人参皂苷 Rb3 通过调控 miR-513a-5p/ZBTB20 轴减轻 ox-LDL 诱导的人主动脉内皮细胞损伤。

Ginsenoside Rb3 reduces ox-LDL-induced injury in human aortic endothelial cells by regulating the miR-513a-5p/ZBTB20 axis.

机构信息

Department of Agency Office, First Affiliated Hospital of Hunan University of Traditional Chinese Medicine, Changsha, China.

Innovation Experiment Center, First Affiliated Hospital of Hunan University of Traditional Chinese Medicine, Changsha, China.

出版信息

Adv Clin Exp Med. 2023 Nov;32(11):1291-1298. doi: 10.17219/acem/161804.

DOI:10.17219/acem/161804

PMID:37093091

Abstract

BACKGROUND

Atherosclerosis (AS) is a common vascular disease, and its main influencing factor is endothelial damage caused by oxidized low-density lipoprotein (ox-LDL). As one of the main active ingredients of ginseng, ginsenoside Rb3 has anti-inflammatory and anti-oxidative effects. However, the role of ginsenoside Rb3 in endothelial injury induced by ox-LDL is not clear.

OBJECTIVES

This study aimed to evaluate the effect and potential mechanism of ginsenoside Rb3 action on ox-LDL-treated human aortic endothelial cells (HAECs).

MATERIAL AND METHODS

The HAECs treated with ox-LDL were used to establish an in vitro AS model. The viability of the HAECs was analyzed with Cell Counting Kit-8 (CCK-8). Flow cytometry was performed to assess the apoptosis. Oxidative stress, inflammation and endothelial dysfunction were evaluated using enzyme-linked immunosorbent assay (ELISA) and western blotting. The levels of miR-513a-5p were assessed using quantitative real-time polymerase chain reaction (qPCR). A dual-luciferase assay was performed to analyze the relationship between miR-513a-5p and a zinc finger and BTB domain-containing protein (ZBTB20).

RESULTS

Exposure of HAECs to ox-LDL (50 μg/mL) reduced cell viability, superoxide dismutase (SOD) activity and endothelial nitric oxide synthase (eNOS) expression, while increasing the levels of malondialdehyde (MDA), interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-α), and soluble intercellular adhesion molecule-1 (sICAM-1). The pretreatment with Rb3 markedly enhanced cell viability and decreased ox-LDL-induced oxidative stress, inflammation and endothelial dysfunction in HAECs. The ox-LDL decreased the level of miR-513a-5p, which was reversed by Rb3 pretreatment. The ZBTB20 was a target of miR-513a-5p in HAECs, and ox-LDL upregulated ZBTB20 expression, which was reversed by Rb3 pretreatment. The protective effect of Rb3 on ox-LDL-induced HAECs was diminished by miR-513a-5p inhibition, which was reversed by ZBTB20 knockdown.

CONCLUSIONS

Ginsenoside Rb3 reduces the effects of ox-LDL on HAECs by regulating the miR-513a-5p/ZBTB20 axis, which provides a theoretical basis for the treatment of AS.

摘要

背景

动脉粥样硬化（AS）是一种常见的血管疾病，其主要影响因素是氧化型低密度脂蛋白（ox-LDL）引起的内皮损伤。人参的主要活性成分之一，人参皂苷 Rb3 具有抗炎和抗氧化作用。然而，人参皂苷 Rb3 在 ox-LDL 诱导的内皮损伤中的作用尚不清楚。

目的

本研究旨在评估人参皂苷 Rb3 对 ox-LDL 处理的人主动脉内皮细胞（HAEC）的作用及其潜在机制。

材料与方法

用 ox-LDL 处理 HAECs 建立体外 AS 模型。用细胞计数试剂盒（CCK-8）分析 HAECs 的活力。采用流式细胞术评估细胞凋亡。用酶联免疫吸附试验（ELISA）和 Western blot 评估氧化应激、炎症和内皮功能障碍。用实时定量聚合酶链反应（qPCR）评估 miR-513a-5p 的水平。采用双荧光素酶报告基因分析技术分析 miR-513a-5p 与锌指和 BTB 结构域蛋白（ZBTB20）之间的关系。

结果

ox-LDL（50μg/ml）暴露降低了 HAECs 的细胞活力、超氧化物歧化酶（SOD）活性和内皮型一氧化氮合酶（eNOS）的表达，同时增加了丙二醛（MDA）、白细胞介素 6（IL-6）、肿瘤坏死因子-α（TNF-α）和可溶性细胞间黏附分子-1（sICAM-1）的水平。Rb3 预处理显著增强了细胞活力，并降低了 ox-LDL 诱导的 HAECs 氧化应激、炎症和内皮功能障碍。ox-LDL 降低了 miR-513a-5p 的水平，而 Rb3 预处理则逆转了这一现象。ZBTB20 是 HAECs 中 miR-513a-5p 的靶标，而 ox-LDL 上调了 ZBTB20 的表达，而 Rb3 预处理则逆转了这一现象。miR-513a-5p 抑制减弱了 Rb3 对 ox-LDL 诱导的 HAECs 的保护作用，而 ZBTB20 的敲低则逆转了这一现象。