The Effect Of Hepatitis B Virus X Protein To Hnf-4α And NF-κB On Human HepG2Hepatocarcimona Cell

1PurposeThis study investigated the HepG2liver cancer cells which are transected by X gene of liposome, then explore the expression of X gene, HNF-4a and NF-κB through cell culture, cell flow, fluorescence microscopy and PCR, Western Blot. It is investigated the relation of X gene, HNF-4a and NF-kappa B in liver cancer cells。2MethodsThe HepG2adult liver cancer cells are divided into three groups:transfected with HBx cells Group, cells transfected with empty vector group and untransfected HepG2cells as control group. Contrast to liver cancer cells growth by microscopic observation, the proliferation of hepatoma cells by cell-flow and fluorescence microscopy to detect different groups. The expression of X gene, HNF-4a and NF-κB nuclear RNA are detected by PCR in each group. The expression of NF-κB nuclear protein is detected by Western blot to explore the relationship between the X gene and NF-κB.3Results3.1GeneralHepG2cells were cultured in medium containing10%fetal bovine serum. The growth of liver cancer cells in transfected X gene group growth better than the other two groups at37℃10%fetal bovine serum for72hours.3.2each group of cells streaming proliferation analysis and fluorescence microscopy imagesThe flow cell proliferation assay:The HepG2cells which is in the transfected HBx group are detected the case of proliferation under37℃in6well plates and incubated for72hours. The rate of proliferation is that Dip G2is5.68%at192.21in the hepatoma cells. Fluorescence microscopy images:the rate of proliferation is that transfected HepG2cells is0.47incubated at37℃for72hours. It is more than other two groups.3.3The expression of HNF-4a and NF-κB detected by RT-PCR method in liver cancer cells.The expression value of HBx transfected group is0.24, and the expression of NF-κB is1.13. The expression value of HNF-4a in transfected with empty vector cell group is0.89, while the expression value of the NF-κB is0.59. The expression of HNF-4a in untransfected HepG2cells group is0.36, while the expression of NF-κB is0.46. This shows that the expression of HNF-4a in the transfected HBx cells group is less than that of the other two groups. And the NF-κB expression in the transfected HBx cells group is more than the other two groups.In the same group, the HNF-4a is negatively correlated with NF-κB expression.3.4The detection of NF-κB protein levels in transfected liver cells by Western-blot.The expression of NF-kB protein in the Transfected HBx cells group is1.019. The expression of HNF-4a in transfected with empty vector cell group is0.294. That of untransfected HepG2cells group is0.487. It shows that the NF-κB protein expression in the HBx cells transfected group is significantly higher than the other two groups. The expression of NF-kB RNA PCR results is a positive correlation with X gene expression.4ConclusionThe X gene was successfully transfected into HepG2cells, and the expression of X gene protein in transfected with HBx cells can inhibit the expression of HNF-4a. Then it promotes the activity of NF-κB and the occurrence of hepatocellular carcinoma.