间充质干细胞逆转皮质类固醇激素抗炎作用的机制研究

Mesenchymal stem cells （MSCs）are proved to exert therapeutic effects on various inflammatory disorders through their specific chemotaxis toward sites of inflammation and great immunosuppressive capacity. However, the outcomes of MSC therapy in clinical trials are variable, especially when corticosteroids are co-administrated. Previously, we found that concurrent administration of MSCs and Dexamethasone (Dex), a widely used anti-inflammatory corticosteroid drug, could abrogate their both immunosuppressive effects in treatment of mouse liver fibrosis. Dex could abolish immunosuppression of MSCs through interfering STAT1 phosphorylation-prompted expression of inducible nitric oxide synthase (iNOS). But the mechanism by which MSCs reversed the immunosuppressive effect of Dex has not been uncovered. Our preliminary data showed that co-administration of MSCs with Dex could reverse their immunosuppressive effects on T cell proliferation. In addition, a significant increase of VEGF-c production by MSCs was observed. Remarkably, administration of VEGF-R3 inhibitor could regain the immunosuppressive effects of Dex on T cell proliferation. In this project, we aim to further investigate the effects on immune system mediated by the interaction between MSCs and Dex. To define the critical role of VEGF-C/VEGFR-3 pathway in MSC-mediated elimination of immunosuppressive effect of Dex in vivo, we will neutralize VEGF-C and block VEGF-C/VEGFR-3 signal transduction specifically in mouse model. We will examine the effects of MSC-derived VEGF-C on proliferation and anti-apoptosis function associated PI3K/AKT pathway activation in both CD4+ and CD8+ T cells in the presence of Dex, which will help us to obtain an optimizing strategy for MSC-based therapy. This project will provide lots of experimental evidence for identifying the immunoregulatory property of VEGF-C and optimizing the clinical application of MSCs.

间充质干细胞（MSCs）可在炎症部位发挥免疫抑制作用，已用于多种疾病的治疗。临床实验发现MSCs与皮质类固醇激素同时使用，两者疗效大受影响。我们证明MSCs与地塞米松（Dex）同时使用无法控制肝纤维化小鼠体内炎症；Dex可通过抑制iNOS的表达，消除MSCs的免疫抑制能力。然而，MSCs如何逆转Dex的抗炎作用尚不明确。前期实验已发现，MSCs与Dex同时使用无法抑制T细胞增殖，此时，MSCs产生VEGF-C；而VEGF-R3抑制剂的使用，可恢复Dex对于T细胞增殖的抑制能力。本课题将通过阻断VEGF-C/VEGF-R3信号传导，明确MSCs通过VEGF-C在体内逆转Dex抗炎作用；探究Dex作用下，MSCs 通过VEGF-C/VEGF-R3信号通路对于T细胞亚群增殖、抗凋亡信号活化的影响，形成MSCs治疗疾病的优化新策略，为明确VEGF-C免疫调控特性，优化MSCs临床应用提供理论依据。

间充质干细胞（Mesenchymal stem cells, MSCs）是一类极具临床应用潜能的多能干细胞。大量研究证明，MSCs可趋化至损伤局部，发挥免疫调节功能，治疗多种炎症性疾病。前期工作已发现MSCs的免疫调节作用具有可塑性，而以糖皮质类固醇激素（Glucocorticoids, GCs）为代表的免疫抑制剂可消除MSCs的免疫抑制作用，但机制尚不明确，此研究中，我们在肝脏纤维化和GvHD小鼠中同时输注地塞米松（Dexamethasone, Dex）以及MSCs，两者的免疫抑制作用被阻断，同时CD8+T细胞介导的免疫反应加剧，促进GvHD病程。基因芯片分析提示在与活化的CD8+T细胞共培养体系中加入Dex前后MSCs表达血管内皮细胞生长因子C（Vascular Endothelial Growth Factor C, VEGFC）水平显著提高。在混合淋巴细胞反应体系及anti-CD3/28激活的CD8+T细胞增殖体系内，VEGFC能剂量依赖性地提高CD8+T淋巴细胞增殖水平。同时， VEGFC对应的受体VEGFR3的酪氨酸激酶抑制剂MAZ51可特意性阻断这种促进作用。此外，条件性敲除VEGFR3的CD8+T细胞对VEGFC的响应明显减弱。向GvHD小鼠体内注射重组VEGFC蛋白的GvHD小鼠病程显著加速，外周血和多种组织中的炎性浸润增加以及CD8+T细胞上调。综上所述，本研究明确了GCs调节MSCs产生免疫促进作用的分子机制，并证明这种免疫促进效果在不同的疾病中会带来正反两个方面效应，为临床上糖皮质激素及间充质干细胞的合理使用提供理论依据，对特异性提高MSCs的免疫调节功能提供潜在靶点，也为有效打破免疫抑制剂抵抗提高免疫紊乱性疾病治疗疗效提供了新思路。