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Nuclear reprogramming of somatic tissue enables derivation of induced pluripotent stem (iPS) cells from an autologous, non-embryonic origin. The purpose of this study was to establish efficient protocols for lineage specification of human iPS cells into functional glucose-responsive, insulin-producing progeny. We generated human iPS cells, which were then guided with recombinant growth factors that mimic the essential signaling for pancreatic development. Reprogrammed with four stemness factors, human fibroblasts were here converted into authentic iPS cells. Under feeder-free conditions, fate specification was initiated with activin A and Wnt3a that triggered engagement into definitive endoderm, followed by priming with fibroblast growth factor 10 (FGF10) and KAAD-cyclopamine. Addition of retinoic acid, boosted by the pancreatic endoderm inducer indolactam V (ILV), yielded pancreatic progenitors expressing pancreatic and duodenal homeobox 1 (PDX1), neurogenin 3 (NGN3) and neurogenic differentiation 1 (NEUROD1) markers. Further guidance, under insulin-like growth factor 1 (IGF-1), hepatocyte growth factor (HGF) and N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT), was enhanced by glucagon-like peptide-1 (GLP-1) to generate islet-like cells that expressed pancreas-specific markers including insulin and glucagon. Derived progeny demonstrated sustained expression of PDX1, and functional responsiveness to glucose challenge secreting up to 230 pM of C-peptide. A pancreatogenic cocktail enriched with ILV/GLP-1 offers a proficient means to specify human iPS cells into glucose-responsive hormone-producing progeny, refining the development of a personalized platform for islet-like cell generation.

Indolactam V/GLP-1-mediated differentiation of human iPS cells into glucose-responsive insulin-secreting progeny