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Induction of pluripotent stem cells from primary human fibroblasts with only Oct4 and Sox2

Nature Biotechnology volume 26, pages 1269–1275 (2008)Cite this article

Abstract

Ectopic expression of defined sets of genetic factors can reprogram somatic cells to induced pluripotent stem (iPS) cells that closely resemble embryonic stem (ES) cells. The low efficiency with which iPS cells are derived hinders studies on the molecular mechanism of reprogramming, and integration of viral transgenes, in particular the oncogenes c-Myc and Klf4, may handicap this method for human therapeutic applications. Here we report that valproic acid (VPA), a histone deacetylase inhibitor, enables reprogramming of primary human fibroblasts with only two factors, Oct4 and Sox2, without the need for the oncogenes c-Myc or Klf4. The two factor–induced human iPS cells resemble human ES cells in pluripotency, global gene expression profiles and epigenetic states. These results support the possibility of reprogramming through purely chemical means, which would make therapeutic use of reprogrammed cells safer and more practical.

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Figure 1: Efficient generation of iPS cells with three transcription factors (Oct4, Sox2, and Klf4).
Figure 2: Generation of iPS cells with two factors (Oct4 and Sox2) and VPA.
Figure 3: In vitro differentiation of two factor–induced iPS cells.
Figure 4: Teratomas from two factor–induced iPS cells.
Figure 5: Methylation analysis of OCT4 and NANOG promoter regions in two factor–induced iPS cells.
Figure 6: The gene expression profile of two factor–induced iPS cells closely resembles that of human ES cells.

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Acknowledgements

D.H. and D.A.M. conceived the experiments and wrote the manuscript. D.H., K.O., R.M., W.G., A.E., S.C. and W.M. performed experiments.

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Authors and Affiliations

  1. Department of Stem Cell and Regenerative Biology, Howard Hughes Medical Institute, Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, 02138, Massachusetts, USA

    Danwei Huangfu, Kenji Osafune, René Maehr, Astrid Eijkelenboom, Shuibing Chen, Whitney Muhlestein & Douglas A Melton

  2. ICORP Organ Regeneration Project, Japan Science and Technology Agency (JST), 3-8-1 Komaba, Meguro-ku, 153-8902, Tokyo, Japan

    Kenji Osafune

  3. The Whitehead Institute for Biomedical Research, 9 Cambridge Center, Cambridge, 02142, Massachusetts, USA

    Wenjun Guo

  4. Biomedical Sciences, Utrecht University, The Netherlands

    Astrid Eijkelenboom

Authors
  1. Danwei Huangfu
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  7. Whitney Muhlestein
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  8. Douglas A Melton
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Contributions

D.A.M. is a Howard Hughes Medical Institute Investigator. D.H. is funded by the Helen Hay Whitney Foundation and Novartis Institutes for BioMedical Research. S.C. is supported by the Juvenile Diabetics Research Foundation. The authors thank Anastasie Kweudjeu for assistance with micoarray analysis, Konrad Hochedlinger for providing probes for Southern blot analysis, Shinya Yamanaka for providing viral vectors through Addgene and Robert Weinberg for support of this study. Some monoclonal antibodies were obtained from the Developmental Studies Hybridoma Bank, which was developed under the auspices of the National Institute of Child Health and Human Development and is maintained by The University of Iowa, Department of Biological Sciences.

Corresponding author

Correspondence to Douglas A Melton.

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Huangfu, D., Osafune, K., Maehr, R. et al. Induction of pluripotent stem cells from primary human fibroblasts with only Oct4 and Sox2. Nat Biotechnol 26, 1269–1275 (2008). https://doi.org/10.1038/nbt.1502

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