基于T细胞分化可塑性研究温阳散寒除湿法治疗类风湿关节炎的作用机理

Recently, the balance of regulatory T cell(Treg) and type 17 helper 17 cell (Th17) was broadly reported to be critical to maintain the immunotolerance of the immune system, while the brokendown of this balance will resulted in the development of autoimmune disease. Treg cells has potent immunosuppressive function. However, recent reports showed that Treg cels could not suppress everything, and even that Treg cells could be polarized into pro-inflammatory Th17 cells in the autoimmune disease. Thus,the reciprocal suppression and transition between Treg and Th17 cells really make the pathogenesis of autoimmune disease complicated, and also represent the typical Ying and Yang theory in the traditional chinese medicine. Previous studies have shown that WenYangSanHanChushi method has a dramatic clinical efficacy in treating rheumatoid arthritis (RA),significantly inhibited collagen (CII)-induced arthritis (CIA) in mice and promoted Treg cells immune suppressive cytokine IL-10 production. In this study, we will focus on the the Developmental Plasticity of Th17 and Treg cells to elucidate the mechanisms of WenYangSanHanChuShi method to treat RA.Briefly,through investigating the amplification of T cell, cytokine secretion, the expression of specific transcription factor in the animal model of collagen-induced arthriti in DBA/1 mice,in vitro differentiation of Th17 and Treg cells and the Treg cells converting into Th17 cells，in depth to study the WenYangSanHanChushi method regulates Th17/Treg cell balance mediated mechanism for treatment of RA. From this pointcut, we resolve the scientific connotation of "WenYangSanHanChushi method treats Bi Syndrome (HanShiBiZu Syndrome)",lay a theoretical foundation to improve the clinical efficacy of the treatment with RA, drug research and QianYaoZuFang.

最新研究表明，介导免疫耐受的调节性T细胞（Treg）与介导炎症反应的Th17细胞都来源于初始T细胞，两者的功能和分化过程相互对抗、转化，类似于中医阴阳对立统一关系，其平衡失调影响着类风湿关节炎（RA）的发生与转归。 前期研究表明，温阳散寒湿除法对RA具有较好临床疗效，显著抑制胶原诱导性小鼠关节炎，并能促进Treg免疫细胞分泌免疫抑制性细胞因子IL-10。基于此，本课题拟以Th17细胞/ Treg分化可塑性为切入点，利用DBA/1小鼠胶原关节炎、iTreg/Th17体外诱导分化和转化细胞模型，从T细胞的扩增、细胞因子的分泌、特异转录因子的表达、分化和相关转化等层面，深入研究温阳散寒除湿法调控Th17/Treg细胞平衡而介导治疗类风湿关节炎的作用机理，并以此为切入点解析"温阳散湿除湿法治疗痹证（寒湿痹阻）"科学内涵，为临床遣药组方、提高疗效和新药研究奠定理论基础。

研究温阳散寒除湿法（蠲痹颗粒）调控致病性Th17细胞/ 免疫抑制功能Treg之间分化可朔性而介导治疗类风湿关节炎的作用机理。以Th17细胞/ Treg分化可塑性为切入点，利用DBA/1小鼠胶原关节炎、iTreg/Th17体外诱导分化和转化细胞模型，从T细胞的扩增、细胞因子的分泌、特异转录因子的表达、分化和相关转化等层面，深入研究温阳散寒除湿法调控Th17/Treg细胞平衡而介导治疗类风湿关节炎的作用机理。结果显示，蠲痹颗粒能显著降低CIA小鼠胶原关节炎的临床评分，明显改善CIA小鼠局部关节炎的症状（P＜0.05），减轻炎症细胞浸润，抑制CIA模型小鼠脾淋巴细胞内IL-17细胞因子和Th17细胞相关基因的表达，上调免疫抑制功能Treg的比例，从而维持Treg/Th17正常平衡。蠲痹颗粒醇提物体外给药显著抑制T细胞增殖活性，显著降低IFN-gamma、细胞因子IL-17A和促炎因子IL-6产生水平，上调IL-10的表达水平，抑制MAPK通路ERK 磷酸化水平。蠲痹颗粒醇提物药物干预组显著抑制初始CD4细胞向致病性Th17细胞分化，减少培养上清Th17细胞因子IL-17A的分泌；Treg细胞比例有一定程度的提高，显著增加Treg/Th17比值；蠲痹颗粒醇提物50 ug/ml显著促进iTreg体外分化培养上清中TGF-beta 产生水平，显著上调IL-10和TGF-beta表达。温阳散寒除湿方显著抑制转化Th17细胞比例，差异有显著性意义，一定程度增加Treg细胞比例，提示温阳散寒除湿方干预一定程度阻断Treg细胞转化成致病性Th17细胞。.温阳散寒除湿法可能通过调控T细胞分化可塑性，即有效抑制初始CD4+T细胞向高致病的Th17细胞分化，诱导Treg细胞扩增，同时防止已分化Treg细胞再转化成高致病性Th17细胞而介导治疗类风湿关节炎。这一作用机制可能为扶阳学派及吴生元教授应用“温阳散寒除湿法治疗RA（寒湿痹阻证）理论提供科学基础，丰富和发展中医治疗痹证理论的科学内涵。