氯喹对脂多糖诱导的人脐静脉内皮细胞损伤的影响  

The dose dependent bidirectional effect of chloroquine on lipopolysaccharide-induced injury to human umbilical vein endothelial cells

作　　者：王宁[1] 朱莹莹[1] 张姣姣 周罗成[1] 康健[1] 王海嵘[1] WANG Ning;ZHU Yingying;ZHANG Jiaojiao;ZHOU Luocheng;KANG Jian;WANG Hairong(Department of Emergency,Xinhua Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai 200092,China)

机构地区：[1]上海交通大学医学院附属新华医院急诊医学科,上海200092

出　　处：《内科理论与实践》2020年第2期111-115,共5页Journal of Internal Medicine Concepts & Practice

基　　金：上海市卫生计生系统重要薄弱学科建设计划(项目编号:2016ZB0203);上海市卫生和计划生育委员会科研课题(项目编号:201540110);国家自然科学基金青年科学基金项目(项目编号:81601658)。

摘　　要：目的:探究氯喹(chloroquine,CQ)对脂多糖(lipopolysaccharide,LPS)诱导的人脐静脉内皮细胞(human umbilical vein endothelial cell,HUVEC)损伤的作用及机制。方法:用对数生长期的HUVEC进行实验,分为5组,对照组细胞不经药物处理,LPS组用10μg/mL LPS刺激细胞,低、中、高浓度CQ组用10μg/mL LPS和低、中、高浓度CQ(0.1、1和10μmol/L)共同作用细胞。培养24 h后用细胞计数试剂盒(cell counting kit-8,CCK-8)检测细胞活力,用流式细胞仪检测细胞凋亡水平,蛋白质印迹法检测BCL-2、BAX、裂解的胱天蛋白酶3(cleaved caspase 3)、微管相关蛋白1轻链3(microtubule-associated protein 1 light chain 3,LC3)蛋白的表达。结果:LPS刺激细胞后导致HUVEC活力降低、凋亡增加,同时BCL-2/BAX蛋白比值降低,裂解的胱天蛋白酶3蛋白表达增高(P<0.05),LC3-Ⅱ/LC3-Ⅰ比值无明显变化(P>0.05);加入CQ作用后,低、中浓度CQ改善了LPS所致的细胞损伤,表现为细胞活力升高和细胞凋亡减少、BCL-2/BAX蛋白比值升高、裂解的胱天蛋白酶3蛋白表达降低(均P<0.05),LC3-Ⅱ/LC3-Ⅰ比值仍无明显变化(P>0.05),而高浓度CQ对LPS所致的细胞活力抑制和细胞凋亡无明显改善作用,BCL-2/BAX蛋白比值无明显变化(P>0.05),裂解的胱天蛋白酶3蛋白表达及LC3-Ⅱ/LC3-Ⅰ比值则有所升高(P<0.05)。结论:低、中浓度CQ可通过抑制细胞凋亡途径减轻LPS对HUVEC造成的损伤,低、中浓度CQ对于自噬没有影响;高浓度CQ可通过激活自噬、促进细胞凋亡途径加重LPS对HUVEC的损伤。Objective To investigate the effects of chloroquine(CQ)on the injury of human umbilical vein endothelial cells(HUVECs)induced by lipopolysaccharide(LPS)and its potential mechanism.Methods HUVECs in the logarithmic growth phase were stimulated with 10μg/mL LPS and treated with CQ of various concentration,0.1,1 and 10μmol/L.After 24 h,the cell viability was assessed by cell counting with kit-8(CCK-8)assay,cell apoptosis was measured by flow cytometry and the expression of BCL-2,BAX,cleaved caspase 3 and microtubule-associated protein 1 light chain 3(LC3)were examined by Western blotting.Results The LPS stimulation impaired the viability of HUVECs and induced apoptosis.Consistently,LPS was found to decrease the ratio of BCL-2/BAX and increased expression of cleaved caspase(P<0.05),however,it had no impact on LC3-Ⅱ/LC3-Ⅰratio(P>0.05).The CQ ameliorated cell injury inflicted by LPS at low and medium concentrations(P<0.05),improving the viability of HUVECs and suppressing the apoptosis.The low and medium dose CQ treatment increased ratio of BCL-2/BAX and inhibited expression of cleaved caspase 3,but had negligible impact on the expression of LC3(P>0.05).Interestingly,CQ at high concentration could not rescue the HUVECs from LPS induced apoptosis and had no significant impact on the cell viability,the BCL-2/BAX ratio(P>0.05),instead,it elevated the expression of cleaved caspase 3 and the ratio of LC3-Ⅱ/LC3-Ⅰ(P<0.05).Conclusions CQ at low concentrations could attenuate the LPS induced HUVECs injury by suppressing autophagy independent apoptosis,while CQ at high concentrations might enhance apoptosis and aggravate cell injury through autophagy activation.

关 键 词：氯喹 脂多糖 人脐静脉内皮细胞 凋亡 

分 类 号：R631[医药卫生—外科学]