Inhibition of hepatitis B virus expression and replication by RNA interference

AIM: To s tudy the RNA inter ference on hepatitis B virus (HBV) replication by a reverse transcription virus vector which can express short hairpin RNA inside cells. METHODS: pSIREN vectors with inserted oligonucleotides targeting on reverse transcriptase (RT) regions of HBV genome were constructed. These plasmids were co-transfected with pHBV3.8 into Huh-7 cells. Viral antigens were measured by enzyme-linked immunosorbent assay (ELISA). HBV core particle DNA was measured and quantified by real-time fluorescence quantitative polymerase chain reaction (RFQPCR) and Southern blot. Viral RNA was analyzed by Northern blot. RESULTS: Three RNA interfering targets were identified, and three corresponding retrovirus vectors, named 154i, 312i and 734i, were obtained. It was found that 312i markedly inhibited the expression of pHBV3.8, and the levels of HBsAg and HBeAg were 39% and 41% of those in the negative control group (P = 0.001, P = 0.000). RFQ-PCR showed that the level of HBV core particle DNA was significantly lower in 312i group than that in the negative control group (21.3% ± 1.1% vs 100.0% ± 10.6%, P = 0.0046). Southern and Northern blot demonstrated a lowest replication and transcription level of HBV in 312i group (10.5%, 12.0%). CONCLUSION: A new RNAi system is identified in the RT regions of HBV genome, and the corresponding retrovirus vectors, which can remarkably inhibit the replication and expression of HBV, are also constructed.