Inhibition Of Hepatitis C Virus Gene Expression By Hammerhead Ribozymes

Hepatitis C , caused by hepatitis C virus (HCV) , is a worldwide infectious disease transmitted mainly by transfusion and blood products. It has been estimated that more than 1% of world population is infected with HCV, while 3.2% in China. The individuals infected with HCV will become candidate of chronic hepatitis in most of cases, closely related to the pathogenesis of liver cirrhosis and hepatocellular carcinoma. Studies on HCV genome revealed that it is a highly variable positive-strand RNA virus. Currently, there is no vaccine for HCV infection, and the only therapy for chronic hepatitis C is largely ineffective.Ribozymes are catalytic RNA molecules that specifically cleave target RNA substrates based on sequence-specific recognition. Hammerhead ribozyme, as a potential therapeutic agent, is used to inhibit specifically gene expression for many of viruses based on its small RNA with simple construction. In this studies, two hammerhead ribozymes were designed with computer and synthesized by DNA synthesizer, the activity of ribozyme-mediated cleavage was analysized in vitro, and inhibition of HCV gene expression by two ribozyms was studied in HCV-transfected human hepatocellular carcinoma. And also the fussion genes of the HCV C-E1(336-1358nt) DNA with a luciferase reporter gene were cloned in tetracycline-regulated retroviral vector to establish a stable cellular model with the expression of HCV-luc genes.The studies consisited of:1. The secondary structure of 5'noncodening region (5'NCR) and core region of HCV-H strain as target sequences was calculated with the help of computer, the ribozymes against HCV were designed according to the "hammerhead structure" suggested by...