Differentiation Efficiency of Induced Pluripotent Stem Cells Depends on the Number of Reprogramming Factors§

Authors

  • Matthias Löhle,

    1. Division of Neurodegenerative Diseases, Department of Neurology,Dresden University of Technology, Dresden, Germany
    2. Center for Regenerative Therapies Dresden (CRTD)Dresden University of Technology, Dresden, Germany
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  • Andreas Hermann,

    1. Division of Neurodegenerative Diseases, Department of Neurology,Dresden University of Technology, Dresden, Germany
    2. Center for Regenerative Therapies Dresden (CRTD)Dresden University of Technology, Dresden, Germany
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  • Hannes Glaß,

    1. Division of Neurodegenerative Diseases, Department of Neurology,Dresden University of Technology, Dresden, Germany
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  • Andrea Kempe,

    1. Division of Neurodegenerative Diseases, Department of Neurology,Dresden University of Technology, Dresden, Germany
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  • Sigrid C. Schwarz,

    1. Department of Neurology, University of Leipzig, Leipzig, Germany
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  • Jeong Beom Kim,

    1. Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany
    2. School of Nano-Biotechnology and Chemical Engineering, Ulsan National Institute of Science and Technology (UNIST), Ulsan, Republic of Korea
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  • Claire Poulet,

    1. Institute of Pharmacology and Toxicology, Dresden University of Technology, Dresden, Germany
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  • Ursula Ravens,

    1. Institute of Pharmacology and Toxicology, Dresden University of Technology, Dresden, Germany
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  • Johannes Schwarz,

    1. Department of Neurology, University of Leipzig, Leipzig, Germany
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  • Hans R. Schöler,

    1. Department of Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany
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  • Alexander Storch

    Corresponding author
    1. Division of Neurodegenerative Diseases, Department of Neurology,Dresden University of Technology, Dresden, Germany
    2. Center for Regenerative Therapies Dresden (CRTD)Dresden University of Technology, Dresden, Germany
    3. German Center for Neurodegenerative Diseases (DZNE), Research Site Dresden, Dresden, Germany
    • Division of Neurodegenerative Diseases, Department of Neurology, Dresden University of Technology, Fetscherstrasse 74, 01307 Dresden, Germany
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    • Author contributions: M.L.: conception and design, collection and assembly of data, data analysis and interpretation, drafting and critical revision of manuscript, and fund raising; A.H.: conception and design, collection and assembly of data, data analysis and interpretation, critical revision of manuscript, and fund raising; H.G., A.K., S.C.S., C.P., U.R., and J.S.: collection and assembly of data and critical revision of manuscript; J.B.K.: provision of study material and critical revision of manuscript; H.R.S.: provision of study material, critical revision of manuscript, and fund raising; A.S.: conception and design, data analysis and interpretation, drafting and critical revision of manuscript, and fund raising.

    • Telephone: 49-351-458-2532; Fax: 49-351-458-4352


  • Disclosure of potential conflicts of interest is found at the end of this article.

  • §

    First published online in STEM CELLSEXPRESS December 1, 2011.

Abstract

Reprogramming of somatic cells into induced pluripotent stem cells (iPSCs) by retroviral overexpression of the transcription factors Oct4, Sox2, Klf4, and c-Myc holds great promise for the development of personalized cell replacement therapies. In an attempt to minimize the risk for chromosomal disruption and to simplify reprogramming, several studies demonstrated that a reduced set of reprogramming factors is sufficient to generate iPSC, albeit at lower efficiency. To elucidate the influence of factor reduction on subsequent differentiation, we compared the efficiency of neuronal differentiation in iPSC generated from postnatal murine neural stem cells with either one (Oct4; iPSC1F-NSC), two (Oct4, Klf4; iPSC2F-NSC), or all four factors (iPSC4F-NSC) with those of embryonic stem cells (ESCs) and iPSC produced from fibroblasts with all four factors (iPSC4F-MEF). After 2 weeks of coculture with PA6 stromal cells, neuronal differentiation of iPSC1F-NSC and iPSC2F-NSC was less efficient compared with iPSC4F-NSC and ESC, yielding lower proportions of colonies that stained positive for early and late neuronal markers. Electrophysiological analyses after 4 weeks of differentiation identified functional maturity in neurons differentiated from ESC, iPSC2F-NSC, iPSC4F-NSC, and iPSC4F-MEF but not in those from iPSC1F-NSC. Similar results were obtained after hematoendothelial differentiation on OP9 bone marrow stromal cells, where factor-reduced iPSC generated lower proportions of colonies with hematoendothelial progenitors than colonies of ESC, iPSC4F-NSC, and iPSC4F-MEF. We conclude that a reduction of reprogramming factors does not only reduce reprogramming efficiency but may also worsen subsequent differentiation and hinder future application of iPSC in cell replacement therapies. STEM CELLS 2012;30:570–579

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