The effect of hepatitis B virus X protein on the expression of CtIP in HepG2 Cells

Objective To investigate the effect of hepatitis B virus X protein(HBx) on CtBP-interacting protein(CtIP) which is an important repair factor of DNA double strand break damage in HepG2 cells induced by bleomycin. Methods A HBx stably expressing HepG2 cell line and a control HepG2 cell line with empty vector transfected were established. After the double strand break (DSB) damage occurred, the mRNA and protein levels of CtIP were detected by Real-time PCR and Western blot assay respectively, cell cycle profiles and apoptotic cell death were determined by a flow cytometry, and the position of CtIP in cells was observed by confocal laser scanning microscopy. Results It showed that HepG2 cells transfected with hepatitis B virus X gene could stably express HBx protein. After being induced by bleomycin, the percentage of apoptotic cell was 16.90% ± 0.89% in HBx stably expressing HepG2 cell line and 15.30% ± 0.86% in control cell line, respectively( q = 2.074, P ＞ 0.05). While the percentage of death cell was 8.71% ± 0.74% in HBx stably expressing HepG2 cell line and 4.90% ± 0.46% in control cell line, respectively( q = 7.126, P ＜ 0.01). The two cell lines manifested the increase of G2/M arrest and significant difference existed between the two cell lines.HBx down regulated the expression levels of CtIP and its mRNA. The CtIP level was 0.66 ± 0.04 in HepG2-HBx cell and 0.73 ± 0.05 in HepG2-vec cell, respectively( t = 2.314, P ＜ 0.05). The relative mRNA level was 1.00 ± 0.06 in HepG2-HBx cell and 1.23 ± 0.08 in HepG2-vec cell, respectively( t = 2. 732, P ＜ 0.05). We also found that CtIP was concentrated in the cell nucleus. Conclusion The research suggests that HBx may affect DNA-repair pathways by disrupting the expression of CtIP. Key words: Carcinoma,hepatocellular; Hepatitis B virus X protein; DNA double-strand break; Repair