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目的探讨TLR2/TLR4介导的信号通路在大肠杆菌麦芽糖结合蛋白诱导Th1活化中的调控作用。方法免疫磁珠分选方法获取纯的CD4+T细胞,经CD3/CD28抗体活化后的CD4+T细胞分别用MBP、MBP+anti-TLR2、MBP+anti-TLR4刺激。ELISA检测培养上清中IFN-γ和IL-4的分泌;RT-PCR检测CD4+T细胞IFN-γ、MyD88、TRIF、TRAF3和TRAF6的mRNA表达;Western blotting方法检测CD4+T细胞MyD88、TRIF、TRAF3和TRAF6的蛋白表达。结果 MBP组IFN-γ水平升高、IFN-γ、MyD88、TRIF、TRAF3的mRNA上调、TRAF6的mRNA下调;MBP+anti-TLR2组IFN-γ水平降低、TRAF6的mRNA上调、MyD88的mRNA下调、TRIF、TRAF3的mRNA无影响;MBP+anti-TLR4组IFN-γ水平升高、TRAF6的mRNA上调、MyD88、TRIF、TRAF3的mRNA下调,Western blotting实验也出现了相似的结果。结论 TLR2介导的MyD88依赖途径和TLR4介导的TRIF/TRAF3依赖途径在MBP诱导的Th1活化过程中发挥了重要的调控作用。
Objective To investigate the role of TLR2 / TLR4-mediated signaling pathway in the induction of Th1 activation by E. coli maltose binding protein. Methods CD4 + T cells were obtained by immunomagnetic bead sorting. CD4 + T cells activated by CD3 / CD28 antibody were stimulated with MBP, MBP + anti-TLR2 and MBP + anti-TLR4 respectively. The secretion of IFN-γ and IL-4 in the culture supernatant was detected by ELISA. The mRNA expression of IFN-γ, MyD88, TRIF, TRAF3 and TRAF6 in CD4 + T cells was detected by RT-PCR. , TRAF3 and TRAF6 protein expression. Results The mRNA level of IFN-γ, MyD88, TRIF and TRAF3 were up-regulated and the mRNA of TRAF6 was down-regulated in MBP group. The level of IFN-γ in MBP + anti-TLR2 group was decreased, TRIF mRNA and TRAF3 mRNA. The level of IFN-γ in MBP + anti-TLR4 group was up-regulated, the mRNA of TRAF6 was up-regulated, the mRNA of MyD88, TRIF and TRAF3 was down-regulated. Conclusion TLR2-mediated MyD88-dependent pathway and TLR4-mediated TRIF / TRAF3-dependent pathway play an important regulatory role in MBP-induced Th1 activation.