香菇多糖通过TLR4激活STUB1/TFEB通路诱导自噬防治糖尿病血管病变的作用及机制研究

Vascular endothelial cell (VEC) injury is the main inducing factor of diabetic angiopathy. The efficacy of Lentinan (LNT) conforms to the traditional Chinese medicine treatment principle of diabetic angiopathy, but its actual effect and mechanism are still unclear. Adaptive autophagy can alleviate diabetic angiopathy. STUB1/TFEB pathway is the latest key pathway to regulate autophagy level, and is a potential important target to regulate autophagy in the treatment of diseases. However, the role of STUB1/TFEB pathway in diabetic angiopathy has not been reported. Previous studies have found that LNT can affect TLR4 level to induce autophagy and protect VEC, while TLR4 has regulatory effect on STUB1/TFEB pathway. Based on the research progress of the autophagy mechanism and the preliminary results, we hypothesize that LNT can activate STUB1/TFEB pathway through TLR4, regulate autophagy to reduce VEC damage and improve diabetic angiopathy. This topic clarifies the role of STUB1/TFEB pathway in diabetic angiopathy, and reveals the mechanism of LNT regulating autophagy through activating STUB1/TFEB pathway through TLR4 to prevent and treat diabetic angiopathy, providing new ideas for the prevention and treatment of the disease, expanding the indications of LNT, and finding potential drug targets.

血管内皮细胞（VEC）损伤是糖尿病血管病变的主要诱发因素，香菇多糖（LNT）的功效符合糖尿病血管病变的中医治疗法则，然而其实际效应和机制尚不明确。适应性自噬可缓解糖尿病血管病变，STUB1/TFEB通路是调节自噬的最新关键性通路，可作为调控自噬以治疗疾病的潜在重要靶点。但STUB1/TFEB通路在糖尿病血管病变中的作用还未见报道。课题组前期研究发现LNT能影响TLR4水平诱导自噬保护VEC，而TLR4对STUB1/TFEB通路有调控效应。结合上述自噬机制的研究进展及预实验结果，我们提出假说：LNT通过TLR4激活STUB1/TFEB通路，调控自噬减轻VEC损伤从而改善糖尿病血管病变。本课题明确STUB1/TFEB通路在糖尿病血管病变中的作用；揭示LNT通过TLR4激活STUB1/TFEB通路调控自噬防治糖尿病血管病变的机制，为该疾病的防治提供新思路，扩大LNT的适应症，找到潜在的药物靶点。

背景：血管内皮细胞（VEC）损伤是糖尿病血管病变的主要诱发因素，香菇多糖（LNT）的功效符合糖尿病血管病变的中医治疗法则，然而其实际效应和机制尚不明确。适应性自噬可缓解糖尿病血管病变，STUB1/TFEB通路是调节自噬的最新关键性通路，可作为调控自噬以治疗疾病的潜在重要靶点。但STUB1/TFEB通路在糖尿病血管病变中的作用还未见报道。主要研究内容：通过动物实验和细胞实验探讨香菇多糖通过促进自噬保护内皮细胞改善糖尿病血管病变的作用及机制。重要结果：香菇多糖通过作用于TLR4调控STUB1/TFEB通路诱导自噬，改善氧化应激，逆转血管内皮细胞损伤，治疗糖尿病血管病变。关键数据：体内实验结果显示香菇多糖能降低糖尿病大鼠血糖水平，增加糖尿病大鼠体重；改善糖化血红蛋白、尿素、肌酐、甘油三酯等生化指标及腹主动脉血管病变；体外实验结果显示，香菇多糖能提高HUVEC细胞在高糖下的细胞活力，降低细胞黏附、vWF、E-Selectin等水平，改善高糖下的细胞功能障碍；体内体外实验均显示香菇多糖能减少血管内皮细胞凋亡，提高LC3、Beclin-1水平，降低iNOS、P38MAPK、ROS含量，改善血管组织氧化应激情况；抑制TFEB活化，促进STUB1蛋白表达，激活STUB1/TFEB自噬信号通路。在加入3-MA自噬抑制剂后，香菇多糖对血管内皮细胞的保护作用消失；抑制香菇多糖作用靶点TLR4后，香菇多糖对细胞的保护作用、自噬水平、氧化应激等指标的影响消失。科学意义：本研究揭示LNT通过TLR4激活STUB1/TFEB通路调控自噬防治糖尿病血管病变的机制，为该疾病的防治提供新思路，扩大LNT的适应症，找到潜在的药物靶点。