Evaluation of a combinatorial RNAi lentivirus vector targeting foot-and-mouth disease virus in vitro and in vivo.
Evaluation of a combinatorial RNAi lentivirus vector targeting foot-and-mouth disease virus in vitro and in vivo.
复制标题
一种针对口蹄疫病毒的组合RNAi慢病毒载体的体外和体内评价
DOI：
10.3892/mmr.2015.4246
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发表时间：
2015-11
影响因子：
3.4
通讯作者：
Shi D
中科院分区：
文献类型：
--
作者：
Zhang X;Zheng H;Xu M;Zhou Y;Li X;Yang F;Liu Q;Shi D
口蹄疫病毒（Foot-and-mouth disease virus，FMDV）是一种引起偶蹄类动物高度接触性传染病，给人类造成严重的经济损失。FMDV不能通过疫苗接种或生物安全措施得到充分控制。为了产生经遗传修饰的FMDV抗性动物，使用慢病毒（LV）载体LV-3shRNA构建产生三个短发夹（sh）RNA的组合表达盒。这三个shRNA分别在来自布法罗和牛的DNA聚合酶III启动子的调控下表达，其中一个靶向FMDV的非结构蛋白3B，另外两个靶向FMDV的病毒聚合酶蛋白3D。在BHK-21细胞和乳鼠中测定LV-3shRNA在抑制FMDV复制中的作用。结果显示，当用107倍于组织培养感染剂量（TCID 50）/ml的O血清型FMDV攻击细胞时，LV-3shRNA使病毒生长减少3倍（感染后24小时）。用LV-3shRNA预处理的乳鼠在施用5倍于FMDV剂量的FMDV时完全受到保护，否则足以杀死50%的实验动物（LD 50）。这些结果表明，LV介导的三个FMDV特异性shRNA的双重表达提供了一种对抗FMDV的新策略，这有助于将来将新的抗病性状永久引入到布法罗和牛基因组中。
Foot-and-mouth disease virus (FMDV) causes a highly contagious disease of cloven-hoofed animals, which leads to serious economical losses. FMDV is not adequately controlled by vaccination or biosecurity measures. To generate genetically modified FMDV-resistant animals, a combinatorial expression cassette producing three short hairpin (sh) RNAs was constructed using the lentivirus (LV) vector, LV-3shRNA. The three shRNAs were expressed under the regulation of DNA polymerase III promoters from a buffalo and a bovine source, with one targeted to the non-structural protein 3B, and the other two targeted to the viral polymerase protein 3D of FMDV, respectively. The role of LV-3shRNA in the inhibition of the replication of FMDV was determined in BHK-21 cells and in suckling mice. The results revealed that LV-3shRNA reduced viral growth 3-fold (24 h post-infection) when the cells were challenged with 107-times the tissue culture infective dose (TCID50)/ml of O serotype FMDV. The suckling mice pretreated with LV-3shRNA were completely protected on administration of 5-times the dose of FMDV otherwise sufficient to kill 50% of the experimental animals (LD50). These results demonstrated that the LV-mediated dual expression of three FMDV-specific shRNAs provided a novel strategy towards combating FMDV, which facilitates the permanent introduction of novel disease-resistance traits into the buffalo and bovine genomes in the future.