Variation of hypervariable regions of the genome of hepatitis C virus in EBV transforming PBMC from patients with hepatitis C / 中华传染病杂志

ABSTRACT

Objective To study the persistence and replication of hepatitis C virus (HCV) RNA in Epstein Barr virus (EBV) transforming peripheral blood mononuclear cell (PBMC) from patients with he patitis C as well as variation of hypervariable regions (HVR) of the HCV genome. Methods PBMC from one patient with hepatitis C was infected by EBV and then transformed into lymphoblasts capable of being propagated indefinitely. Then, HCV RNA of the cultured cells and supernatants was detected by reverse transcriptase polymerase chain reaction (RT PCR) every month and in situ PCR to identify the location of HCV RNA in the cells. The HVR genome sequences of HCV in the first and the ninth month subculture cells were identified by sequence analysis. Results HCV plus strand RNA could be detected in the cultured cells for as long as 1 year. The HCV plus strand RNA could be identified in supernatants and the minus strand RNA were also observed in the cultured cells intermittently. In situ PCR showed that the blue black positive signals of HCV RNA located mainly in cytoplasmas of EBV transforming B cell but negative in nucleous. The positive signal was not found in negative control cells by in situ RT PCR. HCV HVR genome sequence after half year was found to have two regions of a high degree of variability in 1491 to 1583 nucleotide (384 to 414 amino acid) and 1761～1781 nucleotide (473 to 479 amino acid). But, compared the HCV HVR genome sequence in first month subcultured cells with that in ninth month, there was not significant difference. Conclusion HCV may exist in the cultured cell line for a prolonged period wihtout HVR genome sequence changed.