ORIGINAL ARTICLE

Cell Research (2007): 333-344
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In vitro derivation of functional insulin-producing cells from human embryonic stem cells

Wei Jiang^1,*, Yan Shi^1,5,*, Dongxin Zhao¹, Song Chen¹, Jun Yong¹, Jing Zhang², Tingting Qing¹, Xiaoning Sun^1,3, Peng Zhang¹, Mingxiao Ding¹, Dongsheng Li⁴ and Hongkui Deng^1,2,3

Correspondence: Hongkui Deng Tel: 86-10-6275-6954; Fax: 86-10-6275-6954 E-mail: hongkui_deng@pku.edu.cn Dongsheng Li Tel: 86-719-8801418; Fax: 86-719-8801418 E-mail: dsli@yymc.edu.cn *These two authors contributed equally to this work 5Present address: The Scripps Research Institute, Chemistry Department, SP3130, 10550 North Torrey Pines Road, La Jolla, CA, USA.

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The capacity for self-renewal and differentiation of human embryonic stem (ES) cells makes them a potential source for generation of pancreatic beta cells for treating type I diabetes mellitus. Here, we report a newly developed and effective method, carried out in a serum-free system, which induced human ES cells to differentiate into insulin-producing cells. Activin A was used in the initial stage to induce definitive endoderm differentiation from human ES cells, as detected by the expression of the definitive endoderm markers Sox17 and Brachyury. Further, all-trans retinoic acid (RA) was used to promote pancreatic differentiation, as indicated by the expression of the early pancreatic transcription factors pdx1 and hlxb9. After maturation in DMEM/F12 serum-free medium with bFGF and nicotinamide, the differentiated cells expressed islet specific markers such as C-peptide, insulin, glucagon and glut2. The percentage of C-peptide-positive cells exceeded 15%. The secretion of insulin and C-peptide by these cells corresponded to the variations in glucose levels. When transplanted into renal capsules of Streptozotocin (STZ)-treated nude mice, these differentiated human ES cells survived and maintained the expression of beta cell marker genes, including C-peptide, pdx1, glucokinase, nkx6.1, IAPP, pax6 and Tcf1. Thirty percent of the transplanted nude mice exhibited apparent restoration of stable euglycemia; and the corrected phenotype was sustained for more than six weeks. Our new method provides a promising in vitro differentiation model for studying the mechanisms of human pancreas development and illustrates the potential of using human ES cells for the treatment of type I diabetes mellitus.

Cell Research (2007) 17: 333–344. doi: 10.1038/cr.2007.28; published online 10 April 2007

Keywords: human embryonic stem cell, direct differentiation, insulin-producing cell, diabetes