Analysis of intracellular signaling status for local immune tolerance of mucosal T cells

粘膜T细胞的特性是对正常的腔菌群和饮食抗原的耐受性，以维持肠道内环境的稳定。为了阐明局部耐受的机制，我们研究了CD25^+CD4^+调节性T细胞在粘膜T细胞对T细胞受体(TCR)刺激的低反应性中的作用。我们已经发现，与外周血T细胞(PBT)相比，LPT依赖CD3激活的酪氨酸磷酸化模式明显减少。因此，我们从固有层T细胞(LPT)中剔除了调节性T细胞。然而，CD3刺激后LPT的酪氨酸磷酸化模式仍然减少。这一结果提示粘膜T细胞对TCR刺激的低反应性可能是其基本性质。然后，我们研究了被认为是TCR激活后重要的MAPK级联反应。PBT和LPT中均检测到相同数量的MAPK蛋白。刺激CD3后，PBT和LPT的MAPK磷酸化水平均升高。这些结果表明，上游TCR信号分子中酪氨酸磷酸化模式的降低并不影响MAPK级联，而Plcy1或PKC级联可能更重要。酪氨酸磷酸酶在TCR介导的激活中也起重要作用。接下来，我们研究了SHP-1和SHP-2蛋白的表达。SHP-2在PBT和LPT中的表达程度相同。虽然SHP-1在PBT和LPT中均有表达，但SHP-1的两个较小分子仅在LPT中表达。如果LPT中的SHP-1蛋白在功能上也发生了改变，这可能是解释TCR激活后LPT低反应性的重要线索。有必要进行进一步的研究，以解决SHP-1在LPT中的作用。

Natures of mucosal T cells are tolerant to normal luminal flora and dietary antigens to maintain the intestinal homeostasis. To clarify the mechanisms of the local tolerance, we investigated the involvement of CD25^+CD4^+ regulatory T cells in the hyporesponsiveness of mucosal T cells to the stimulation via T cell receptor (TCR). We already revealed that the CD3 activation- dependent pattern of tyrosine phosphorylation of LPT was markedly reduced compared to that of peripheral blood T cells (PBT). Therefore, we eliminated the regulatory T cells from lamina propria T cells (LPT). However, the pattern of tyrosine phosphorylation of LPT after CD3 stimulation remained still reduced. The result suggested that the hyporesponsiveness of mucosal T cells to the stimulation via TCR might belong to its fundamental nature.Then, we investigated the MAPK cascade, which was thought to be important after TCR activation. The comparable amounts of MAPK protein were detected both in PBT and LPT. After CD3 stimulation, phosphorylated MAPK was increased both in PBT and LPT. These results indicated that decreased pattern of tyrosine phosphorylation in the up-stream TCR signaling molecules did not affect the MAPK cascade, and the PLCy1 or PKC cascade might be more important.Tyrosine phosphatases are also important in TCR-mediated activation. Next, we investigated the expression of SHP-1 and SHP-2 proteins. SHP-2 was expressed to the same extent both in PBT and LPT. Although SHP-1 was expressed in both PBT and LPT, two smaller sized molecules of SHP-1 were expressed only in LPT. If the SHP-1 protein in LPT also had been altered in its function, it might be an important clue to explain the hyporesponsiveness of LPT after TCR activation. Further investigation should be necessary to address the role of SHP-1 in LPT.