Qin Bing, Xiao Bo, Jiang Ting, Yang Huan
Department of Neurology, Central South University, Changsha, China.
Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2012 Jul;37(7):675-81. doi: 10.3969/j.issn.1672-7347.2012.07.005.
To investigate the effect of miR-590-5p on the expression of lectin-like oxidized low density lipoprotein receptor 1 (LOX-1) in apoptotic human umbilical vein endothelial cells (HUVECs) induced by ox-LDL, and to explore the role of miR-590-5p in modulating HUVECs apoptosis.
HUVECs were exposed to ox-LDL (50 μg/mL) for 0 to 48 h. Apoptosis was detected by Annexin V-FITC stain and was distinguished from necrosis by propidium iodide (PI) staining. The relative expression level of miR-590-5p in HUVECs was analyzed using real-time quantitative PCR (RT-qPCR). HUVECs were transfected with miR-590-5p mimics or miRNA mimics control followed by 50 μg/mL ox-LDL stimulation for 48 h. LOX-1 mRNA and protein were measured by RT-qPCR and Western blot, and apoptosis in HUVECs was analyzed by flow ctyometry after Annexin V-FITC/PI double stain.
Incubation of HUVECs with 50 μg/mL ox-LDL for 0 to 48 h resulted in a time-dependent induction of apoptotic cell death and down-regulation of miR-590-5p. Transfection of miR-590-5p mimics suppressed LOX-1 expression at both mRNA and protein levels, leading to a reduction of ox-LDL-induced apoptosis in HUVECs.
MiR-590-5p protects endothelial cells from ox-LDL induced apoptosis by inhibiting the expression of LOX-1.
研究miR - 590 - 5p对氧化型低密度脂蛋白(ox - LDL)诱导的人脐静脉内皮细胞(HUVECs)凋亡过程中凝集素样氧化型低密度脂蛋白受体1(LOX - 1)表达的影响,并探讨miR - 590 - 5p在调节HUVECs凋亡中的作用。
将HUVECs暴露于50 μg/mL的ox - LDL中0至48小时。通过Annexin V - FITC染色检测细胞凋亡,并通过碘化丙啶(PI)染色与坏死细胞区分开来。使用实时定量PCR(RT - qPCR)分析HUVECs中miR - 590 - 5p的相对表达水平。将miR - 590 - 5p模拟物或miRNA模拟物对照转染至HUVECs,随后用50 μg/mL的ox - LDL刺激48小时。通过RT - qPCR和蛋白质印迹法检测LOX - 1的mRNA和蛋白质水平,并在Annexin V - FITC/PI双重染色后通过流式细胞术分析HUVECs中的细胞凋亡情况。
将HUVECs与50 μg/mL的ox - LDL孵育0至48小时导致凋亡细胞死亡呈时间依赖性诱导,且miR - 590 - 5p表达下调。转染miR - 590 - 5p模拟物在mRNA和蛋白质水平上均抑制了LOX - 1的表达,导致ox - LDL诱导的HUVECs凋亡减少。
MiR - 590 - 5p通过抑制LOX - 1的表达保护内皮细胞免受ox - LDL诱导的凋亡。