血小板调控CD4+CD25+Foxp3+T细胞分化和功能在肝移植排斥反应中的作用

Recently accumulative evidents suggest that platelet, upon activation, have been demonstrated to inhibite the CD4+CD25+Foxp3+T lymphocyte function depending on PF4,meanwhile CD4+CD25+Foxp3+T lymphocyte play an important role in inducing the transplant immune tolerance. Piror work from our experiment has found the reinforce of platelet activation during the rejection reaction after liver transplantation,which were accompany with the increase of PF4 levels and decreases of CD4+CD25+Foxp3+T lymphocyte proportion. Based on these findings, we supposed that activated platelet affected the rejection reaction after liver transplantation by which PF4 inhibited the CD4+CD25+Foxp3+T lymphocyte differentiation and function.For realize the hypothesis we designed this study, liver transplantation recipents were rolled in and a various of experimental methods including FCM,Immunofluorescence,CFSE,MACS were choosed in our study. Fist, we explored the status of activated platelet in peripheral blood and liver tissue.Second,the peripheral blood platelet were isolated and the regulation effect on CD4+CD25+Foxp3+T lymphocyte differentiation and function were analysed. Finally, we study how the immunosuppressive agents and antiplatelet activity drugs affect the platelet activation and immune function. we expect to find the new mechanim of rejection reaction in liver transplantation and novel immune regulated function of platelet in transplantation immunity, at the same time provid the proofs of new mechanism of action of immunosuppressive agents.

近年研究表明血小板活化后可通过PF4抑制CD4+CD25+Foxp3+T细胞功能，而CD4+CD25+Foxp3+T细胞在诱导移植免疫耐受中发挥重要作用。我们前期研究发现肝移植术后排斥反应时血小板活化增强,且伴随PF4增加和CD4+CD25+Foxp3+T细胞比例降低。因此，推测血小板活化后可通过PF4抑制CD4+CD25+Foxp3+T细胞分化和功能从而影响肝移植排斥反应的进程。本课题以肝移植受者为研究对象，利用流式细胞术、免疫荧光技术、CFSE稀释实验、磁性细胞分选等方法，首先研究排斥反应时外周血和肝组织血小板活化情况；然后分离外周血小板，研究血小板对CD4+CD25+Foxp3+T细胞分化和功能的调控作用；最后研究免疫抑制剂和抗血小板活性药物对血小板活化和免疫功能的影响。通过上述研究以期发现肝移植排斥反应新机制和血小板在移植免疫中新的免疫调控功能，同时提供免疫抑制剂新作用机理的证据。

众多证据表明血小板活化后可通过PF4抑制CD4+CD25+Foxp3+T细胞功能，从而促进肝移植术后排斥反应的发生。本课题主要探索肝移植患者术后血清PF4浓度、CD4+CD25+Foxp3+T细胞功能、TH1/TH2/TH17-T 细胞相关因子表达量以及术后免疫抑制剂维持浓度等与排斥反应密切相关因素之间的相关性与相互作用。研究结果证实了血小板活化参与了肝移植术后排斥反应的发生，并可通过PF4抑制CD4+CD25+Foxp3+T细胞分化和功能从而影响肝移植排斥反应进程。此外，通过回顾性分析，验证了肝移植术后早期降低他克莫司使用剂量不仅可降低术后并发症，还有助于移植物功能的长期维持。再者，首次探索了肝移植术后远期他克莫司维持浓度降至3ng/ml以下的安全性与有效性，发现了CD8+CD28- T细胞比例可作为监测肝移植术他克莫司浓度减量的生物学指标。通过本课题的研究，证实了血小板在肝移植排斥反应中具有重要的免疫调控功能，同时提供了免疫抑制剂最小化的新依据。