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. 1996 Aug;87(8):787–792. doi: 10.1111/j.1349-7006.1996.tb02101.x

Replication of Hepatitis C Virus in Cultured Non‐neoplastic Human Hepatocytes

Nobuyuki Kato 1,✉, Masanori Ikeda 1, Tetsuya Mizutani 1, Kazuo Sugiyama 1, Masayuki Noguchi 2, Setsuo Hirohashi 2, Kunitada Shimotohno 1
PMCID: PMC5921176  PMID: 8797883

Abstract

We established a replication system for hepatitis C virus (HCV) using the PH5CH non‐neoplastic human hepatocyte line that had been immortalized with simian virus 40 large T antigen. In cells inoculated with sera derived from two HCV‐positive blood donors, positive‐stranded HCV RNA was detected up to 30 days postinoculation (p.i.). Semi‐quantitative analysis of HCV RNA revealed that HCV multiplied during the period of culture. Sequence analysis of the HCV hypervariable region 1 (HVR1) in both cases indicated that HVR1 populations from the cells at 8 days p.i. were apparently different from those of the original inocula. HVR1 populations in infected cells became homogeneous or just a few species were selected over time. These results suggest that HCV is replicating in the human hepatocyte PH5CH cells. This culture system will be useful for detailed studies of the biological effects of HCV in human hepatocytes.

Keywords: Hepatitis C virus, Human hepatocyte, HCV replication system, Hypervariable region 1

