Detection of hepatitis C virus in the supernatant of human placenta trophoblastic cells and cell lysates

Objective To investigate HCV infection and replication in the primarily cultured human trophoblastic cells which were cultured by improved separation and purification methods and co-cultured with HCV infection serum.Methods Human placenta was digested with trypsin-DNase and then centrifuged with percoll density gradient in order to obtain the trophoblastic cells, which were incubated in HCV positive serum (the quantification of HCV RNA was 5.9×107copies/ml) .The expression of HCV non-structure protein 5 (HCV-NS5) was evaluated by immunofluorescence.The infected cells were lysed.HCV RNA in HCV infected trophoblast cells was quantitated by RT-PCR.Results The culture model of trophoblastic cells from human placenta was established and cytokerat was observed in positive cells.Observation of the morphology of trophoblastic cells showed that no significant cytopathic effect was found when co-cultured with HCV positive serum.The antigen of HCV NS5 could be detected at 48 hours after infection.The HCV RNA was detected in the supernatent of the culture medium at 24, 48, 72, 96, 120 hours after infection respectively.HCV RNA was detected in HCV infected cells.Washing liquid was not contaminated by HCV positive serum.Cell lysates which contained HCV RNA was detected by RT-PCR.Conclusion Our study indicates that trophoblastic cells cultured in vitro can be infected by HCV.The replication of HCV in trophoblastic cells was detected.