海藻糖激活核因子E2相关因子2改善缺氧复氧诱导的H9C2心肌细胞损伤的研究  

Trehalose protects H9C2 cardiomyocytes against cardiac ischemia/reperfusion injury by activating Nrf2

作　　者：卢衍羽 张丽娟[1] 毛艺锟 李瑜[1] Lu Yanyu;Zhang Lijuan;Mao Yikun;Li Yu(Department of Anesthesiology,Affiliated Hospital of Qingdao University,Qingdao 266555,Shandong Province,China)

机构地区：[1]青岛大学附属医院麻醉科,266555

出　　处：《中华老年心脑血管病杂志》2024年第8期954-959,共6页Chinese Journal of Geriatric Heart,Brain and Vessel Diseases

基　　金：青岛大学附属医院“临床医学+X”科研项目(QDFY+X202101062)。

摘　　要：目的通过建立氧糖剥夺-缺氧复氧(oxygen-glucose deprivation/reoxygenation,OGD/R)损伤模型模拟心肌缺血再灌注损伤,探讨海藻糖对OGD/R大鼠H9C2心肌细胞损伤的影响及其作用机制。方法H9C2细胞分为对照组、OGD/R组、海藻糖组(OGD/R+海藻糖)、联合组(OGD/R+海藻糖+ML385)。四甲基偶氮唑盐法检测细胞增殖能力,并通过检测乳酸脱氢酶及Hoechst/丙啶碘化物染色检测细胞膜受损情况。Western blot检测核因子E2相关因子2(nuclear factor erythroid 2-related factor 2,Nrf2)及其下游相关蛋白表达;活性氧、线粒体膜电位检测氧化应激水平;Western blot检测凋亡相关蛋白表达。结果与对照组比较,OGD/R组细胞活力明显降低。与OGD/R组比较,不同浓度海藻糖干预能显著提升细胞活力,与海藻糖浓度呈正相关(P<0.01);与OGD/R组比较,海藻糖组线粒体膜电位(mitochondrial membrane potential,MMP)、谷胱甘肽(glutathione,GSH)、Nrf2、血红素加氧酶1和烟酰胺腺嘌呤二核苷酸磷酸醌氧化还原酶1、Bcl-2、半胱氨酸天冬氨酸蛋白酶3(cysteinyl aspartate specific proteinase-3,Caspase-3)表达明显增高,活性氧、丙二醛、应答元素结合蛋白1、Bax、Bax/Bcl-2、裂解型Caspase-3表达明显降低,差异有统计学意义(P<0.05,P<0.01)。与海藻糖组比较,联合组活性氧、丙二醛及肿瘤坏死因子α、白细胞介素(interleukin,IL)1βmRNA、IL-6 mRNA表达明显增高,MMP、GSH水平明显降低,差异有统计学意义(P<0.05,P<0.01);联合组Bax、Bax/Bcl-2、裂解型Caspase-3表达明显高于海藻糖组(1.77±0.08 vs 1.20±0.20,3.41±1.45 vs 0.99±0.15,4.10±1.05 vs 1.79±0.52,P<0.01),Bcl-2、Caspase-3表达明显低于海藻糖组(0.58±0.21 vs 1.23±0.25,0.87±0.25 vs 1.45±0.31,P<0.01)。结论海藻糖可以被视为一种Nrf2激活剂,通过激活Nrf2抑制氧化应激和凋亡,改善OGD/R诱导的心肌细胞损伤。Objective To investigate the effect of trehalose(Tre)H9C2 cardiomyocytes under oxygen-glucose deprivation/reoxygenation(OGD/R)injury and its mechanism of action with the cellular model simulating the process of myocardial ischemia/reperfusion injury.Methods H9C2 cells were divided control group,OGD/R group,Tre group(OGD/R+Tre),and combination group(OGD/R+Tre+ML385).MTT assay was used to observe cell proliferation,and lactate dehydrogenase(LDH)release and Hoechst/propidium iodide staining were employed to evaluate cell membrane damage.Western blot analysis was utilized to detect the expression of nuclear factor erythroid 2-related factor 2(Nrf2)and its downstream related proteins.The generation of reactive oxygen species(ROS)and mitochondrial membrane potential(MMP)were measured to quantify the level of oxidative stress.The expression of apoptosis-related proteins was determined by Western blot analysis.Results In comparison to the control group,the OGD/R group exhibited a significantly reduced cell viability.When compared with the OGD/R group,the intervention of varying concentrations of Tre obviously improved cell viability in a dose-dependent manner(P<0.01),increased MMP,and up-regulated the expression of glutathione(GSH),Nrf2,hemeoxygenase 1,and nicotinamide adenine dinucleotide phosphate quinone oxidoreductase 1(NQO1),Bcl-2,and cysteinyl aspartate specific proteinase 3(Caspase-3)(P<0.05,P<0.01),and decreased the production of ROS and MDA and the expression of responding element binding protein 1,Bax,Bax/Bcl-2,and cleaved Caspase-3(P<0.05,P<0.01).What's more,the combined group exhibited significantly higher production of ROS,MDA,increased mRNA levels of TNF-α,IL-1β,and IL-6,and reduced MMP and GSH levels(P<0.05,P<0.01),as well as,enhanced expression of Bax,Bax/Bcl-2,and cleaved Caspase-3(1.77±0.08 vs 1.20±0.20,3.41±1.45 vs 0.99±0.15,4.10±1.05 vs 1.79±0.52,P<0.01),and decreased expression of Bcl-2 and Caspase-3(0.58±0.21 vs 1.23±0.25,0.87±0.25 vs 1.45±0.31,P<0.01)in comparison with the Tre group.Con

关 键 词：海藻糖 NF-E2相关因子2 低氧 氧化性应激 细胞凋亡 大鼠H9C2心肌细胞 

分 类 号：R542.22[医药卫生—心血管疾病]