摘要目的 探讨联合靶向特异性小干扰RNA(siRNA)对疱疹病毒(HSV)2的体外特异抑制作用.方法 将体外合成的靶向HSV-2编码包膜糖蛋白gB的UL27.2基因、靶向DNA结合蛋白UL29.2基因的siRNA和该两种联合靶向特异性siRNA制剂,共转染Vero细胞并感染HSV-2,观察靶基因的表达情况、Vero细胞病变、空斑减数实验和子代病毒滴度并进行各组间比较:结果特异性UL27.2siRNA、UL29.2 siRNA和联合siRNA转染Vero细胞后,均能不同程度抑制各HSV-2临床株感染所导致的细胞病变,对病毒增殖抑制率分别为63.9%、86.7%和93.3%.实时定量PCR检测各组siRNA分别对UL27.2和UL29.2两个基因的表达,结果显示UL27.2 siRNA和UL29.2 siRNA对各自的靶向基因均有抑制,而联合靶向siRNA制剂对两个基因的抑制率最高.结论 联合靶向特异性siRNA制剂能有效抑制HSV-2的感染和复制.

abstractsObjective To investigate the inhibition of herpes simplex virus (HSV)2 by combined targeting small interfering RNAs(siRNAs) in vitro. Methods Two different siRNAs,which target HSV-2 UL27.2 gene encoding envelope glycoprotein gB and UL29.2 gene encoding DNA binding protein, were synthesized chemically in vitro. Combined siRNAs composed of these two target-specific siRNAs were transcripted and transfected into Vero cells to infect HSV-2. Real-time PCR was used to measure the effect of siRNA on target gene expression, and estimate the anti-HSV-2 effect with pathological changes, plaque reduce test and the concentrations of filial generation virus. The results were compared among groups. Results Specific U L27.2 siRNA, UL29.2 siRNA and the combined siRNA transfected into Veto cells could inhibit the cytopathic effect due to clinical strains of HSV-2 infection in varying degrees, siRNAs could effectively inhibit virus growth by 63.9%, 86.7% and 93.3%, respectively. Real-time PCR detection of the UL27.2 and UL29.2 gene expression showed that UL27.2 siRNA and UL29.2 siRNA inhibited the expression of the respective targeting genes, while combined siRNA showed the highest inhibition rate of expression. Conclusion Combined target-specific siRNAs may obviously inhibit the replication and infection of HSV-2.