Study On Traditional Chinese Medicine Compound Polysaccharides With Different MHC B-L?? Genotype Of Chicken Immunomodulatory Effects On TLR4 Signaling Pathway And Immunity

Object:The experimental medicine in this study is a Chinese medicine compound polysaccharide?c CHMPS?,which has been proved to have immune enhancement in the early stage of our laboratory,and its purity is77.1%.The cCHMPS were applied to the peripheral blood lymphocyte of different genotypes of MHC B-L beta II gene.The changes of TLR4 signaling pathway,immune signal molecule and cytokine content in lymphocytes of chicken with different genotypes of MHC B-L?II gene were detected by cCHMPS,to explore the effects of MHC B-L beta II gene polymorphism on cCHMPS regulation of immunity dose,to elucidate the molecular mechanism of traditional Chinese medicine polysaccharide regulating the immune function of chicken,for the research of Chinese medicine polysaccharides in improving disease resistance of chicken and polysaccharide of Chinese herb used in poultry farming to provide scientific and theoretical basis.Method:1.The peripheral blood lymphocytes of different genotypes of MHC B-L?II were separated and added cCHMPS with the final concentration of 100?g/mL,75?g/mL,50?g/mL,0?g/m L?high,middle,low?to support 16h,24h,32h and 48h.,respectively.The expression of mRNA of TLR4,MYD88,TRAF-6,TRIFR3 and IFN-?in lymphocytes was detected by Real-time PCR assay.2.The peripheral blood lymphocytes of different genotypes of MHC B-L?II were separated and added cCHMPS with the final concentration of 100?g/mL?75?g/mL?50?g/mL?0?g/mL to support 24h,the expression of mRNA of IL-1?,IL-2,IL-4,IL-6,IL-8,IL-12,TNF-?,NF-?B in lymphocytes was detected by Real-time PCR assay.3.The peripheral blood lymphocytes of different genotypes of MHC B-L?II were separated and added cCHMPS with the final concentration of 200?g/mL,100?g/mL,75?g/mL,50?g/mL,25?g/mL,0?g/mL to support 24h,the contents of cAMP,Ca²⁺,NO,iNOS and cGMP in lymphocytes were detected by ELISA.Result:1.At each detection point,compared with the control group,different doses of cCHMPS could significantly enhance the expression of mRNA of TLR4,MYD88,TRAF-6,TRIFF,IRF3 and IFN-??P<0.05?in chicken with different genotypes of MHC B-L?II,but the expression of mRNA was different at different time points.The expression of mRNA of MYD88,TRAF-6,TRIF and IRF3 gene in AA genotype chicken was significantly higher than that in high dose group.The expression of mRNA ofTLR4 and IFN-?gene in BB genotype chickenwas significantly higher than that in high dose group?P<0.05?at 48 h.The expression of mRNA of MYD88,TRAF-6,TRIF and IRF3 gene in high dose group was significantly higher than that in other dose groups.The expression of mRNA of TLR4 gene was significantly higher than that of other dose groups at 16 h and 24 h,and the expression of mRNA of IFN-?was significantly higher than that of other dose groups at 24,32,and 48h?P<0.05?.Among BC genotypes,the expression of mRNA of MYD88,TRAF-6,TRIFN and IRF3 gene in low dose group was significantly higher than that in other dose groups,TLR4 and IFN-?was significantly higher than other dose groups?P<0.05?at 24,32and 48 h.2.The expression of mRNA of IL-1??IL-2?IL-4?IL-6?IL-8?IL-12?NF-?B?TNF-?was significantly increased by cCHMPS treatment on the lymphocytes of different genotypes of chicken?P<0.05?.When the dose of cCHMPS was 50?g/mL,the expression of the gene detected in the chicken was significantly higher than that in the other groups?P<0.05?,and when the dose of cCHMPS was 100?g/m L,the expression of the gene detected in the BB genotype chicken was significantly higher than that of other groups?P<0.05?.3.Compared with the control group,the cCHMPS group could significantly increase the content of cAMP?cGMP?Ca2+?NO?iNOS in the supernatant of cell culture medium?P<0.05?.When the dose of cCHMPS was 50?g/mL,the content of c GMP?cAMP?Ca2+?NO?iNOS in the supernatant of lymphocyte culture was significantly higher than that in other groups?P<0.05?,and when the dose of cCHMPS was 100?g/mL,the content of cGMP?cAMP?Ca2+?NO?iNOS in the supernatant of lymphocyte culture was significantly higher than that in other groups?P<0.05?.Comclusion:1.CCHMPS can increase the expression of genes related to TLR4 signaling pathway in chicken lymphocytes,that is,cCHMPS can bind to TLR4 receptor on lymphocyte surface and activate MYD88dependent signal transduction pathway and TRIF dependent signal transduction pathway.The immune status of chicken lymphocytes was optimized and the molecular mechanism of c CHMPS regulating chicken immune function was further elucidated from the point of view of receptor pattern and signal transduction on chicken lymphocytes in vitro.2.CCHMPS can promote the secretion of lymphocyte-associated cytokines and enhance the immunity of the body,and the polymorphism of MHC B-L?II gene has a certain effect on the optimal dose of cCHMPS to promote the secretion of lymphocyte-associated cytokines.3.CCHMPS can promote the secretion of lymphocyte immune signal molecules and enhance the immunity of the body,and the polymorphism of MHC B-L?II gene has a certain effect on the optimal dose of cCHMPS to promote the secretion of lymphocyte immune signal molecules.