Nrf2介导的TGFβ-Smad通路在氧化应激致肺纤维化中的作用及当归补血总苷的调控机制

Idiopathic pulmonary fibrosis (IPF) has caused serious consequences, while the exact mechanisms underlying have not been clarified. Oxidative stress and the occurrence of IPF are closely related. （Nuclear Factor-Erythroid 2-Related Factor 2）Nrf2 is the key transcription factor that links oxidative stress to pulmonary fibrosis. Previous studies have indicated that the total glycosides of DangGui BuXue Decoction may attenuates bleomycetin induced pulmonary fibrosis via regulating the remodeling process of ECM by adjusting TGFβ/Smad signaling pathway. However, there has been no reported about the relationship links TGFβ- Smad and Nrf2 to pulmonary fibrosis at home and abroad. Based on literatures and our previous research foundation, we speculate that TGFβ- Smad mediated by Nrf2 could control the process of pulmonary fibrosis. To prove this hypothesis, this project try to adopt Nrf2 gene knockout and wild type mice and cell model in PF, then use the method of RT-PCR and Western Blot to detect Nrf2 and related antioxidant enzymes, as well as the changes of the key genes in TGFβ and the protein expression. So we can approach the role of TGFβ-Smad that Nrf2 mediated in pulmonary fibrosis and the regulation mechanisms of DBTG from the global, cell and molecular standard, to explore the prevention and therapy's new targets in IPF, that could provide new thinking and new methods to prevent and cure pulmonary fibrosis in TCM.

特发性肺纤维化(IPF)危害严重，其确切机制尚未完全阐明。氧化应激与IPF发生密切相关，Nrf2是氧化应激中起关键保护作用的转录因子，可能在氧化应激致肺纤维化中起调节作用。课题组前期研究发现，当归补血总苷通过TGFβ-Smad通路调控ECM重塑发挥抗肺纤维化作用。但Nrf2与TGFβ-Smad通路在肺纤维化发生发展中的关系，迄今国内外尚无报道。结合文献和既往研究基础，我们推测Nrf2可能介导TGFβ-Smad信号通路调控肺纤维化进程。为证实这一假说，本项目拟在Nrf2基因敲除、野生型小鼠和细胞PF模型上，采用RT-PCR和Western Blot法检测Nrf2及相关抗氧化酶和TGFβ通路中蛋白表达的变化，在整体、细胞和分子水平上探讨Nrf2介导的TGFβ-Smad通路在肺纤维化进程中的调控作用及当归补血总苷抗肺纤维化的作用机制，探索IPF防治新靶点，为中医药防治肺纤维化提供新思路和新方法。

前期研究表明，当归补血总苷（DBTG）能通过TGFβ-Smad 通路调控ECM 重塑发挥抗肺纤维化作用。但Nrf2 与TGFβ-Smad 通路在肺纤维化发生发展中的关系尚不明确。本项目拟在Nrf2 基因敲除、野生型小鼠和细胞PF 模型上，采用RT-PCR 和Western Blot 法检测Nrf2 及相关抗氧化酶和TGFβ通路中蛋白表达的变化，在整体、细胞和分子水平上探讨Nrf2 介导的TGFβ-Smad 通路在肺纤维化进程中的调控作用及当归补血总苷抗肺纤维化的作用机制。结果发现，DBTG能通过下调NOX4表达，减少ROS生成，调节体内氧化应激水平，进而抑制TGF-β1激活，影响其下游蛋白COL-Ⅰ和α-SMA的合成，阻碍ECM沉积，最终影响PF发展。HMGB1可通过激活TGF-β1/Smad2/3信号通路促进EMT的发生。进一步研究发现，Nrf2能通过抑制TGF-β1-Smad信号通路抑制肺肌成纤维细胞增殖，且Nrf2可通过调节snail蛋白的表达从而调节肺纤维化发生进程。以上研究结果揭示了Nrf2 介导的TGFβ-Smad 通路在肺纤维化进程中的调控作用及当归补血总苷抗肺纤维化的作用机制，为中医药防治肺纤维化提供新思路和新方法。