猪TLR5基因表达水平与F18大肠杆菌侵染关系分析

摘要

In order to explore the relationship between Toll-like receptor 5(TLR5)gene expres-sion and resistance to E.coli F18,the bacterial stimulation experiments were conducted in this study.Intestinal epithelial cells(IPEC-J2)were induced by lipopolysaccharide(LPS)and E.coli F18 with different serotypes(F18ab and F18ac),the mRNA expression level of TLR5 gene was detected by Real-time PCR,and the protein expression was detected by Western blotting.The re-sults showed that the expression level of TLR5 gene was extremely significantly up-regulated (P<0.01)after IPEC-J2 cells were infected with different serotypes of E.coli(F18ab and F18ac),which was also extremely significantly up-regulated after IPEC-J2 cells were induced by LPS for 4 and 8 h(P<0.01).After IPEC-J2 cells were induced by LPS for 8 h,the expression level of TLR5 gene increased more significantly than induction for 4 h.After LPS and bacterial stimulation,the expression level of TLR5 protein in cells was extremely significantly up-regulated (P<0.01),which was consistent with the results of mRNA expression.This study further con-firmed that the expression level of porcine T L R5 gene played an important role in the process of cell resistance to E.coli F18 infection by studying the relationship between the expression level of T L R5 gene and E.coli.It also laid the foundation for future research on T L R5 gene function and its application in genetic breeding of E.coli diarrhea.%为了探究猪Toll样受体(Toll-like receptor 5,TLR5)基因表达水平与F18大肠杆菌抗性的关系,试验通过不同血清型产肠毒素大肠杆菌(F18ab和F18ac)侵染猪小肠上皮细胞(IPEC-J2),同时通过脂多糖(LPS)分别诱导IPEC-J2细胞4和8 h,利用实时荧光定量PCR检测 TLR5基因表达水平变化,并利用Western blotting进行蛋白表达分析.结果显示,不同血清型大肠杆菌(F18ab和F18ac)菌体侵染IPEC-J2细胞后,TLR5基因表达水平均极显著上调(P<0.01);LPS诱导IPEC-J2细胞4和8 h后,TLR5基因表达水平均极显著上调(P<0.01),且在LPS诱导IPEC-J2细胞8 h后,TLR5基因表达水平明显高于诱导4 h.与对照组相比,细胞中 TLR5蛋白的表达水平极显著上调(P<0.01),与LPS诱导及F18大肠杆菌菌体刺激IPEC-J2细胞后mRNA表达水平结果相一致.本研究在细胞水平上分析了TLR5表达水平和F18大肠杆菌侵染的相关性,进一步证实猪 TLR5基因的表达水平在细胞抵抗F18大肠杆菌的侵染过程中发挥了重要的调控作用,为今后关于 TLR5基因功能及其在大肠杆菌腹泻遗传育种应用的研究奠定基础.