晚期糖基化终产物诱导人主动脉内皮细胞氧化应激损伤及线粒体功能紊乱

Advanced glycation end products induce oxidative stress and mitochondrial dysfunction in human aortic endothelial cells

目的:观察晚期糖基化终产物(advanced glycation end products,AGEs)对内皮细胞氧化应激水平和线粒体功能的影响,以探讨糖尿病血管内皮细胞功能障碍可能的发生机制。方法:以不同质量浓度(50,100μg/mL)的AGE修饰的牛血清白蛋白(AGE-bovine serum albumin,AGE-BSA)作用于人主动脉内皮细胞(human aortic endothelial cells,HAECs)48 h,CCK-8法测定HAECs的增殖能力,黄嘌呤氧化酶法测定超氧化物歧化酶(superoxide dismutase,SOD)活力,应用DCFH-DA探针检测细胞内活性氧(reactive oxygen species,ROS)的水平;JC-1法检测线粒体膜电位(mitochondrial membrane potential,MMP),荧光素-荧光素酶法和Clark氧电极法测定细胞中ATP含量及细胞耗氧率。结果:AGE-BSA能显著性地抑制HAECs增殖,增加细胞内ROS水平,降低SOD活力,且高质量浓度作用更加明显。同时,AGE-BSA还能使MMP下降、ATP生成及耗氧减少。结论:AGE-BSA诱导HAECs产生氧化应激损伤,其机制与其造成线粒体功能紊乱相关。

Objective:To investigate whether advanced glycation end products (AGEs) can induce cell injury and mitochondrial dysfunction in human aortic endothelial cells (HAECs).Methods:HAECs were treated with increasing concentrations (50,1 00μg/mL) of AGE-bovine serum albumin (AGE-BSA) for 48 h.The proliferative inhibition of HAECs was measured by CCK-8 method.Contents of ATP and activity of superoxide dismutase (SOD) were determined by the luciferase assay and SOD kits.Reactive oxygen species (ROS) were determined by DCFH-DA staining.Mitochondrial membrane potential (MMP) was observed with JC-1 staining.Oxygen utilization was measured polarographically with a Clark oxygen electrode.Results:Compared with control group,AGE-BSA (50,100 μg/mL) significantly reduced HAECs proliferation.AGE-BSA significantly increased the levels of ROS,while decreased the activity of SOD compared to the control group.Importantly,AGE-BSA-mediated oxidative stress was followed by a collapse of mitochondrial membrane potential (MMP),the inhibition of ATP generation,and the down-regulation of oxygen utilization.Conclusion:AGE-BSA induced oxidative stress in HAECs.Moreover,AGE-BSA-induced HAECs injury was related to mitochondrial dysfunction.