Hypermethylation Of P16 And Biological Effects Mediated By HBx In Early HCC

Hepatocellular carcinoma is one of the most common malignant tumor. The incidence of hepatocellular carcinoma is related with many factors. Infection of hepatitis B virus is a worldwide problem. The process leading to liver cancer is not clear, but the long-term carrier of Hepatitis B virus has a significant relationship with the occurrence of Hepatocellular carcinoma. HBV expresses four proteins, and HBx protein (hepatitis B virus X protein) plays an important role in HCC. HBx greatly promotes the development of hepatocellular carcinoma in chronic hepatitis, but its specific mechanism of carcinogenesis is still controversial. For early diagnosis, risk assessment and preventive treatment, we should find and understand the molecular mechanisms of HBx led to liver cancer. Hypermethylation of p16 which HBx induce may be a important reason in HCC occurrence.Work Objective:To get the cell of Chang-HBx expressing HBx and Chang-vector not expressing HBx, confirm that HBx induces p16 promoter hypermethylation and reduce the expression of p16.Hypermethylation of p16 promoter caused by HBx introduce some biological effects, which are important events in early HCC.Methods:PcDNA3.1B-HBx and pcDNA3.1B plasmids were introduced into Chang cells by liposome transfection method. Get Chang-HBx cell expressing HBx and Chang-vector not expressing HBx. Monitor p16 mRNA transcription and protein expression levels of Chang, Chang-vector and Chang-HBx by RT-PCR and Western blot methods. Know the level of p16 promoter methylation by MSP, draw the proliferation curves by CCK-8, and observe S phase in cell cycle and apoptosis by flow cytometry.Results:Hypermethylation of p16 can be mediated by HBx, which decreases the expression of mRNA and protein of p16. Chang-HBx cells which express HBx growth faster, and their phase of S in cell cycle increases and apoptosis decreases.