中华医学杂志
中華醫學雜誌
중화의학잡지
National Medical Journal of China
2009年
5期
347-351
,共5页
边中启%崔志磊%陈维灶%刘明秋%严维耀%郑兆鑫
邊中啟%崔誌磊%陳維竈%劉明鞦%嚴維耀%鄭兆鑫
변중계%최지뢰%진유조%류명추%엄유요%정조흠
肝炎病毒,乙型%RNA干扰%小干扰RNA%病毒复制
肝炎病毒,乙型%RNA榦擾%小榦擾RNA%病毒複製
간염병독,을형%RNA간우%소간우RNA%병독복제
Hepatitis B virus%RNAi%miRNA%Virus replication
目的 探讨靶向HBV S基因的siRNA表达质粒在HepG2.2.15细胞中对HBV病毒的复制与表达的抑制效应及其抗病毒活性.方法 设计并构建了两个靶向HBV S基因的siRNA表达质粒S1和S2,随机设计的用于对照的非同源siRNA表达质粒S3.首先在HepG2.2.15细胞中通过实时荧光定量PCR评估该siRNA表达质粒对HBV mRNA表达的抑制效应,接着在HepG2.2.15细胞中通过ELISA方法进一步枪测它们的抗病毒活性细胞上清液中HBV标志性蛋白HBsAg、HBeAg的表达水平.结果 在siRNA表达质粒转染HepG2.2.15细胞后48 h,HBV S基因mRNA表达量下降64%～88%,HBsAg和HBeAg的表达水平分别降低了60%～82%和56%～78%.S1+S2联合使用转染细胞中显著抑制HBV病毒的复制与表达的效率,而对照组S3无抑制效果.结论 发现靶向HBV S基因的siRNA表达质粒能够在HepG2.2.15细胞中有效的抑制HBV病毒的复制与表达,并能够降低细胞上清液中HBsAg和HBeAg的表达水平.S1+S2联合使用转染细胞中可以显著抑制HBV的复制与表达的效率.RNAi可能成为防治HBV感染有效的全新的抗病毒防御策略.
目的 探討靶嚮HBV S基因的siRNA錶達質粒在HepG2.2.15細胞中對HBV病毒的複製與錶達的抑製效應及其抗病毒活性.方法 設計併構建瞭兩箇靶嚮HBV S基因的siRNA錶達質粒S1和S2,隨機設計的用于對照的非同源siRNA錶達質粒S3.首先在HepG2.2.15細胞中通過實時熒光定量PCR評估該siRNA錶達質粒對HBV mRNA錶達的抑製效應,接著在HepG2.2.15細胞中通過ELISA方法進一步鎗測它們的抗病毒活性細胞上清液中HBV標誌性蛋白HBsAg、HBeAg的錶達水平.結果 在siRNA錶達質粒轉染HepG2.2.15細胞後48 h,HBV S基因mRNA錶達量下降64%～88%,HBsAg和HBeAg的錶達水平分彆降低瞭60%～82%和56%～78%.S1+S2聯閤使用轉染細胞中顯著抑製HBV病毒的複製與錶達的效率,而對照組S3無抑製效果.結論 髮現靶嚮HBV S基因的siRNA錶達質粒能夠在HepG2.2.15細胞中有效的抑製HBV病毒的複製與錶達,併能夠降低細胞上清液中HBsAg和HBeAg的錶達水平.S1+S2聯閤使用轉染細胞中可以顯著抑製HBV的複製與錶達的效率.RNAi可能成為防治HBV感染有效的全新的抗病毒防禦策略.
목적 탐토파향HBV S기인적siRNA표체질립재HepG2.2.15세포중대HBV병독적복제여표체적억제효응급기항병독활성.방법 설계병구건료량개파향HBV S기인적siRNA표체질립S1화S2,수궤설계적용우대조적비동원siRNA표체질립S3.수선재HepG2.2.15세포중통과실시형광정량PCR평고해siRNA표체질립대HBV mRNA표체적억제효응,접착재HepG2.2.15세포중통과ELISA방법진일보창측타문적항병독활성세포상청액중HBV표지성단백HBsAg、HBeAg적표체수평.결과 재siRNA표체질립전염HepG2.2.15세포후48 h,HBV S기인mRNA표체량하강64%～88%,HBsAg화HBeAg적표체수평분별강저료60%～82%화56%～78%.S1+S2연합사용전염세포중현저억제HBV병독적복제여표체적효솔,이대조조S3무억제효과.결론 발현파향HBV S기인적siRNA표체질립능구재HepG2.2.15세포중유효적억제HBV병독적복제여표체,병능구강저세포상청액중HBsAg화HBeAg적표체수평.S1+S2연합사용전염세포중가이현저억제HBV적복제여표체적효솔.RNAi가능성위방치HBV감염유효적전신적항병독방어책략.
Objective To study the inhibitive effects of transfection of siRNA expressing plasmids targeting S gene, one of the 4 open reading frames of hepatitis B virus (HBV) , on the rephcation and expression of HBV. Methods Two plasmids expressing 2 siRNAs targeting S gene, one of the 4 open reading frames of HBV (S1 and S2) and one nonspecific plasmid (siRNA-S3), as negative control, with the length of 21 m heterologous to the HBV/U95551 genome were constructed, and then transfected into the hepatic cancer cells of the line HepG2. 2.15. 48 hours later, real-time PCR was used to evaluate the gene silencing efficiency and ELISA was used to detect the expression of HBsAg and hepatitis B e antigen (HBeAg), protein markers of HBV, in the supernatants. Results The inhibition rates of HBsAg and HBeAg expression of the HepG2.2.15 cells transfected with S1 were 60% and 56% respectively, those of the HepG2.2.15 cells transfected with S2 were 73% and 70% respectively, those of the HepG2.2.15 cells transfected with S1 + S2 were 82% and 78% respectively, and those of the HepG2.2.15 cells transfected with S3 were not significantly different from those of the blank control group. RT-PCR showed that the mRNA expression rates of HBsAg and HBeAg in the HepG2.2.15 cells transfected with S1, S2, and S1+S2 were inhibited by 64%-88% t respectively. Conclusion Transfection of the vector plasmids expressing the siRNAs targeting S gene inhibits the expression of HBsAg and HBeAg in hepatic cancer cells. RNAi may provide a viable strategy against viruses for the prevention and treatment of HBV infection.
