基于PI3K-Akt-mTOR自噬通路探讨益气活血法治疗硬皮病硬化期纤维化的效应机制

Qi deficiency and blood stasis is the main pathogenesis of scleroderma. Yiqi Huoxue therapy can delay the process of fibrosis in hardening of scleroderma. Modern medical research suggests it is the key to the progress of fibrosis in scleroderma that PI3K-Akt-mTOR pathway activated by ROS can mediate inhibited autophagy in in fibroblast. Insufficient autophagy is considered to be the microscopic material foundation of qi deficiency and blood stasis. Our previous studies showed that Yiqi Huoxue Chinese Compound (Huoxue Chubi Decoction) could inhibit the process of fibrosis by inhibiting oxidative stress. So we propose hypothesis that Yiqi Huoxue therapy maybe fight against oxidative stress, regulate PI3K-Akt-mTOR pathway and promote autophagy to eliminate blood stasis, which plays a therapeutic role in scleroderma. In this study, a model of mouse scleroderma will be proposed and a model of human fibroblasts under oxidative stress will be established to simulate the autophagy inhibition of fibroblasts under oxidative stress induced by scleroderma. The models will be treated with Huoxu Chubi Decoction and its pharmacological serum. And then a variety of experimental methods will be used to elucidate the molecular biological mechanism of Yiqi Huoxue Therapy against oxidative stress, regulating autophagy of human fibroblasts and treating scleroderma from different aspects such as whole body, tissue, cell and molecule, in order that improve the objective clinical diagnosis and treatment of scleroderma.

气虚血瘀是硬皮病硬化期的基本病机，益气活血法治疗硬皮病硬化期纤维化疗效确切，但其效应机制尚未完全阐明。最近研究发现，氧化应激条件下PI3K-Akt-mTOR通路介导的成纤维细胞自噬抑制，是硬皮病硬化期纤维化的重要机制。我们前期研究证实益气活血中药复方“活血除痹汤”能够通过对抗氧化应激改善硬皮病硬化期纤维化，结合自噬不足是气虚血瘀的微观物质基础、益气活血药可以促进自噬的研究进展，据此提出“益气活血法可能是通过对抗氧化应激，调控PI3K-Akt-mTOR通路，促进自噬，转化、消除瘀血实邪，对硬皮病硬化期纤维化起到治疗作用”的科学假说。本研究拟构建小鼠硬皮病硬化期模型和人成纤维细胞氧化应激模型，分别以益气活血中药复方“活血除痹汤”及其含药血清进行干预，探讨益气活血法对氧化应激条件下PI3K-Akt-mTOR通路介导的自噬抑制的调控机制，为阐明中医药治疗硬皮病硬化期纤维化的效应机制提供实验依据。

气虚血瘀是硬皮病硬化期的基本病机，益气活血法治疗硬皮病硬化期纤维化疗效确切，但其效应机制尚未完全阐明。本研究中我们构建小鼠硬皮病硬化期模型和人成纤维细胞氧化应激模型，分别以益气活血中药复方“活血除痹汤”及其含药血清进行干预，探讨益气活血法对氧化应激条件下PI3K-Akt-mTOR通路介导的自噬抑制的调控机制。结果发现小鼠硬皮病皮损中存在异常氧化应激反应，且氧化应激与PDGF、TGF-β在硬皮病发病机制中存在一定的关联。硬皮病成纤维细胞中存在的氧化应激可通过抑制PI3K-Akt-mTOR通路活性进而抑制成纤维细胞自噬相关因子beclin-1、LCI/II的表达。应用活血除痹汤干预后，成纤维细胞中促纤维因子TGF-β、PDGF降低，ROS受到抑制，PI3K-Akt-mTOR通路活性恢复，自噬相关因子Akt、mTOR、LC3-I/II活性及表达增强。上述结果表明，活血除痹汤可能通过降低组织氧化应激进而促进自噬，从而加强对胶原的降解能力，同时可下调促纤维化因子，从而减少皮肤胶原合成，进而发挥其对硬皮病的治疗作用。