Identification of host genes showing differential expression profiles with cell-based long-term replication of hepatitis C virus RNA

Hiroe Sejima, Kyoko Mori, Yasuo Ariumi, Masanori Ikeda, Nobuyuki Kato

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Persistent hepatitis C virus (HCV) infection frequently causes hepatocellular carcinoma. However, the mechanisms of HCV-associated hepatocarcinogenesis and disease progression are unclear. Although the human hepatoma cell line, HuH-7, has been widely used as the only cell culture system for robust HCV replication, we recently developed new human hepatoma Li23 cell line-derived OL, OL8, OL11, and OL14 cells, in which genome-length HCV RNA (O strain of genotype 1b) efficiently replicates. OL, OL8, OL11, and OL14 cells were cultured for more than 2 years. We prepared cured cells from OL8 and OL11 cells by interferon-γ treatment. The cured cells were also cultured for more than 2 years. cDNA microarray and RT-PCR analyses were performed using total RNAs prepared from these cells. We first selected several hundred highly or moderately expressed probes, the expression levels of which were upregulated or downregulated at ratios of more than 2 or less than 0.5 in each set of compared cells (e.g., parent OL8 cells versus OL8 cells cultured for 2 years). From among these probes, we next selected those whose expression levels commonly changed during a 2-year culture of genome-length HCV RNA-replicating cells, but which did not change during a 2-year culture period in cured cells. We further examined the expression levels of the selected candidate genes by RT-PCR analysis using additional specimens from the cells cultured for 3.5 years. Reproducibility of the RT-PCR analysis using specimens from recultured cells was also confirmed. Finally, we identified 5 upregulated genes and 4 downregulated genes, the expression levels of which were irreversibly altered during 3.5-year replication of HCV RNA. These genes may play roles in the optimization of the environment in HCV RNA replication, or may play key roles in the progression of HCV-associated hepatic diseases.

Original languageEnglish
Pages (from-to)74-85
Number of pages12
JournalVirus Research
Volume167
Issue number1
DOIs
Publication statusPublished - Jul 2012

Fingerprint

Hepacivirus
RNA
Genes
Hepatocellular Carcinoma
Cultured Cells
Virus Replication
Polymerase Chain Reaction
Down-Regulation
Genome
Cell Line
Virus Diseases
Oligonucleotide Array Sequence Analysis
Interferons
Disease Progression
Cell Culture Techniques
Genotype
Gene Expression
Liver

Keywords

  • ACSM3
  • ALXR
  • ANGPT1
  • ANXA1
  • AREG
  • BASP1
  • CDKN2C
  • CIDEC
  • CPB2
  • Downregulated host genes
  • E1
  • EGF
  • HCC
  • HCV
  • HCV RNA replication system
  • HSPA6
  • ICAM-1
  • IFN
  • Li23 cells
  • Long-term RNA replication
  • PI3
  • PLA1A
  • RT-PCR
  • SEL1L3
  • SLC1A3
  • SLC39A4
  • TBC1D4
  • THSD4
  • Upregulated host genes
  • WISP3

ASJC Scopus subject areas

  • Virology
  • Infectious Diseases
  • Cancer Research

Cite this

Identification of host genes showing differential expression profiles with cell-based long-term replication of hepatitis C virus RNA. / Sejima, Hiroe; Mori, Kyoko; Ariumi, Yasuo; Ikeda, Masanori; Kato, Nobuyuki.

In: Virus Research, Vol. 167, No. 1, 07.2012, p. 74-85.

Research output: Contribution to journalArticle

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AB - Persistent hepatitis C virus (HCV) infection frequently causes hepatocellular carcinoma. However, the mechanisms of HCV-associated hepatocarcinogenesis and disease progression are unclear. Although the human hepatoma cell line, HuH-7, has been widely used as the only cell culture system for robust HCV replication, we recently developed new human hepatoma Li23 cell line-derived OL, OL8, OL11, and OL14 cells, in which genome-length HCV RNA (O strain of genotype 1b) efficiently replicates. OL, OL8, OL11, and OL14 cells were cultured for more than 2 years. We prepared cured cells from OL8 and OL11 cells by interferon-γ treatment. The cured cells were also cultured for more than 2 years. cDNA microarray and RT-PCR analyses were performed using total RNAs prepared from these cells. We first selected several hundred highly or moderately expressed probes, the expression levels of which were upregulated or downregulated at ratios of more than 2 or less than 0.5 in each set of compared cells (e.g., parent OL8 cells versus OL8 cells cultured for 2 years). From among these probes, we next selected those whose expression levels commonly changed during a 2-year culture of genome-length HCV RNA-replicating cells, but which did not change during a 2-year culture period in cured cells. We further examined the expression levels of the selected candidate genes by RT-PCR analysis using additional specimens from the cells cultured for 3.5 years. Reproducibility of the RT-PCR analysis using specimens from recultured cells was also confirmed. Finally, we identified 5 upregulated genes and 4 downregulated genes, the expression levels of which were irreversibly altered during 3.5-year replication of HCV RNA. These genes may play roles in the optimization of the environment in HCV RNA replication, or may play key roles in the progression of HCV-associated hepatic diseases.

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