the organizational principles of islets would advance our ability both to reconstitute stem-cell derived islets as a cure for type 1 diabetes (T1D) and to block the progression of events that lead to the loss of beta cells during the progression of diabetes. Therefore, the goal of this proposal is twofold: first, to identify and experimentally validate the critical organizational principles of the islet in general and the beta cell niche in particular, and second, to leverage these organizational principles to engineer more functional islets as a cure for T1D. Towards the first goal, we have developed a custom, semi-automated, 3D imaging and analysis pipeline that permits quantification of the statistical properties of the beta cell niche at sub-micron resolution and across hundreds of individual beta cells. Preliminary analyses of healthy mouse and human islets revealed that (1) in both species beta and delta cells maintain at least one physical contact with a source of basement membrane, whereas alpha cells do not, and (2) beta cells in engineered islets that contact sources of vascular basement membrane have dramatically elevated insulin expression. We hypothesize that beta cell contact with basement membrane is a conserved element of islet structure that must be incorporated into engineered islets to optimize beta cell function. Towards the second goal, we have demonstrated that reconstituting stem cell- derived beta cells into pseudo-islets in a manner that maximizes their contact with basement membrane improves their response to glucose by at least two-fold in vitro and further extends their functionality in vivo. Building on these preliminary findings, we first aim to dramatically expand this analysis across tens of thousands of individual cells in human and mouse islets, incorporating all endocrine cell types along with immune cells, vascular cells, and nerves. This will result in the first quantitative assessment of the endocrine cell structural niche that acknowledges the structural heterogeneity of islets and aims to identify conserved structural motifs. Second, we aim to determine if conserved features of the beta cell niche are necessary and sufficient for optimal beta cell function. We will test this hypothesis using in vitro reconstituted islets, primary human islets cultured ex vivo, and engineered human islets transplanted into mice in vivo. Finally, we will use genome editing techniques to test the necessity of specific pericyte-derived basement membrane molecules for glucose homeostatic function in engineered islets. Taken together, our study will provide the first quantitative structural blueprint for the pancreatic islet, will identify features of the beta cell niche that are conserved and divergent across humans and mice, and will demonstrate a strategy for reconstituting more functional human tissues from stem cells that uses a structural blueprint to guide tissue engineering.