[Effect of cell cycle on telomerase activity of hepatoma cells and its relationship with replication of hepatitis B virus]

Abstract

Background & objective: Previous studies have shown that the expression of telomerase activity is closely correlated with the formation and development of tumor cells. Furthermore, the cell cycle is associated with hepatitis B virus (HBV) replication level and telomerase activity. For further study their relationship, this experiment was designed to investigate the effect of serum deprivation or all-trans-retinoic acid(RA)on cell cycle of human hepatoma cells transfected by HBV DNA (HepG2 cell line) and the associations of cell cycle with telomerase activity and HBV replication.

Methods: Human hepatoma HepG2 cells were respectively treated with serum deprivation or RA. Cell cycle was analyzed using flow cytometry. Telomerase activity was determined quantitatively by TRAP-PCR-ELISA. HBV-DNA in culture media was determined using quantitative PCR and semiquantitative dot blot hybridization assay. HBsAg and HBeAg in cell culture media were measured using quantitative ELISA.

Results: RA treatment or serum deprivation inhibited the proliferation of HepG2 cells and the cells were arrested at G(0)/G(1) phase. The percentages of G(0)/G(1) phase of RA group and serum deprivation were 68.3% and 65.2%, respectively, while that of control group was 43.1% (P< 0.01). The levels of telomerase activity also significantly decreased. The absorbance values that represented the telomerase activity of RA group and serum deprivation group were 0.32 and 0.41, respectively, while that of control group was 1.34(P< 0.01). In addition,HBV replication of HepG2 cells remarkably increased, which was shown as high products of HBV-DNA, HBsAg and HBeAg in culture media of RA group and serum deprivation group. The contents of HBV DNAs were 4.4x10(6), 5.1x10(6), and 1.2x10(6) copies/ml in RA group, serum deprivation group, and control group, respectively(P< 0.01). The values of P/N of HBsAg were 3.5, 3.7, and 1.3 in RA group, serum deprivation group, and control group, respectively (P< 0.01). The values of P/N of HBeAg were 19.8, 22.5, and 13.4 in RA group, serum deprivation group, and control group, respectively (P< 0.01).

Conclusion: Telomerase expression was associated with cell cycle in HepG2 cells. Telomerase was mainly expressed in S phase of cell cycle. HBV replication was also closely correlated with cell cycle, which increased in quiescent hepatocytes and decreased in proliferating hepatocytes.