The Influence Of Pig IRF3 And IRF7 On Expression Of Key Genes In TLR4 Signaling Pathway

Type I Interferon(IFN)can inhibit a variety of viruses,parasites,and bacteria and viruses mixed infection,attracting attention in pig diseasees prevention and control.Interferon regulatory factor(IRF)3 and IRF7 play an important regulatory role on the expression and secretion of type I interferons.both of them can be activated by LPS and in phosphorylation,regulating the expression and secretion of Type I IFN.At present the research about IRF3 and IRF7 focuse on human and mouse,but relatively few study on pigs.In this study we use PB vector system build pig IRF3 and IRF7 eukaryotic expression vector,transfected PK15 cells,screened by puromycin,observed green fluorescence with fluorescence microscope,real time quantitative PCR detected IRF3 and IRF7 gene m RNA expression.screening IRF3 and IRF7 gene stablely transfection cell line;LPS and Licl processing cells,activate and block TLR4 signaling pathway,with real-time quantitative PCR detected the expression of key genes m RNA in TLR4 signaling pathways.Ained at studying the effect of IRF3 and IRF7 to TLR4 signaling pathway,in order to provide preliminary experimental basis for further revealing IRF3 and IRF7 mechanism of action in pig disease prevention and control.Results show that,we can observed green fluorescence of stable transfection of cells under the fluorescence microscope,real time quantitative PCR results show that,the IRF3 and IRF7 m RNA expression level in transfection PB-IRF3 and PB-IRF3 IRF7 cells higher than that in empty vector transfection group,extremely significant difference(P < 0.001),indicating that IRF3 and IRF7 genes overexpression in PK15 cells;My D88,TRAF6,TBK1,NF-?B and IFN-? m RNA expression level in IRF3 and IRF7 transfection group of cells higher than that in empty vector transfection cells,IL-6 m RNA expression level lower than that in empty vector transfection cells.and My D88,TBK1 and IL-6 m RNA expression in IRF3 transfection group cells significant difference(P < 0.05),TRAF6 and IFN? m RNA expression in IRF7 transfection group significant difference(P < 0.05),the rest of the gene expression difference was not significant;after LPS processing,all genes m RNA expression in the empty vector transfection group and IRF3 and IRF7 gene transfection group increased,IFN? and IL-6 m RNA expression in IRF3 transfection group cells significant difference(P < 0.05),NF-?B and IFN? m RNA expression in IRF7 transfection group significant difference(P < 0.05),the rest of the gene expression difference was not significant;Licl inhibited activity of TBK1 activated by LPS,IRF3 and IRF7 expression level downgraded.My D88,TRAF6,TBK1,NF-?B and IFN-? m RNA expression declined in empty vector transfection cells and IRF3 and IRF7 transfection cells,IL-6 m RNA expression level rise.My D88,TBK1,IFN? and IL-6 m RNA expression in IRF3 transfection group cells significant difference(P < 0.05).TRAF6,TBK1,and IFN? m RNA expression in IRF7 transfection group significant difference(P < 0.05).The results showed that pig IRF3 and IRF7 genes can improve m RNA expression level of their upstream molecular TBK1 and downstream gene IFN? in PK15 cells,reduce IL-6 m RNA expression,improve My D88,TRAF6 and NF-?B m RNA expression in the TLR4 mediated signaling pathway.Prompting that IRF3 and IRF7 not only involved in the body's antiviral processs,but also involved in inflammatory reaction process mediated by TLR4 signaling pathway that activated by LPS.