Abstract:   Background and purpose: Hepatitis B virus X protein (HBx) and hypoxia inducible factor-1α (HIF-1α) play key roles in hepatocarcinogenesis and the development of hepatocellular carcinoma. Positive correlation on the expression of these 2 proteins in hepatocellular carcinoma tissues has been found, whereas the underlying mechanisms have not been fully elucidated. This study focused on the role of HBx in regulating HIF-1α and the underlying mechanisms in hepatocellular carcinoma cells. Methods: The expression plasmids were transfected into Huh7 cells with Lipofectemine^TM 2000. Western blot analysis was applied to detect the expressions of HIF-1α and HIF- 1β protein. The transcriptional activity of HIF-1α was detected by the commercial analysis kits. The mRNA levels of HIF-1α and its target genes, including vascular endothelial growth factor (VEGF) and multi-drug resistance gene 1 (MDR1), were detected by quantitative real-time PCR (qRT-PCR). Immunoprecipitation analysis was applied to detect the interaction of HIF-1α, HBx and protein von Hippel-Lindau (pVHL). Results: Huh7 cells transfected with HBx plasmid led to sharp increase of HIF-1α protein and transcriptional activity, as well as the mRNA of VEGF and MDR1 (P<0.05). However, the mRNA level of HIF-1α was not obviously changed after HBx transfection (P>0.05). Meanwhile, HBx also significantly impaired the function of pVHL in mediating the degradation of HIF-1α by ubiquitin hydrolase. This finding was further confirmed by the immunoprecipitation analysis, which showed that HBx could directly bind to pVHL, but not to HIF-1α. Conclusion: HBx may inhibit the inter-activation between pVHL and HIF-1α through directly binding to pVHL, and thus enhance the stability and transcriptional activity of HIF-1α.

Key words: Hepatocellular carcinoma, Hypoxia inducible factor-lα, Hepatitis B virus X protein, Protein von Hippel-Lindau