VitD调节SLE滤泡辅助性T细胞的功能及机制研究

Systemic lupus erythematosus (SLE) is a systemic autoimmune disorder common articipation of T cells, B cells and so on. Abnormal activation of B cells causes increasing antibody-producing cells populations, high gamma globulin hematic disease, autoantibody production and immune complex formation. 1, 25-Dihydroxyvitamin D3 [VitD] supplementation can induce a decrease of B cell numbers and part of autoantibodies, improve some symptoms of SLE. B cell activation needs the auxiliary of Tfh cells, and the precise mechanisms of VitD on Tfh cell is not clear. VitD deficiency is a prominent feature of patients with SLE. However, only high level VitD has obvious effect on B cells. Therefore, it is reasonable to speculate that VitD may inhibit Tfh cells which subquently can reduce excessive B cell activation. In this study, with cultured Tfh cells in vitro and SLE animal model of Peyer's Patches in vivo, we investigate the effects of VitD on the Tfh cells function and possible mechanism to activate nuclear factor of activated T cells using flow cytometry, VDR knockout, VDR induced and siRNA interference technology, and discuss the influence of VitD on B cells activation and function, as well as providing the experimental and clinical data for VitD to treat autoimmune disease.

系统性红斑狼疮(SLE)是T细胞、B细胞等共同参与的全身性自身免疫疾病，B细胞异常活化导致抗体产生细胞数目增多、高丙种球蛋白血症、自身抗体形成。1,25-二羟维生素D3[VitD]有免疫调节功能，补充VitD可诱导B细胞数目及部分自身抗体减少，部分改善SLE的症状。B细胞的活化需要滤泡辅助性T细胞（Tfh）的辅助，目前VitD对Tfh的作用尚不清楚。SLE患者中VitD普遍较低，而其仅在较高水平才对B细胞有明显作用，由此推测VitD可能通过抑制Tfh从而减少B细胞的活化。本研究从离体和在体两方面，应用离体Tfh培养及在体SLE动物模型Peyer小结，采用流式细胞仪、VDR基因敲除、VDR诱导和siRNA干扰等技术，观察VitD对Tfh及其活化T细胞核因子的作用，对B细胞活化及产生自身抗体的影响，探索Tfh的体外培养条件，为临床使用VitD治疗自身免疫性疾病提供理论和实验依据。

系统性红斑狼疮是经典而常见的自身免疫性疾病，Tfh细胞在其发病过程中起重要作用，VitD有免疫调节作用。本课题观察VitD调节SLE滤泡辅助性T细胞的功能及机制，通过Tfh细胞的培养分选、流式细胞仪、VDR慢病毒质粒和siRNA干扰、RT-PCR、Western blot等技术，探讨VitD对系统性红斑狼疮Tfh的作用及机制。结果显示：1.VitD可降低外周血炎症及免疫指标，上调VDR、Bcl-6、Pax-5的表达，降低Peyer小结Tfh细胞百分率及细胞凋亡水平，下调IL-21、IL-4、Blimp-1、XBP-1、BAFF的表达，抑制B细胞的发育及浆细胞生成，且呈剂量依赖性，VitD与VDR激动剂联用较单用VitD效应更加明显。2.VitD可下调Tfh细胞IL-21的表达，抑制Blimp-1、XBP-1、BAFF的表达，降低自身免疫抗体的生成，且呈剂量依赖性。3.VitD通过抑制Tfh细胞内Ca-CaN-NFAT通路抑制Tfh的功能，VitD受体激动剂或干扰质粒可增加或拮抗上述变化。4.VitD抑制狼疮组Tfh细胞的表达，给予慢病毒质粒下调Tfh细胞VDR的表达，则VitD抑制Tfh表达明显减弱。结果提示：VitD与其受体结合，通过抑制Tfh细胞内的Ca-CaN-NFAT通路，抑制Tfh细胞及相关因子及微环境，抑制自身免疫抗体的生成，从而对SLE有治疗作用。