Transfection and expression of HCV-NS5B gene in Huh-7 cells

To study the mechanism of hepatitis C virus (HCV) replication and to test gene therapy for hepatitis C, a human liver cell line expressed HCV RNA polymerase has been established.Methods NS5B gene has been transfected into Huh 7 cells by lipofectamine The results of transfection were confirmed by PCR and Southern blot analysis, and the level of the non structural protein 5B (NS5B) in Huh 7 cells was detected by Western blot analysis Results There were NS5B gene fragments and the expression of NS5B protein in Huh 7c cells transfected with pTeT NS5B or pcDNA NS5B plasmid Conclusions We have established a HCV RNA polymerase expression system in Huh 7 cells which can be further used to analyze the mechanism of HCV replication and provide a cell model for gene therapy in vitro

Effects and mechanisms of silibinin on human hepatoma cell lines

AIM： To investigate in vitro effects and mechanisms of silibinin on hepatocellular carcinoma （HCC） cell growth, METHODS： Human HCC cell lines were treated with different doses of silibinin. The effects of silibinin on HCC cell growth and proliferation, apoptosis, cell cycle progression, histone acetylation, and other related signal transductions were systematically examined. RESULTS： We demonstrated that silibinin significantly reduced the growth of HUH7, HepG2, Hep3B, and PLC/PRF/5 human hepatoma cells. Silibinin-reduced HuH7 cell growth was associated with significantly up- regulated p21/CDK4 and p27/CDK4 complexes, down- regulated Rb-phosphorylation and E2F1/DP1 complex. Silibinin promoted apoptosis of HuH7 cells that was associated with down-regulated survivin and upregulated activated caspase-3 and -9. Silibinin＇s antiangiogenic effects were indicated by down-regulated metalloproteinase-2 （MMP2） and CD34. We found that silibinin-reduced growth of HuH7 cells was associated with increased activity of phosphatase and tensin homolog deleted on chromosome ten （PTEN） and decreased p-Akt production, indicating the role of PTEN/ PI3K/Akt pathway in silibinin-mediated anti-HCC effects. We also demonstrated that silibinin increased acetylation of histone H3 and H4 （AC-H3 and AC-H4）, indicating a possible role of altered histone acetylation in silibininreduced HCC cell proliferation. CONCLUSION： Our results defined silibinin＇s in vitro anti-HCC effects and possible mechanisms, and provided a rationale to further test silibinin for HCC chemoprevention.