Analysis of replication of Hepatitis C Virus using highly efficient replication system

丙型肝炎病毒(丙型肝炎病毒)是一个主要的公共卫生问题，全世界估计有1.7亿人感染。目前丙型肝炎病毒相关性慢性肝炎的治疗是基于干扰素的使用。然而，病毒清除率还不够。由于缺乏病毒培养系统，开发抗病毒疗法或了解这种病毒的生命周期的研究一直受到阻碍。我们从一例重型肝炎患者中分离到JFH-1毒株，JFH-1亚基因组复制子可以在培养细胞中高效复制，而不需要适应性突变。利用该JFH-1毒株，我们建立了高效的丙型肝炎病毒培养细胞复制系统。将JFH-1全长RNA导入Huh7细胞。随后，病毒RNA在转基因细胞中有效复制，并分泌病毒颗粒。经电子显微镜分析，病毒分泌物颗粒呈球形，分离出的病毒颗粒密度为1.15g/ml。此外，分泌型病毒对未成熟的HuH7细胞具有感染性。此外，我们还开发了利用JFH-1全基因组的复制子系统。这两个系统都为研究病毒生命周期和构建抗病毒策略提供了强大的工具。

Hepatitis C virus (HCV) is a major public health problem, infecting an estimated 170 million people worldwide. Current therapy for HCV-related chronic hepatitis is based on the use of interferon. However, virus clearance rates are insufficient. Investigations to develop the anti-viral therapy or to understand the life cycle of this virus have been hampered by the lack of viral culture systems. We isolated the JFH-1 strain from a patient with fulminant hepatitis, and the JFH-1 subgenomic replicon could replicate efficiently in culture cell without adaptive mutation. Using this JFH-1 strain, we developed highly efficient HCV replication system in culture cells. The full-length JFH-1 RNA was transfected into Huh7 cells. Subsequently, viral RNA efficiently replicated in transfected cells and viral particles were secreted. Secreted viral particle exhibited spherical shape by electron-microscopic analysis and was separated in the fraction eith 1.15g/ml density. Furthermore, secreted virus displayed infectivity for naive Huh7 cells. In addition, we have also developed an replicon system using full-genomic JFH-1 genome. Both systems provide powerful tools for studying the viral life cycle and constructing anti-viral strategies.