AIM: To investigate the inhibitory effects of hepatitis B virus (HBV) replication and expression by transfecting artificial microRNA (amiRNA) into HepG2.2.15 cells. METHODS: Three amiRNA-HBV plasmids were constructed and transfected into HepG2.2.15 cells. HBV antigen secretion was detected in the cells with transient and stable transfection by time-resolved fluoroimmunoassays (TRFIA). HBV DNA replication was examined by fluorescence quantitative PCR, and the level of HBV S mRNA was measured by semi-quantitative RT-PCR. RESULTS: The efficiency of transient transfection of the vectors into 2.2.15 cells was 55%-60%. All the vectors had significant inhibition effects on HBsAg and HBeAg at 72 h and 96 h after transfection (P < 0.01 for all). The secretion of HBsAg and HBeAg into the supernatant was inhibited by 49.8% +/- 4.7% and 39.9% +/- 6.7%, respectively, at 72 h in amiRNA-HBV-S608 plasmid transfection group. The copy of HBV DNA within culture supernatant was also significantly decreased at 72 h and 96 h after transfection (P < 0.01 for all). In the cells with stable transfection, the secretion of HBsAg and HBeAg into the supernatant was significantly inhibited in all three transfection groups (P < 0.01 for all, vs negative control). The copies of HBV DNA were inhibited by 33.4% +/- 3.0%, 60.8% +/- 2.3% and 70.1% +/- 3.3%, respectively. CONCLUSION: In HepG2.2.15 cells, HBV replication and expression could be inhibited by artificial microRNA targeting the HBV S coding region. Vector-based artificial microRNA could be a promising therapeutic approach for chronic HBV infection.
AIM: To investigate the inhibitory effects of hepatitis B virus (HBV) replication and expression by transfecting artificial microRNA (amiRNA) into HepG2.2.15 cells. METHODS: Three amiRNA-HBV plasmids were constructed and transfected into HepG2.2.15 cells. HBV antigen secretion was detected in the cells with transient and stable transfection by time-resolved fluoroimmunoassays (TRFIA). HBV DNA replication was examined by fluorescence quantitative PCR, and the level of HBV S mRNA was measured by semi-quantitative RT-PCR. RESULTS: The efficiency of transient transfection of the vectors into 2.2.15 cells was 55%-60%. All the vectors had significant inhibition effects on HBsAg and HBeAg at 72 h and 96 h after transfection (P < 0.01 for all). The secretion of HBsAg and HBeAg into the supernatant was inhibited by 49.8% +/- 4.7% and 39.9% +/- 6.7%, respectively, at 72 h in amiRNA-HBV-S608 plasmid transfection group. The copy of HBV DNA within culture supernatant was also significantly decreased at 72 h and 96 h after transfection (P < 0.01 for all). In the cells with stable transfection, the secretion of HBsAg and HBeAg into the supernatant was significantly inhibited in all three transfection groups (P < 0.01 for all, vs negative control). The copies of HBV DNA were inhibited by 33.4% +/- 3.0%, 60.8% +/- 2.3% and 70.1% +/- 3.3%, respectively. CONCLUSION: In HepG2.2.15 cells, HBV replication and expression could be inhibited by artificial microRNA targeting the HBV S coding region. Vector-based artificial microRNA could be a promising therapeutic approach for chronic HBV infection.
Authors: Carl D Novina; Michael F Murray; Derek M Dykxhoorn; Paul J Beresford; Jonathan Riess; Sang-Kyung Lee; Ronald G Collman; Judy Lieberman; Premlata Shankar; Phillip A Sharp Journal: Nat Med Date: 2002-06-03 Impact factor: 53.440
Authors: Joyce A Wilson; Sumedha Jayasena; Anastasia Khvorova; Sarah Sabatinos; Ian Gaël Rodrigue-Gervais; Sudha Arya; Farida Sarangi; Marees Harris-Brandts; Sylvie Beaulieu; Christopher D Richardson Journal: Proc Natl Acad Sci U S A Date: 2003-02-19 Impact factor: 11.205
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