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Astragaloside IV alleviates atherosclerosis through targeting circ_0000231/miR-135a-5p/CLIC4 axis in AS cell model in vitro

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Abstract

Non-coding RNAs (ncRNAs) have shown to act as crucial mediators in atherosclerosis (AS) development. The purpose of our study was to explore the role of Astragaloside IV (ASV) and circular RNA_0000231 (circ_0000231) in AS using AS cell model. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry were used to analyze cell viability and apoptosis. Migration ability was assessed by transwell migration assay and wound healing assay. The inflammatory response was evaluated via enzyme-linked immunosorbent assay (ELISA). Oxidative status was assessed via matching commercial kits. Western blot assay was conducted to detect the expression of monocyte chemoattractant protein 1 (MCP1), intercellular adhesion molecule 1 (ICAM1), and chloride intracellular channel 4 (CLIC4). The levels of circ_0000231, its linear form Rho GTPase activating protein 12 (ARHGAP12), microRNA-135a-5p (miR-135a-5p), and CLIC4 messenger RNA (mRNA) were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Circ_0000231–miRNA interactions were established using Starbase and Circbank softwares, while the targets of miR-135a-5p were explored by Starbase software. Dual-luciferase reporter assay and RNA-pull down assay were used to verify these target interactions. ASV suppressed the apoptosis, inflammation, and oxidative stress while recovered the viability and migration ability of HUVECs which were mediated by oxidized low-density lipoprotein (ox-LDL). Circ_0000231 overexpression antagonized the protective role of ASV in ox-LDL-induced HUVECs. MiR-135a-5p was verified as a direct target of circ_0000231, and circ_0000231 contributed to ox-LDL-induced cell injury of HUVECs through down-regulating miR-135a-5p. MiR-135a-5p directly interacted with the 3′ untranslated region (3′-UTR) of CLIC4 mRNA in HUVECs, and miR-135a-5p protected HUVECs against ox-LDL-induced injury through down-regulating CLIC4. ASV protected HUVECs against ox-LDL-induced injury through targeting circ_0000231/miR-135a-5p/CLIC4 axis. Targeting circ_0000231/miR-135a-5p/CLIC4 axis might provide a novel insight to develop effective strategy for AS treatment.

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Abbreviations

ASV:

Astragaloside IV

AS:

Atherosclerosis

Non-coding:

RNAs

ELISA:

Enzyme-linked immunosorbent assay

MCP1:

Monocyte chemoattractant protein 1

ICAM1:

Intercellular adhesion molecule 1

CLIC4:

Chloride intracellular channel 4

ox-LDL:

Oxidized low-density lipoprotein

ROS:

Reactive oxygen species

SOD:

Superoxide dismutase

MDA:

Malondialdehyde

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Authors and Affiliations

  1. Changchun University of traditional Chinese medicine, Changchun city, Jilin Province, China

    Xiao Shao

  2. Department of Cardiology, Affiliated Hospital of Changchun University of traditional Chinese medicine, Changchun city, Jilin Province, China

    Zhaozheng Liu & Yue Deng

  3. Department of Pediatrics, The Second Hospital of Fushun, Fushun city, Liaoning Province, China

    Shanshan Liu

  4. Department of Cardiology and rehabilitation, Affiliated Hospital of Changchun University of traditional Chinese medicine, Changchun city, Jilin Province, China

    Na Lin

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  1. Xiao Shao
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Correspondence to Yue Deng.

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Shao, X., Liu, Z., Liu, S. et al. Astragaloside IV alleviates atherosclerosis through targeting circ_0000231/miR-135a-5p/CLIC4 axis in AS cell model in vitro. Mol Cell Biochem 476, 1783–1795 (2021). https://doi.org/10.1007/s11010-020-04035-8

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