微小RNA-142-3p靶向Rictor诱导动脉粥样硬化相关的内皮细胞凋亡

MicroRNA-142-3p modulates atherosclerosis-associated endothelial cell apoptosis via targeting Rictor

目的:探讨微小RNA-142-3p(miR-142-3p)对动脉粥样硬化(atherosclerosis,AS)进程中人主动脉内皮细胞(human aortic endothelial cells,HAECs)凋亡的影响及作用机制。方法:HAECs经氧化型低密度脂蛋白(oxi⁃dized low-density lipoprotein,ox-LDL)作用后,采用RT-qPCR检测miR-142-3p的表达水平。Annexin V-FITC/PI双染法流式细胞术(flow cytometry,FCM)和caspase-3活性检测试剂盒检测细胞凋亡。使用TargetScan等生物信息学软件预测miR-142-3p的潜在靶基因,双萤光素酶报告基因实验验证miR-142-3p与Rictor mRNA 3′-UTR直接靶向结合。结果:在ox-LDL诱导HAECs凋亡过程中miR-142-3p表达显著上调(P<0.05,P<0.01)。过表达miR-142-3p促进HAECs凋亡,相反,抑制miR-142-3p的表达水平可以部分程度缓解ox-LDL介导的内皮细胞(endothelial cells,ECs)凋亡。进一步研究发现,Rictor为miR-142-3p的下游靶基因,沉默Rictor基因抑制了miR-142-3p抑制物(miR-142-3p inhibitor)的抗凋亡作用。Akt/eNOS信号通路参与了miR-142-3p对ECs凋亡的调控。结论:抑制miR-142-3p表达可促进靶基因Rictor表达并激活Akt/eNOS信号通路从而发挥抗凋亡的内皮保护作用。

AIM:To investigate the role of microRNA-142-3p(miR-142-3p)in endothelial cell apoptosis dur⁃ing atherosclerosis(AS)and the underlying mechanism.METHODS:Human aortic endothelial cells(HAECs)were treated with oxidized low-density lipoprotein(ox-LDL).The expression level of miR-142-3p was detected by RT-qPCR.Apopto⁃sis was determined via flow cytometry(FCM)and caspase-3 activity assay.Prediction of the binding site between miR-142-3p and 3’-UTR of Rictor mRNA was performed by bioinformatics analysis and confirmed by dual-luciferase reporter assay.RE⁃SULTS:The expression of miR-142-3p was substantially up-regulated during the ox-LDL-elicited apoptosis in HAECs(P<0.05,P<0.01).Forced expression of miR-142-3p exacerbated apoptosis in HAECs whereas inhibition of miR-142-3p partly al⁃leviated apoptotic cell death mediated by ox-LDL.Further analysis identified Rictor as a direct target gene of miR-142-3p,and Rictor knock-down abolished the anti-apoptotic effect of miR-142-3p inhibitor.Moreover,the Akt/endothelial nitric oxide syn⁃thase(eNOS)signaling pathway was found to mediate the beneficial effect of miR-142-3p inhibitor on endothelial cells apopto⁃sis.CONCLUSION:Down-regulation of miR-142-3p inhibits endothelial cell apoptosis and atherosclerotic development by upregulating the expression of Rictor and activating the Akt/eNOS signaling pathway.