异常血流动力对HUVEC胆固醇代谢的影响及其致AS的机制

Effect of abnormal hemodynamics on cholesterol metabolism of HUVEC and its mechanism in inducing atherosclerosis

目的通过研究异常血流动力对人脐静脉内皮细胞(HUVEC)摄取、合成及外排胆固醇的影响,进一步探讨异常血流动力和胆固醇代谢在动脉粥样硬化(AS)形成机制中的作用。方法待细胞生长良好后,换用新的含有低密度脂蛋白(LDL)和三磷酸腺苷结合盒转运体A1(ABCA1)的培养液,同时施加作用力。按照细胞所受作用力不同,随机分为应力组、壁面压力组、对照组A和对照组B。在各自作用力的作用下培养24 h。收集细胞和培养液待用。用酶法检测4组培养液中LDL浓度和胆固醇浓度。用高效液相色谱法检测HUVEC内和细胞膜中胆固醇的含量。分别用实时定量RT-PCR法和蛋白免疫印迹法检测低密度脂蛋白受体(LDLR)、羟-甲基戊二酸单酰辅酶A还原酶(HMG-CR)、ABCA1的mRNA和蛋白表达水平。结果与对照组B相比较,应力组培养液中LDL含量明显减少,细胞内胆固醇含量明显增高,而且LDLR和HMG-CR的mRNA和蛋白表达明显增高。壁面压力组培养液中胆固醇含量明显增高,细胞膜中胆固醇含量明显降低,而且ABCA1的mRNA和蛋白表达明显增高。对照组A培养液中LDL含量明显降低,胆固醇含量明显增高,而且LDLR、HMG-CR和ABCA1的mRNA和蛋白表达均明显增高。结论应力通过上调LDLR和HMG-CR的mRNA和蛋白的表达促进VEC摄取LDL和合成胆固醇,壁面压力通过上调ABCA1的mRNA和蛋白的表达促进内皮细胞膜外排胆固醇。异常血流动力使血管内皮细胞胆固醇代谢平衡紊乱进而引起AS。

Objective To investigate the effects of abnormal hemodynamics on the uptake,synthesis and efflux of cholesterol in human umbilical vein endothelial cells(HUVECs)and explore the role of abnormal hemodynamics and cholesterol metabolism in the formation of atherosclerosis.Methods When the cells grew well,a new culture solution containing low-density lipoprotein(LDL)and ATP binding cassette transporter A1(ABCA1)was used,and pressure was added.According to the pressure used,the cells were randomly divided into four groups:a stress group,a wall pressure group,a control group A and a control group B.They were cultured for 24 h under corresponding pressures.Then,the cells and culture media were collected for later use.The LDL and cholesterol concentrations in the culture media of the four groups were detected by enzymatic method.The content of intracellular cholesterol and the content of cholesterol within cell membrane were determined by HPLC.The expression of low density lipoprotein receptor(LDLR),3-hydroxy-3-methylglutaryl-CoA reductase(HMG-CR)and ABCA1 mRNA and protein were detected by real-time quantitative RT-PCR and Western blot,respectively.Results Compared with the control group B,the content of LDL in the culture media of the stress group significantly de-creased and the content of intracellular cholesterol increased.Moreover,the expression of LDLR and HMG-CR mRNA and protein significantly increased.In the wall pressure group,the cholesterol content in the culture media significantly increased,the cholesterol content within the cell membrane significantly decreased,and the expres-sion of ABCA1mRNA and protein significantly increased.In the control group A,the LDL content significantly decreased and the cholesterol content significantly increased in the culture media,and the expression of LDLR,HMG-CR and ABCA1 mRNA and protein significantly increased.Conclusions Stress promotes the uptake of LDL and cholesterol synthesis by up-regulating the expression of LDLR and HMG-CR mRNA and protein,while wall pressure promotes cholesterol efflux by up-regulating the expression of ABCA1 mRNA and protein.Abnormal he-modynamics disturbs the cholesterol metabolism balance in vascular endothelial cells and causes AS.