蛭龙活血通瘀胶囊通过miR-30b-5p/NLRP3减轻氧化型低密度脂蛋白诱导的血管内皮细胞焦亡。
Zhilong Huoxue Tongyu Capsule Alleviated the Pyroptosis of Vascular Endothelial Cells Induced by ox-LDL through miR-30b-5p/NLRP3.
作者信息
Liu Mengnan, Luo Gang, Liu Tianzhu, Yang Tingfu, Wang Raoqiong, Ren Wei, Liu Ping, Lai Xiaoling, Zhou Hua, Yang Sijin
机构信息
National Traditional Chinese Medicine Clinical Research Base and Department of Cardiovascular, Hospital (T.C.M) Affiliated to Southwest Medical University, Luzhou, Sichuan, China.
Faculty of Chinese Medicine and State Key Laboratory of Quality Research in Chinese Medicine, Macau University of Science and Technology, Taipa, Macao, China.
出版信息
Evid Based Complement Alternat Med. 2022 Jan 27;2022:3981350. doi: 10.1155/2022/3981350. eCollection 2022.
BACKGROUND
Our previous studies have demonstrated a protective role of Zhilong Huoxue Tongyu capsule in atherosclerosis (AS); however, the molecular mechanisms are unclear.
METHODS
Human coronary artery endothelial cells (HCAECs) were induced with oxidized low-density lipoprotein (ox-LDL) to obtain cellular AS models. Then, the medicated serum of Zhilong Huoxue Tongyu capsule was obtained and used for treatment with ox-LDL-induced HCAECs. The cell viability was detected by CCK-8 assay. Besides, the binding between miR-30b-5p and NLRP3 was determined by the dual-luciferase reporter gene system assay. Furthermore, ox-LDL-induced HCAECs were transfected with miR-30b-5p mimic or miR-30b-5p inhibitor. The pyroptosis of HCAECs was assessed by flow cytometry, LDH content detection, and qRT-PCR assays.
RESULTS
10% medicated serum of Zhilong Huoxue Tongyu capsule was the maximum nontoxic concentration and it was used in subsequent assays. The rate of pyroptosis, LDH content, and the mRNA expression level of pyroptosis-related genes including NLRP3, ASC, Caspase 1, IL-1, and IL-18 were prominently enhanced after HCAECs were induced by ox-LDL, which were markedly rescued with medicated serum of Zhilong Huoxue Tongyu capsule. In addition, the medicated serum of Zhilong Huoxue Tongyu capsule significantly enhanced the ox-LDL-induced reduction of miR-30b-5p level. NLRP3 could bind to miR-30b-5p and was negatively corrected with miR-30b-5p. Moreover, all the rates of pyroptosis, LDH content, and the mRNA expression levels of pyroptosis-related genes including NLRP3, ASC, Caspase 1, IL-1, and IL-18 were further observably decreased after ox-LDL-induced HCAECs treated with medicated serum were transfected with miR-30b-5p mimic, while these were significantly rescued with transfection of miR-30b-5p inhibitor.
CONCLUSION
Zhilong Huoxue Tongyu capsule alleviated the pyroptosis of vascular endothelial cells induced by ox-LDL through miR-30b-5p/NLRP3.
背景
我们之前的研究已证明蛭龙活血通瘀胶囊在动脉粥样硬化（AS）中具有保护作用；然而，其分子机制尚不清楚。
方法
用氧化型低密度脂蛋白（ox-LDL）诱导人冠状动脉内皮细胞（HCAECs）以获得细胞AS模型。然后，制备蛭龙活血通瘀胶囊含药血清并用其处理ox-LDL诱导的HCAECs。通过CCK-8法检测细胞活力。此外，采用双荧光素酶报告基因系统检测法确定miR-30b-5p与NLRP3之间的结合。此外，用miR-30b-5p模拟物或miR-30b-5p抑制剂转染ox-LDL诱导的HCAECs。通过流式细胞术、乳酸脱氢酶（LDH）含量检测和qRT-PCR检测评估HCAECs的焦亡情况。
结果
蛭龙活血通瘀胶囊10%含药血清为最大无毒浓度，并用于后续实验。ox-LDL诱导HCAECs后，焦亡率、LDH含量以及包括NLRP3、凋亡相关斑点样蛋白（ASC）、半胱天冬酶1（Caspase 1）、白细胞介素-1（IL-1）和白细胞介素-18（IL-18）在内的焦亡相关基因的mRNA表达水平显著升高，而蛭龙活血通瘀胶囊含药血清可明显改善上述情况。此外，蛭龙活血通瘀胶囊含药血清显著提高了ox-LDL诱导的miR-30b-5p水平降低。NLRP3可与miR-30b-5p结合，且与miR-30b-5p呈负相关。此外，在用含药血清处理的ox-LDL诱导的HCAECs转染miR-30b-5p模拟物后，所有焦亡率、LDH含量以及包括NLRP3、ASC、Caspase 1、IL-1和IL-18在内的焦亡相关基因的mRNA表达水平进一步明显降低，而转染miR-30b-5p抑制剂后这些情况得到显著改善。
结论
蛭龙活血通瘀胶囊通过miR-30b-5p/NLRP3减轻ox-LDL诱导的血管内皮细胞焦亡。
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