NRF2激动剂衣康酸4-辛酯对猪伪狂犬病病毒的抑制作用研究  

NRF2 agonist 4-octyl itaconate inhibits the proliferation of porcine pseudorabies virus

作　　者：方春艳 杨发誉 朱紫祥[2] 郑海学[2] 包世俊[1] 魏衍全 张石磊 FANG Chunyan;YANG Fayu;ZHU Zixiang;ZHENG Haixue;BAO Shijun;WEI Yanquan;ZHANG Shilei(College of Veterinary Medicine,Gansu A gricultural University,Lanzhou 730070,China;State Key Laboratory for Animal Disease Control and Prevention/College of Veterinary Medicine of Lanzhou University/Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Lanzhou 730000,China;Gansu Province Research Center for Basic Disciplines of Pathogen Biology,Lanzhou730046,China)

机构地区：[1]甘肃农业大学动物医学院,甘肃兰州730070 [2]中国农业科学院兰州兽医研究所兰州大学动物医学与生物安全学院动物疫病防控全国重点实验室,甘肃兰州730070 [3]甘肃省病原生物学基础学科研究中心,甘肃兰州730046

出　　处：《中国兽医科学》2024年第11期1421-1429,共9页Chinese Veterinary Science

基　　金：甘肃省科技重大专项(22ZD6NA001);甘肃省基础研究创新群体项目(23JRRA561)。

摘　　要：为筛选高效抑制猪伪狂犬病病毒(pseudorabies virus,PRV)的小分子抑制剂,首先利用流式细胞分析技术结合小分子药物库筛选出对PRV具有高效抑制活性的小分子药物,并利用CCK-8法检测其对PK15细胞活力的影响。其次,利用Western-blot检测PRV g B蛋白、TCID_(50)测定PRV滴度等方法检测小分子药物对PRV的抑制作用。最后,利用Western-blot检测小分子药物抑制PRV生命周期的具体阶段。结果显示,流式细胞术分析筛选出的对PRV抑制最明显的3种小分子药物中,Cyanosafracin B、Staurosporine对PK15细胞有很强的细胞毒性,而衣康酸4-辛酯(4-OI)在300μmol/L的浓度下对PK15细胞基本无毒性。4-OI对不同MOI的PRV均表现出较强的抑制作用,表现为PRV子代病毒滴度的降低、PRV g B蛋白的表达水平下降,并呈剂量依赖性。4-OI及其靶蛋白Nuclear factor erythroid 2-related factor 2(NRF2)能够抑制PRV在细胞内的复制,而不影响PRV吸附PK15细胞及其内化过程。本研究表明4-OI是一种潜在的高效抑制PRV的小分子药物,为PRV防控提供了新的参考依据。College of Veterinary Medicine,Gansu Agricultural University;State Key Laboratory for Animal Disease Control and Prevention/College of Veterinary Medicine of Lanzhou University/Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences;Gansu Province Research Center for Basic Disciplines of Pathogen Biology;To identify small molecules that can effectively inhibit porcine pseudorabies virus(PRV),we performed a high-throughput screening of a small molecule library by flow cytometry analysis technology and evaluated the effect of identified small molecules on PK15 cell viability by CCK-8 assay.The inhibitory effects of small molecules on PRV were investigated by detecting PRV g B protein expression and determining viral titer by TCID_(50).The specific stages of the PRV life cycle that are hindered by small molecules were performed by Western-blot assays.The results showed that the three small molecules with the most obvious inhibitory effects on PRV were identified by flow cytometry analysis.Cyanosafracin B and Staurosporine showed high cytotoxicity,while 4-Octl itaconate(4-OI)is non-toxic even at a concentration of 300μmol/L.4-OI showed strong antiviral activity upon PRV infection of different MOIs,as evidenced by a dose-dependent decrease in PRV progeny virus titer and PRV g B protein expression.4-OI and its target protein NRF2 can inhibit the replication of PRV in cells without affecting the adsorption of PRV to PK15 cells and its internalization process.This study shows that 4-OI is a potentially efficient small-molecule drug that inhibits PRV,providing a new reference for PRV prevention and control.

关 键 词：药物筛选 衣康酸4-辛酯(4-OI) 猪伪狂犬病病毒(PRV) 抗病毒作用 

分 类 号：S852.659.1[农业科学—基础兽医学]