Abstract
The ectopic expression of several transcription factors can restore embryonic cell fate to cultured somatic cells and generate induced pluripotent stem cells (iPSCs), revealing a previously unknown pathway to pluripotency. However, this technology is currently limited by low efficiency, slow kinetics and multi-factorial requirement. Here we show that reprogramming can be improved and dramatically accelerated by optimizing culture conditions. First, we developed an optimized defined medium, iCD1, which allows Oct4/Sox2/Klf4 (OSK)-mediated reprogramming to achieve ultra-high efficiency (∼10% at day 8). We also found that this optimized condition renders both Sox2 and Klf4 dispensable, although the elimination of these two factors leads to lower efficiency and slower kinetics. Our studies define a shortened route, both in timing and factor requirement, toward pluripotency. This new paradigm not only provides a rationale to further improve iPSC generation but also simplifies the conceptual understanding of reprogramming by defined factors.
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Acknowledgements
We acknowledge Dr Guoliang Xu (Institute of Biochemistry and Cell Biology, Shanghai Institute for Biological Sciences, Chinese Academy of Sciences) for discussion. We are grateful to Jian Zhang and Xia Kuang for assistance with Southern blotting. We are also grateful to Ronghui Li, Jieying Zhu, Jing Li, Keyu Lai, Hengpeng Guo, Shaoxian Sun, Haixiang Zhang, Yi Zheng and Hongwen Pang for their technical assistance. We also thank all members of our laboratory for supporting our work. This work is supported by the National Natural Science Foundation of China (90813033), Key Technologies R&D Program: the 973 Program of China (2009CB941102 and 2011CB965200), Ministry of Science and Technology International Technology Cooperation Program (2010DFB30430), Chinese Academy of Sciences/SAFEA International Partnership Program for Creative Research Teams, Major Scientific and Technological Special Project: New Drug Innovation and Development (2011X09102-010-01), and “Strategic Priority Research Program” of the Chinese Academy of Sciences (XDA01020401).
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( Supplementary information is linked to the online version of the paper on the Cell Research website.)
Supplementary information
Supplementary information, Figure S1
The effect of reported protocols for generation of iPSCs. (PDF 58 kb)
Supplementary information, Figure S2
Efficient reprogramming in iCD1. (PDF 169 kb)
Supplementary information, Figure S3
iCD1 is optimized for OKS mediated somatic reprogramming. (PDF 91 kb)
Supplementary information, Figure S4
iCD1 is optimized for OKS mediated somatic reprogramming. (PDF 84 kb)
Supplementary information, Figure S5
Application of iCD1 on inducible system based on lentivirus. (PDF 227 kb)
Supplementary information, Figure S6
iCD1 supports efficient reprogramming in other cell types. (PDF 118 kb)
Supplementary information, Figure S7
Effect of iCD1 on reprogramming factor expression. (PDF 73 kb)
Supplementary information, Figure S8
iPSCs induced by Oct4/Klf4 (OK) or Oct4/Sox2 (OS) are pluripotent. (PDF 508 kb)
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Chen, J., Liu, J., Chen, Y. et al. Rational optimization of reprogramming culture conditions for the generation of induced pluripotent stem cells with ultra-high efficiency and fast kinetics. Cell Res 21, 884–894 (2011). https://doi.org/10.1038/cr.2011.51
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DOI: https://doi.org/10.1038/cr.2011.51
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