Notoginsenoside R1 alleviates oxidized low-density lipoprotein-induced apoptosis, inflammatory response, and oxidative stress in HUVECS through modulation of XIST/miR-221-3p/TRAF6 axis.,Cellular Signalling

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Notoginsenoside R1 alleviates oxidized low-density lipoprotein-induced apoptosis, inflammatory response, and oxidative stress in HUVECS through modulation of XIST/miR-221-3p/TRAF6 axis.
Cellular Signalling ( IF 4.8 ) Pub Date : 2020-09-15 , DOI: 10.1016/j.cellsig.2020.109781
Jian Zhao ₁ , Lin Cui ₂ , Jing Sun ₁ , Zhouliang Xie ₁ , Leilei Zhang ₁ , Zhiwei Ding ₁ , Xiaoqiang Quan ₁

Affiliation

Background:

Atherosclerosis is a common reason for acute cardio-cerebral vascular diseases. The purpose of this study was to clarify the functional effects of Notoginsenoside R1 (NGR1) on atherosclerosis.

Methods

HUVECS were exposed to oxidized low-density lipoprotein (ox-LDL) in the current study. The proliferation ability of HUVECS was measured by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-2H-tetrazol-3-ium bromide (MTT). Flow cytometry assays were performed to evaluate the apoptosis of HUVECS. Ox-LDL caused inflammatory response and oxidative stress were assessed by checking pro-inflammatory cytokines and intracellular reactive oxygen species (ROS), malondialdehyde (MDA), and superoxide dismutase (SOD). The regulatory roles of NGR1 in HUVECs were measured by real-time quantitative polymerase chain reaction (RT-qPCR) and western blot assays. The interaction relationship between miR-221-3p and X-inactive specific transcript (XIST) or TNF-receptor-associated factor 6 (TRAF6) was predicted by bioinformatics tools. Dual-luciferase reporter and RNA pull-down assays were used to confirm the interaction relationship.

Results

Currently, ox-LDL inhibited proliferation and induced apoptosis, inflammatory response, and oxidative stress in HUVECs, which were alleviated by treatment with NGR1. Importantly, the increase of XIST in ox-LDL-induced HUVECs was abolished by NGR1. In addition, the gain-of-functional experiment suggested that the upregulation of XIST neutralized the protection effects of NGR1 in HUVECs treated with ox-LDL. In addition, miR-221-3p was a target of XIST in HUVECs as confirmed by dual-luciferase reporter and RNA pull-down assays. Furthermore, miR-221-3p interacted with TRAF6, and NGR1 regulated proliferation, apoptosis, inflammatory response, and oxidative stress in HUVECs exposed to ox-LDL by regulation of the XIST/miR-221-3p/TRAF6 axis through Nuclear Factor Kappa B (NF-κB) pathway.

Conclusion

NGR1 could exert regulatory functions in ox-LDL-induced HUVECS by regulation of XIST/miR-221-3p/TRAF6 axis, which provided valuable insights into the development of potential therapeutic strategy for atherosclerosis.

中文翻译：

三七皂苷 R1 通过调节 XIST/miR-221-3p/TRAF6 轴减轻 HUVECS 中氧化低密度脂蛋白诱导的细胞凋亡、炎症反应和氧化应激。

背景：

动脉粥样硬化是急性心脑血管疾病的常见原因。本研究的目的是阐明三七皂苷 R1 (NGR1) 对动脉粥样硬化的功能影响。

方法

在当前的研究中，HUVECS 暴露于氧化低密度脂蛋白 (ox-LDL)。HUVECS的增殖能力通过3-(4, 5-二甲基噻唑-2-基)-2, 5-二苯基-2H-四唑-3-鎓溴化物(MTT)测定。进行流式细胞术测定以评估 HUVECS 的细胞凋亡。通过检查促炎细胞因子和细胞内活性氧 (ROS)、丙二醛 (MDA) 和超氧化物歧化酶 (SOD) 来评估 Ox-LDL 引起的炎症反应和氧化应激。NGR1 在 HUVEC 中的调节作用通过实时定量聚合酶链反应 (RT-qPCR) 和蛋白质印迹分析进行测量。通过生物信息学工具预测 miR-221-3p 与 X 失活特异性转录物 (XIST) 或 TNF 受体相关因子 6 (TRAF6) 之间的相互作用关系。

结果

目前，ox-LDL 抑制 HUVECs 的增殖并诱导细胞凋亡、炎症反应和氧化应激，这些通过 NGR1 治疗得到缓解。重要的是，NGR1 消除了 ox-LDL 诱导的 HUVEC 中 XIST 的增加。此外，功能获得实验表明 XIST 的上调中和了 NGR1 对 ox-LDL 处理的 HUVEC 的保护作用。此外，双荧光素酶报告基因和 RNA 下拉分析证实，miR-221-3p 是 HUVEC 中 XIST 的靶标。此外，miR-221-3p 与 TRAF6 相互作用，NGR1 通过核因子 Kappa B 调节 XIST/miR-221-3p/TRAF6 轴来调节暴露于 ox-LDL 的 HUVEC 的增殖、凋亡、炎症反应和氧化应激(NF-κB) 通路。