Circ_0068087 Silencing Ameliorates Oxidized Low-Density Lipoprotein-Induced Dysfunction in Vascular Endothelial Cells Depending on miR-186-5p-Mediated Regulation of Roundabout Guidance Receptor 1.

Shuanghong Li,Tao Huang,Luchang Yin,Limin Qin

doi:10.3389/fcvm.2021.650374

Abstract

Background: 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.Methods: 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).Results: 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.Conclusion: Circ_0068087 interference alleviated ox-LDL-induced dysfunction in HUVECs partly by reducing ROBO1 expression via upregulating miR-186-5p.

Highlights

Atherosclerosis (AS) is a chronic inflammatory disorder that contributes to the progression of various cardiovascular diseases (CVDs)
We find that Oxidized low-density lipoprotein (ox-LDL) exposure reduced the viability of Human umbilical vein endothelial cells (HUVECs) in a dose- and time-dependent manner (Figures 1A,B). ox-LDL (40 mg/L; 24 h) was chosen for further experiments. ox-LDL treatment significantly upregulated circ_0068087 expression in HUVECs (Figure 1C)
Cell apoptosis was triggered by ox-LDL stimulation, and the interference of circ_0068087 restrained the apoptosis of oxLDL-induced HUVECs (Figure 2C)

Summary

Introduction

Atherosclerosis (AS) is a chronic inflammatory disorder that contributes to the progression of various cardiovascular diseases (CVDs). There are three mainstream theories regarding the pathogenesis of AS, including the lipid infiltration theory, endothelial injury theory, and vascular smooth muscle cell migration/proliferation theory [2]. Endothelial cells are widely distributed in vascular networks, and endothelial dysfunction is identified as a vital factor in AS pathogenesis [3]. Oxidized low-density lipoprotein (ox-LDL) is a critical inducer for the dysfunction of endothelial cells [4, 5]. We explored the mechanism underlying the aberrant transformation of endothelial cells in ox-LDL-induced AS cell model. 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

Methods

Results

Conclusion