This application to join the Consortium on Beta Cell Death and Survival (CBDS) within the HIRN outlines new technologies, of value to all HIRN investigators and specifically requested in the RFA, to provide unprecedented detail in our understanding of the development, plasticity, and molecular signatures of the human pancreatic islet. While it is increasingly clear that rodent and human islets and ? cells have major structurl and expression differences, new data from our groups and others indicate that young (juvenile) human islets are substantially different from adult human islets. New data in humans suggests that the juvenile period (<5 years of age) is an incredibly important developmental period of substantial islet plasticity, but our knowledge of the human pancreatic islet in this tim period is quite limited. For example, the fetal and juvenile determinants of an individual's ?-cel mass, which varies by three- to five-fold in adult humans, are unknown, yet a person's initial ?-cell mass is likely deterministic of if or when one develops T1D. We know little about how the human islet becomes vascularized and innervated, but both are essential for normal islet and ?-cell microenvironment and function. Recent studies demonstrate that ?-cell-directed autoimmunity in individuals with genetic susceptibility to type 1 diabetes (T1D) appears within this same period of the first 5 years of life. The temporal overlap of these two processes generates our over-arching hypothesis that the onset of ?-cell- directed autoimmunity is causally related to ongoing alterations in islet or pancreas architecture, or individual expression states o endocrine cells, including ? cells, during this period of maturation. Previous limitations in the analysis of juvenile tissue result partly from difficulty in procuring human pancreas tissue, but also because standard methods are limited in their resolution and number of markers that can be concurrently analyzed. To address these limitations, our interdisciplinary team brings advanced tissue-clearing technology and multiplexed RNA in situ tissue-analysis approaches, plus substantial experience in studying the human pancreas over the fetal to human juvenile periods, wherein we made essential discoveries about human- specific aspects of the birth and differentiation of endocrine cells, and their assembly into functional islets. The assembled research group incorporates expertise in human and mouse pancreas developmental biology, islet function, and brings in superior methods of analysis towards the production of an atlas of structure and function of the human pancreas over the juvenile period. We propose to: (1) Define the expression of key cell-surface, physiological markers, transcription factors, and other factors to define the sequence of events in human juvenile pancreatic islet development; (2) Define the intra- and inter-islet vascular and neural networks, and these 3D structure-function relationships at the cellular and organ-wide level; (3) Integrate data from Aims #1 and #2 to create a comprehensive profile of the human juvenile pancreatic islet during the first 5 years of life.
This proposal responds to the HIRN RFA for using advanced analytical techniques to produce a high- resolution atlas, at the cellular and organ-wide level, of the differentiation process and molecular signatures of islets and endocrine Beta cells, focusing on the developmentally plastic juvenile period (0-5 years of life). Recent evidence suggests that this period involves the finalization or maturation of numerous essential functional connections with the neural and vascular networks, and substantial alteration of the intra-islet number and organization of endocrine cells. We target the juvenile period because of recent findings that autoimmunity developed within this period is strongly correlated with a greatly enhanced predisposition towards overt Type 1 diabetes (T1D).
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