摘要
目的:本研究旨在探讨黄芩苷(Baicalin)对氯化钴(CoCl_(2))诱导H9c2心肌细胞缺氧损伤的作用机制。方法:采用CoCl_(2)建立心肌细胞缺氧损伤模型,并分别加入不同浓度的黄芩苷培养。将正常氧培养设置为对照组,CoCl_(2)培养H9c2心肌细胞设置为CoCl_(2)组;Baicalin、Y27632(Rho激酶抑制剂)预处理的H9c2缺氧心肌细胞设置为CoCl_(2)+Baicalin组、CoCl_(2)+Y27632组、CoCl_(2)+Baicalin+Y27632组。通过细胞毒性检测试剂盒(CCK8)检测细胞活性;荧光探针测定细胞中活性氧(ROS)的表达;WST-8检测心肌细胞超氧化歧化酶(SOD)活力;TBA法测定丙二醇(MDA)浓度;同时采用WesternBlot方法分析RhoA、ROCK1、ROCK2、TNF-α、IL-1β蛋白表达情况。结果:处理H9c2细胞24 h后,1 000μmol/L的CoCl_(2)和75μmol/L黄芩苷对治疗心肌细胞缺氧损伤具有较好的细胞活力。CoCl_(2)组细胞活性明显低于对照组,加入黄芩苷后可显著提高细胞活性(P<0.05);与对照组相比,CoCl_(2)组心肌细胞可上调ROS、MDA表达,下调SOD活力,升高RhoA、ROCK1、ROCK2、TNF-α、IL-1β蛋白表达水平,加入Baicalin后可逆转上诉蛋白的表达水平;与CoCl_(2)组相比,CoCl_(2)+Baicalin可抑制ROS、MDA的表达,上调SOD含量,下调RhoA、ROCK1、ROCK2、TNF-α、IL-1β蛋白表达水平,而加入Y27632(Rho激酶抑制剂)后则可显著增强Baicalin带来的保护作用。结论:Baicalin可减轻CoCl_(2)诱导H9c2心肌细胞缺氧损伤的炎症、氧化应激反应,其机制与抑制ROS依赖性RhoA/ROCK通路有关。
Objective:To investigate the mechanism of Baicalin on the hypoxic injury of H9c2 cardiomyocytes induced by co-balt chloride(CoCl_(2)).Methods:Cobalt chloride(CoCl_(2))was used to establish a hypoxic injury model in cardiomyocytes,and dif-ferent concentrations of Baicalin were added for culturing,respectively.The normoxic culture was set as the control group,and the cobalt chloride cultured H9c2 cardiomyocytes were set as the CoCl_(2) group.The hypoxic H9c2 cardiomyocytes pretreated with Ba-icalin and/or Y27632(Rho kinase inhibitor)were set as the CoCl_(2)+Baicalin group,CoCl_(2)+Y27632 group and CoCl_(2)+Baicalin+Y27632 group.Cell proliferation was assessed using the cytotoxicity assay kit(CCK8)assay,and reactive oxygen species(ROS)levels were detected using a fluorescence method.The activity of superoxide dismutase(SOD)in cardiomyocytes was detected by WST-8.The concentration of propylene glycol(MDA)was determined by TBA method.Meanwhile,the expression of RhoA,ROCK1,ROCK2,TNF-αand IL-1βwere analyzed by Western blot as well.Results:After treatment of H9c2 cells for 24 h,1000μmol/L CoCl_(2) and 75μmol/L Baicalin showed better cell viability in the treatment of hypoxic injury of cardiomyocytes.The cell activity of the CoCl_(2) group was significantly lower than that of the control group,and the pre-protective effect of Baicalin on hypoxia-induced cells could significantly increase the cell activity.Compared with the control group,ROS expression level and MDA content in cardiomyocytes of the CoCl_(2) group increased,protein expression levels of RhoA,ROCK1,ROCK2,TNF-αand IL-1βup-regulated,and SOD activity decreased.Compared with the CoCl_(2) group,CoCl_(2)+Baicalin could inhibit the expression of ROS and MDA content,up-regulate SOD activity,and down-regulate the expression levels of RhoA,ROCK1,ROCK2,TNF-αand IL-1β,while adding Y27632(Rho kinase inhibitor)significantly enhanced the protective effect of Baicalin.Conclusion:Ba-icalin could alleviate inflammatory and oxidative stress responses to CoCl_(2)-induced injury in H9c2 cardiomyocytes,and its mecha-nism might be related to the inhibition of ROS-dependent RhoA/ROCK pathway.
作者
沈艳玲
刘承红
王世魁
徐尧
张云波
顾申红
SHEN Yanling;LIU Chenghong;WANG Shikui;XU Yao;ZHANG Yunbo;GU Shenhong(Hainan Medical University,Key Laboratory of Hainan Trauma and Disaster Rescue,Haikou 571199,China;Department of General Medicine,the First Affiliated Hospital of Hainan Medical University,Haikou 570102,China)
出处
《海南医学院学报》
CAS
北大核心
2024年第7期481-488,共8页
Journal of Hainan Medical University
基金
海南省重点研发项目(EPC-MVs富集miR-126作用于靶基因抑制RhoA/ROCK通路对高血压肾病作用及机制、ZDYF2022SHFZ101)。
作者简介
沈艳玲,E‑mail:499318100@qq.com;通讯作者:顾申红,教授,硕士生导师,E‑mail:1574063710@qq.com。
