环状 RNA 0000231 通过调控 miR-590-5p/PDCD4 轴抑制氧化型低密度脂蛋白诱导的人脐静脉内皮细胞凋亡、自噬及炎症反应。

Inhibition of circ_0000231 suppresses oxidized low density lipoprotein-induced apoptosis, autophagy and inflammation in human umbilical vein endothelial cells by regulating miR-590-5p/PDCD4 axis.

机构信息

Department of Cardiology, Ningbo Medical Treatment Center Lihuili Hospital, Ningbo University, Ningbo, China.

Department of Cardiology, Zhejiang Provincial People's HospitalHangzhou, China.

出版信息

Clin Hemorheol Microcirc. 2024;87(3):283-299. doi: 10.3233/CH-231696.

DOI:10.3233/CH-231696

PMID:37066904

Abstract

BACKGROUND

Circular RNAs (circRNAs) are the emerging informative RNAs, involved in cardiovascular diseases including atherosclerosis (AS). Endothelial injury is the initial qualitative change of AS. Thus, the objective of this study was to confirm the dysregulation and mechanism of circ_0000231 in cell model of AS at early stage in human umbilical vein endothelial cells (HUVECs) induced by oxidized low-density lipoprotein (ox-LDL).

METHODS

The expression of circ_0000231, miR-590-5p and programmed cell death 4 (PDCD4) was detected using real-time quantitative PCR and western blot. Cell injury was measured with MTT, flow cytometry, caspase-3 activity assay and enzyme-linked immunosorbent assay (ELISA). The interaction among circ_0000231, miR-590-5p and PDCD4 was validated by dual-luciferase reporter assay, RNA immunoprecipitation (RIP) and pull-down assays.

RESULTS

Stress ox-LDL decreased cell viability, and increased apoptosis rate and caspase-3 activity in HUVECs in a dose- and time-dependent manner in concomitant with promotions of interleukin-6, interleukin-1β, tumor necrosis factor-α, LC3-II/I and Beclin-1 levels. Besides, circ_0000231 and PDCD4 expressions were upregulated, and miR-590-5p was downregulated in ox-LDL-stimulated HUVECs. Functionally, knockdown of circ_0000231 and overexpression of miR-590-5p could suppress ox-LDL-elicited above effects on apoptosis, autophagy and inflammatory response, accompanied with PDCD4 downregulation. Physically, miR-590-5p could directly interact with circ_0000231 and PDCD4.

CONCLUSION

Downregulation of circ_0000231 suppresses HUVECs from ox-LDL-induced injury partially through regulating miR-590-5p/PDCD4 axis via competing endogenous RNA mechanism, showing a novel potential target for the pathology and treatment of endothelial injury in AS.

摘要

背景

环状 RNA（circRNAs）是新兴的信息 RNA，参与包括动脉粥样硬化（AS）在内的心血管疾病。内皮损伤是 AS 的初始质变。因此，本研究的目的是在人脐静脉内皮细胞（HUVEC）中，在氧化低密度脂蛋白（ox-LDL）诱导的 AS 早期细胞模型中，证实 circ_0000231 的失调及其机制。

方法

采用实时定量 PCR 和 Western blot 检测 circ_0000231、miR-590-5p 和程序性细胞死亡因子 4（PDCD4）的表达。用 MTT、流式细胞术、caspase-3 活性测定和酶联免疫吸附试验（ELISA）测定细胞损伤。采用双荧光素酶报告基因检测、RNA 免疫沉淀（RIP）和下拉实验验证 circ_0000231、miR-590-5p 和 PDCD4 之间的相互作用。

结果

应激 ox-LDL 呈剂量和时间依赖性地降低 HUVECs 的细胞活力，并增加细胞凋亡率和 caspase-3 活性，同时促进白细胞介素-6、白细胞介素-1β、肿瘤坏死因子-α、LC3-II/I 和 Beclin-1 水平升高。此外，ox-LDL 刺激的 HUVECs 中 circ_0000231 和 PDCD4 的表达上调，miR-590-5p 的表达下调。功能上，circ_0000231 敲低和 miR-590-5p 过表达可抑制 ox-LDL 诱导的细胞凋亡、自噬和炎症反应，同时下调 PDCD4 表达。实际上，miR-590-5p 可以直接与 circ_0000231 和 PDCD4 相互作用。