PD-L1协同人胎盘间充质干细胞诱导T细胞免疫耐受减轻GVHD机制研究

The major obstacle to allogeneic hematopoietic stem cell transplantation is the occurrence of acute graft versus host disease (GVHD) induced by activated T cells from graft. To induce T cell immune tolerance is significance for overcoming GVHD. We have found that PD-L1 expressed in human placenta derived mesenchymal stem cells (HPMSCs) played a key role in the immunosuppressive effects of HPMSCs on T cells. In this research we will focus on, in vitro and in vivo, whether PD-L1 expressed in HPMSCs will promote the inhibitory effects of HPMSCs on the response of T cells, which were from GVHD patients and healthy donors, by changing the property of T cell TCRCDR3 and regulating the differentiation of regulatory T cells and the secretion of some soluble cytokins, et al. The signal regulatory mechanism of IFN-γ, TNF-α and IL-1β up regulating the expression of PD-L1 in HPMSCs and promoting inhibitory effects of HPMSCs on T cells will also be explored in this study. Previously, we have found that PD-L1 could adjust the capacity of HPMSCs along SDF-1α/CXCR4 axial migration. Here, the possible mechanism of PD-L1 regulating the distribution of HPMSCs in the GVHD model mice as well as the preventing effects of HPMSCs on GVHD will be investigated. All the results in this research will demonstrate the role, specially related to PD-1/PD-L1 signal pathway, of HPMSCs in controlling GVHD and reveal the molecular and cellular mechanism of HPMSCs mediating T cell immune tolerance, which will enhance the process of HPMSCs in clinical application.

移植物中T细胞活化增殖诱发的移植物抗宿主病（GVHD）是异基因造血干细胞移植成功的主要障碍。诱导T细胞耐受是克服GVHD发生的关键。项目拟在发现人胎盘间充质干细胞（HPMSCs）通过其表达的PD-L1分子增强对T细胞抑制作用的基础上，通过体内外实验证明PD-L1在HPMSCs上表达可通过改变GVHD患者T细胞TCRCDR3特性，调节Treg功能及细胞因子分泌等方式协同HPMSCs抑制T细胞应答。分析IFN-γ、TNF-α及IL-1β调节PD-L1在HPMSCs上表达的信号通路及其对HPMSCs抑制T细胞增殖的调节作用。并在已发现PD-L1可调节HPMSCs沿SDF-1α/CXCR4轴向迁移的基础上，证明PD-L1可调节HPMSCs在GVHD模型体内的分布及其对GVHD的防治效果。结果将揭示PD-L1协同HPMSCs诱导T细胞耐受的细胞与分子机制，推进HPMSCs治疗GVHD的临床应用进程。

移植物抗宿主病（graft-versus-host disease, GVHD）的产生是造血干细胞移植成功的主要障碍，目前尚无理想的解决方案。T细胞活化增殖是引起GVHD的主要原因，间充质干细胞(mesenchyma stem cells, MSCs)可通过抑制T细胞的增殖减弱GVHD，但其具体机制不明。项目以共刺激分子程序性死亡蛋白配体1/2(programed death ligand 1/2, PDL1/2)为切入点通过体内外实验研究了人胎盘间充质干细胞(human placenta-derived mesenchymal stromal cells, hPMSCs)对T细胞亚群重塑的作用机制及其对GVHD模型鼠、实验性自身免疫性脑脊髓炎(experimental autoimmune encephalomyelitis, EAE)模型鼠和卵巢早衰（premature ovarian failure, POF）模型鼠的治疗效果。结果发现（1）PDL1/2可增强hPMSCs对T细胞向IL-10+T细胞亚群转化形成的能力，促进hPMSCs对Th17、Th1和Th2亚群的平衡作用；（2）hPMSCs可促进CD4+CXCR5+Foxp3+调节性T细胞亚群的形成；（3）IFN-γ、TNF-α、IL-1β及IL-27均可调节PDL1/2在hPMSCs上的表达，IFN-γ可通过JAK/STAT/IRF-1通路上调PDL1/2的表达；发现hPMSCs上表达IL-27Rα，IL-27通过JAK/STAT1通路上调PDL1在hPMSCs上的表达；（4）IFN-γ、TNF-α、IL-1β不仅能够调节hPMSCs迁移、黏附和增殖，而且能促进hPMSCs对IL-10+T细胞亚群的诱导作用，IL-27增强hPMSCs对活化的PBMC向Th1、Th2亚群转化的调节作用，促进IL-10+T细胞亚群的形成。（5）体内实验结果显示hPMSCs可通过诱导IL-10+T细胞的形成减弱GVHD、缓解EAE症状；通过诱导Foxp3+Treg亚群的形成明显减弱POF小鼠炎症反应。. 研究结果初步揭示了hPMSCs与活化T细胞之间的双向调节作用特点，为进一步研究hPMSCs免疫调节机理及hPMSCs在临床细胞治疗中的应用提供理论支持。