激活Galectin-7/MMP-9途径协同诱导移植免疫耐受

Our research group found that galectin-7 protein (gal-7) was mainly located in the membrane of the graft alloreactive T cells.There were indications that the protein can reduce the activity of T cells through combination with the CD3 molecule.In addition, we found that gal-7 can upregulate the expression of matrix metalloproteinase-9 (MMP-9) of lymphocytes.It has been reported that MMP-9 can cause T cells inactivation by hydrolyzing α chain of IL-2R..Therefore, we speculated that gal-7/MMP-9 pathway may synergistically specifically induce anergy of alloreactive T cells..To verify the hypothesis,our group plan to transfect grafts with rAAV-gal-7 and induce grafts to express stably gal-7 for long term, so as to activate gal-7/MMP-9 pathway, then to chronically reduce the activity of alloreactive T cells and achieve the purpose of immune tolerance in allograft recipients,in which gal-7/MMP-9 pathway is taken as the target.Moreover,we explore the role of the differentiation of Th1,regulatory T cells, the mechanism of apoptosis and cell cycle regulation in this program in order to clarify its mechanism..This project proposed a new solution on how to induce immune tolerance.Completion of the study would be the supplement of the theory of clearance of peripheral T cells and may provide an new idea for exploring the strategies of donor-specific immune suppression.

本课题组研究发现，galectin-7蛋白（gal-7）主要定位于移植物同种反应性T细胞的胞膜；有迹象表明，该蛋白可通过结合CD3分子而降低T细胞活性。此外，我们发现gal-7可上调淋巴细胞MMP-9的表达；已有研究证明MMP-9可水解IL-2R的α链而导致T细胞失活。因此，我们推测，gal-7/MMP-9途径协同作用，可以特异性的诱导同种反应性T细胞失能。为验证假设，本课题以gal-7/MMP-9途径为干预靶点，用rAAV-gal-7转染移植物，使其长期稳定表达gal-7，激活gal-7/MMP-9途径，持续性特异性抑制同种反应性T细胞，以期达到诱导免疫耐受的目的；并探讨Th1的分化、调节性T细胞、凋亡机制及细胞周期调节等在本方案中的作用，以阐明其作用机制。本课题提出了一种全新的免疫耐受诱导方案，该项目的完成将对外周T细胞清除理论进行补充，并可能为探索供者特异性免疫抑制策略提供新的思路。

通过构建小鼠心脏移植模型，我们发现Galectin-7分子在急性排斥反应及T细胞应答和增殖中具有重要作用；接着进一步研究了Galectin-7对T淋巴细胞增殖和Th1/Th2平衡的影响。在接下来的研究探讨Galectin-7蛋白促进活化的T淋巴细胞增殖并诱导其向Th1细胞优势应答偏移的作用中，我们发现Galectin-7分子通过对TGFβ/Smad3通路的抑制，进而促进活化的T淋巴细胞增殖并诱导其向Th1细胞优势应答偏移。通过噬菌体展示筛选实验、DNA抽提、测序及Blast软件对比，找到了与Galectin-7匹配的3个短肽序列，用这三个短肽的碱基序列在小鼠GeneBank中对比，找到了与Galectin-7匹配的8个蛋白，并验证是否与Galectin-7蛋白相互作用。.本项目利用Galectin-7干预达到了预期目的，初步证实了Galectin-7蛋白通过对TGFβ/Smad3通路的抑制，进而促进活化的T淋巴细胞增殖并诱导其向Th1细胞优势应答偏移。由于筛选出的蛋白相关研究很少，只是通过ELISA初步验证是否与Galectin-7结合，需要进一步验证。