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Journal of Molecular Cell Biology Advance Access published online on November 12, 2009
Journal of Molecular Cell Biology, doi:10.1093/jmcb/mjp037
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© The Author (2009). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS. All rights reserved.

Generation of Homogeneous PDX1+ Pancreatic Progenitors from Human ES Cell-derived Endoderm Cells

Jun Cai1,{dagger}, Chen Yu1,{dagger}, Yanxia Liu1, Song Chen1, Yuxuan Guo1, Jun Yong1, Wei Lu1, Mingxiao Ding1 and Hongkui Deng1,2,*

1 The MOE Key Laboratory of Cell Proliferation and Differentiation, College of Life Sciences, Peking University, Beijing, China
2 Laboratory of Chemical Genomics, Shenzhen Graduate School of Peking University, Shenzhen, China

* Correspondence to: Hongkui Deng, Fax: +86 10 6275 6954; E-mail: hongkui_deng{at}pku.edu.cn


  Abstract

One key step in producing insulin-secreting cells from human embryonic stem (hES) cells is the generation of pancreatic and duodenal homeobox gene 1 (PDX1)-expressing pancreatic progenitor cells. All-trans retinoic acid (RA) has important roles in pancreas development and is widely used to induce pancreatic differentiation of ES cells. When RA was added directly to the activin A-induced hES cells, <20% cells were positive for the pancreatic marker PDX1, whereas the other cells were mainly hepatic cells. We found that when the activin A-induced hES cells were replated and seeded at low cell densities, the addition of RA induced significant pancreatic differentiation and over 70% of cells in culture expressed PDX1. When the endodermal cells were isolated with the surface marker CXCR4 from the activin A-induced culture and further differentiated with RA, a homogeneous PDX1+ cell population (over 95% pure) was generated. The PDX1+ cells could further differentiate into cells that expressed pancreatic transcription factors and pancreatic endocrine or exocrine markers. We also found that RA inhibited the hepatic differentiation of endodermal cells that were seeded at low cell densities, and this inhibition may have been through the inhibition of Smad1/5/8 activity. Thus, we present a highly efficient and reproducible protocol for generating PDX1+ pancreatic progenitor cells from hES cells.

Keywords: embryonic stem cell, retinoic acid, PDX1, pancreatic progenitor, hepatic, Smad

Received September 21, 2009; Revised September 29, 2009; Accepted October 10, 2009

{dagger} These authors contributed equally to this work.


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