Study on cellular factors facilitating HCV replication

• Main Authors: Min-ching Lin, 林旻靜
• Other Authors: Shih-Yen Lo
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/90881780253858667447

碩士 === 慈濟大學 === 醫學生物技術研究所 === 97 === Hepatitis C Virus (HCV) infects 170 million individuals around the world. Infection with HCV frequently causes chronic hepatitis and progresses to liver cirrhosis and even hepatocellular carcinoma (HCC). HCV belongs to the genus Hepacivirus, of the family Flaviviridae. HCV is an enveloped virus. Its genome is a single-stranded (9.6 kb) RNA. HCV isolates are further classified into six genotypes (1–6) and over 100 subtypes. Presently, no safe and effective vaccine is available to prevent HCV infection. Conventional treatment, using the combination of pegylated interferon-α (PEG-IFN-α) and ribavirin is only effective in about 50% of the patients and is associated with important side-effects. Cyclosporin A ( CsA ) , an immunosuppressive agent widely used in the management of liver transplant recipients, has been reported to suppress the hepatitis C virus replication. Suppression of HCV replication by cyclosporin A is mediated by blockade of its cellular targets, cyclophilins. Cyclophilin (CyP) B, a cellular target of CsA, regulates the function of HCV RNA polymerase NS5B. Therefore, cellular factors could affect hepatitis C virus replication and could be the anti-viral targets. In this study, we are going to search for the cellular factors that can facilitate HCV replication. Microarray analysis was used to screen for the genes differentially expressed between HCV replicon cell line and the cell line without HCV replicon(Re*). We them individually knocked-down selected genes over-expressed in HCV replicon by shRNA technology and determined their effects on the HCV replication by measuring the expression of HCV NS5A protein. PLCG2、ELOVL4 and PLA1A were identified as candidate HCV replication facilitators through this screening system and all of these genes are involved in lipid metabolic pathways. Moreover, PLCG2 inhibitor U73122 and PLA1A inhibitor could also inhibit HCV replication. The other genes involved in the metabolic pathways, which these three genes were participated in, were also analyzed. Knock-down of these selected genes involved in the metabolic pathways could also repress HCV replication. Therefore, the end-products rather than the proteins participate in these metabolic pathways could facilitate HCV replication. We are going to confirm these results in the system using JFH-1 infectious clone and the ex vivo system using liver biopsy samples.