Low Oxygen Tension Enhances Hepatitis C Virus Replication

Abstract

; Applied Molecular Virology, Institut Pasteur Korea, Seongnam-si, South Korea g Low oxygen tension exerts a significant effect on the replication of several DNA and RNA viruses in cultured cells. In vitro propagation of hepatitis C virus (HCV) has thus far been studied under atmospheric oxygen levels despite the fact that the liver tissue microenvironment is hypoxic. In this study, we investigated the efficiency of HCV production in actively dividing or differentiating human hepatoma cells cultured under low or atmospheric oxygen tensions. By using both HCV replicons and infectionbased assays, low oxygen was found to enhance HCV RNA replication whereas virus entry and RNA translation were not affected. Hypoxia signaling pathway-focused DNA microarray and real-time quantitative reverse transcription-PCR (qRT-PCR) analyses revealed an upregulation of genes related to hypoxic stress, glycolytic metabolism, cell growth, and proliferation when cells were kept under low (3% [vol/vol]) oxygen tension, likely reflecting cell adaptation to anaerobic conditions. Interestingly, hypoxia-mediated enhancement of HCV replication correlated directly with the increase in anaerobic glycolysis and creatine kinase B (CKB) activity that leads to elevated ATP production. Surprisingly, activation of hypoxia-inducible factor alpha (HIF-␣) was not involved in the elevation of HCV replication. Instead, a number of oncogenes known to be associated with glycolysis were upregulated and evidence that these oncogenes contribute to hypoxia-mediated enhancement of HCV replication was obtained. Finally, in liver biopsy specimens of HCV-infected patients, the levels of hypoxia and anaerobic metabolism markers correlated with HCV RNA levels. These results provide new insights into the impact of oxygen tension on the intricate HCVhost cell interaction. H epatitis C virus (HCV) infection causes a wide range of clinical manifestations, from a healthy carrier state to acute and chronic hepatitis that can lead to fibrosis, cirrhosis, and hepatocellular carcinoma. Nearly 3% of the world's population is chronically infected with HCV (1, 2), and current therapeutic approaches are not broadly effective (3).HCV is a positive-strand RNA virus with a 9.6-kb genome that is flanked at both termini by conserved, nontranslated regions (NTRs), required for RNA translation and replication. The 5= NTR comprises an internal ribosome entry site (IRES) that directs the expression of a polyprotein precursor (4, 5). The polyprotein is cleaved into structural (core, E1, E2) and nonstructural (p7, NS2, NS3, NS4A, NS4B, NS5A, NS5B) proteins that, in association with cellular factors, form a membrane-associated replicase complex. This copies the viral positive-strand RNA into a negative-strand intermediate that serves as the template for the synthesis of progeny genomes. The alternative reading frame (ARFP) or coreϩ1 and minicore proteins, with as-yet-unknown functions, appear to be synthesized from the core region by alternative translation mechanisms (6, 7).Studies of the...

Evaluation of the fidelity of immunolabelling obtained with clone 5D8/1, a monoclonal antibody directed against the enteroviral capsid protein, VP1, in human pancreas

“…This implies that CKB might become upregulated under such conditions. To our knowledge, such upregulation has not been noted previously in cells infected with enteroviruses, although it has been reported that CKB is required for the replication of certain strains of hepatitis C, another positive-strand RNA virus [25,26]. Thus, it seems possible that CKB might play a hitherto unrecognised role in the development of a sustained infection of islet cells.…”

Hepatitis C virus utilizes VLDLR as a novel entry pathway

“…It has been shown that hypoxic conditions enhance HCV replication (12). We analyzed HCV infection in Huh7.5 cells under hypoxic conditions and observed increased infectivity of JFH1 (HCVcc JFH1 ), an infectious HCV clone (Fig.…”

“…Recent studies have demonstrated that HCV RNA replication in Huh7.5 cells is enhanced under hypoxic conditions (12). Because the oxygen content in liver tissue in vivo is estimated to be lower (with a gradient of 9-3%) than the oxygen content under in vitro culture conditions (13), the HCV life cycle may differ significantly from that observed using in vitro culture systems.…”

“…In vitro analysis has shown that during the early stage of infection HCV recognizes lipoprotein receptors such as SR-BI and LDLR on target cells via the association of the virus with apolipoprotein E (ApoE) or other related ligands (18). However, the cell lines that have been widely used for the analysis of HCV infection/replication (i.e., Huh7 and its derivatives) do not express VLDLR under conventional culture conditions (12), thereby preventing analysis of the role of VLDLR in HCV infection.…”

Stemness-Related Transcriptional Factors and Homing Gene Expression Profiles in Hepatic Differentiation and Cancer

“…in Huh7.5 cells, which was associated with increased activity of anaerobic cell energy production and proliferation (59). Thus, the interplay between hypoxia-induced stemness and cell energy machinery activation by HCV, and through that enhancement of HCV replication, may promote HCV spread and tumorigenesis in chronically infected individuals (60).…”