LNAzyme抑制HCV 5'端非编码区内源性核糖体进入位点基因表达

LNAzyme targeting the HCV internal ribosome entry site inhibits HCV RNA expression in HepG2.9706 cells

目的:观察针对丙型肝炎病毒(HCV)5’非编码区(NCR)内源性核糖体进入位点(IRES)的锁核酸核酶对HepG2.9706细胞中HCVRNA表达的抑制作用.方法:设计合成针对HCV5'-NCR区IRES位点的DNAzyme、硫代修饰DNAzyme及LNAzyme.实验设对照组与实验组.对照组包括空白对照组、脂质体对照组、无关DNAzyme对照组、裸DNAzyme对照组.实验组包括脂质体-DNAzyme组、脂质体-硫代修饰DNAzyme组、脂质体-LNAzyme组.观察用药后1、3、5、7d时,对HepG2.9706细胞的HCVRNA及荧光素酶基因表达的抑制效应.结果:脂质体包裹的DNAzyme、硫代修饰DNAzyme及LNAzyme对HCVRNA表达均有抑制作用,平均抑制率分别为28.10%、35.25%和44.58%.对荧光素酶基因表达也有抑制作用,平均抑制率分别为31.18%、40.69%和52.33%.与对照组比较均有显著性差异(P<0.05).对HepG2.9706细胞未见明显细胞毒性作用.结论:LNAzyme对HepG2.9706细胞内HCVRNA的表达具有显著抑制作用,且优于硫代修饰的DNAzyme,是一类特异的高效抗HCV分子药物.

AIM：To investigate the inhibitory effects of LNAzyme targeting the hepatitis C virus（HCV） internal ribosome entry site（IRES） on HCV RNA expression in HepG2.9706 cells. METHODS：The sequences encoding DNAzyme,thiolmodificated DNAzyme and LNAzyme directing against the HBV IRES（located in the 5＇ non-coding region） were designed and synthesized.The following experimental groups were set up：Lipofectamine-DNAzyme group,Lipofectamine-S-DNAzyme group,LipofectamineLNAzyme group,blank control group,empty Lipofectamine group,DNAzyme group,and random DNAzyme group.On days 1,3,5 and 7 after transfection,the expression of HCV RNA and luciferase gene in HepG2.9706 cells was detected. RESULTS：Signif icant down-regulation of HCV RNA and luciferase gene expression was noted in the Lipofectamine-DNAzyme,LipofectamineS-DNAzyme and Lipofectamine-LNAzyme groups when compared with other groups（all P〈0.05）.The reduced rates of HCV RNA expression in the above three groups were 28.10%,35.25% and 44.58%,respectively.The reduced rates of luciferase gene expression were 31.18%,40.69% and 52.33%,respectively.LNAzyme did not exert cytotoxicity in HepG2.9706 cells. CONCLUSION：LNAzyme has a significant inhibitory effect on HCV RNA expression in HepG2.9706 cells.The inhibitory effect of LNAzyme on HCV RNA expression is stronger than that of thiolmodif icated DNAzyme.