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Homogeneous Differentiation of Functional Hepatocytes from Human Induced Pluripotent Stem Cells

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Stem Cell Assays

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2429))

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Abstract

Hepatocyte-like cells (HLCs) generated from human induced pluripotent stem cells (iPSCs) could provide an unlimited source of liver cells for regenerative medicine, disease modeling, drug screening, and toxicology studies. Here we describe a stepwise improved protocol that enables highly efficient, homogeneous, and reproducible differentiation of human iPSCs into functional hepatocytes through controlling all three stages of hepatocyte differentiation, starting from a single cell (non-colony) culture of iPSCs, through homogeneous definitive endoderm induction and highly efficient hepatic specification, and finally arriving at matured HLCs. The final population of cells exhibits morphology closely resembling that of primary human hepatocytes, and expresses specific hepatic markers as evidenced by immunocytochemical staining. More importantly, these HLCs demonstrate key functional characteristics of mature hepatocytes, including major serum protein (e.g., albumin, fibronectin, and alpha-1 antitrypsin) secretion, urea synthesis, glycogen storage, and inducible cytochrome P450 activity.

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Acknowledgments

The authors would like to thank Dr. Marianne Miliotis-Solomotis and Dr. Mary E. Torrence for critical review of the manuscripts as well as for their support of this work. The authors would also like to thank Dr. Seung Bum Park at the National Institute of Diabetes and Digestive and Kidney Diseases for helpful discussions during the development of this protocol. This work was supported by internal funds of the U.S. Food and Drug Administration. The statements made in this article are those of the authors and do not necessarily represent views, opinions, or policies of the U.S. Food and Drug Administration.

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Authors and Affiliations

  1. Division of Toxicology, Office of Applied Research and Safety Assessment, Center for Food Safety and Applied Nutrition, U.S. Food and Drug Administration, Laurel, MD, USA

    Rong Li, Yang Zhao, Jeffrey J. Yourick, Robert L. Sprando & Xiugong Gao

Authors
  1. Rong Li
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  2. Yang Zhao
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  3. Jeffrey J. Yourick
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  4. Robert L. Sprando
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  5. Xiugong Gao
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Corresponding author

Correspondence to Xiugong Gao .

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Editors and Affiliations

  1. Stem Cell and Cancer Biology Laboratory, Division of Experimental Medicine and Pathology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, USA

    Nagarajan Kannan

  2. Hull York Medical School, York, UK

    Philip Beer

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Li, R., Zhao, Y., Yourick, J.J., Sprando, R.L., Gao, X. (2022). Homogeneous Differentiation of Functional Hepatocytes from Human Induced Pluripotent Stem Cells. In: Kannan, N., Beer, P. (eds) Stem Cell Assays. Methods in Molecular Biology, vol 2429. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1979-7_9

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  • DOI: https://doi.org/10.1007/978-1-0716-1979-7_9

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1978-0

  • Online ISBN: 978-1-0716-1979-7

  • eBook Packages: Springer Protocols

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