肺痹汤调控BRP-39/IL-17与TGF-β/ Smad3的交互作用以改善肺纤维化免疫炎性病理损伤

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive deteriorating lung disease whose incidence increases year by year, and mortality rate is extremely high. There is still no effective treatment for it. .The pathogenesis of this disease is multifactorial. According to the latest research, immune inflammatory reaction is the most important part of the damage of pulmonary fibrosis..The pre-test study proved that Feibitang can effectively inhibit the overexpression of IL-6/TGF-β1/P38MAPK cell signaling pathway, reduce pulmonary fibrosis of the bleomycin model rat. .Therefore deduce the biological mechanisms of Feibitang suppressing immunopathological damage of pulmonary fibrosis may function by regulating the interaction of immune inflammatory and fibrosis factor. It is found that chitinase is the first cytokine released after macrophage activation, as well as the upstream factor of pulmonary fibrosis cytokine network, which participates in immune inflammatory pathological damage of pulmonary fibrosis. .On the basis of the pre-project, this study focuses on gene and protein level, combining vivo experiment of cells with in vitro experiment of CC10-rtTA-tTS-TGFβ1 triple transgene model mice, aiming to study the impact of Feibitang on the signaling pathway and interaction of immune inflammatory and fibrosis factor, and target of its intervention to immunopathological injury of pulmonary fibrosis..The project discusses the molecular mechanisms of Feibitang in suppressing the immunopathological damage of pulmonary fibrosis, intend to provide evidences to enrich the treatment for pulmonary fibrosis, and build a foundation in the study of biological mechanism of interaction of immune inflammatory and fibrosis factor in immunopathological injury of pulmonary fibrosis.

特发性肺纤维化是一种进行性恶化的肺系疾病，死亡率高，目前仍缺乏有效治疗手段。最新研究认为，免疫炎性反应是介导及参与肺纤维化病理损伤的核心环节。本课题前期试验证明：肺痹汤通过下调IL-6/TGFβ1/P38MAPK信号通路相关因子的过表达，减轻大鼠肺纤维化程度。由此推论，肺痹汤可能通过抑制免疫炎性及纤维化途径的交互作用，改善肺纤维化病理损伤。预实验发现，BRP39/YKL40作为介导肺组织免疫炎性反应的上游机制，在肺纤维化损伤中扮演重要角色。故本项目在原工作基础上，在基因蛋白质水平，采用CC10-rtTA-tTS-TGFβ1三重转基因模型小鼠体内与体外实验相结合方式，研究该方对BRP39/IL-17与TGFβ1/Smad3 信号途径的影响及作用环节，探讨肺痹汤抑制肺纤维化免疫炎性损伤的分子机制，为肺纤维化的治疗寻找新突破；为研究BRP39/IL-17在纤维化损伤领域中的生物学机制奠定基础。

特发性肺纤维化是疑难病，缺乏有效治疗手段。探究其关键病理机制及有效干预靶点是亟待解决的问题。本课题组据前期研究推论，肺痹汤可能通过抑制免疫炎性及纤维化途径的交互作用，改善肺纤维化病理损伤。本项目研究肺痹汤对BRP-39/IL-17与TGF-β1/Smad3通路的影响，探讨肺痹汤抑制肺纤维化免疫炎性损伤的作用靶点。结果示：体外：（1）肺痹汤含药血清抑制THP-1巨噬细胞和MRC-5共培养体系YKL40及受体IL13-R2、TGF-β1、Smad3表达，上调Smad7水平，减少ColⅠ/Ⅲ、α-SMA合成。（2）肺痹汤含药血清通过降低TGF-β1、Smad3、ColⅠmRNA表达，上调Smad7而抑制IL-17诱导的MRC-5增殖。（3）薯蓣皂苷元抑制LPS诱导MRC-5中TGF-β1水平及JNK、P38磷酸化，提示薯蓣皂苷元可能通过抑制TGF-β1激活的JNK和P38 MAPK通路来发挥抗纤维化作用。体内：（1）肺痹汤抑制博来霉素诱导肺纤维化小鼠BRP-39、IL-17、IL-6、CXCL-13、Icam-1表达而减轻博来霉素诱导的免疫炎症反应，抑制TGF-β1/Smad通路激活，达到减轻肺纤维化的作用。（2）肺痹汤可抑制AdTGF-β1诱导的肺纤维化小鼠TGF-β1/Smad通路激活，降低BRP-39、IL-17、IL-13表达，减弱Th17细胞分泌IL-17强度而减轻AdTGF-β1诱导的小鼠肺纤维化。综上，无论在博来霉素诱导的小鼠肺纤维化炎症模型，或AdTGF-β1致TGF-β1高表达的小鼠肺纤维化模型，以及体外实验中，炎症因子BRP-39/YKL-40、IL-17均参与免疫炎性病理损伤过程，与纤维化因子交互作用，诱导纤维化发生发展。而肺痹汤可能是通过调节BRP-39/IL-17与TGF-β1/Smad信号通路的交互作用，改善了免疫炎性病理损伤，从而发挥抗肺纤维化作用。