SGLT2i通过HIF1α/NRF2/HO-1通路对缺氧-复氧心肌细胞老化的保护作用  

Sodium-glucose Cotransport Protein 2 Inhibitor Protects Cardiomyocytes from Hypoxia-reoxygenation Injury via HIF1α/NRF2/HO1 Pathway

作　　者：张瑶 杨少娟 薛佳佳 江燕丽 刘侃玲 Zhang Yao;Yang Shaojuan;Xue Jiajia(Endocrine Department of Liyuan Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430077,China)

机构地区：[1]华中科技大学同济医学院附属梨园医院内分泌科,武汉430077 [2]华中科技大学同济医学院附属梨园医院心血管临床医学中心,武汉430077

出　　处：《华中科技大学学报（医学版）》2023年第3期301-306,共6页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：国家自然科学基金重大研究计划项目(No.91949205);湖北省自然科学基金资助项目(No.2021CFB337)。

摘　　要：目的探讨钠-葡萄糖共转运蛋白2抑制剂(SGLT2i)卡格列净在缺氧心肌细胞修复损伤中的作用及机制。方法应用H9C2大鼠心肌细胞构建缺氧-复氧模型,将心肌细胞分为对照组、单纯SGLT2i组、缺氧-复氧组及缺氧-复氧+SGLT2i组。采用蛋白免疫印迹和荧光定量PCR检测心肌细胞老化相关因子SA-β-gal、p16、p21、p53及抗氧化应激相关核因子2相关因子(NRF2)和血红素氧合酶1(HO1)以及具有转录调节作用的低氧诱导因子1α(HIF1α)表达水平,采用微量酶标法检测超氧化物歧化酶(SOD)、丙二醛(MDA)、过氧化氢酶(CAT)及谷胱甘肽过氧化物酶(GSH-Px),免疫荧光检测HIF1α和NRF2核定位,双荧光素酶报告基因实验检测HIF1α与NRF2基因的相互作用。结果与对照组相比,缺氧-复氧组心肌细胞老化相关因子SA-β-gal、p16、p21和p53表达水平明显上升,氧化应激水平增强。给予SGLT2i预处理后,老化相关因子表达水平下降,氧化应激水平下调。进一步检测发现,缺氧-复氧组抗氧化应激相关分子NRF2和HO1蛋白表达下降,并伴随HIF1α表达上升;给予SGLT2i后NRF2与HIF1α表达恢复至正常水平;而下调NRF2基因表达则消除了SGLT2i对缺氧心肌细胞的保护作用。双荧光素酶报告基因检测发现,HIF1α可直接与NRF2基因启动子区域结合,并转录调控其表达。结论SGLT2i通过抑制低氧诱导的HIF1α激活,上调NRF2抗氧化通路,改善缺氧-复氧心肌细胞的氧化应激损伤和老化,对心肌细胞起保护作用。Objective To investigate the role and mechanism of sodium-glucose cotransport protein 2 inhibitor(SGLT2i)in hypoxia-reoxygenation injury of cardiomyoc ytes.Methods A hypo xia-reoxygenation model was constructed using H9C2 rat cardiomyocytes.The cells were divided into control group,SGLT2i-only group,hypoxia group and hypoxia+SGLT2i group.W estern blotting and fluorescence quantitative PCR were used to detect aging-related factors.Biochemical assays were used to detect the level of malondialdehyde(MDA),and the activities of superoxide dismutase(SOD),catalase(CAT)and gluta-thione pero xidase(GSH-Px).Immunofluorescent staining and dual luciferase were respectively employed to detect hypoxia-in-ducible factor(HIF la)and nuclear factor 2 correlation factor(NRF2)nuclear localization,and the regulatory effect of HIF1α on NRF2.Results Compared with the control group,levels of aging factors(such as SA-B-gal,p16,p21 and p53)and oxidative stress were significantly increased in the cardiomyocytes of the hy poxia/reoxygenation-treated group.After pretreatment with SGLT2i,the aging-associated factors and oxida tive stress were down-regulated.We also found that hypoxia/reoxygenation treat-ment decreased the total protein level of NRF2(an anti-oxidative stress factor)accompanied by the increased HIFla,while SGLT2i pretreatment restored the levels of NRF2 and HIF la.Furthermore,downregulation of NRF2 expression abolished the protective effect of SGLT2i on the hypoxic cardiomyocytes.Dual luciferase reporter gene assay revealed that HIF1α could bind to the NRF2 gene NFE2L2 promoter region and transcriptionally regulated NFE2L2 expression.Conclusion SGLT2i plays a protective role against hypoxia-repair injury in cardiomyocytes by inhibiting hypoxia-induced HIF1α activation and upregulating the NRF2/HO1 antioxidant stress pathway to inhibit cardiomyocyte aging and oxidative stress.

关 键 词：钠-葡萄糖共转运蛋白2抑制剂 老化 心肌细胞 低氧诱导因子 核因子2相关因子 氧化应激 

分 类 号：R542.2[医药卫生—心血管疾病]