Li Shuanghong, Huang Tao, Qin Limin, Yin Luchang
Department of Emergency, Weifang Hospital of Traditional Chinese Medicine, Weifang, China.
Department of Cardiovascular Medicine, Affiliated Hospital of Weifang Medical University, Weifang, China.
Front Cardiovasc Med. 2021 May 26;8:650374. doi: 10.3389/fcvm.2021.650374. eCollection 2021.
Circular RNAs (circRNAs) are endogenous non-coding RNAs involved in the progression of atherosclerosis (AS). We investigated the role of circ_0068087 in AS progression and its associated mechanism. The 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay, flow cytometry, and enzyme-linked immunosorbent assay (ELISA) were performed to analyze the viability, apoptosis, and inflammatory response of HUVECs, respectively. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and the Western blot assay were performed to measure the expression of RNA and protein. Cell oxidative stress was analyzed using commercial kits. The dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were conducted to verify the interaction between microRNA-186-5p (miR-186-5p) and circ_0068087 or roundabout guidance receptor 1 (ROBO1). Oxidized low-density lipoprotein (ox-LDL) exposure upregulated the circ_0068087 level in HUVECs. ox-LDL-induced dysfunction in HUVECs was largely attenuated by the silence of circ_0068087. Circ_0068087 negatively regulated the miR-186-5p level by interacting with it in HUVECs. Circ_0068087 knockdown restrained ox-LDL-induced injury in HUVECs partly by upregulating miR-186-5p. ROBO1 was a downstream target of miR-186-5p in HUVECs. Circ_0068087 positively regulated ROBO1 expression by sponging miR-186-5p in HUVECs. MiR-186-5p overexpression exerted a protective role in ox-LDL-induced HUVECs partly by downregulating ROBO1. Circ_0068087 interference alleviated ox-LDL-induced dysfunction in HUVECs partly by reducing ROBO1 expression via upregulating miR-186-5p.
环状RNA（circRNAs）是参与动脉粥样硬化（AS）进展的内源性非编码RNA。我们研究了circ_0068087在AS进展中的作用及其相关机制。分别采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐（MTT）法、流式细胞术和酶联免疫吸附测定（ELISA）分析人脐静脉内皮细胞（HUVECs）的活力、凋亡和炎症反应。采用逆转录定量聚合酶链反应（RT-qPCR）和蛋白质免疫印迹法检测RNA和蛋白质的表达。使用商业试剂盒分析细胞氧化应激。进行双荧光素酶报告基因测定和RNA免疫沉淀（RIP）测定以验证微小RNA-186-5p（miR-186-5p）与circ_0068087或轴突导向受体1（ROBO1）之间的相互作用。氧化型低密度脂蛋白（ox-LDL）暴露上调了HUVECs中circ_0068087的水平。circ_0068087的沉默在很大程度上减轻了ox-LDL诱导的HUVECs功能障碍。在HUVECs中，circ_0068087通过与miR-186-5p相互作用负向调节其水平。circ_0068087基因敲低部分通过上调miR-186-5p抑制ox-LDL诱导的HUVECs损伤。ROBO1是HUVECs中miR-186-5p的下游靶点。在HUVECs中，circ_0068087通过吸附miR-186-5p正向调节ROBO1表达。miR-186-5p过表达部分通过下调ROBO1对ox-LDL诱导的HUVECs发挥保护作用。circ_0068087干扰部分通过上调miR-186-5p降低ROBO1表达减轻ox-LDL诱导的HUVECs功能障碍。
