摘要
肠黏膜屏障的完整性与猪流行性腹泻病毒(Porcine epidemic diarrhea virus,PEDV)的致病性密切相关。黏蛋白2(Mucin 2,Muc2)作为肠黏膜屏障的重要组成蛋白,在阻止病原体、毒素和异物入侵等方面发挥重要作用。本文旨在探究黏蛋白Muc2对PEDV复制的拮抗作用。通过观察PEDV感染仔猪空肠黏膜屏障情况,利用RNA干扰策略和分子病毒学检测方法在细胞水平上研究分析Muc2表达水平与PEDV复制的关系,并进一步探讨猪源Muc2天然蛋白发挥抗PEDV作用的具体阶段。结果显示:PEDV感染仔猪空肠段进行PAS染色发现,病毒感染后肠绒毛表面黏液层变薄,结构被破坏,杯状细胞明显减少;与对照组相比,干扰Muc2基因能够上调PEDV⁃N的mRNA及蛋白水平;Muc2蛋白添加后显著抑制PEDV⁃N蛋白的表达,且病毒mRNA水平显著下降(P<0.05);在病毒感染前使用50μg/mL Muc2蛋白预处理1 h后,PEDV⁃N mRNA相对表达量极显著下降(P<0.001),而在病毒吸附、入侵和复制阶段无显著差异(P>0.05),表明Muc2具有降低PEDV侵染宿主细胞的作用。综上所述,本研究验证了Muc2在PEDV感染中的拮抗作用,发现了Muc2主要作用于病毒侵染宿主细胞阶段,为深入探讨Muc2与PEDV间的作用机制奠定了基础,也为制定PEDV新防控策略提供了思路。
The integrity of the intestinal mucosal barrier is closely related to the pathogenicity of porcine epidemic diarrhea virus(PEDV).As an important component protein of intestinal mucosal barrier,Mucin 2(Muc2)plays an important role in preventing the invasion of pathogens and toxins.The purpose of this study was to explore the antagonistic effect of Muc2 on PEDV replication.The mucosal barrier of the jejunum of piglets infected with PEDV was observed,and the relationship between the expression of Muc2 and PEDV replication was analyzed at the cellular level using RNA interference strategy and molecular virological detection methods.Finally,the specific stage of the anti⁃PEDV effect of porcine natural Muc2 protein was further explored.The results showed as follows:1.PAS staining of the jejunum segment of piglets infected with PEDV revealed that the mucous layer on the surface of intestinal villi became thinner,and the structure was damaged.Also,the number of goblet cells were significantly decreased after virus infection;2.Compared with control group,interference with Muc2 gene could up⁃regulate PEDV⁃N mRNA and protein levels;3.The expression of PEDV⁃N protein was significantly inhibited and the viral mRNA level was significantly decreased(P<0.05)after Muc2 protein supplementation;4.The PEDV⁃N mRNA expression level was significantly decreased(P<0.001)after pretreatment with 50μg/mL Muc2 protein,and there was no significant difference in virus adsorption,entry and replication phases(P>0.05),suggesting that Muc2 pretreatment of PEDV could reduce PEDV infection in host cells.In conclusion,this study validated the antagonistic role of Muc2 in PEDV infection and revealed that Muc2 plays an antiviral role mainly in the pretreatment stage.This study laid a foundation for further study on the regulation mechanism of Muc2,and also provide new ideas for the prevention strategy of PEDV with a new perspective.
作者
张雪
王传红
钱嘉莉
徐红
宋旭
宝梅英
刘诗雨
胡朝阳
张雪寒
郭容利
贾斌
范宝超
李彬
ZHANG Xue;WANG Chuanhong;QIAN Jiali;XU Hong;SONG Xu;BAO Meiying;LIU Shiyu;HU Zhaoyang;ZHANG Xuehan;GUO Rongli;JIA Bin;FAN Baochao;LI Bin(College of Animal Science and Technology,Shihezi University,Shihezi 832000,China;Institute of Veterinary Medicine,Jiangsu Academy of Agricultural Sciences,Nanjing 210014,China;Jiangsu University,Zhenjiang 212013,China;Nanjing Agricultural University,Nanjing 210095,China;Hebei Agricultural University,Baoding 071000,China)
出处
《病毒学报》
CAS
CSCD
北大核心
2023年第4期1084-1092,共9页
Chinese Journal of Virology
基金
国家自然科学基金(项目号:32272996),题目:PEDV调控基质金属酶13促进病毒吸附的分子机制研究
江苏省种业振兴揭榜挂帅项目(项目号:JBGS〔2021〕024),题目:猪特色种质资源关键基因挖掘与育种新材料创制。
关键词
猪流行性腹泻病毒
MUC2
肠道屏障
Porcine epidemic diarrhea virus
Mucin 2
Intestinal barrier
