PD-1/PD-L1信号通路调控Treg/Th17平衡在子痫前期发病机制中的研究

Our previous study found that the percentage of regulatory T (Treg) cells in peripheral blood of patients with preeclampsia (PE) was obviously decreased, which was related to the increase of Treg cell apoptosis. However, whether the quantitative deficeincy of Treg cells is associated with the decrease of Treg cell generation and what the underlying mechanism is remain unclear. Treg cells and Th17 cells are conditioned by each other. But they have the same origin. Both are differenciated from naive CD4+ T cells. Treg/Th17 banlance plays a key role in maternal-fetal immune tolerance, which is regulated by the negative signal of programmed death factor-1 (PD-1) and its ligand (PD-L1). From the above, we speculate that the quantitative decrease of Treg cells might be associated with the imbanlance of Treg/Th17 cell differenciation, which might be regulated by PD-1/PD-L1 pathway. In the present study, by using in-vivo, in-vitro study and animal model, the cause of the decrease of Treg cell generation, its molecular mechanism and the treatment efficacy for PE animal model by targeting PD-1/PD-L1 pathway were evaluated. In view of PD-1/PD-L1 regulating Treg/Th17 banlance, we aim to disclose the molecular immune mechanism of PE and to find the potential treatment target, which will provide the new concept for making treatment pathway for PE.

课题组前期研究发现子痫前期(PE)患者外周调节性T细胞(Treg)数量明显降低，与其凋亡增加有关。然而，其中是否伴有Treg产生减少、调节机制如何尚不明确。Treg和Th17细胞是一对制衡细胞，但均由初始CD4+T细胞分化而来。Treg/Th17细胞平衡是母胎免疫耐受的关键，此平衡受程序性死亡分子1(PD-1)及其配体(PD-L1)负性信号调控。我们推测：PE中Treg数量减少可能与Treg/Th17分化失衡有关，且此过程受PD-1/PD-L1信号调控。因此，本项目立足于课题组现有的研究基础，通过在体、体外研究和动物模型三个方面，从PD-1/PD-L1信号通路调控Treg/Th17平衡的视角，延续性探讨PE中Treg数量减少的原因、分子机制以及以PD-1/PD-L1信号通路为靶点治疗PE动物模型的效果。以期解析PE的分子免疫学发病机制和发现潜在的治疗靶点，为制定该疾病的治疗路径提供新思路。

本项目探讨了PD-1/PD-L1通路调控Treg/Th17细胞免疫平衡在PE中的作用、机制及干预效果。首先，分析了正常晚孕妇女和子痫前期（PE）患者外周血和胎盘局部Treg和Th17细胞百分比及 PD-1、PD-L1的表达；结果表明：PE患者外周和胎盘局部均存在Treg/Th17平衡向Th17偏移，且PD-1和PD-L1在PE中的表达存在明显差异。揭示了 PD-1/PD-L1 信号通路可能影响 PE患者外周和母胎界面局部Treg/Th17 细胞平衡的现象。然后，通过体外实验从细胞增殖、分化和转化等方面探讨PE中PD-1/PD-L1信号通路对Treg/Th17失衡的调节机制；研究发现：(i) PD-1/PD-L1通路减弱能促进Th17细胞增殖，抑制naïve CD4+T细胞向Treg细胞分化，促进Th17细胞产生，并伴有PI3K/AKT/m-TOR mRNA表达降低，PTEN mRNA表达增加；(ii) IL-17+Treg细胞数量在PE中明显增加，并与Th17细胞数量呈正相关；(iii) 阻断PD-1/PD-L1通路下调Treg细胞特异性核转录因子Foxp3和IL-10的表达，促进Th17细胞特异性核转录因子RORγt和IL-17的表达。结果表明：PE中存在PD-1/PD-L1信号通路减弱，且通过调节T细胞增殖、分化和转化，导致Treg/Th17免疫失衡，表现为促进Th17细胞增殖，抑制Treg细胞分化和促进Treg细胞向Th17细胞转化。最后，通过建立PE样大鼠模型探讨PD-L1：Fc 活化 PD-1/PD-L1 信号通路对PE样大鼠的治疗效应和分子机制。研究发现：采用PD-L1:Fc活化PD-1/PD-L信号通路，不仅可逆转PE样大鼠外周及胎盘局部的Treg/Th17细胞失衡，且对PE样大鼠及其子代均有一定的保护作用。与传统治疗 PE 的药物硫酸镁相比，PD-L1:Fc 降低PE样大鼠血压的效果持续时间更长。进一步的分子机制研究表明 PD-L1:Fc 对PE样大鼠的治疗效应与抑制PI3K/AKT/m-TOR信号通路和促进PTEN分子表达从而扭转Treg/Th17细胞失衡有关。本项目进一步解析了PE的分子免疫学发病机制、发现了潜在的治疗靶点，为开创以PD-1/PD-L1信号通路为靶点探索在PE中的治疗策略提供了新思路和理论依据。