Human serum leads to differentiation of human hepatoma cells, restoration of very-low-density lipoprotein secretion, and a 1000-fold increase in HCV Japanese fulminant hepatitis type 1 titers

Authors

  • Rineke H.G. Steenbergen,

    1. Department ofMedical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
    2. Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada
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  • Michael A. Joyce,

    1. Department ofMedical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
    2. Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada
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  • Bradley S. Thomas,

    1. Department ofMedical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
    2. Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada
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  • Daniel Jones,

    1. Division of BioMedical Sciences, Memorial University of Newfoundland, St. John's, Newfoundland, Canada
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  • John Law,

    1. Department ofMedical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
    2. Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada
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  • Rodney Russell,

    1. Division of BioMedical Sciences, Memorial University of Newfoundland, St. John's, Newfoundland, Canada
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  • Michael Houghton,

    1. Department ofMedical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
    2. Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada
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  • D. Lorne Tyrrell

    Corresponding author
    1. Department ofMedical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
    2. Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada
    • Address reprint requests to: Lorne Tyrrell, M.D., Ph.D., Department of Medical Microbiology and Immunology, Li Ka Shing Institute of Virology, University of Alberta, 6-010 Katz Center for Pharmacy and Health Research, Edmonton, Alberta, T6G 2S2, Canada. E-mail: lorne.tyrrell@ualberta.ca; fax: +-1-780-492-5304.

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  • Potential conflict of interest: Nothing to declare.

  • The authors thank Dr. Rice for the kind gift of Huh7.5 cells and Dr. Wakita for the JFH-1 construct. This research was funded by the Li Ka Shing Foundation, Canadian Institutes for Health Research, and Alberta Advanced Education and Technology (AAET).

Abstract

In this study, we differentiated the human hepatoma cell line Huh7.5 by supplementing tissue culture media with human serum (HS) and examined the production of hepatitis C virus (HCV) by these cells. We compared the standard tissue culture protocol, using media supplemented with 10% fetal bovine serum (FBS), to media supplemented with 2% HS. Cells cultured in HS undergo rapid growth arrest, have a hepatocyte-like morphology, and increase the expression of hepatocyte differentiation markers. In addition, expression of cell adhesion proteins claudin-1, occludin, and e-cadherin are also increased. The lipid droplet content of these cells is highly increased, as are key lipid metabolism regulators liver X receptor alpha, peroxisome proliferator-activated receptor (PPAR)-α, and PPAR-γ. Very-low-density lipoprotein secretion, which is absent in FBS-grown cells, is restored in Huh7.5 cells that are cultured in HS. All these factors have been implicated in the life cycle of HCV. We show that viral production of Japanese fulminant hepatitis type 1 increases 1,000-fold when cells are grown in HS, compared to standard FBS culture conditions. The virus produced under these conditions is associated with apolipoprotein B, has a lower density, higher specific infectivity, and has a longer half-life than virus produced in media supplemented with FBS. Conclusion: We describe a convenient, cost-effective method to produce hepatocyte-like cells, which produce large amounts of virus that more closely resemble HCV present in serum of infected patients. (Hepatology 2013; 58:1907–1917)

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