二氢杨梅素调控自噬保护高糖诱导血管内皮细胞损伤的作用与机制

Diabetic vascular complications is the leading cause of morbidity and mortality among people suffering from diabetes,and high glucose-induced vascular endothelial cells injury is a major initiator of diabetic vascular complications.Dihydromyricetin is extremely rich content of flavonoids in ampelopsis grossedentata,our preliminary study showed that dihydromyricetin significantly protected human umbilical vein endothelial cells from high glucose-induced injury,and counteracted inhibition of autophagy and AMPK activity triggered by high glucose.In this project,we hypothesize that autophagy impairment through AMPK inhibition is implicated in high glucose-induced vascular endothelial cells injury,and dihydromyricetin could protect vascular endothelial cells from high glucose-induced damage through promotion of autophagy via activation of AMPK.Based on this hypothesis,we will investigate the role of autophagy in high glucose-induced vascular endothelial cells damage and the protective effect of dihydromyricetin,the role of AMPK in regulation of autophagy by high glucose and dihydromyricetin and its signaling pathway,and the underlying mechanisms of high glucose and dihydromyricet modulate AMPK activity in vitro, and then futher evaluate the protective effect of dihydromyricetin on the vascular endothelium injury through AMPK-dependent autophagy in streptozotocin-induced type 1 diabetic rats.This study has important significance in clarifying the mechanisms of vascular endothelial cells injury induced by high glucose,and provides scientific basis for application of dihydromyricetin or drinking ampelopsis grossedentata in the prevention of diabetic vascular complications among patients with diabetes in the future.

血管并发症(DVC)是糖尿病致残和致死的首要原因，而高糖诱导血管内皮细胞(VEC)损伤是DVC发生的主要启动因子。二氢杨梅素(DHM)是藤茶中含量极高的黄酮类化合物，我们前期初步研究发现DHM对高糖诱导VEC损伤具有明显保护作用，并抑制高糖诱导的AMPK活性和自噬活性下降。本项目提出"AMPK活性抑制引起自噬功能受损参与高糖诱导VEC损伤，DHM可通过激活AMPK促进自噬从而保护高糖诱导VEC损伤"的研究假说，首先采用体外损伤模型研究自噬在高糖诱导VEC损伤中的作用及DHM的干预效应、AMPK在高糖和DHM调控VEC自噬中的作用及其信号通路、高糖和DHM调节AMPK活性的机制，然后利用1型糖尿病大鼠模型验证DHM通过促进AMPK介导的自噬对高血糖损伤血管内皮的保护效应。本研究对阐明高糖诱导VEC损伤的作用机制具有重要理论意义，为将来在糖尿病患者中应用DHM干预或饮用藤茶防治DVC奠定基础。

糖尿病血管并发症是糖尿病致残和致死的首要原因，而高糖诱导血管内皮细胞损伤是糖尿病血管并发症发生的主要启动因子。业已证实腺苷酸活化蛋白激酶（AMP-activated protein kinase, AMPK）介导的细胞自噬功能障碍在肿瘤、神经退行性疾病等多种疾病的发生发展过程中发挥重要作用，但其在高糖诱导血管内皮细胞损伤中的意义尚未见研究报道。二氢杨梅素是藤茶中含量极高的黄酮类化合物，具有抗氧化、抗肿瘤等多种药理学功效，我们前期研究发现二氢杨梅素是AMPK的天然激动剂。本项目首先采用体外损伤模型研究自噬在高糖诱导血管内皮细胞损伤中的作用及二氢杨梅素的干预效应、AMPK在高糖和二氢杨梅素调控血管内皮细胞自噬中的作用及其信号通路、高糖和二氢杨梅素调节AMPK活性的机制，然后利用1型糖尿病大鼠模型验证二氢杨梅素通过促进AMPK介导的自噬对高血糖损伤血管内皮的保护效应。体外实验研究发现，高糖通过调控Akt/GSK3β/GS通路促进血管内皮细胞糖原合成继而抑制AMPK/mROR和AMPK/ULK1信号通路、降低细胞自噬功能，从而导致血管内皮细胞损伤；而二氢杨梅素干预则可通过抑制血管内皮细胞F1F0-ATP合成酶活性降低其ATP含量、升高AMP/ATP比值继而增强AMPK/mROR和AMPK/ULK1信号通路、促进自噬功能，进而减轻高糖诱导的血管内皮细胞损伤效应。体内实验研究结果与体外细胞实验结果一致，进一步验证了二氢杨梅素可通过调控AMPK介导的自噬信号通路对糖尿病大鼠血管内皮损伤发挥保护效应。本研究结果不仅初步阐明了自噬功能障碍在高糖损伤血管内皮细胞中的作用，同时也为在糖尿病患者中应用二氢杨梅素干预或饮用藤茶预防糖尿病血管并发症提供了实验证据。