Isolation and characterization of an Huh.7.5.1-derived cell clone highly permissive to hepatitis C virus

An efficient cell culture and infection system for hepatitis C virus (HCV) facilitates analyses of the complete virus life cycle. Human hepatic Huh7.5.1 cells and an HCV-JFH1 strain have been widely employed in infection experiments. In the present study, cultured Huh7.5.1 cells exhibited heterogeneous phenotypes of HCV infection. Using single-cell cloning of Huh7.5.1 cells, we isolated a clone highly permissive to HCV (Huh7.5.1-8) and a CD81-defective clone nonpermissive to HCV (Huh7.5.1-5). Expression of CD81 in Huh7.5.1-5 cells restored permissiveness to HCV, indicating that CD81 is essential for HCV infection and a defect in CD81 causes nonpermissiveness to HCV in Huh7.5.1-5 cells. Huh7.5.1-8 cells had approximately 10-fold higher HCV replication rates, with cellular HCV RNA copy numbers of >10(9) copies/μg of cellular RNA and viral titers of >10(6) infectious units/ml of culture supernatant. Permissiveness of Huh7.5.1-8 cells to HCV infection was phenotypically very stable because there was no difference in permissiveness after more than 100 passages (1-year culture). This efficient cell culture system for HCV using Huh7.5.1-8 cell provides a powerful tool for studying the HCV life cycle and constructing antiviral strategies.