Propagation of hepatitis 8 virus in a rat hepatoma cell line stably transfected with human annexin-V

ABSTRACT

Hepatitis B virus [HBV] displays a distinct hepatotropism and a narrow host range in vivo. However, very little is known about the interaction of HBV with its host cells, mainly because of difficulties in the development of suitable tissue culture system. We present here confirmatory evidence of a putative role of annexin-V in HBV infection. HBV from both human sera and from culture supernatants from HepG2 2.15 cells were used to infect FTO9.1 cells [a rat hepatoma cell line transfected with a construct containing human annexin-V]. Cells and culture supernatants were assayed at various times post-infection by immunofluorescent microscopy [HBcAg staining in nucleus], and by HBV cccDNA-specific PCR. Supernatants from these initially infected cells were then used to infect fresh FTO9.1 cells with a similar outcome to primary infection. Core and surface gene PCRs were positive on days 2, 5 and following transfer experiments. cccDNA-specific PCR confirmed internalisation of the virus into the nucleus. HBcAg fluorescence showed nuclear staining on days 2, 5 and following transfer experiments. Addition of recombinant annexin-V and DMSO to the cell culture medium resulted in a greater efficiency of infection. Later washes were negative for HBV-DNA, ruling out contamination of the cells by external HBV particles. This cell line does appear to be useful in the study of the early stages of HBV infection, but requires further evaluation