大肠杆菌麦芽糖结合蛋白激活TLR2介导的MyD88依赖途径和TLR4介导的TRIF/TRAF3依赖途径诱导Th1活化

Maltose binding protein induces Th1 activation by activating MyD88-dependent pathway of TLR2 and TRIF/TRAF3-dependent pathway of TLR4

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。

To study the regulatory effect of single pathway of TLR2/TLR4 on the activation of Thl induced by maltose binding protein （MBP）, CD4＋T cells pretreated with anti-CD3 and anti-CD28 antibody were stimulated with MBP, MBP ＋ anti-TLR2 and MBP ＋ anti-TLR4. ELISA was used to detect the production of IFN-γ and IL-4 in supernatant of CD4＋T cells from different groups. The mRNA levels of IFN-γ, MyD88, TRIF, TRAF3 and TRAF6 in CD4＋T cells from different groups were examined with RT-PCR, while the protein levels of MyD88, TRIF, TRAF6 and TRAF3 were detected by Western blotting. Data showed that MBP elevated the level of IFN-γ, production in supernatant of CD4＋T cells and enhanced the mRNA level of IFN-γ. MBP increased the mRNA levels of MyD88, TRIF and TRAF3 expressed in CD4＋T cells, but decreased the mRNA level of TRAF6. The production of IFN-γ decreased in CD4＋T cells treated with MBP and anti-TLR2 antibody; the mRNA level of MyD88 decreased but TRAF6 increased in CD4＋T cells treated with MBP and anti-TLR2 antibody. The mRNA levels of TRIF and TRAF3 in CD4＋T cells treated with anti-TLR2 antibody had been less affected by the MBP. In addition, anti-TLR4 antibody decreased the mRNA levels of MyD88, TRIF and TRAF3 but increased the mRNA levels of TRAF6. The results of MyD88, TRIF, TRAF6 and TRAF3 detected by Western blotting were similar with the mRNA results. In conclusion,MyD88-dependent pathway of TLR2 and TRIF/ TRAF3-dependent pathway of TLR4 are involved in the mechanism of Thl activation induced by MBP.