钝化“多环芳香烃-环境感受器AhR轴”调控肺泡巨噬细胞活化和肺部炎症的作用研究

Haze is a serious environmental issue and harmful to the health, so this issue is drawing widespread attention. The harmful polycyclic aromatic hydrocarbons (PAHs) are highly enriched in the fine particulate matter with a diameter of ≤2.5 μm (PM2.5), which is the main component of the haze. Inhalation of PAHs-containing PM2.5 may induce lung inflammation, and the study of its specific mechanism has become the research focus.. Our research is based on the facts that PAHs are the ligands for aryl hydrocarbon receptor (AhR) in alveolar macrophages, and that activated AhR is involved in the regulatory mechanism of inflammatory response. Furthermore, our preliminary experiment found the elevated levels of AhR protein and inflammatory factor (TNF-α) in the lung of mice treated with vehicle exhaust, and some possible non-proinflammatory ligands of AhR in bioinformatics analysis. Therefore, the corresponding hypothesis is proposed: the binding of PAHs to AhR can induce the imbalance between proinflammatory and anti-inflammatory responses after PAHs-containing PM2.5 is ingested by alveolar macrophages. In this study, we plan to adopt the mice treated with vehicle exhaust and airway instillation, take AhR as the cut-in point of the research, and use AhR-/- mice and the corresponding alveolar macrophages as experimental objects. We would like to clarify: (i) the role of AhR on the inflammatory response of the lung induced by haze-associated PAHs; (ii) the process of lung inflammation and the imbalanced pattern between the proinflammatory and anti-inflammatory response modulated by alveolar macrophages; and (iii) the discovery of non-proinflammatory ligands or truncation protein of AhR which can passivate AhR of alveolar macrophages. This study may provide theoretical evidence and novel therapy for lung inflammation induced by haze-associated PAHs.

目前雾霾对健康的危害日益受到关注，其主要成分PM2.5颗粒含危害健康的多环芳香烃（PAHs），后者入肺后可导致肺部炎症，其具体机制已成为研究热点。基于“PAHs是肺泡巨噬细胞内芳香烃受体（AhR）的配体”，“AhR激活后参与炎症反应调节”，以及前期实验结果（汽车尾气干预的小鼠肺组织高表达AhR和炎症因子TNF-α，生信分析发现部分可能的AhR非促炎配体），提出假说：含有PAHs的PM2.5被肺泡巨噬细胞吞噬后，PAHs与胞内AhR结合，诱发促炎-抗炎反应平衡紊乱。本项目拟采用汽车尾气和气道滴注小鼠模型，以AhR为切入点，以AhR-/-小鼠及肺泡巨噬细胞为研究对象，拟明确①AhR在雾霾相关PAHs导致肺部炎症中的作用，②肺部炎症进程和巨噬细胞调控的促炎-抗炎反应的失衡模式，③寻找钝化巨噬细胞AhR的非促炎配体或截断型蛋白。该研究将为雾霾相关PAHs导致的肺部炎症提供理论依据和新的治疗思路。

PM2.5颗粒中的多环芳香烃（PAHs）导致肺部炎症的机制及应对策略已成为研究热点。肺泡巨噬细胞（AMs）芳香烃受体（AhR）作为环境感受器，是PAHs的高亲和受体。本项目揭示AMs内AhR参与了PAHs导致的肺部炎症。与对照组相比，经PAHs暴露的小鼠AMs的TLR2、TLR4、TNF-α基因表达显著升高（p<0.05）。下调AhR（shRNA干扰）的小鼠AMs，AhR基因回补可恢复PAHs引发的TNF-α高表达（p<0.05）。我们运用生物信息学方法从临床药物、天然化合物等数据库筛选AhR配体候选化合物；再以AhR下游CYP1A1作为活性筛选指标，结果显示筛选出的37个候选化合物中9个呈现抑制效应，2个配体化合物呈现激动剂效应。文献报道香烟烟雾中含多种PAHs。因此用香烟烟雾造模，干预小鼠（体内）和小鼠AMs（体外），制造PAHs相关的肺部炎症动物和细胞模型，选取具有潜在抗炎作用的脂联素作为阻断剂。实验证实脂联素能剂量依赖性降低小鼠肺部炎症细胞聚集，抑制AMs向促炎表型（M1）极化，抑制促炎因子（IL-1β、IL-6、TNF-α）表达（p<0.05），提升抗炎因子IL-10的表达水平（p<0.05），表明脂联素能缓解香烟烟雾造成的肺部炎症。以香烟烟雾暴露AMs为细胞模型，基因表达谱、GO富集分析、WB实验等方法证实脂联素能作用于香烟烟雾暴露AMs内的两个途径抑制肺部炎症：1）通过抑制toll样受体（TLR2/4）触发胞内NF-κB、JNK/p38通路，2）借助COX-2/PGE2通路抑制AMs向促炎表型（M1）极化。总之，本项目揭示了AhR抑制性配体的潜在应用价值，阐明了脂联素能通过抑制AMs的活化及促炎因子分泌进而抑制PAHs相关的肺部炎症，具有潜在的临床应用前景。