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Long-term expansion of functional pancreatic islets on a laminin-521 matrix and curative transplantation into diabetic mice

ABSTRACT: In this study we show, for the first time, that whole islets can be cultured and significantly expanded long-term in vitro on specific basement membrane (BM) laminin LN-521 which is a normal component of islet BMs. Isolated islets adhere and flatten to form 2-3 cell layer entities with ~10 % of all endocrine cell types proliferating after 2-5 weeks in culture. Time-lapse imaging of freshly isolated islets in suspension labeled with hypoxia and DNA degradation indicators reveals that spherical islets on LN-111 rapidly develop hypoxia and central necrosis, in contrast to islets attached to LN-521, which normal b-cell glucose responsiveness. RNA sequencing analyses reveal that freshly isolated islets express LN-521 binding integrins. Compared to other matrices, islets grown on LN-521 develop a unique expression signature, including expression of specific integrins, involved in cell adhesion and matrix production as well as proliferation. The results reveal specific LN-521-mediated effects into islet cells and suggest that use of in vitro expanded islets may significantly enhance the efficacy of islet transplantation treatment of T1D.

ORGANISM(S): Mus musculus

PROVIDER: GSE89677 | GEO | 2018/11/09

REPOSITORIES: GEO

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Project description:Pancreatic islet transplantation as a cure for type 1 diabetes (T1D) cannot be scaled up due to a scarcity of human pancreas donors. In vitro expansion of beta cells from mature human pancreatic islets provides an alternative source of insulin-producing cells. The exact nature of the expanded cells produced by diverse expansion protocols, and their potential for differentiation into functional beta cells, remain elusive. We performed a large-scale meta-analysis of gene expression in human pancreatic islet cells, which were processed using three different previously described protocols for expansion and attempted re-differentiation. All three expansion protocols induced dramatic changes in the expression profiles of pancreatic islets; many of these changes are shared among the three protocols. Attempts at re-differentiation of expanded cells induce a limited number of gene expression changes. Nevertheless, these fail to restore a pancreatic islet-like gene expression pattern. Comparison with a collection of public microarray datasets confirmed that expanded cells are highly comparable to mesenchymal stem cells. Genes induced in expanded cells are also enriched for targets of transcription factors important for pluripotency induction. The present data increases our understanding of the active pathways in expanded and re-differentiated islets. Knowledge of the mesenchymal stem cell potential may help development of drug therapeutics to restore beta cell mass in T1D patients. Experiment Overall Design: In this study, we have tested three different protocols to expand human pancreatic islets in monolayer and after attempted maneuvers to re-differentiate the expanded cells back to islets. We have characterized the resulting cells in detail by performing microarray analyses with fresh pancreatic islets, expanded islet cells and re-differentiated cells. Genes modified by either of three protocols have 70 to 80% overlap with the genes changed by the other two protocols. Although there are promising changes in the right direction, none of the three protocols could achieve a return to a functional islet state. The expanded cells highly resemble Mesenchymal Stem Cells (MSC), and similar gene regulatory networks seem to be active in both cell types. On the other hand, the expanded islet cells are different from MSC in that they seem to retain activity of some islet gene modules. The current results highlight the importance of designing new strategies that take into account the MSC potential of expanded cells.

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Long-term expansion of functional pancreatic islets on a laminin-521 matrix and curative transplantation into diabetic mice

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