Inhibition of hepatitis B virus gene expression and replication by artificial microRNA |
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Authors: | Gao Yu-Feng Yu Li Wei Wei Li Jia-Bin Luo Qing-Li Shen Ji-Long |
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Institution: | 1. Institute of Clinical Pharmacology, Anhui Medical University, Hefei 230032, Anhui Province, China;Anhui Key Laboratory of Zoonoses, Hefei 230032, Anhui Province, China;The Key Laboratory of Gene Resource Utilization for Severe Diseases, The Ministry of Education of China and Anhui Province, Hefei 230032, Anhui Province, China;Department of Infectious Diseases, The First Affiliated Hospital of Anhui Medical University, Hefei 230032, Anhui Province, China 2. Institute of Clinical Pharmacology, Anhui Medical University, Hefei 230032, Anhui Province, China;Anhui Key Laboratory of Zoonoses, Hefei 230032, Anhui Province, China;The Key Laboratory of Gene Resource Utilization for Severe Diseases, The Ministry of Education of China and Anhui Province, Hefei 230032, Anhui Province, China 3. Institute of Clinical Pharmacology, Anhui Medical University, Hefei 230032, Anhui Province, China 4. Institute of Clinical Pharmacology, Anhui Medical University, Hefei 230032, Anhui Province, China;Department of Infectious Diseases, The First Affiliated Hospital of Anhui Medical University, Hefei 230032, Anhui Province, China |
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Abstract: | 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 HBeAginto the supernatant was in hibited by 49.8% 4.7%and 39.9% ± 6.7%, respectively, at 72 h in amiRNA-HBV-S608 plasmid transfection group. The copy of HBVDNA within culture supernatant was also significantlydecreased 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. |
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Keywords: | Hepatitis B virus RNA interference Artificial microRNA HepG2 2 15 cell |
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