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Mol. Hum. Reprod. Advance Access published online on October 15, 2008
Molecular Human Reproduction, doi:10.1093/molehr/gan058
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© The Author 2008. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org

Chemically defined sequential culture media for TH+ cell derivation from human embryonic stem cells

Tianran Song1,2, Guian Chen1,2,3, Yue Wang4, Genhong Mao1,2, Yanxia Wang1,2 and Haiyan Bai1,2

1Reproductive Medicine Center, Peking University, Beijing 100083, China 2Stem Cell Research Center, Peking University Third Hospital, Beijing 100083, China 4Neuroscience Research Institute, Peking University, Beijing 100083, China

3 To whom correspondence should be addressed at: Reproductive Medicine Center, Peking University Third Hospital, Beijing 100083, China. E-mail: chenguian2008{at}bjmu.edu.cn

During the past few years several differentiation protocols to derive midbrain dopamine (DA) neurons from human embryonic stem (hES) cells have been developed, but the production of sufficient amounts of the "right" therapeutic DA cells has not yet been accomplished. The aim of this study was to efficiently generate tyrosine hydroxylase (TH)-positive cells in vitro from our hES cells using a chemically defined culture system. At the end of differentiation, the vast majority of cells (> 90%) were positive for both TH and β-tubulin isotype III (TuJ1). Other markers of dopaminergic cells, like dopamine transporter (DAT) and Nurr1 were also detected by immunofluorescence or RT-PCR. The functions of these cells were confirmed by measurements of DA release in vitro and by transplantation of derived cells into Parkinson's disease (PD) rats in vivo. We found these cells were able to release dopamine when depolarized by high K+. Moreover, four weeks after transplantation, the hES-derived cells could survive and reduce the apomorphine-induced rotation behaviour of the rats. In conclusion, the experimental system presented here provided a reliable protocol to produce a large number of hES-derived TH+ cells which may be used in cell therapy for PD in future.

Key Words: Human embryonic stem cell/in vitro differentiation/TH-positive cells/transplantation

Submitted on August 6, 2008; resubmitted on September 11, 2008; accepted on October 1, 2008.


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