基于“心痛治肝”理论探讨从肝治心方调控心肌缺血再灌注损伤中铁死亡的机制  

Protective mechanism of regulating ferroptosis by Conggan Zhixin recipein myocardial ischemia-reperfusion injury based on the theory ofheartache governing liver

作　　者：陈亚[1] 曾阳 何飘[2] 谢丽华 张程程[2] 王瑾茜 袁华[1] 刘莹莹[1] 朴美虹 汪辛强 CHEN Ya;ZENG Yang;HE Piao;XIE Lihua;ZHANG Chengcheng;WANG Jinxi;YUAN Hua;LIU Yingying;PIAO Meihong;WANG Xinqiang(the First Affiliated Hospital of Hunan University of Chinese Medicine,Changsha 410007,China.;Hunan University of Chinese Medicine,Changsha 410208)

机构地区：[1]湖南中医药大学第一附属医院,长沙410007 [2]湖南中医药大学,长沙410208

出　　处：《中国实验动物学报》2023年第8期999-1006,共8页Acta Laboratorium Animalis Scientia Sinica

基　　金：湖南省自然科学基金项目(2021JJ40418);湖南省中医药科研项目(2021179,2021172);长沙市科技计划项目(kq2014222,kq2014224);湖南省卫健委科研计划项目(202103010864);湖南中医药大学校级科研基金项目(2019XJJJ057,2022XYLH005)。

摘　　要：目的本研究拟基于“心痛治肝”理论探讨从肝治心方调控心肌缺血再灌注损伤(myocardial ischemia-reperfusion injury,MIRI)大鼠模型铁死亡的机制。方法SD大鼠按随机数字表分为空白组(n=10)、假手术组(n=10)、模型组(n=10)、从肝治心方组(CGZX组)(n=10)、麝香保心丸组(SXBX组)(n=10)。除空白组、假手术组外,各组均采用“冠状动脉前降支结扎+缺血后再灌注”构建MIRI模型,造模24 h后灌胃给药,灌胃周期为14 d,频率为每天1次。于第16天经腹主动脉采血,ELISA分析血清中肌酸激酶同工酶(CK-MB)、肌钙蛋白T(cTnT)、超氧化物歧化酶(SOD)、丙二醛(MDA)、多不饱和脂肪酸(PUFA)含量;另剪心脏进行HE染色观察心肌组织病理形态、普鲁士蓝染色观察心肌细胞铁沉积情况;电镜下观察心室肌细胞铁死亡后线粒体的形态;Western Blot及PCR测试心肌组织中Nrf2、ACSL4的蛋白及mRNA表达量。结果与空白组相比,模型组大鼠存在明显心肌组织损伤、电镜下心肌超微结构受损明显,出现肌丝断裂或疏松及心肌组织线粒体肿胀现象,心肌细胞存在大量黑褐色铁沉积。血清中CK-MB、cTnT、MDA及PUFA含量升高(P<0.01),SOD含量降低(P<0.01),心肌组织中Nrf2、ACSL4的mRNA及蛋白表达量显著增加(P<0.01);与模型组相比,从肝治心方能明显改善心肌组织水肿变性,减少心肌细胞铁沉积及细胞嵴突疏松程度,且显著下调血清中MDA、PUFA、CK-MB、cTnT的含量(P<0.01),上调SOD含量(P<0.01),并可上调心肌组织Nrf2、下调ACSL4的mRNA及蛋白表达(P<0.01)。结论从肝治心方对MIRI大鼠起到心肌保护作用,可能与调控Nrf2-ACSL4通路从而抑制心肌细胞铁死亡有关。Objective To explore the mechanism of ferroptosis in a rat model of myocardial ischemia-reperfusion injury(MIRI),basing on the theory of heartache governing liver.Methods SD rats were divided into a blank group(n=10),sham operation group(n=10),model group(n=10),Conggan Zhixin recipe group(CGZX group)(n=10),and Shexiang Baoxin Pill group(SXBX group)(n=10)in accordance with a random number table.Except in blank and sham operation groups,all groups underwent ligation of the anterior descending coronary artery and ischemia-reperfusion to establish a myocardial ischemia-reperfusion injury model.The drug was administered by gavage at 24 h after model establishment.The gavage cycle was 14 days,and the frequency was once a day.Blood was collected from the abdominal aorta on day 16,and serum contents of creatine kinase isoenzyme(CK-MB),cardiac troponin T(cTnT),superoxide dismutase(SOD),malondialdehyde(MDA),and polyunsaturated fatty acid(PUFA)were analyzed by ELISA.The harvested heart was stained with HE to observe pathological morphology of myocardial tissue and with Prussian blue to observe iron deposition of myocardial cells.Mitochondrial morphology after ferroptosis was observed in ventricular muscle by electron microscopy.mRNA and protein expression of Nrf2 and ACSL4 in the myocardium was measured by PCR and Western Blot,respectively.Results Compared with the normal group,rats in the model group had obvious myocardial tissue and ultrastructure damage under the electron microscope,broken or loose myofilaments,swollen mitochondria in myocardial tissue,and a large amount of dark brown iron deposition in myocardial cells.The serum contents of CK-MB,cTnT,MDA,and PUFA were increased(P<0.01),while the SOD content was decreased(P<0.01).Moreover,mRNA and protein expression of Nrf2 and ACSL4 in the myocardium was increased significantly(P<0.01).Compared with the model group,Conggan Zhixin recipe significantly improved edema and degeneration of myocardial tissue,reduced the iron deposition of myocardial cells and the degree of c

关 键 词：从肝治心方 心肌缺血再灌注损伤 铁死亡 心痛治肝 

分 类 号：Q95-33[生物学—动物学]