胆固醇对人脐静脉内皮细胞活性氧生成及细胞凋亡的影响

Effect of cholesterol on ROS generation and cell apoptosis in human umbilical vein endothelial cells

目的:探讨胆固醇对人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)活性氧生成及细胞凋亡的影响。方法:分别用12.5,25,50,100 mg/L的胆固醇与HUVECs作用24 h,用50 mg/L胆固醇与HUVECs分别作用6,12,24,48 h,流式细胞术(FCM)检测细胞内活性氧(ROS)的生成量。分别用12.5,25,50,100 mg/L的胆固醇以及50mg/L胆固醇+N-乙酰半胱氨酸(NAC)与HUVECs孵育24 h,MTT法测定细胞活力,FCM检测细胞凋亡率。结果:12.5,25,50,100 mg/L胆固醇作用HUVECs 24 h后,均可诱导HUVECs中ROS升高,与对照组(不加胆固醇)比较差异有统计学意义(P<0.01);50 mg/L范围内组间差异明显(P<0.01),呈剂量依赖性。50 mg/L胆固醇作用HUVECs 6,12,24,48h后细胞内ROS生成量均明显高于对照组(P<0.01);24 h内组间差异明显(P<0.01),呈时间依赖性。MTT结果显示,12.5,25,50,100 mg/L胆固醇作用HUVECs 24 h后均能降低细胞的存活率,加入抗氧化剂NAC后,细胞存活率明显升高(P<0.01)。凋亡率分析结果表明,50,100 mg/L胆固醇作用HUVECs 24 h后,凋亡率比对照组明显上升(P<0.01),加入抗氧化剂NAC后凋亡率明显下降(P<0.01)。结论:胆固醇可通过诱导HUVECs中ROS的升高,促进HUVECs的凋亡。

Objective: To explore the effect of cholesterol on reactive oxygen species( ROS) generation and cell apoptosis in human vascular endothelial cells. Methods: HUVECs were treated with different cholesterol concentrations( 12. 5,25,50,100 mg / L) for 24 hours and were stimulated with 50 mg / L cholesterol at different times( 0,6,12,24,48 h). Flow cytometry detected ROS generation. HUVECs were treated with different cholesterol concentrations and Nac. HUVECs survival rate were examined by MTT colorimetric assay and apoptosis rate were examined by flow cytometry. Results: The results showed that the level of ROS will all be elevated when HUVECs were treated with 12. 5,25,50,100 mg / L cholesterol for 24 h respectively. Compared with the control group,there was a significant difference( P < 0. 01). It had a significant difference between each groups( P < 0. 01) in the range of 50 mg / L,and appeared dose-dependent manners. HUVECs were processed by 50 mg / L cholesterol for 6,12,24 and 48 hours,the level of ROS generation were significantly higher than control group( P < 0. 01); within 24 h emerge significant difference between each groups( P < 0. 01),and appeared the time-dependent manners. The results of MTT showed that12. 5,25,50,100 mg / L cholesterol could reduce the cell survival rate,and adding NAC could enhance survival rate. After HUVECs were processed by 50 mg / L or 100 mg / L cholesterol for 24 h,apoptosis rate was higher than control group( P < 0. 01),and the apoptosis rate were decline by adding antioxidants NAC.Conclusion: Cholesterol can enhance the level of ROS in HUVECs,and increase the HUVECs apoptosis.