Inhibition of West Nile virus replication in cells stably transfected with vector-based shRNA expression system
Inhibition of West Nile virus replication in cells stably transfected with vector-based shRNA expression system
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DOI：
10.1016/j.virusres.2008.04.014
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发表时间：
2008-08-01
期刊：
影响因子：
5
通讯作者：
Ng, M. L.
中科院分区：
文献类型：
--
作者：
Ong, S. P.;Chu, J. J. H.;Ng, M. L.
本研究研究了短发夹rna （short hairpin rna, shRNAs）靶向西尼罗病毒（WNV） Sarafend基因组不同区域的效果。将靶向Capsid、NS2B和NS4B基因的短发夹rna克隆到pSilencer (TM) 3.1-H1 neo中，分别命名为pshCapsid、pshNS2B和pshNS4B。选择阳性转染的Vero细胞构建稳定的表达shrna的细胞系。这些细胞以10的感染倍数（M.O.I.）感染西尼罗病毒。pshCapsid稳定转染的细胞在12和24 h的沉默效果最好。与未转染的wnv感染细胞相比，pshCapsid稳定转染的细胞病毒转录物减少超过4 log（10）个单位，病毒产量减少超过3 log（10）个单位。与pshCapsid稳定转染的细胞相比，稳定转染pshNS2B的细胞没有表现出强烈的抑制作用，病毒滴度仅降低了2 log（10）单位。pshns4b稳定转染的细胞没有抑制西尼罗河病毒的复制。因此，从本研究来看，pshCapsid有可能成为西尼罗河病毒感染的有效抗病毒策略。(C) 2008 Elsevier B.V.版权所有
In this study, the efficacies of short hairpin RNAs (shRNAs) targeting different regions of West Nile virus (WNV) strain Sarafend genome were investigated. Short hairpin RNAs targeting Capsid, NS2B and NS4B genes were cloned into pSilencer (TM) 3.1-H1 neo and designated as pshCapsid, pshNS2B and pshNS4B, respectively. Vero cells that were positively transfected were selected for creating stable cell lines expressing shRNAs constitutively. These cells were subjected to West Nile virus at multiplicity of infection (M.O.I.) of 10. The cells stably transfected with pshCapsid gave the best silencing effect among the three stable cell lines (transfected with pshCapsid, pshNS2B and pshNS4B) at both 12- and 24 h p.i. When compared to the non-transfected WNV-infected cells, pshCapsid stably transfected cells showed more than 4 log(10) unit reduction in viral transcripts and greater than 3 log(10) unit reduction in virus production. Cells stably transfected with pshNS2B did not exhibit as strong an inhibition when compared to the pshCapsid stably transfected cells having only 2 log(10) unit reduction in virus titre. The pshNS4B-stably transfected cells did not suppress WNV replication. Hence, from this study, pshCapsid has the potential to be developed into effective antiviral strategy for WNV infection. (C) 2008 Elsevier B.V. All rights reserved.