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REFERENCE

  • 1.Choo , Q.‐L. , Kuo , G. , Weiner , A. J. , Overby , L. R. , Bradley , D. W. and Houghton , M.Isolation of a cDNA clone derived from a blood‐borne non‐A, non‐B viral hepatitis genome . Science , 244 , 359 – 362 ( 1989. ). [DOI] [PubMed] [Google Scholar]
  • 2.Kuo , G. , Choo , Q.‐L. , Alter , H. J. , Gitnick , G. L. , Redeker , A. G. , Purcell , R. H. , Miyamura , T. , Dienstag , J. L. , Alter , M. J. , Stevens , C. E. , Tegtmeier , G. E. , Bonino , F. , Colombo , W.‐S. , Lee , W.‐S. , Kuo , C. , Berger , K. , Shuster , J. R. , Overby , L. R. , Bradley , D. W. and Houghton , M.An assay for circulating antibodies to a major etiologic virus of human non‐A, non‐B hepatitis . Science , 244 , 362 – 364 ( 1989. ). [DOI] [PubMed] [Google Scholar]
  • 3.Ohkoshi , S. , Kojima , H. , Tawaraya , H. , Miyajima , T. , Kamimura , T. , Asakura , H. , Satoh , A. , Hirose , S. , Hijikata , M. , Kato , N. and Shimotohno , K.Prevalence of antibody against non‐A, non‐B hepatitis virus in Japanese patients with hepatocellular carcinoma . Jpn. J. Cancer Res ., 81 , 550 – 553 ( 1990. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4.Saito , I. , Miyamura , T. , Ohbayashi , A. , Harada , H. , Katayama , T. , Kikuchi , S. , Watanabe , Y. , Koi , S. , Onji , M. , Ohta , Y. , Choo , Q.‐L. , Houghton , M. and Kuo , G.Hepatitis C virus infection is associated with the development of hepatocellular carcinoma . Proc. Natl. Acad. Sci. USA , 87 , 6547 – 6549 ( 1990. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Kato , N. , Hijikata , M. , Ootsuyama , Y. , Nakagawa , M. , Ohkoshi , S. , Sugimura , T. and Shimotohno , K.Molecular cloning of the human hepatitis C virus genome from Japanese patients with non‐A, non‐B hepatitis . Proc. Natl. Acad. Sci. USA , 87 , 9524 – 9528 ( 1990. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6.Houghton , M. , Weiner , A. , Han , J. , Kuo , G. and Choo , Q.‐L.Molecular biology of the hepatitis C viruses: implications for diagnosis, development and control of viral disease . Hepatology , 14 , 381 – 388 ( 1991. ). [PubMed] [Google Scholar]
  • 7.Bukh , J. , Miller , R. H. and Purcell , R. H.Genetic heterogeneity of hepatitis C virus: quasispecies and genotypes . Sem. Liver Dis ., 15 , 41 – 63 ( 1995. ). [DOI] [PubMed] [Google Scholar]
  • 8.Hijikata , M. , Kato , N. , Ootsuyama , Y. , Nakagawa , M. and Shimotohno , K.Gene mapping of the putative structural region of the hepatitis C virus genome by in vitro processing analysis . Proc. Natl. Acad. Sci. USA , 88 , 5547 – 5551 ( 1991. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9.Hijikata , M. , Mizushima , H. , Tanji , Y. , Komoda , Y. , Hirowatari , Y. , Akagi , T. , Kato , N. , Kimura , K. and Shimotohno , K.Proteolytic processing and membrane association of putative nonstructural proteins of hepatitis C virus . Proc. Natl. Acad. Sci. USA , 90 , 10773 – 10777 ( 1993. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Muraiso , K. , Hijikata , M. , Ohkoshi , S. , Cho , M.‐J. , Kikuchi , M. , Kato , N. and Shimotohno , K.A structural protein of hepatitis C virus expressed in E. coli facilitates accurate detection of hepatitis C virus . Biochem. Biophys. Res. Commun ., 172 , 511 – 516 ( 1990. ). [DOI] [PubMed] [Google Scholar]
  • 11.Kato , N. , Sekiya , H. , Ootsuyama , Y. , Nakazawa , T. , Hijikata , M. , Ohkoshi , S. and Shimotohno , K.Humoral immune response to hypervariable region 1 of the putative envelope glycoprotein (gp70) of hepatitis C virus . J. Virol ., 67 , 3923 – 3930 ( 1993. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Kato , N. , Ootsuyama , Y. , Sekiya , H. , Ohkoshi , S. , Nakazawa , T. , Hijikata , M. and Shimotohno , K.Genetic drift in hypervariable region 1 of the viral genome in persistent hepatitis C virus infection . J. Virol ., 68 , 4776 – 4784 ( 1994. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Shimizu , Y. K. , Purcell , R. H. and Yoshikura , H.Correlation between the infectivity of hepatitis C virus in vivo and its infectivity in vitro. Proc. Natl. Acad. Sci. USA , 90 , 6037 – 6041 ( 1993. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Zibert , A. , Schreier , E. and Roggendorf , M.Antibodies in human sera specific to hypervariable region 1 of hepatitis C virus can block viral attachment . Virology , 208 , 653 – 661 ( 1995. ). [DOI] [PubMed] [Google Scholar]
  • 15.Cribier , B. , Schmitt , C. , Bingen , A. , Kirn , A. and Keller , F.In vitro infection of peripheral blood mononuclear cells by hepatitis C virus . J. Gen. Virol ., 76 , 2485 – 2491 ( 1995. ). [DOI] [PubMed] [Google Scholar]
  • 16.Lanford , R. E. , Sureau , C. , Jacob , J. R. , White , R. and Fuerst , T. R.Demonstration of in vitro infection of chimpanzee hepatocytes with hepatitis C virus using strand‐specific RT/PCR . Virology , 202 , 606 – 614 ( 1994. ). [DOI] [PubMed] [Google Scholar]
  • 17.Kato , N. , Nakazawa , T. , Mizutani , T. and Shimotohno , K.Susceptibility of human T‐lymphotropic virus type I infected cell line MT‐2 to hepatitis C virus infection . Biochem. Biophys. Res. Commun ., 206 , 863 – 869 , 1995. . [DOI] [PubMed] [Google Scholar]
  • 18.Mizutani , T. , Kato , N. , Hirota , M. , Sugiyama , K. , Murakami , A. and Shimotohno , K.Inhibition of hepatitis C virus replication by antisense oligonucleotide in culture cells . Biochem. Biophys. Res. Commun ., 212 , 906 – 911 ( 1995. ). [DOI] [PubMed] [Google Scholar]
  • 19.Noguchi , M. and Hirohashi , S.Cell lines from non‐neo‐plastic liver and hepatocellular carcinoma tissue from a single patient . In Vitro Cell. Dev. Biol ., 32 , 135 – 137 ( 1996. ). [DOI] [PubMed] [Google Scholar]
  • 20.Nakazawa , T. , Kato , N. , Ootsuyama , Y. , Sekiya , H. , Fujioka , T. , Shibuya , A. and Shimotohno , K.Genetic alteration of the hepatitis C virus hypervariable region obtained from an asymptomatic carrier . Int. J. Cancer , 56 , 204 – 207 ( 1994. ). [DOI] [PubMed] [Google Scholar]
  • 21.Gunji , T. , Kato , N. , Hijikata , M. , Hayashi , K. , Saitoh , S. and Shimotohno , K.Specific detection of positive and negative stranded hepatitis C viral RNA using chemical RNA modification . Arch. Virol ., 134 , 293 – 302 ( 1994. ). [DOI] [PubMed] [Google Scholar]
  • 22.Kato , N. , Hijikata , M. , Ootsuyama , Y. , Nakagawa , M. , Ohkoshi , S. and Shimotohno , K.Sequence diversity of hepatitis C viral genomes . Mol. Biol. Med ., 7 , 495 – 501 ( 1990. ). [PubMed] [Google Scholar]
  • 23.Nei , M. , Gojobori , T.Simple methods for estimating the numbers of synonymous and nonsynonymous nucleotide substitutions . Mol. Biol. Evol ., 3 , 418 – 426 ( 1986. ). [DOI] [PubMed] [Google Scholar]
  • 24.Hijikata , M. , Mizuno , K. , Rikihisa , T. , Shimizu , Y. K. , Iwamoto , A. , Nakajima , N. , Yoshikura , H.Selective transmission of hepatitis C virus in vivo and in vitro. Arch. Virol ., 140 , 1623 – 1628 ( 1995. ). [DOI] [PubMed] [Google Scholar]

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