基于TGF-β1/Snail和Wnt/β-catenin信号通路介导上皮细胞间质转化探讨补肾通络方对肺纤维化大鼠的抑制作用

Abstract:Idiopathic pulmonary fibrosis (IPF) may be related to alveolar epithelial cells with damage and mesenchymal transformation of epithelial cells (EMT),TGF-β-1/Snail andWnt/β-catenin signaling pathway activation is key part of EMT.Based on our previous study of buqi tongluo and bu zongqi tong luo，we proposed that pathogenesis of "pulmonary collateral stasis and deficiency of" is the core hypothesis in IPF.Clinical research had proven bushen tongluo decoction in treating IPF has a good effect,we speculate that it may be related to inhibition of TGF-β-1/Snail and Wnt/β-catenin signaling pathway.In support of this hypothesis,we plan through the lung tissue of rats with pulmonary fibrosis and interstitial cell markers, TGF-β 1, Snail, Wnt, β--catenin changes in gene expression,to study bushen tongluo decoction on TGF-β 1/Snail and Wnt/β-catenin effects of signaling pathway-mediated EMT, through the gene silencing and overexpression to confirm TGF-β1/Snail and Wnt/β-catenin signaling pathways have interaction.Clarify bushen tongluo decoction on pulmonary fibrosis in rats with role of EMT,revealing its mechanisms and targets.

特发性肺纤维化（IPF）发病可能与肺泡上皮细胞损伤后的上皮细胞间质转化（EMT）有关，TGF-β1/Snail和Wnt/β-catenin信号通路的激活是EMT的关键环节。我们在前期补气通络、补宗气通络基础上进一步提出“肺络痹阻、肺肾亏虚”是IPF核心病机的假说，临床已证实补肾通络方治疗IPF有良好疗效，推测其机理可能与抑制TGF-β1/Snail和Wnt/β-catenin信号通路有关。为了证实这一假说，我们拟从肺纤维化大鼠肺组织上皮及间质细胞标记物、TGF-β1、Snail、Wnt、β-catenin基因蛋白表达水平的变化，探讨补肾通络方对TGF-β1/Snail和Wnt/β-catenin信号通路介导EMT的影响，并通过基因沉默及过表达，证实TGF-β1/Snail和Wnt/β-catenin信号通路存在交互作用，阐明补肾通络方对肺纤维化大鼠EMT的调节作用，揭示其作用机理及靶点。

本课题组首先基于网络药理学探讨了补肾通络方治疗间质性肺病的作用机制，然后开展了体内及体外实验研究。进一步验证补肾通络方对肺纤维化大鼠肺功能下降的延缓作用；观察补肾通络方对肺纤维化大鼠血清及支气管肺泡灌洗液细胞因子的影响、补肾通络方对肺纤维化大鼠肺组织的保护作用、补肾通络方对肺纤维化大鼠细胞外基质重建的抑制作用及补肾通络方对肺纤维化大鼠 EMT 的抑制作用，探讨 TGF-β1/Snail、Wnt3a/β-catenin 信号通路的交互作用在大鼠肺纤维化形成过程中的作用，在转录、翻译水平分别验证补肾通络方对肺纤维化大鼠 TGF-β1/Snail及 Wnt3a/β-catenin 信号通路相关蛋白的调节作用。.研究证实，TGF-β1/Snail和Wnt3a/β-catenin信号通路的激活能促进大鼠肺纤维化过程中EMT，从而促进肺纤维化的发生发展。补肾通络方对于肺纤维化大鼠肺功能FVC的下降具有延缓作用；进一步研究表明，补肾通络方通过干预肺泡上皮细胞-间质细胞转化，从而抑制细胞外基质重建，达到治疗肺纤维化的目的；分析其分子生物学作用机制，可能与补肾通络方调控TGF-β1/Snail及Wnt3a/β-catenin信号通路的激活，从而抑制大鼠EMT过程，一定程度上延缓纤维化进程。