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
Introduction
Atherosclerosis, as an inflammatory disease, is the main contributor to cardiovascular diseases-related vascular diseases, leading to myocardial infarction, stoke, and heart failure [1]. Inflammation response and infiltration of leukocytes were major characterization at whole stages of the atherosclerotic process [2]. Consistently, high levels of inflammatory cytokines were closely correlated with plaque progression in atherosclerosis [3]. Previous studies well clarified that Notoginsenoside R1 (NGR1) has beneficial effects on animal models of cardiovascular diseases [[4], [5], [6]]. Importantly, NGR1, a major active ingredient of Radix notoginseng, was reported to alleviate oxidized low-density lipoprotein (ox-LDL)-caused inflammatory response by decreasing the cytokines production in endothelial cells through modulation of Nuclear Factor Kappa B (NF-κB) signal path [7], while ox-LDL was reported to be an important risk factor contributing to the progression of atherosclerosis [8]. However, the detailed mechanism by which NGR1 involved in atherosclerosis development remained largely elusive.
It was widely accepted that long non-coding RNAs (lncRNAs, longer than 200 nucleotides) act predominant roles in various pathologies, including atherosclerosis [9,10]. For instance, lncRNA H19 was increased in the blood sample of atherosclerosis compared with control, and it could be a potential target for treating atherosclerosis [11]. The lncRNA X-inactive specific transcript (XIST) was transcribed from the XIST gene and locate on Xq13.2 (1–19,296). According previous reports, XIST was closely related to the development of many human cancers, including pancreatic cancer and breast cancer [12,13]. In addition, dysregulation of XIST could act as a biomarker for poor prognosis of cancer patients [14]. Nevertheless, the expression pattern and function of XIST in atherosclerosis were scarcely documented.
Furthermore, microRNAs (miRNAs) have been demonstrated as vital regulators in atherosclerosis [15]. Moreover, miR-221-3p has attracted attention for its prognostic value in multiple human diseases, such as colon cancer [16], triple-negative breast cancer [17], and acute myocardial infarction [18]. Furthermore, research by Xue et al. claimed that miR-221 was involved in the dysfunction of endothelial cells [19]. Nevertheless, the regulatory of miR-221 in the pathogenesis of atherosclerosis still require further investigation.
TNF-receptor-associated factor 6 (TRAF6), a member of TNF-receptor-associated factor family, mediated multiple signaling signals transduction, including TNF receptor (TNFR) family, IL-18 receptor (IL-18R), and Toll-like receptors [20]. A study indicated that TRAF6 participated in the development of atherosclerosis [21].
According previous researches, NGR1 exerted its anti-inflammation function in atherosclerosis and other inflammation-related diseases. Therefore, the investigation for the detailed mechanism by which NGR1 regulated atherosclerosis development was meaningful for atherosclerosis treatment. Thus, we explored the effects and underlying mechanism of NGR1 on apoptosis and inflammation in endothelial cells stimulated by ox-LDL.
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Cell lines
Human umbilical vein endothelial cells (HUVECs) were derived from the Chinese Academy of Sciences (Shanghai, China). HUVECs were cultured in RPMI 1640 medium (Wisent, Shanghai, China) contained with 10% (v/v) fetal bovine serum (FBS; Hyclone, South Logan, UT, USA) and 1% penicillin/streptomycin in a humidified atmosphere containing 5% CO2 at 37 °C. In addition, ox-LDL was purchased from Yiyuan Biotechnologies (Guangzhou, China). NGR1 (HPLC≥98%; Yiyuan Biotechnologies) was dissolved in dimethyl
NGR1 mitigated ox-LDL-induced apoptosis, oxidative stress, and inflammatory response in HUVECs
To begin with, ox-LDL inhibited the proliferation of HUVECs, which was weakened by treatment with 30 μM of NGR1 (Fig. 1A). Additionally, NGR1 protected HUVECs from ox-LDL-induced apoptosis (Fig. 1B). Inflammatory factors were upregulated in HUVECs after treatment with ox-LDL, including TNF-α, IL-6, and IL-1β, which was overturned in HUVECs by treatment with NGR1 (Fig. 1C-E). We also noticed that the enhancement of ROS induced by ox-LDL in HUVECs was reduced by NGR1 (Fig. 1F). Analysis results
Discussion
Although the inducement and pathogenesis mechanisms of atherosclerosis remain unclear thoroughly, injury and apoptosis of endothelial cell is a critical step during the development of atherosclerosis [23]. Therefore, ox-LDL was used to trigger the injury of endothelial cells and aggravate the process of plaque development and instability, which was widely reported in atherosclerosis-related researches [24]. As for pathogenesis mechanisms of atherosclerosis, apoptosis of vascular endothelial
Funding
This work was supported by The Project of Science and Technology of Henan Province [Grant No. 162102310031]; and The Project of Science and Technology of Zhengzhou Science and Technology Bureau [Grant No. 153PKJGG058].
Declaration of Competing Interest
The authors declare that they have no financial conflicts of interest.
Acknowledgement
None.
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Jian Zhao and Lin Cui contributed equally to this work.