红景天苷调控SIRT1/FoxO1通路对缺氧复氧人冠状内皮细胞的作用及机制探讨

Protective Effect and Mechanism of Salidroside on Hypoxia-Reoxygenation-Treated Human Coronary Endothelial Cells Based on SIRT1/FoxO1

目的探讨红景天苷对缺氧复氧人冠状内皮细胞(HCAEC)的保护效应及机制。方法通过缺氧复氧构建HCAEC缺血/再灌注损伤模型,给予不同剂量红景天苷(10、20、40μmol·L^(-1))进行干预。采用CCK-8法检测细胞活力,流式细胞术检测细胞凋亡和活性氧(ROS)水平,分光光度法检测超氧化物歧化酶(SOD)、谷胱甘肽(GSH)活性、丙二醛(MDA)含量,蛋白免疫印迹法检测二价金属离子转运体1(DMT1)、膜铁转运蛋白(FPN)、沉默信息调节因子1(SIRT1)、叉头框转录因子1(FoxO1)乙酰化表达水平。利用siRNA沉默SIRT1后,给予高剂量红景天苷干预,检测SOD、GSH和MDA含量。结果与空白对照组比,缺氧复氧模型组中细胞活力、SOD、GSH活性和FoxO1乙酰化水平降低,细胞凋亡率、MDA含量、ROS水平、SIRT1、DMT1、FPN表达升高(P<0.05,P<0.01)。与缺氧复氧模型组比,低、中、高剂量红景天苷干预后,各组上述指标均呈现逆转,中、高剂量组差异有统计学意义(P<0.05),且具有一定的剂量依赖性(P<0.05)。SIRT1特异性siRNA明显下调SIRT1表达(P<0.01)。沉默SIRT1后,红景天苷对SOD、GSH活性的上调作用及对MDA含量的下调作用均被逆转。结论红景天苷能够通过减少细胞凋亡,改善铁代谢,并通过SIRT1/FoxO1信号通路抑制氧化应激,从而改善缺氧复氧对HCAEC的损伤。

Objective To explore the protective effect and mechanism of salidroside on hypoxia-reoxygenationtreated human coronary endothelial cells(HCAEC).Methods The HCAEC ischemia/reperfusion injury model was established by hypoxia-reoxygenation,and different doses of salidroside(10,20,40 μmol·L^(-1))were given for intervention. Cell viability was detected by CCK-8,cell apoptosis and reactive oxygen species(ROS)levels were detected by flow cytometry. Superoxide dismutase(SOD),glutathione(GSH)activity,malondialdehyde(MDA)content were detected by spectrophotometry. The acetylation levels of divalent metal ion transporter 1(DMT1),ferroportin(FPN),silent information regulator 1(SIRT1),and forkhead box transcription factor 1(FoxO1)were detected by Western Blot. After siRNA was used to silence SIRT1, a high-dose of salidroside was given to intervene,and the contents of SOD,GSH and MDA were detected.Results Compared with the control group,cell viability, SOD and GSH activities and FoxO1 acetylation level decreased in the hypoxia-reoxygenation model group,and the apoptosis rate,MDA content,ROS level,SIRT1,DMT1,FPN expression increased(P<0.05,P<0.01). Compared with the hypoxia-reoxygenation group,the above indicators in each group were reversed after the intervention of low-,medium-and high-doses of salidroside,and the difference between the medium-and high-dose groups was statistically significant(P<0.05),and there was a certain dose dependence(P<0.05).SIRT1-specific siRNA significantly down-regulated the expression of SIRT1(P<0.01). After SIRT1 was silenced,the up-regulating effect of salidroside on SOD and GSH activities and the down-regulating effect of MDA content were all reversed.Conclusion Salidroside could improve hypoxia-reoxygenation-induced damage in HCAEC by reducing apoptosis, improving iron metabolism, and inhibiting oxidative stress through SIRT1/FoxO1 signaling pathway.