Rapid generation of mature hepatocyte-like cells from human induced pluripotent stem cells by an efficient three-step protocol

Authors

  • Yu-Fan Chen,

    1. Department of Medical Research and Education, Taipei Veterans General Hospital, Taipei, Taiwan
    2. Stem Cell Research Center, Immunology, National Yang-Ming University, Taipei, Taiwan
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    • These authors contributed equally to this study.

  • Chien-Yu Tseng,

    1. Stem Cell Research Center, Immunology, National Yang-Ming University, Taipei, Taiwan
    2. Institute of Clinical Medicine, Immunology, National Yang-Ming University, Taipei, Taiwan
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    • These authors contributed equally to this study.

  • Hsei-Wei Wang,

    1. Institute of Microbiology and Immunology, National Yang-Ming University, Taipei, Taiwan
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  • Hung-Chih Kuo,

    1. Genomics Research Center, Organismic Biology, Academia Sinica, Taipei, Taiwan
    2. Institute of Cellular and Organismic Biology, Academia Sinica, Taipei, Taiwan
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  • Vincent W. Yang,

    1. Department of Medicine, Stony Brook University School of Medicine, Stony Brook, NY
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  • Oscar K. Lee

    Corresponding author
    1. Department of Medical Research and Education, Taipei Veterans General Hospital, Taipei, Taiwan
    2. Stem Cell Research Center, Immunology, National Yang-Ming University, Taipei, Taiwan
    3. Institute of Clinical Medicine, Immunology, National Yang-Ming University, Taipei, Taiwan
    • Department of Medical Research and Education, Taipei Veterans General Hospital; Institute of Clinical Medicine, National Yang-Ming University, 201, Sec. 2, Shi-Pai Road, Taipei 11217, Taiwan===

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    • fax: +886-2-2875-7824


  • Potential conflict of interest: Nothing to report.

  • This work was supported in part by the University System of Taiwan–University of California San Diego International Center of Excellence in Advanced Bio-engineering sponsored by the Taiwan National Science Council I-RiCE Program under grant NSC-99-2911-I-009-101. The authors also acknowledge financial support from the Taipei Veterans General Hospital (VGH100E1-010, VGH100C-056, VN100-05, and VGH100D-003-2), the National Science Council, Taiwan (NSC100-2120-M-010-001, NSC100-2314-B-010-030-MY3, NSC100-2321-B-010-019, NSC99-3111-B-010-002, NSC98-2314-B-010-001-MY3, NSC 99-2911-I-010-501, and NSC 99-3114-B-002-005). This work was technically assisted, in part, by the Division of Experimental Surgery of the Department of Surgery, Taipei Veterans General Hospital. This study was also supported by a grant from the Ministry of Education, Aim for the Top University Plan.

Abstract

Liver transplantation is the only definitive treatment for end-stage cirrhosis and fulminant liver failure, but the lack of available donor livers is a major obstacle to liver transplantation. Recently, induced pluripotent stem cells (iPSCs) derived from the reprogramming of somatic fibroblasts, have been shown to resemble embryonic stem (ES) cells in that they have pluripotent properties and the potential to differentiate into all cell lineages in vitro, including hepatocytes. Thus, iPSCs could serve as a favorable cell source for a wide range of applications, including drug toxicity testing, cell transplantation, and patient-specific disease modeling. Here, we describe an efficient and rapid three-step protocol that is able to rapidly generate hepatocyte-like cells from human iPSCs. This occurs because the endodermal induction step allows for more efficient and definitive endoderm cell formation. We show that hepatocyte growth factor (HGF), which synergizes with activin A and Wnt3a, elevates the expression of the endodermal marker Foxa2 (forkhead box a2) by 39.3% compared to when HGF is absent (14.2%) during the endodermal induction step. In addition, iPSC-derived hepatocytes had a similar gene expression profile to mature hepatocytes. Importantly, the hepatocyte-like cells exhibited cytochrome P450 3A4 (CYP3A4) enzyme activity, secreted urea, uptake of low-density lipoprotein (LDL), and possessed the ability to store glycogen. Moreover, the hepatocyte-like cells rescued lethal fulminant hepatic failure in a nonobese diabetic severe combined immunodeficient mouse model. Conclusion: We have established a rapid and efficient differentiation protocol that is able to generate functional hepatocyte-like cells from human iPSCs. This may offer an alternative option for treatment of liver diseases. (Hepatology 2012)

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