Fu Wei, Liu Zidong, Zhang Jing, Shi Yuxue, Zhao Ruiyao, Zhao Heng
Department of Cardiology, The Third Affiliated Hospital of Jinzhou Medical University, Jinzhou, Liaoning 121000, P.R. China.
Exp Ther Med. 2020 Mar;19(3):1817-1823. doi: 10.3892/etm.2019.8369. Epub 2019 Dec 23.
The purpose of the present study was to investigate the effect of microRNA (miR)-144-5p on human umbilical vein endothelial cells (HUVECs) to explore the role of miR-144-5p in atherosclerosis. miR-144-5p expression was upregulated in HUVECs using miR-144-5p mimics. The relative expression level of miR-144-5p in HUVECs was detected using reverse transcription-quantitative PCR (RT-qPCR). Cell proliferation was detected by performing an MTT assay. Apoptosis was determined via flow cytometry. Cell migration ability was detected by a wound-healing assay. Cell invasion was determined by a transwell assay. The protein levels of phosphorylated (p)-PI3K, p-Akt and endothelial nitric oxide synthase (eNOS) were detected using western blot analysis. The binding sites between miR-144-5p and 3'-untranslated region of rapamycin-insensitive companion of mTOR (RICTOR) mRNA were predicted by TargetScan and confirmed by a dual luciferase reporter assay. The present study showed that miR-144-5p mimics significantly inhibited cell proliferation and induced apoptosis in HUVECs. In addition, miR-144-5p mimics could suppress migration and invasion of HUVECs. Further analysis identified that RICTOR was a direct target gene of miR-144-5p. Moreover, miR-144-5p upregulation decreased the protein level of p-PI3K, p-Akt and eNOS. In conclusion, miR-144-5p regulated HUVEC proliferation, migration, invasion, and apoptosis through affecting the PI3K-Akt-eNOS signaling pathway by altering the expression of RICTOR. These results indicated that miR-144-5p may be a potential target for the prevention and treatment of atherosclerosis.
本研究旨在探讨微小RNA（miR）-144-5p对人脐静脉内皮细胞（HUVECs）的影响，以探究miR-144-5p在动脉粥样硬化中的作用。使用miR-144-5p模拟物上调HUVECs中miR-144-5p的表达。采用逆转录定量PCR（RT-qPCR）检测HUVECs中miR-144-5p的相对表达水平。通过MTT法检测细胞增殖。通过流式细胞术测定细胞凋亡。通过伤口愈合试验检测细胞迁移能力。通过Transwell试验测定细胞侵袭能力。采用蛋白质印迹分析检测磷酸化（p）-PI3K、p-Akt和内皮型一氧化氮合酶（eNOS）的蛋白水平。通过TargetScan预测miR-144-5p与雷帕霉素不敏感的mTOR伴侣（RICTOR）mRNA的3'-非翻译区之间的结合位点，并通过双荧光素酶报告基因试验进行验证。本研究表明，miR-144-5p模拟物显著抑制HUVECs的细胞增殖并诱导其凋亡。此外，miR-144-5p模拟物可抑制HUVECs的迁移和侵袭。进一步分析确定RICTOR是miR-144-5p的直接靶基因。此外，miR-144-5p上调降低了p-PI3K、p-Akt和eNOS的蛋白水平。总之，miR-144-5p通过改变RICTOR的表达影响PI3K-Akt-eNOS信号通路，从而调节HUVECs的增殖、迁移、侵袭和凋亡。这些结果表明，miR-144-5p可能是预防和治疗动脉粥样硬化的潜在靶点。
