羟基红花黄色素A通过靶向MD2减轻LPS诱导的心肌损伤  

Hydroxysafflor yellow A attenuates LPS-induced myocardial injury by targeting MD2

作　　者：袁林[1] 游丽娇 欧阳冰清 齐璐瑶 楼丽虹 杨海南 雷鸣[1] Yuan Lin;You Lijiao;Ouyang Bingqing;Qi Luyao;Lou Lihong;Yang Hainan;Lei Ming(Department of Critical Care Medicine,Seventh People's Hospital Affiliated to Shanghai University of Traditional Chinese Medicine,Shanghai 200137,China)

机构地区：[1]上海中医药大学附属第七人民医院重症医学科,上海200137

出　　处：《保健医学研究与实践》2024年第8期9-15,共7页Health Medicine Research and Practice

基　　金：国家自然科学基金项目(82174189,82304920);国家中医优势专科建设(重症医学科);浦东新区科技发展基金民生科研专项(PKJ2020-Y65)。

摘　　要：目的 探讨羟基红花黄色素A(HSYA)能否通过靶向髓样分化蛋白2(MD2)减轻小鼠心肌细胞炎症损伤及细胞凋亡。方法 利用不同浓度的HSYA(0.1、1、10μmol/L)预处理小鼠心室肌细胞2 h,脂多糖(LPS)(5μg/m L)干预4 h后进行检测,进一步通过分子对接技术明确HSYA与MD2的结合方式及结合位点,免疫沉淀检测HSYA对TLR4/MD2复合物生成的影响,并通过Western blotting进行作用机制探究。结果 与对照组相比,LPS能够诱导心肌细胞凋亡,促使细胞内及线粒体内活性氧(ROS)的释放,促使炎症因子白介素6(IL-6)的释放,但白介素1β(IL-1β)、肿瘤坏死因子α(TNF-α)并没有明显变化。与LPS诱导组相比,HSYA能够有效抑制LPS诱导的心肌细胞凋亡、ROS释放及炎症因子损伤;分子对接结果提示HSYA能够与MD2的疏水口袋结合,HSYA能够剂量依赖性地减少TLR4/MD2复合物的生成,提示HSYA与TLR4竞争性结合MD2蛋白。采用Westrn blotting检测TLR4、MD2、NF-κB p65磷酸化蛋白的表达,结果显示HSYA以浓度依赖的方式降低了LPS诱导的TLR4和MD2、NF-κB p65磷酸化蛋白表达。结论 HSYA能够通过靶向MD2(TLR4/MD2复合物介导),减轻LPS诱导的炎症损伤,发挥降低心肌细胞炎症损伤及凋亡的作用。Objective To investigate whether Hydroxysafflor yellow A(HSYA)can target myeloid differentiation factor 2(MD2)to alleviate inflammation and apoptosis in mouse cardiomyocytes induced by lipopolysaccharide(LPS).Methods Mouse ventricular myocytes were pretreated with different concentrations of HSYA(0.1μmol/L,1μmol/L,10μmol/L)for 2 hours,followed by LPS(5μg/mL)intervention for 4 hours before detection.Molecular docking technology was further used to determine the binding mode and binding site of HSYA with MD2.The effect of HSYA on the generation of TLR4/MD2 complexes was detected by immunoprecipitation,and the mechanism of action was explored by Western blotting.Results Compared with the control group,LPS induced apoptosis in cardiomyocytes,promoted the release of reactive oxygen species(ROS)within the cells and the mitochondria,and facilitated the release of the inflammatory factor interleukin-6(IL-6),but without significant changes in interleukin-1β(IL-1β)and tumor necrosis factorα(TNF-α).Compared with the LPS-induced group,HSYA effectively reduced LPS-induced cardiomyocyte apoptosis,ROS release,and inflammatory factor damage.Molecular docking results suggested that HSYA could bind to the hydrophobic pocket of MD2,and HSYA could dose-dependently reduce the generation of TLR4/MD2 complexes,indicating that HSYA competitively binds to MD2 protein with TLR4.The results of TLR4,MD2,and phosphorylated NF-κB p65 protein expressions via Western blotting showed that HSYA lowered the expression of LPS-induced TLR4 and MD2,phosphorylated NF-κB p65 proteins in a concentration-dependent manner.Conclusion HSYA can target MD2 and alleviate LPS-induced inflammatory damage through mediation of the TLR4/MD2 complex,playing a role in reducing inflammatory damage and apoptosis in cardiomyocytes.

关 键 词：羟基红花黄色素A 髓样分化蛋白2 心肌损伤 

分 类 号：R965[医药卫生—药理学]