[Expression and subcellular localization of APOBEC3G in peripheral blood mononuclear cells and liver tissues of chronic HBV patients].

OBJECTIVE: To study the expression level and intracellular localization of APOBEC3G in peripheral blood mononuclear cells (PBMCs) and liver tissues of chronic HBV patients. METHODS: The expression level and intracellular localization of APOBEC3G in PBMCs and liver tissues were detected using the western blot and confocal laser scanning microscope (CLSM). RESULTS: Western-blot showed that the expression level of APOBEC3G in PBMCs of healthy controls was very low. The relative expression levels of APOBEC3G in PBMC of patients with chronic hepatitis B, chronic severe hepatitis, liver cirrhosis, or liver cancer were 4.12+/-0.21, 4.07+/-0.28, 4.16+/-0.36 or 4.21+/-0.39 respectively, which were higher than that in the healthy controls. However, there was no significant difference in APOBEC3G expression among different chronic HBV patients (q = 0.931, 0.744, 1.675, 1.675, 2.606 or 0.931, respectively, all P values more than 0.05). In addition, there was no significant difference on APOBEC3G in liver tissues between chronic hepatitis B patients and hepatocellular carcinoma patients (4.40+/-0.34 vs 4.34+/-0.43, q = 0.588, P more than 0.05). CLSM indicated that the localization of APOBEC3G protein was in cytoplasm of PBMCs and hepatocytes. CONCLUSION: APOBEC3G is upregulated in the PBMCs of chronic hepatitis B patients.

Effects of cell cycle on telomerase activity and on hepatitis B virus replication in HepG2 2.2.15 cells.

BACKGROUND: It has been shown that telomerase activity and hepatitis B virus (HBV) replication are closely associated with cell cycle. This study aimed to further investigate the effects of cell cycle on telomerase activity and on HBV replication. METHODS: Human hepatoma cells transfected with HBV DNA (HepG2 2.2.15 cell line) were treated respectively with serum deprivation, all-trans retinoic acid (RA), dimethyl sulfoxide (DMSO), or sodium butyrate. The cell cycle of HepG2 2.2.15 cells was analyzed by flow cytometry. The telomerase activities of the cells were detected by TRAP-PCR-ELISA. HBV DNA in culture medium was assayed by a fluorescent quantitative PCR assay and a semiquantitative dot blot hybridization technique. HBsAg and HBeAg in culture media were quantitatively examined by an ELISA assay. RESULTS: Treatments with serum deprivation, RA, DMSO, or sodium butyrate inhibited the proliferation of HepG2 2.2.15 cells and led to cell arrest in the G0/G1 phase of cell cycle. The percentage of the G0/G1 phase in the groups of sodium butyrate, DMSO, RA and serum-free was 85.2%, 71.9%, 68.3% and 65.2%, respectively, but in the control group, 43.1% (P<0.01). The activities of telomerase of the cells were also significantly inhibited by 82.8%, 74.6%, 76.1% and 69.4% respectively. In addition, HBV replication of the HepG2 2.2.15 cells remarkably increased as shown by the contents of HBV DNA, HBsAg and HBeAg in the culture media of the cells treated with sodium butyrate, DMSO, RA or serum deprivation (P<0.01). The amounts of HBV DNA in the groups of sodium butyrate, DMSO, RA, serum deprivation and control were 6.7X10(6), 4.8X10(6), 4.4X10(6), 5.1X10(6) and 1.2X10(6) copies/ml, respectively (P<0.01). Telomerase was expressed mainly in the cells in S phase. HBV replication increased in quiescent cells (G0/G1 phase), and decreased in proliferating phase (S phase). CONCLUSION: The current data approve that HBV replication is associated with the cellular proliferative activity.

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

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.