麻黄细辛附子汤干预ILC2s介导2型免疫应答治疗AR机制及其拆方研究

Ephedra Asarum Aconite Decoction warms yang to relieve exterior syndrome and treats both interior and exterior syndromes, so it has superior effect in the treatment of allergic rhinitis (AR). The newly discovered ILC2s are a key group of cells expressing type 2 cytokines, which provide a new perspective to explain the mechanisms of the allergic reactions in AR. Based on previous studies, we hypothesize that ILC2s are important targets for Ephedra Asarum Aconite Decoction to intervene the type 2 immune responses in AR, and a potential mechanism for the efficacy of the Traditional Chinese Medicine (TCM). In this project, we focused on ILC2s, and studied DCs and Th1/Th2, linking innate immunity and adaptive immunity. AR model was established in wild-type and gene knockout animals, and early initiation of ILC2s was stimulated in vitro to study its effects on DCs and T cells. Classical molecular biology, mass spectrometry, as well as the latest transcriptome sequencing and transmission electron microscopy techniques were used to elucidate the mechanisms of action of this formula in intervening type 2 immune responses, from the aspects of ultrastructure, activation, metabolism, differentiation, migration, gene transcription and signal pathway of the cells. The use of decomposed recipe in this study enables screening of effective components on each target and the preferred target of each herb, as well as interaction among the three herbs, i.e., Ephedra, Asarum and Aconite. This will reveal the rule of compatibility and specific efficacy, which will inspire and set example for TCM studies, and improve clinical treatment for AR.

麻黄细辛附子汤温阳解表、表里兼顾，临证治疗AR优势突出。新发现ILC2s是表达2型细胞因子的关键细胞，是阐释AR变态反应机理新视角。结合前期研究，我们提出“ILC2s是麻黄细辛附子汤治疗AR干预2型免疫应答的重要机制并是体现其中医功效的潜在途径”科学假说。本项目拟以ILC2s为核心，兼及DCs、Th1/Th2，对接固有免疫与适应性免疫，对野生型和基因缺陷型动物诱导AR造模，并体外模拟ILC2s早期启动及其对DCs、T细胞影响，利用经典分子生物学方法和质谱流式、转录组测序、透射电镜等新技术，从细胞超微结构、激活、代谢、分化、迁移、基因转录及信号通路等角度，深入研究本方对2型免疫应答的干预作用，阐明其药理机制。并利用拆方研究，筛选本方在各节点的重点起效部位和各拆方组的抗过敏优势靶点，还能明确麻辛附三药相互作用，揭示本方配伍规律和功效特色的科学内涵，以启发和示范同领域研究，推动AR临床治疗水平。

本项目在实施过程中基本按照计划执行。.本研究表明，麻黄细辛附子汤全方与各拆方配伍均能够治疗OVA致敏小鼠发挥抗过敏疗效，全方效果最优，证明了全方配伍的科学意义。相比于抑制下游IgE释放，麻黄细辛附子汤全方在抑制ILC2s增殖，调节整个Th2偏移环节效果更显著；在影响Th2偏移的关键转录因子GATA3的基因层面，全方及各拆方组配伍中，以附子单用的干预效应最为突出，同时，在下游IgE释放环节中，附子单用、辛附合用也显示了最显著的抑制效应，提示温阳扶正法在Th2偏移型免疫疾病中的运用至为关键。麻黄细辛附子汤可以通过缓解线粒体氧化应激损伤，保护线粒体结构，保证其能量代谢功能从而发挥温补脾肾经阳气治疗变应性鼻炎的作用。麻黄细辛附子汤能够有效提升非变应性鼻炎中ILC2细胞的含量，该作用可能参与炎症消退、粘膜修复等过程。.麻黄细辛附子汤全方可以减少CD4+T细胞升高IL-4、STAT6mRNA的表达，麻黄拮抗细辛、附子的作用，升高IL-4水平，细辛、附子配伍升高IL-5水平，促进T细胞向Th2分化，从而进行免疫调节。辛附的配伍或者细辛、附子单用可通过降低STAT6mRNA水平发挥免疫调节作用，而全方、麻附、麻辛配伍则相反。麻黄细辛附子汤可以影响上游抑制树突状细胞的迁移，使其无法诱导初始T细胞向Th2分化。.本研究确证了麻黄细辛附子汤对变应性鼻炎发病2型免疫应答的有效干预、阐明麻黄细辛附子汤干预ILC2s作用机制，挖掘了本方抗过敏效应靶点，为麻黄细辛附子汤临床治疗变应性鼻炎提供了科学依据，开拓了变应性鼻炎临床论治思路，初步筛选了麻黄细辛附子汤治疗变应性鼻炎的物质基础在一定程度上揭示了麻黄细辛附子汤组方特色及配伍机制的科学意义。