SiRNA Targeting S Gene Inhibits Hepatitis B Virus Expression And Replication In HepG2.2.15 Cells

The hepatitis B virus(HBV),as a major cause of acute and chronic hepatitis in humans,contains a partial double-stranded circular DNA genome of 3.2 kb that is transcribed into the 3.5-,2.4-,2.1-,and 0.7-kb viral transcripts by the host RNA polymeraseâ…¡.HBV infection may leads to severe hepatitis,liver cirrhosis and hepatocellular carcinoma.HBV is a member of the Hepadnaviriade family.S gene,one of four encoding protein genes of HBV,is essential to encode for hepatitis B surface antigen(HBsAg) during the life cycle of the virus and plays a key role in virus attachment to susceptible cells.RNA interference(RNAi)is a mechanism that inhibits gene expression by causing the degradation of specific RNA molecules or hindering the transcription of specific genes.In this study,we construct two plasmids expressing siRNAs(S1 and S2)targeting S gene of HBV.Also,one nonspecific siRNA(S3)with the length of 21 nt heterologous to the U95551 genome was designed randomly for negative control.These expression plasmids are transfected into HepG2215 (2215)cells.First,we evaluate the gene silencing efficiency of both plasmids in 2215 cells.Then,the antiviral potentials of both plasmids in 2215 cells are investigated.It was found by real time PCR(RT-PCR)and enzyme-linked immunosorbent assay(ELISA)detection conducted after cotransfection at intervals of 48h that the expression of either groups S1 or S2 and S1+S2 were reduced significantly in 2215 ceils at the 48h,in comparison with negative control group(transfected with S3).The results indicated by an ELISA that transfection of siRNA-expressing vectors S1 and/or S2 caused an 60%-82%reduction of HBsAg and 56%-78%reduction of hepatitis B e antigen(HBeAg)in 2215 cells,respectively.The HBV mRNA was significantly reduced by 64%-88%based on RT-PCR.Furthermore,we found that RNAi induced by siRNA targeting HBV S gene is continous and stable inhibition of HBV expression and replication in 2215 cells.Our data suggest that RNAi may provide a viable preventive strategy against viruses for the prevention and treatment HBV infection.