从肝治心组方调控SLC7A11/GPX4介导的铁死亡对大鼠心肌缺血再灌注损伤的影响

Effects of Conggan Zhixin Prescription on Myocardial Ischemia Reperfusion Injury by Regulating the Ferroptosis Mediated by SLC7A11/GPX4 in Rats

目的基于SLC7A11/GPX4介导的铁死亡途径探讨从肝治心组方对心肌缺血再灌注损伤的保护作用及机制。方法45只SD大鼠随机分为空白组、假手术组、模型组、从肝治心组方组和地尔硫?组,制备心肌缺血再灌注大鼠模型,并予相应药物灌胃14 d。ELISA检测血清丙二醛(MDA)、肌钙蛋白T(c TnT)、超氧化物歧化酶(SOD)含量和肌酸激酶同工酶(CK-MB)活性,生化法检测血清谷胱甘肽(GSH)含量,HE染色观察心肌组织病理形态,透射电镜观察心肌细胞超微结构,普鲁士蓝染色观察心肌细胞铁沉积水平,免疫组化检测谷胱甘肽过氧化物酶4(GPX4)阳性表达,Western blot和RT-PCR检测心肌组织溶质载体家族7成员11(SLC7A11)、谷胱甘肽合成酶(GSS)、GPX4蛋白及m RNA表达。结果与假手术组比较,模型组大鼠血清MDA、cTnT含量和CK-MB活性显著升高,SOD、GSH含量显著降低(P<0.01),心肌纤维断裂、排列紊乱,细胞肿胀伴炎性细胞浸润,胞质空泡样变性,胞膜破裂,线粒体嵴断裂、排列不规则,铁沉积显著增加(P<0.01),心肌组织SLC7A11、GSS、GPX4蛋白和mRNA表达显著降低(P<0.01);与模型组比较,从肝治心组方组和地尔硫?组大鼠血清MDA、cTnT含量和CK-MB活性显著降低,SOD、GSH含量显著升高(P<0.05,P<0.01),心肌纤维排列较规则,细胞肿胀减轻,胞膜相对完整,线粒体结构改善,铁沉积显著减少(P<0.01),心肌组织SLC7A11、GPX4蛋白和mRNA表达显著升高(P<0.01)。结论从肝治心组方可改善心肌缺血再灌注大鼠心肌损伤,减少心肌组织铁沉积,其机制可能与调控SLC7A11/GPX4信号通路从而抑制铁死亡有关。

Objective To explore the protective effects and mechanism of Conggan Zhixin Prescription on myocardial ischemia reperfusion injury based on the ferroptosis mediated by SLC7A11/GPX4.Methods Totally 45 SD rats were randomly divided into the blank group,sham-operation group,model group,Conggan Zhixin Prescription group and diltiazem group.The myocardial ischemia reperfusion injury model in rats was established.The rats were given corresponding drug intervention for 14 consecutive days.The content of MDA,cTnT,SOD and CK-MB activity were detected by ELISA.The serum content of GSH was detected by biochemical test.HE was used to observe the histopathological changes in myocardial tissue.Prussian blue staining was used to detect the level of iron deposition in cardiomyocytes.The positive expression rate of GPX4 was detected by immunohistochemistry.Western blot and RT-PCR were used to detect the protein and mRNA expression of SLC7A11,GSS and GPX4 in myocardial tissue.Results Compared with the sham-operation group,the content of MDA,cTnT and the activity of CK-MB in serum of the model group significantly increased,the content of SOD and GSH significantly reduced(P<0.01),myocardial fiber rupture,disordered arrangement,cell swelling accompanied by inflammatory cell infiltration,vacuolar degeneration of cytoplasm,membrane rupture,mitochondrial cristae rupture,irregular arrangement,and significant increase in iron deposition(P<0.01),and the expressions of SLC7A11,GSS,GPX4 protein and mRNA in myocardial tissue significantly decreased(P<0.01).Compared with the model group,the contents of MDA,cTnT and the activity of CK-MB in serum of Conggan Zhixin Prescription group and diltiazem group significantly reduced,while the contents of SOD and GSH significantly increased(P<0.05,P<0.01),the arrangement of myocardial fibers was relatively regular,cell swelling was reduced,cell membrane was relatively intact,mitochondrial structure was improved,and iron deposition significantly reduced(P<0.01),the expressions of SLC7A11,GPX4 protein and mRNA in myocardial tissue significantly increased(P<0.01).Conclusion Conggan Zhixin Prescription can improve myocardial ischemia reperfusion injury and alleviate iron deposition in myocardial tissue.The mechanism may be related to regulating SLC7A11/GPX4 signaling pathway to inhibit ferroptosis.