Xiaoguang Li1,2, Yuan Hong1, Qi Wang1, Shunai Liu1, Hongshan Wei1, Jun Cheng3. 1. Institute of Infectious Diseases, Beijing Ditan Hospital, Capital Medical University, No. 8 Jingshun East Street, Chaoyang District, Beijing, 100015, China. 2. Department of Infectious Diseases, The Second Affiliated Hospital of Harbin Medical University, No. 246 Xuefu Road, Harbin, 150086, Heilongjiang, China. 3. Institute of Infectious Diseases, Beijing Ditan Hospital, Capital Medical University, No. 8 Jingshun East Street, Chaoyang District, Beijing, 100015, China. jun.cheng.ditanhospital@gmail.com.
Abstract
PURPOSE: To assess the functions of mismatched short hairpin RNAs (shRNAs) that inhibit replication and the expression of hepatitis B virus (HBV), two shRNAs possessing a 2- or 3-base mismatch that targeted HBV were studied. METHODS: shRNAs and pHY106-HBV were cotransfected into HepG2 cells. The culture supernatants were collected and used in hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) assays. The levels of HBsAg and HBcAg mRNA were detected by reverse-transcriptase PCR (RT-PCR). HBV DNA replication intermediates were extracted for Southern blot hybridization. RESULTS: The results demonstrate that mismatched shRNA-458 and shRNA-635 can significantly inhibit HBsAg and HBeAg protein expression, and the maximal inhibition ratio for both proteins was found at 72 h after cotransfection: 80 and 50 %, respectively. Similar inhibitory effects were found on HBsAg and HBcAg mRNA levels and HBV DNA replication intermediates at 72 h after cotransfection, and the inhibition ratio was found to be approximately 70 and 90 %, respectively. CONCLUSIONS: Despite the 2- or 3-base mismatch between the shRNAs and the HBV target sequences, shRNA-458 and shRNA-635 exerted a significant inhibitory effect on HBsAg and HBeAg expression and HBV replication. This indicates that mismatched shRNAs could be a promising therapy for HBV.
PURPOSE: To assess the functions of mismatched short hairpin RNAs (shRNAs) that inhibit replication and the expression of hepatitis B virus (HBV), two shRNAs possessing a 2- or 3-base mismatch that targeted HBV were studied. METHODS: shRNAs and pHY106-HBV were cotransfected into HepG2 cells. The culture supernatants were collected and used in hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) assays. The levels of HBsAg and HBcAg mRNA were detected by reverse-transcriptase PCR (RT-PCR). HBV DNA replication intermediates were extracted for Southern blot hybridization. RESULTS: The results demonstrate that mismatched shRNA-458 and shRNA-635 can significantly inhibit HBsAg and HBeAg protein expression, and the maximal inhibition ratio for both proteins was found at 72 h after cotransfection: 80 and 50 %, respectively. Similar inhibitory effects were found on HBsAg and HBcAg mRNA levels and HBV DNA replication intermediates at 72 h after cotransfection, and the inhibition ratio was found to be approximately 70 and 90 %, respectively. CONCLUSIONS: Despite the 2- or 3-base mismatch between the shRNAs and the HBV target sequences, shRNA-458 and shRNA-635 exerted a significant inhibitory effect on HBsAg and HBeAg expression and HBV replication. This indicates that mismatched shRNAs could be a promising therapy for HBV.
Authors: C L Lai; R N Chien; N W Leung; T T Chang; R Guan; D I Tai; K Y Ng; P C Wu; J C Dent; J Barber; S L Stephenson; D F Gray Journal: N Engl J Med Date: 1998-07-09 Impact factor: 91.245
Authors: Torgeir Holen; Svein Erik Moe; Jan Gunnar Sørbø; Trine J Meza; Ole Petter Ottersen; Arne Klungland Journal: Nucleic Acids Res Date: 2005-08-19 Impact factor: 16.971