Combination of small interfering RNAs mediates greater suppression on hepatitis B virus cccDNA in HepG2.2.15 cells

AIM: To observe the inhibition of hepatitis B virus (HBV) replication and expression in HepG2.2.15 cells by combination of small interfering RNAs (siRNAs). METHODS: Recombinant plasmid psil-HBV was constructed and transfected into HepG2.2.15 cells. At 48 h,72 h and 96 h after transfection,culture media were collected and cells were harvested for HBV replication assay. HBsAg and HBeAg in the cell culture medium were detected by enzyme-linked immunoadsorbent assay (ELISA). Intracellular viral DNA and covalently closed circular DNA (cccDNA) were quantifi ed by real-time polymerase chain reaction (PCR). HBV viral mRNA was reverse transcribed and quantifi ed by reverse-transcript PCR (RT-PCR). RESULTS: siRNAs showed marked anti-HBV effects. siRNAs could specifically inhibit the expression of HBsAg and the replication of HBV DNA in a dose-dependent manner. Furthermore,combination of siRNAs,compared with individual use of each siRNA,exerted a stronger inhibition on antigen expression and viral replication. More importantly,combination of siRNAs significantly suppressed HBV cccDNA amplifi cation. CONCLUSION: Combination of siRNAs mediates a stronger inhibition on viral replication and antigenexpression in HepG2.2.15 cells,especially on cccDNA amplifi cation.