基于TLRs/MyD88/NF-kB及紧密连接研究健脾养血祛风法调控特应性皮炎皮肤免疫及机械屏障机制

The pathogenesis of atopic dermatitis involving immunologic barrier and mechanic barrier function of mice model, and the two barriers are related and influenced with each other recently. Our previous studies have confirmed that the itching degree was induced and increased ceramide by Jianpi yangxue qufeng method. There are significant clinical effect and repairing skin barrier function with Jianpi yangxue qufeng method. Current studies demonstrated Toll-like receptors and its medicated innate immune plays very important role in skin immunologic barrier. And Toll-like receptors also controlled the tight junction and junctional adhesion molecules which related with skin mechanic barrier function disorder and epidermis water loss closely. Tight junction and junctional adhesion molecules was the key. construction when the skin barrier was damaged in atopic dermatitis patients. We are going to explore the molecular mechanism underlying the therapeutic effects of Jianpi yangxue qufeng method on immunologic barrier and mechanic barrier function throughout in vivo and in vitro models by molecular biological approaches based on our basic studies . Jianpi yangxue qufeng method will contribute to the clinical treatment and prevention in atopic dermatitis patients.

目前特应性皮炎（atopic dermatitis, AD）的发病机制涉及到免疫屏障功能及表皮机械屏障功能的相互用和相互制约。前期研究证实健脾养血祛风法能减轻脾虚血燥模型小鼠瘙痒程度，提高神经酰胺含量；该法治疗特应性性皮炎具有明显的临床疗效和修复皮肤屏障功能的作用。研究显示Toll样受体(Toll-like receptors, TLRs) 及其介导的天然免疫在皮肤免疫屏障中发挥重要作用，并且受TLRs调控下的紧密连接（tight junction, TJ）及黏附结构这些与皮肤机械屏障功能失调和由此引起的经表皮水分丢失密切相关，紧密连接及黏附结构的改变参与AD患者皮肤机械屏障功能的破坏过程。因此本课题拟在前期研究基础上通过体内外实验模型及分子生物学研究方法，进一步研究健脾养血祛风法调节特应性皮炎免疫屏障及改善机械屏障功能的分子机制及作用靶点，对特应性皮炎的防治具有重要现实意义。

特应性皮炎(atopic dermatitis，AD)，又称为异位性湿疹，是与遗传、免疫等因素密切相关且容易复发的炎症性皮肤病。健脾养血祛风方是名医刘复兴教授治疗AD的经验方。二十多年来，健脾养血祛风方在临床治疗中取得显著疗效。皮肤的免疫屏障功能和表皮机械屏障功能两者的相互作用和相互制约，是AD的发病机制。课题组的前期研究证实：健脾养血祛风法能明显降低脾虚血燥小鼠的瘙痒程度，提高小鼠表皮的神经酰胺含量。健脾养血祛风法治疗AD，临床疗效好及皮肤屏障功能修复强。本课题以Toll样受体和Toll样受体介导的免疫为切入点，提出了"健脾养血祛风法可能通过 TLRs 信号通路调控紧密连接从而改善特应性皮炎"的假说。研究内容：第一：构建IL-4转基因小鼠AD模型；第二、通过动物实验，研究健脾养血祛风方对 IL-4 转基因小鼠AD模型的作用；第三、研究健脾养血祛风方对 TLRs/MyD88/NF-kB及紧密连接信号传导通路的相关蛋白表达水平、细胞因子水平、特异转录因子表达的影响。研究结果：第一.健脾养血祛风方的中、高剂量组能降低IL-4转基因小鼠的瘙痒次数、减少皮损面积；第二、健脾养血祛风方可能通过TLRs/MyD88-NF-kB 信号通路和紧密连接（tight junction, TJ）调控特应性皮炎的皮肤免疫功能和机械屏障功能，缓解AD的病情；第三、健脾养血祛风方可促进DCs的TLRs/MyD88/NF-κB信号通路激活，从而调控炎症因子表达，使Th1/Th2细胞因子网络平衡体系向Th1方向倾斜，从而提高HaCat中TJ相关蛋白表达，达到修复机械屏障，缓解AD的作用；第四、阐述了健脾养血祛风方作用于TLRs/MyD88/NF-κB信号通路及其介导的天然免疫在皮肤免疫屏障中发挥的重要作用，和TLRs调控下的紧密连接及黏附结构等与皮肤机械屏障功能失调的关系，和皮肤机械屏障功能失调引起的经表皮水分丢失密切相关，以及TJ及黏附结构的改变参与AD患者皮肤机械屏障功能的破坏过程。本课题提供了名老中医经验方（健脾养血祛风方）防治AD的实验依据，为后续新药研发奠定科学基础。