DEMONSTRATION OF IN-VITRO INFECTION OF CHIMPANZEE HEPATOCYTES WITH HEPATITIS-C VIRUS USING STRAND-SPECIFIC RT PCR/

Analysis of hepatitis C Virus (HCV) replication has been hampered due to the difficulty encountered in in vitro cultivation of the virus in conventional tissue culture systems. In this study, primary chimpanzee hepatocyte cultures maintained in a serum-free medium formulation wer e susceptible to in vitro infection with HCV. In order to document inf ection, two new methods of reverse transcription/polymerase chain reac tion were developed that permit accurate distinction between positive and negative strand HCV RNA. One method relied upon the use of a tagge d cDNA primer, while the second method employed a thermostable reverse transcriptase. Following inoculation of chimpanzee hepatocytes with H CV, intracellular positive and negative strand HCV RNA were detectable 4 days postinfection and throughout the remainder of the experimental period, 25 days. Analysis of HCV-inoculated baboon hepatocytes reveal ed a total absence of negative strand HCV RNA, while residual positive strand RNA from the inoculum could be detected for up to 11 days. The in vitro replication of HCV RNA in chimpanzee hepatocytes could be su ppressed by alpha-interferon. This system should be amenable to the st udy of HCV replication, antiviral compounds, and the development of ne utralization assays. (C) 1994 Academic Press, Inc.