In anticipation of cellular transplantation for diabetes, we sought to enrich and expand pancreatic beta-cell lines derived from iPSCs. iPCSs offer the advantage of "self" origin thereby avoiding alloimmunity. We differentiated iPSCs into insulin-producing beta cells using growth factors and labeled these cells with a red fluorescent protein driven by insulin promoter for isolation and expansion. Beta-cell marker expression was evaluated by immunocytochemistry and qRT-PCR. Labeled cells were sorted by FACS and transplanted under the kidney capsule of SCID mice that do not reject human cells. iPSC-engrafted kidneys were analyzed by immunohistochemistry (IHC). Higher expression of beta cell markers (insulin, PDX-1, Nkx6.1, Kir6.2, Glut2, GKS, SUR1, PC1/3, PC2, and amylin) was found in sorted cells compared to non-sorted cells (P <0.05). Sorted cells were expanded up to ten passages. Human C-peptide was detected in the culture media of sorted cells at 0 glucose (1.14 ± 1.3 pmol/L) and 17 mM glucose concentrations (4.6 ± 2.2, P<0.05). Quinacrine secretion (beta cell degranulation) was observed after glucose stimulation. At 60 days after transplantation, C-peptide blood levels in SCID mice rose upon glucose challenge.

Harvested cells were positive for insulin after IHC analysis. These results indicate that beta cells derived from iPSCs, enriched and expanded in vitro, maintained their functional phenotype in vivo.

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