27nt-miRNA对氧化型低密度脂蛋白诱导的人脐静脉内皮细胞凋亡的影响  

Effect of 27nt-miRNA on apoptosis of human umbilical vein endothelial cells induced by oxidized low-density lipoprotein

作　　者：赵燕姣 颜渊鸳 李丹 罗雪兰 杨鹏 周福隆 杨瑞霞 唐双意[3] 欧和生 ZHAO Yan-jiao;YAN Yuan-yuan;LI Dan;LUO Xue-lan;YANG Peng;ZHOU Fulong;YANG Rui-xia;TANG Shuang-yi;OU He-sheng(Pharmaceutical College of Guangxi Medical University,Nanning 530021,China;Guangxi International Zhuang Medi-cine Hospital Affiliated to Guangxi University of Chinese Medicine,Nanning 530201,China;Department of Pharmacy,First Affiliated Hospital of Guangxi Medical University,Nanning 530021,China)

机构地区：[1]广西医科大学药学院,广西南宁530021 [2]广西中医药大学附属国际壮医医院科技部,广西南宁530201 [3]广西医科大学第一附属医院药学部,广西南宁530021

出　　处：《中国病理生理杂志》2020年第10期1745-1753,共9页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金项目资助(No.81373403);广西创新驱动发展专项基金资助项目(桂科AA17202034);广西自治区南宁市良庆区科学研究与技术开发计划课题(No.201911);广西中医药大学博士科研启动基金(No.2018BS059);广西高校中青年教师科研基础能力提升项目(No.2019KY0330)。

摘　　要：目的:探讨27nt-miRNA(27nt-miR)对氧化型低密度脂蛋白(Ox-LDL)诱导人脐静脉内皮细胞(HU⁃VECs)凋亡的影响及分子机制。方法:体外培养HUVECs,分为正常对照组、Ox-LDL组、27nt-miR+Ox-LDL组、anti-27nt-miR+Ox-LDL组和阴性对照+Ox-LDL组。正常对照组给予正常培养;Ox-LDL组用40 mg/L Ox-LDL作用48 h诱导细胞凋亡;27nt-miR+Ox-LDL组、anti-27nt-miR+Ox-LDL组及阴性对照+Ox-LDL组分别以相同的操作方法转染相应慢病毒质粒后用40 mg/L Ox-LDL作用48 h诱导细胞凋亡。采用CCK-8法检测细胞活力,划痕实验检测细胞的迁移能力,caspase-3活性试剂盒检测细胞内caspase-3活性,RT-qPCR法和Western blot法检测细胞内Bcl-2、Bax和caspase-3 mRNA及蛋白的表达情况。结果:与阴性对照+Ox-LDL组相比,27nt-miR+Ox-LDL组HUVECs的27ntmiR高表达可显著降低细胞活力和迁移能力(P<0.05),显著增加Ox-LDL所致的caspase-3活性和细胞凋亡(P<0.05),亦可显著上调Ox-LDL所致的Bax和caspase-3 mRNA和蛋白表达(P<0.05),下调Bcl-2的mRNA和蛋白表达(P<0.05);而anti-27nt-miR+Ox-LDL组,上述所测指标均表现出相反趋势。结论:27nt-miR可能通过调控Bax、cas⁃pase-3和Bcl-2凋亡/抗凋亡蛋白的表达促进Ox-LDL体外诱导的HUVECs凋亡进而抑制HUVECs活力和迁移。AIM:To investigate the effect of 27nt-miRNA(27nt-miR)on apoptosis of human umbilical vein endothelial cells(HUVECs)induced by oxidized low-density lipoprotein(Ox-LDL)and its underlying mechanism.METHODS:HUVECs were cultured in vitro and grouped as below:normal control group,Ox-LDL group,27nt-miR+Ox-LDL group,anti-27nt-miR+Ox-LDL group and negative control+Ox-LDL group.The cells in Ox-LDL group were treated with Ox-LDL at 40 mg/L for 48 h,while those in normal control group were untreated but cultured normally.The cells in 27nt-miR+Ox-LDL group,anti-27nt-miR+Ox-LDL group and negative control+Ox-LDL group were transfected with their corresponding lentiviral vectors under the same procedure,followed by treatment with Ox-LDL at 40 mg/L for 48 h to in⁃duce apoptosis.The cell viability was measured by CCK-8 assay.The migration capacity was detected by scratch assay.The caspase-3 activity was measured by caspase-3 activity assay kit.The apoptotic rate was analyzed by Hoechst 33258 and flow cytometry.The mRNA and protein expression levels of Bcl-2,Bax and caspase-3 were determined by RT-qPCR and Western blot.RESUITS:Compared with negative control+Ox-LDL group,the cell viability and migration ability were significantly decreased by over-expression of 27nt-miR in the HUVECs(P<0.05),while the activity of caspase-3 and apoptosis induced by Ox-LDL were significantly increased(P<0.05).Furthermore,the mRNA and protein expres⁃sion levels of Bax and caspase-3 were significantly up-regulated(P<0.05),and the mRNA and protein expression level of Bcl-2 was down-regulated in 27nt-miR+Ox-LDL group(P<0.05).Meanwhile,all the above indexes showed an opposite tendency in anti-27nt-miR+Ox-LDL group.CONCLUSION:27nt-miR promotes Ox-LDL-induced apoptosis and inhibits the viability and migration of HUVECs in vitro,possibly through regulating the expression of apoptotic/anti-apoptotic pro⁃teins such as Bax,caspase-3 and Bcl-2.

关 键 词：27nt-miRNA 人脐静脉内皮细胞 氧化型低密度脂蛋白 细胞凋亡 

分 类 号：R363.2[医药卫生—病理学] R329.2+5[医药卫生—基础医学]