Inhibitory effects of vector-based siRNA targeting S gene on hepatitis B virus gene expression in HepG2 2.2.15 cells

Objective To construct pSUPER vectors expressing short interfering RNA(siRNA) targeting HBV S gene sequence and to evaluate inhibitory effect of these siRNA on HBV replication in HepG2 2.2.15 cells. Methods Based on the sequence of HBV in HepG2 2.2.15 cells, two sequences of targeting HBV S gene were cloned into a RNA polymerase Ⅲ based expression vector pSUPER. The recombinant plasmids were cotransfected with pTK-Hyg into HepG2 2.2.15 cells and monoclonal cell strains were screened. HBsAg and HBeAg in the supernatant of the screened cell strains were assayed with Abbott MEIA kits, the expression of intracellular viral antigens was determined by immunofluorescence staining and HBV mRNA were detected by RT-PCR. Results The pSUPER-S1 and pSUPER-S2 siRNA expression vectors were successfully constructed. The siRNA effectively inhibited the secretion of surface antigen and e antigen of HBV in HepG2 2.2.15 cells by 83% and 78% respectively. Immunofluorescence staining showed that intracellular viral antigens were significantly reduced. RT-PCR results showed that the HBV mRNA was markedly decreased. Whereas the irrelevant siRNA expression vector and control did not show any inhibitory effect on the replication and expression of HBV. Conclusion These results demonstrate that vector-based siRNA targeting HBV S gene can consistently inhibit HBV replication with great potency and specificity in vitro.