靶向干预TLR2-MyD88-PJNK/P38-PI3K信号通路调控肺泡巨噬细胞炎症反应参与哮喘发病的分子机制

Bronchial asthma is a chronic inflammatory airway diseases.The alveolar macrophage is a major effector cell in airway inflammatory diseases,but alveolar macrophage phagocytic molecular mechanism regulated by TLR2 has not been reported.Our published papers showed that TLR signal pathway may be involed in asthma,preliminary experiment found that TLR2 mRNA expression was decreased in alveolar macrophages from patients with asthma than from control patients,Intervention PJNK of TLR2 signaling pathway regulate phagocytosis of alveolar macrophage.Our hypothesis is that TLR2-MyD88-PJNK/P38-PI3K may be involved in pathogenesis of asthma by regulation of inflammtory reaction in alveolar macrophages.We will perform three dimensional cell culture simulating airway,and targeting intervent TLR2-MyD88-PJNK/P38-PI3K pathway,studying the molecular mechanism of alveolar macrophage phagocytosis process, the spatial locus and subcellular localization of phagosome by live cell imaging and 3D reconstruction,and detecing the cytokine expression;using TLR2 knockout mice for the asthma model,intervent key proteins in TLR2-MyD88-PJNK/P38-PI3K signal pathway, determine the role of this signaling pathway involved in the regulation of inflammatory response of alveolar macrophages in the pathogenesis of asthma.Hoping to disclose the new signal pathway in the asthma and proposed a new target for drug treatment .

支气管哮喘是气道慢性炎症性疾患。肺泡巨噬细胞是气道炎症性疾病的主要效应细胞之一，TLR2调控肺泡巨噬细胞吞噬的分子机制尚未见报道。我们已发表论文显示TLR信号通路与哮喘间有联系，预试验发现哮喘患者肺泡巨噬细胞TLR2 mRNA表达降低,抑制TLR2下游分子PJNK降低肺泡巨噬细胞吞噬能力。本课题组推测：TLR2-MyD88-PJNK/P38-PI3K信号通路调控肺泡巨噬细胞炎症反应参与哮喘发病。我们将进行三维细胞培养模拟气道并靶向干预TLR2-MyD88-PJNK/P38-PI3K通路关键蛋白，活细胞动态追踪及三维重构研究肺泡巨噬细胞吞噬过程及吞噬的空间位点与亚细胞定位，并检测炎症因子表达；采用TLR2基因敲除鼠诱导哮喘模型，干预TLR2-MyD88-PJNK/P38-PI3K通路关键蛋白，确定此通路参与的肺泡巨噬细胞炎症反应在哮喘发病中的作用。期望揭示哮喘发病新通路并提出药物治疗的新靶点

支气管哮喘是气道慢性炎症性疾患。肺泡巨噬细胞是气道炎症性疾病的主要效应细胞之一，TLR2 调控肺泡巨噬细胞免疫反应参与哮喘发病的分子机制尚未见报道。本项目以TLR2基因缺失小鼠、原代巨噬细胞、巨噬细胞系为研究对象，以信号通路关键蛋白干预为主要手段。① 建立了三维细胞培养模型、并利用该技术研究巨噬细胞的空间运动特点及TLR2在其中的作用；利用该三维模型，靶向信号通路蛋白干预，发现了TLR2通过JNK信号介导了自噬-吞噬联动，参与了巨噬细胞吞噬病原菌的免疫反应；② 利用原代巨噬细胞及巨噬细胞系，发现MerTK受体可能是TLR2介导巨噬细胞诱导炎症反应的负调控机制；③ 利用TLR2基因缺失小鼠诱导哮喘模型，给予信号通路的抑制剂/协同剂，发现TLR2基因缺失或给与JNK抑制剂SP600125、PI3K抑制剂3-MA可以显著减低气道炎症，但是SP600125或3-MA并不进一步降低TLR2-/-小鼠的气道炎症；④ 在完成计划书研究内容同时，额外研究了TLR家族的另一成员TLR9与哮喘发病的关系，发现TLR9基因缺失与哮喘发病的关系，并初步揭示其机制可能与褪黑素有关。总之，本项目为哮喘的非可控性炎症发病提供了一定的理论基础，为哮喘的治疗提供了潜在的新靶点。