摘要
目的探讨高良姜黄素(Cur)增强内皮细胞(ECs)自噬水平及抑制损伤血管内膜增生(IH)的作用机制。方法体外实验中,将人脐血管内皮细胞(human umbilical vein endothelial cells,HUVECs)分为4组:对照组、3-MA组、Cur组、雷帕霉素(Rapa)组;采用吖啶橙(AO)染色和溶酶体荧光探针法(Lyso-TrackerRed)观察细胞酸性区室积累;Western-blot、免疫荧光检测微管相关蛋白轻链3Ⅱ(LC3-Ⅱ)、p62、Akt及mTOR的蛋白水平。活体实验中,用球囊压迫法构建颈动脉内膜损伤大鼠模型;60只健康成年雄性SD大鼠分为假手术组、血管损伤组、3-MA组、Cur组和Rapa组;采用Westernblot法检测各组大鼠颈动脉组织LC3-Ⅱ、p62、Akt及mTOR的蛋白水平,HE染色检测IH厚度及有效的管腔面积。结果体外实验中,与对照组相比,Cur组和Rapa组HUVECs内酸性区室明显增多,同时LC3-Ⅱ蛋白水平增高,p62、Akt及mTOR蛋白水平降低。活体实验中,与对照组相比,损伤血管中的LC3-Ⅱ蛋白水平上升,而p62、Akt及mTOR蛋白水平稍下降,Cur组和Rapa组大鼠血管中的LC3-Ⅱ蛋白水平明显升高,而p62、Akt及mTOR蛋白水平显著下降。HE染色显示,颈动脉内膜损伤后3周,损伤节段血管可见明显的IH及血管腔狭窄,而Cur组和Rapa组抑制了IH及减轻血管狭窄。无论在体外实验还是活体实验中,3-MA干预后均呈现一个相反的结果。结论 Cur能增加ECs的自噬水平及抑制血管损伤后的IH。其可能机制是抑制了ECs中的Akt/mTOR信号通路。
Objective To explore the mechanism of curcumin(Cur) in enhancing the level of autophagy inside endothelial cells(ECs) and inhibiting damaged vascular intimal hyperplasia(IH). Methods In vitro, Human Umbilical Vein Endothelial Cell(HUVECs) was divided into four groups: Control, 3-MA, Cur and Rapa group. Acridine orange(AO) staining and Lyso-Tracker Red method were used to observe the acidic compartment accumulation. Western-blot and immunofluorescence were used to evaluate the protein levels of microtubule-associated protein-Ⅱ(LC3-Ⅱ), p62, Akt and mTOR. In vivo, rat carotid artery endarterium injury model was created by balloon compression, and 60 SD rats were divided into Sham, injured+vehicle, 3-MA, Cur and Rapa group. The protein levels of LC3-Ⅱ, p62, Akt and mTOR in carotid artery tissue were detected by Western blot. HE staining was used to measure the IH thickness and effective lumen area of each group. Results In vitro, Cur and Rapa treatments significantly increased the acidic compartment accumulation, the protein level of LC3-Ⅱ and inhibited the protein expression of p62,Akt and mTOR in HUVECs compared with control group. In vivo, protein level of LC3-Ⅱ markly increased and p62, Akt and mTOR decreased slightly after endometrial injury and reversed by Cur and Rapa treatments. In addition, HE staining presented obvious IH and vascular stenosis was observed after injury in 3 weeks, and Cur and Rapa both markedly suppressed IH and alleviated vascular stenosis. Finally, 3-MA treatment showed an opposite effect both in vitro and in vivo. Conclusion Cur increases the level of ECs autophagy and inhibits IH after vascular injury, possibly by inhibiting the AKt/mTOR signaling pathway in ECs.
作者
汪洋
董永飞
鲍得俊
程传东
齐印宝
魏祥品
牛朝诗
WANG Yang;DONG Yong-fei;BAO De-jun(Department of Neurosurgery,The First Affiliated Hospital of University of Science and Technology of China(Anhui Provincial Hospital),Hefei 230001,China)
出处
《临床神经外科杂志》
CAS
2019年第5期390-396,共7页
Journal of Clinical Neurosurgery
基金
安徽省自然科学基金青年项目(1708085QH174)
