右美托咪定通过激活Nrf2/HO-1通路减轻热应激诱导的人骨骼肌细胞胀亡  

Dexmedetomidine attenuates heat stress-induced oncosis in human skeletal muscle cells by activating the Nrf2/Ho-1 pathway

作　　者：刘洋 贾亿卿[2] 李程程 毛汉丁 刘树元 单毅 LIU Yang;JIA Yiqing;LI Chengcheng;MAO Handing;LIU Shuyuan;SHAN Yi(Department of Emergency Medicine,School of Medicine,South China University of Technology,Guangzhou 510006,China;Department of Emergency Medicine,Sixth Medical Center of PLA General Hospital,Beijing 100048,China)

机构地区：[1]华南理工大学医学院,广东广州510006 [2]解放军总医院第六医学中心急诊医学科,北京100048

出　　处：《南方医科大学学报》2025年第3期603-613,共11页Journal of Southern Medical University

基　　金：166工程项目(231-CXCY-N101-07-07-01)。

摘　　要：目的研究右美托咪定(DEX)对于热打击诱导人骨骼肌细胞(HSKMC)胀亡的保护作用及可能机制。方法体外培养HSKMC,热打击组(HS组)将细胞置于43℃的水浴锅中热打击4 h,构建细胞胀亡模型,设置对照组、HS组、30μmol/L DEX+HS组、ML385+30μmol/L DEX+HS组、si-Nrf2+HS组、si-Nrf2+30μmol/L DEX+HS组。通过CCK-8法检测细胞活力,透射电子显微镜观察细胞超微结构,Annexin V-FITC/PI免疫荧光流式细胞术检测细胞胀亡和细胞凋亡,使用谷胱甘肽(GSH)和GSH-px试剂盒检测细胞中GSH及GSH-px含量,TBA法检测丙二醛(MDA),比色法检测乳酸脱氢酶(LDH)、超氧歧化酶(SOD)、三磷酸腺苷(ATP)表达水平,ELISA法检测肿瘤坏死因子(TNF)-α、白介素(IL)-6、IL-1β水平,qRT-PCR及Western blotting法检测porimin、caspase-3、Nrf2、p-Nrf2、HO-1、NQO1等蛋白表达水平变化,荧光探针法检测细胞内活性氧(ROS),JC-1荧光染色法检测线粒体膜电位损伤。结果与对照组相比,HS组细胞和细胞器肿胀、胞质内空泡化明显,符合胀亡表现,细胞活力下降,细胞双阳性细胞率(Annexin-V+/PI+)、porimin蛋白表达量增加(P<0.05),caspase-3蛋白在各组间的差异无统计学意义(P>0.05),ROS、MDA、LDH、TNF-α、IL-6和IL-1β的含量增多(P<0.05),ATP、线粒体膜电位、GSH、GSH-px和SOD水平降低(P<0.05)。与HS组相比,30μmol/L DEX+HS组细胞损伤减轻,细胞活力升高,细胞双阳性细胞率下降(P<0.05),ROS、MDA、LDH、TNF-α、IL-6和IL-1β的含量下降(P<0.05),ATP、线粒体膜电位、GSH、GSH-px和SOD含量升高(P<0.05),Nrf2、p-Nrf2、HO-1和NQO1蛋白表达升高(P<0.05)。与30μmol/L DEX+HS组相比,经ML385或si-Nrf2干预后,DEX对HSKMC细胞的保护作用减弱(P<0.05)。结论热打击诱导人骨骼肌细胞发生胀亡,DEX抑制热打击诱导的胀亡,可能机制是激活Nrf2/HO-1信号通路减轻HSKMC的氧化损伤以及抑制炎症因子分泌。Objective To investigate the protective effects of dexmedetomidine(DEX)against heat stress(HS)-induced oncosis in human skeletal muscle cells(HSKMCs)and its underlying mechanisms.Methods A HSKMC model of HS-induced oncosis were established by 43℃water bath for 4 h,and the effects of treatments with 30μmol/L DEX,ML385(a Nrf2 inhibitor)+DEX,si-Nrf2+HS,and si-Nrf2+DEX prior to modeling on cell viability was assessed using CCK-8 assay.Oncosis characteristics were evaluated using transmission electron microscopy and Annexin V-FITC/PI flow cytometry.The oxidative stress markers(GSH,GSH-Px,MDA,SOD and ROS),mitochondrial membrane potential,energy metabolism,and inflammatory cytokines(TNF-α,IL-6 and IL-1β)in the cells were quantified using standard kits,and the expressions of porimin,caspase-3 and Nrf2 pathway proteins were analyzed using Western blotting and qRT-PCR.Results HS induced typical oncotic features in HSKMCs including organelle swelling and cytoplasmic vacuolization.DEX pretreatment significantly attenuated these changes,reduced Annexin V+/PI+cell ratio and cellular porimin expression,and lowered the levels of ROS and MDA while restoring GSH and SOD levels.DEX pretreatment also significantly increased the mitochondrial membrane potential and ATP level,upregulated the expressions of Nrf2,p-Nrf2,HO-1 and NQO1,and suppressed the expressions of TNF-α,IL-6 and IL-1β.The protective effects of DEX were obviously attenuated by interventions with ML385 or si-Nrf2.Conclusion DEX mitigates HS-induced HSKMC oncosis by activating the Nrf2/HO-1 pathway to relieve oxidative stress,mitochondrial dysfunction,and inflammatory responses.

关 键 词：右美托咪定 横纹肌溶解症 胀亡 人骨骼肌细胞 Nrf2/HO-1通路 

分 类 号：R28[医药卫生—中药学]