Hepatitis virus infection affects DNA methylation in mice with humanized livers

Gastroenterology. 2014 Feb;146(2):562-72. doi: 10.1053/j.gastro.2013.10.056. Epub 2013 Oct 30.


Background & aims: Cells of tumors associated with chronic inflammation frequently have altered patterns of DNA methylation, including hepatocellular carcinomas. Chronic hepatitis has also been associated with aberrant DNA methylation, but little is known about their relationship.

Methods: Pyrosequencing was used to determine the methylation status of cultured Huh7.5.1 hepatoma cells after hepatitis C virus (HCV) infection. We also studied mice with severe combined immunodeficiency carrying the urokinase-type plasminogen activator transgene controlled by an albumin promoter (urokinase-type plasminogen activator/severe combined immunodeficient mice), in which up to 85% of hepatocytes were replaced by human hepatocytes (chimeric mice). Mice were given intravenous injections of hepatitis B virus (HBV) or HCV, liver tissues were collected, and DNA methylation profiles were determined at different time points after infection. We also compared methylation patterns between paired samples of hepatocellular carcinomas and adjacent nontumor liver tissues from patients.

Results: No reproducible changes in DNA methylation were observed after infection of Huh7.5.1 cells with HCV. Livers from HBV- and HCV-infected mice had genome-wide, time-dependent changes in DNA methylation, compared with uninfected urokinase-type plasminogen activator/severe combined immunodeficient mice. There were changes in 160 ± 63 genes in HBV-infected and 237 ± 110 genes in HCV-infected mice. Methylation of 149 common genes increased in HBV- and HCV-infected mice; methylation of some of these genes also increased in hepatocellular carcinoma samples from patients compared with nontumor tissues. Expression of Ifng, which is expressed by natural killer cells, increased significantly in chimeric livers, in concordance with induction of DNA methylation, after infection with HBV or HCV. Induction of Ifng was reduced after administration of an inhibitor of natural killer cell function (anti-asialo GM1).

Conclusions: In chimeric mice with humanized livers, infection with HBV and HCV appears to activate a natural kill cell-dependent innate immune response. This contributes to the induction and accumulation of aberrant DNA methylation in human hepatocytes.

Keywords: ESR1; Epigenetic; Gene Regulation; HBV; HBx; HCC; HCV; HOXA6; IFN; Inflammatory Response; Liver Cancer; MCAM; NK; PCNA; RASSF1A; ROS; Ras association domain family 1 isoform A; SCID; estrogen receptor 1; hepatitis B virus; hepatitis B virus X; hepatitis C virus; hepatocellular carcinoma; homeobox A6; interferon; methylated CpG island amplification microarray; natural killer; proliferating cell nuclear antigen; reactive oxygen species; severe combined immunodeficient.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adolescent
  • Adult
  • Animals
  • Carcinoma, Hepatocellular / genetics
  • Carcinoma, Hepatocellular / immunology
  • Carcinoma, Hepatocellular / virology
  • Cell Line, Tumor
  • Child
  • Child, Preschool
  • Chimerism
  • CpG Islands
  • DNA Methylation*
  • Epigenesis, Genetic
  • Female
  • Hepatitis B / complications
  • Hepatitis B / genetics*
  • Hepatitis B / immunology
  • Hepatitis C / complications
  • Hepatitis C / genetics*
  • Hepatitis C / immunology
  • Humans
  • Immunity, Innate
  • Killer Cells, Natural / immunology
  • Liver / immunology
  • Liver / virology*
  • Liver Neoplasms / genetics
  • Liver Neoplasms / immunology
  • Liver Neoplasms / virology
  • Lymphocyte Activation
  • Male
  • Mice
  • Mice, SCID
  • Oligonucleotide Array Sequence Analysis
  • Sequence Analysis, DNA
  • Young Adult