Characterization of HCV replication and pathogenesis in animal model

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[Purpose] Previous studies of transmission of hepatitis C virus (HCV) by intrahepatic inoculation with synthetic RNA have been reported. However, rescue of viable HCV particle from these chimpanzees or in vitro culture system had not be successful, so far. In this study, we established reverse genetical system of HCV to characterize genome function. [Methods] We constructed the entire HCV cDNA clone from chronic hepatitis patient plasma and expressed exact mRNA of HCV genome using double ribozyme trimming system. The RNA transcripts from this clone have been transfected into IMY-N9 cells, which was the fused cell line of HepG2 and human primaly hepatocyte. [Results and conclusions] Transfection of the synthetic HCV RNA and cDNA into IMY-N9 cells produced infectios HCV, however in HepG2 cells, they could not support HCV replication. The particle with density of 1.1 g/ml in the culture supernatant was visualized by immunoelectron microscopic study. This particle was considered to be the mature particle. This culture supernatant contained HCV particles even after the DNase and RNase treatment and could re-infect to the naive IMY cells and could cause replication. These results indicated that this cDNA clone rescued the infectious HCV particle and has opened the possibility to the reverse genetical approach of HCV.