RNA interference inhibits hepatitis B virus gene expression and replication in HepG2-N10 cells

OBJECTIVE:  RNA interference (RNAi) refers to the phenomenon of sequence‐specific degradation of homologous mRNA induced by double‐stranded RNA. It has been successfully utilized to down‐regulate endogenous gene expression or suppress the replication of various pathogens in mammalian cells. In this...

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Published in:Chinese journal of digestive diseases 2006-11, Vol.7 (4), p.230-236
Main Authors: REN, Jian Lin, PAN, Jin Shui, CHENG, Tong, DONG, Jing, LU, Ya Pi, HUANG, Song Jie, SHI, Hua Xiu, WANG, Lin, LIAN, Ya Mei
Format: Article
Language:English
Subjects:
Cell Line
DNA, Viral - genetics
Gene Expression Regulation, Viral - drug effects
Genetic Vectors
Hepatitis B e Antigens - drug effects
Hepatitis B Surface Antigens - drug effects
Hepatitis B virus
Hepatitis B virus - genetics
Hepatitis B virus - metabolism
Hepatitis B virus - physiology
Humans
Indicators and Reagents - pharmacology
Lipids - pharmacology
RNA Interference
RNA, Small Interfering - pharmacology
RNA, Viral - metabolism
short hairpin RNA
Transfection
virus replication
Virus Replication - drug effects
Virus Replication - genetics
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Summary:OBJECTIVE:  RNA interference (RNAi) refers to the phenomenon of sequence‐specific degradation of homologous mRNA induced by double‐stranded RNA. It has been successfully utilized to down‐regulate endogenous gene expression or suppress the replication of various pathogens in mammalian cells. In this study, the effect of vector‐based small interfering RNA (siRNA) promoted by pSilencer2.0‐U6 inhibit hepatitis B virus (HBV) replication in cell culture was evaluated. METHODS:  Three fragments of short nucleic acids, respectively, targeting on S, X and C region of HBV genome were inserted into pSilencer vectors after they were annealed with their partly antisense strands. The recombination plasmids were pS, pX and pC. These expression plasmids were transfected into HepG2‐N10 cells, a cell line which stably expresses hepatitis B virus surface antigen (HBsAg), hepatitis B virus e antigen (HBeAg) and adw2 subtype Dane particles. The effect of RNAi was evaluated from the changes of DNA, RNA and protein levels. Viral antigens were measured by ELISA. Viral mRNA was analyzed by RT‐PCR. The covalent closed circular DNA and genome DNA of HBV secreted into the culture media were measured by quantitative real‐time PCR. Analysis of variance was performed for the results. RESULTS:  Vector‐based RNA interference could potently reduce HBsAg (pS vs pN: 47%, pX vs pN: 30%, and pC vs pN: 25%, P 
ISSN:1443-9611
1443-9573
DOI:10.1111/j.1443-9573.2006.00268.x