阿托伐他汀抑制高糖诱导的人脐静脉内皮细胞的氧化应激反应

Atorvastatin suppressed high glucose induced oxidative damage in human umbilical vein endothelial cells

目的探讨阿托伐他汀(Atv)对高糖诱导的人脐静脉内皮细胞(HUVECs)损伤的影响及保护作用、可能作用机制。方法 HUVECs低糖培养贴壁后分为对照A组;持续性高糖B0组、持续性高糖+不同浓度药物(0.1、1.0、10.0μmol/L Atv)B1~B3组;波动性高糖C组。高糖干预、药物培养24 h后检测超氧化物歧化酶(SOD)活性及一氧化氮(NO)含量;细胞计数试剂盒(CCK-8)检测细胞增殖;流式细胞仪检测细胞凋亡率;RT-PCR检测还原型烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶NOX4和NOX2/gp91phox亚基的表达。所有计量数据采用均数±标准差(sx?)表示,SPSS 17.0软件处理分析。结果 (1)与对照组相比,高糖各组细胞增殖均降低(CCK-8 D450值:B0组0.74±0.085 vs.A组1.20±0.045,P<0.01;C组0.56±0.083 vs.A组1.20±0.045,P<0.01),阿托伐他汀可改善高糖对HUVECs增殖的抑制作用,呈浓度依赖性。(2)与对照组比较,高糖组NO含量及SOD活性明显降低、NOX4 m RNA及NOX2/gp91phox m RNA的表达升高,细胞凋亡率显著增高。波动性高糖组变化趋势更明显[早期凋亡率:B0组(23.48±0.73)%vs.C组(27.33±0.89)%,P<0.01]。(3)阿托伐他汀可明显升高持续性高糖诱导的SOD活性及NO的释放,减少细胞凋亡,抑制NOX4 m RNA及NOX2/gp91phox m RNA表达的增加幅度,具有明显的浓度依赖性。结论持续性高糖与波动性高糖对内皮细胞均有损伤作用,其损伤作用可能是通过氧化应激作用来实现的,波动性高糖较持续性高糖对内皮细胞的损伤作用更大。不同浓度的Atv对持续性高糖诱导的内皮细胞损伤有保护作用,呈浓度依赖性。

Objective To explore the protective effect and the influence of atorvastatin(Atv) on the damage induced by high glucose in human umbilical vein endothelial cells(HUVECs) and its possible mechanism. Methods HUVECs were cultivated in low glucose medium and then divided into 6 experimental groups: A: control group, B0: sustained high glucose group, B1-B3: sustained high glucose group+Atv(0.1, 1.0, 10.0 μmol/L) groups, C: fluctuating high glucose group. HUVECs were further cultivated by high glucose and the drug for 24 hours. The SOD activity and NO content were measured, the cell proliferation was examined by CCK-8 kit, the rate of apoptosis was measured via flow cytometry method, subunits expression of NOX4 and NOX2/gp91 phox were measured by RT-PCR. All measuring data were described with mean±standard deviation and analyzed by SPSS 17.0. Results(1)Compared with control group, HUVECs proliferation decreased in all high glucose groups, CCK-8 D450: B0 0.74±0.085 vs. A 1.20±0.045, P<0.01; C 0.56±0.083 vs. A 1.20±0.045, P<0.01. Atv improved the proliferations while were inhibited by high glucose depending on the increasing concentrations of Atv.(2)Compared with the control group, the SOD activity and NO content significantly decreased, while apoptosis rate, the expression of NOX4 m RNA and NOX2/gp91 phox m RNA increased significantly in the high glucose groups. The tendency of the change was more obvious in the fluctuating high glucose group, the rate of early apoptosis: B0(23.48±0.73)% vs. C(27.33±0.89)% P<0.01.(3) In the condition of sustained high glucose, Atv could up-regulate SOD activity and NO content, reduced the apoptosis and down-regulated the expressions of NOX4 m RNA and NOX2/gp91 phox m RNA depending on the increasing concentrations of Atv. Conclusion Endothelial cells were injured in the groups of fluctuating high glucose and sustained high glucose, the mechanism of damage may be associated with its oxidative stress. Compared with the group of sustained high glucose, the damage was more remarkable in the fluctuating high glucose group. Atorvastatin in different concentration could protect endothelial cell against injury induced by sustained high glucose.