Human adipose tissue-derived stem cells (hASCs) are considered an ideal tool for the supply of insulin-producing cells to treat diabetes mellitus, with high differentiation efficiency. Islet neogenesis-associated protein (INGAP) is an initiator of islet neogenesis, and the peptide sequence comprising amino acids 104-118, named INGAP pentadecapeptide (INGAP-PP), has been shown to increase β-cell mass in animals and human pathological states. Here, we report a novel 4-step method to promote hASCs to differentiate into islet-like clusters (ILCs) more efficiently by adding INGAP-PP. The hASCs were isolated, purified and differentiated using a 4-step protocol including trichostatin A, INGAP-PP/scrambled peptide (Scrambled-P), dexamethasone, nicotinamide, glucagon-like peptide-1, transforming growth factor β1 and exendin-4. Results showed that ILCs in the INGAP-PP group were more similar to the fresh islets with regard to both size and morphology and expressed significantly higher levels of both insulin and C-peptide than those in the Scrambled-P group. Moreover, the ILCs from the INGAP-PP group secreted higher levels of insulin and C-peptide than those from the Scrambled-P group in response to both a low (5.6 mM) and high (25 mM) glucose challenge and secreted 6 times more hormones under the high-glucose challenge. Real-time PCR and immunocytochemistry showed that ILCs of the INGAP-PP group expressed human pancreatic endocrine hormones and transcription factors. Transplantation of ILCs into diabetic rats partially reversed diabetes and prolonged their life span. In conclusion, the INGAP-PP protocol can efficiently induce hASCs to differentiate into ILCs in vitro, and thus hASCs could be a promising source of cells for transplantation to treat diabetes mellitus.

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