Inhibitory effects of RNAi on expression of hepatitis B virus antigen in vitro

Objective To construct the vector expressing shRNA that specifically targets the pre-core gene sequence of hepatitis B virus and investigate its inhibitory effect on the expression of hepatitis B virus antigen.MethodsThe recombinant vectors that expressing shRNA was constructed through gene recombination and confirmed with restriction enzyme digestion and DNA sequencing.Then,the eukaryotic expression plasmid pHBV1.5 containing 1.5-fold-overlength genome of HBV and the vectors expressing shRNA were co-transfected into Hela cells.At 72 hours post transfection,the levels of HBsAg and HBeAg in the cell culture medium and cell lysate were determined by microparticle enzyme immunoassay(MEIA) with Abbot kits.Prec/c mRNA was analyzed by semi-quantity PCR.ResultsWe successfully constructed the vectors expressing shRNA that could specifically target the pre-core gene sequence of hepatitis B virus and the vector expressing shRNA that had nothing to do with HBV.psiHBV4 could efficiently inhibit the expression of HBeAg,as well as promotion the expression of HBsAg.Whereas the other two vectors expressing shRNA exhibited less inhibitory effect on the replication and the expression of HBeAg but promoted the expression of HBsAg at different extent.The control vector psiC has no effects on HBV.ConclusionThe psiHBV4 expressing shRNA that specifically targeted the pre-core gene sequence of hepatitis B virus could exert robust inhibitory effects on HBeAg expression,which suggests that RNAi strategy may represent a potentially efficacious approach to the clinical management of HBV infection.