HEPATITIS-B VIRUS-REPLICATION IN HUMAN HEPG2 CELLS MEDIATED BY HEPATITIS-B VIRUS RECOMBINANT BACULOVIRUS

A novel transient mechanism for studying hepatitis B virus (HBV) gene expression and replication using recombinant HBV baculovirus to delive r the HBV genome to HepG2 cells was generated. In HBV baculovirus infe cted HepG2 cells, HBV transcripts, and intracellular and secreted HBV antigens are produced; replication occurs as evidenced by the presence of high levels of intracellular replicative intermediates and protect ed HBV DNA in the medium. Density-gradient analysis of extracellular H BV DNA indicated that the DNA was contained predominantly in enveloped HBV virions. Covalently closed circular (CCC) DNA is present indicati ng that, in this system, HBV core particles are capable of delivering newly synthesized HBV genomes back into the nuclei of infected cells. HBV gene expression is driven exclusively from endogenous promoters. L evels of HBV gene expression and replication can be achieved in HBV ba culovirus-infected HepG2 cells which far exceed levels found in HepG2 2.2.15 cells. HBV baculovirus infection of HepG2 cells lends itself re adily to experimental manipulation as follows: 1) HBV expression can b e initiated any time relative to seeding of HepG2 cells; 2) levels of HBV replication can be regulated over a wide range simply by changing the baculovirus multiplicity of infection; 3) HBV replication is readi ly detectable by one day post infection with HBV baculovirus and persi sts at least through day eleven post infection; and (4) the transient nature of the infection can be extended and/or enhanced by superinfect ing the cultures. We conclude that infection of HepG2 cells by HBV rec ombinant baculovirus represents a simple to use and highly flexible sy stem for studying the effects of antivirals and/or cytokines on HBV pr oduction and for understanding HBV replication and pathogenesis at the molecular level.