红花黄色素对大鼠心梗后心肌重构的影响研究

The Effect of Safflower Yellow Pigment on Myocardial Remodeling after Myocardial Infarction in Rats

目的观察红花黄色素(SY)对急性心肌梗死(AMI)后心肌重构的影响,并探究其与腺苷酸活化蛋白激酶(AMPK)/哺乳动物西罗莫司靶蛋白(mTOR)信号通路的关系。方法利用冠状动脉左前支结扎法构建AMI大鼠模型,随机分为Model组、氯沙坦组(10 mg/kg)、SY低剂量组(8 mg/kg)、SY高剂量组(32 mg/kg)、SY+Compound C组(32 mg/kg SY+10 mg/kg Compound C),并设假手术(Sham)组,每组12只。连续治疗4周后,使用超声心动图仪测定大鼠心功能指标;采用HE、Masson染色观察大鼠心肌组织变化,计算胶原容积积分(CVF)、测定大鼠心肌细胞直径;酶联免疫吸附法(ELISA)测定心肌组织乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)水平;Werstern blot法检测大鼠心肌组织Ⅰ、Ⅲ型胶原(COL-Ⅰ、COL-Ⅲ)、B型钠尿肽(BNP)、心房钠尿肽(ANP)、微管相关蛋白1轻链3Ⅱ/LC3-Ⅰ(LC3-Ⅱ/LC3-Ⅰ)、AMPK、自噬相关蛋白P62、Beclin-1、磷酸化AMPK(p-AMPK)、mTOR、磷酸化mTOR(p-mTOR)蛋白表达水平。结果与Sham组相比,Model组大鼠心肌细胞肥大,心肌纤维排列紊乱,左室短轴缩短率(LVFS)、左室射血分数(LVEF)及LC3-Ⅱ/LC3-I、Beclin-1、p-AMPK、p-mTOR蛋白表达水平及SOD水平降低(P<0.05),左室舒张末期内径(LVEDD)、左室收缩末期内径(LVESD)、心肌细胞直径、CVF、COL-Ⅰ、COL-Ⅲ、BNP、ANP及P62蛋白表达水平、LDH水平升高(P<0.05);与Model组相比,氯沙坦组、SY低、高剂量组大鼠心肌细胞有所缩小,心肌纤维排列趋于正常,LVFS、LVEF及LC3-Ⅱ/LC3-I、Beclin-1、p-AMPK、p-mTOR蛋白表达水平及SOD水平升高(P<0.05),LVEDD、LVESD、心肌细胞直径、CVF、COL-Ⅰ、COL-Ⅲ、BNP、ANP及P62蛋白表达水平、LDH水平降低(P<0.05);与SY高剂量组相比,SY+Compound C组大鼠心肌细胞肥大,心肌纤维排列紊乱,LVFS、LVEF及LC3-Ⅱ/LC3-I、Beclin-1、p-AMPK、p-mTOR蛋白表达水平及SOD水平降低(P<0.05),LVEDD、LVESD、心肌细胞直径、CVF、COL-Ⅰ、COL-Ⅲ、BNP、ANP及P62蛋白表达水平、LDH水平升高(P<0.05)。结论SY可提高AMI心肌组织抗氧化能力,降低心肌肥厚程度,缓解心肌纤维化进程,逆转心肌重构,可能与激活AMPK/mTOR信号通路,上调心肌自噬有关。

Objective To observe the impact of safflower yellow pigment(SY)on myocardial remodeling after acute myocardial infarction(AMI),and explore its relationship with adenosine monophosphate-activated protein kinase(AMPK)/mammalian target of rapamycin(mTOR)signaling pathway.Methods AMI rat models were constructed by ligating the left anterior branch of coronary artery.They were randomly divided into Model group,losartan group(10 mg/kg),SY low-dose group(8 mg/kg),SY high-dose group(32 mg/kg),and SY+Compound C group(32 mg/kg SY+10 mg/kg Compound C),and sham operation(Sham)group were set up,with 12 rats in each group.After 4 weeks of continuous treatment,the indexes of cardiac function were measured by echocardiography;HE and Masson staining were used to observe the changes of myocardial tissue,calculate the collagen volume integral(CVF),and measure the diameter of myocardial cells in rats;the levels of lactate dehydrogenase(LDH)and superoxide dismutase(SOD)in myocardium were measured by enzyme-linked immunosorbent assay(ELISA);the expression levels of collagenⅠ,Ⅲ(COL-I,COL-Ⅲ),B-type natriuretic peptide(BNP),atrial natriuretic peptide(ANP),microtubule associated protein 1 light chain 3Ⅱ/LC3-I(LC3-Ⅱ/LC3-I),AMPK,autophagy associated protein P62,Beclin-1,phosphorylated AMPK(p-AMPK),mTOR,phosphorylated mTOR(p-mTOR)proteins were detected by Werstern blot.Results Compared with Sham group,myocardial cells in Model group were hypertrophy and myocardial fibers were disordered,LVFS,LVEF,the expression levels of LC3-Ⅱ/LC3-I,Beclin-1,p-AMPK,p-mTOR proteins,and the level of SOD decreased(P<0.05),LVEDD,LVESD,myocardial cell diameter,the expression levels of CVF,COL-Ⅰ,COL-Ⅲ,BNP,ANP and P62 proteins,and the level of LDH increased(P<0.05);compared with the Model group,the myocardial cells of the losartan group and SY low and high dose groups were smaller,and the arrangement of myocardial fibers tended to be normal,LVFS,LVEF,the expression levels of LC3-Ⅱ/LC3-I,Beclin-1,p-AMPK,p-mTOR proteins,and the level of SOD increased(P<0.05),LVEDD,LVESD,myocardial cell diameter,the expression levels of CVF,COL-Ⅰ,COL-Ⅲ,BNP,ANP and P62 proteins,and the level of LDH decreased(P<0.05);compared with SY high dose groups,the myocardial cells of rats in SY+Compound C group were hypertrophic and the myocardial fibers were disordered,LVFS,LVEF,the expression levels of LC3-Ⅱ/LC3-I,Beclin-1,p-AMPK,p-mTOR proteins,and the level of SOD decreased(P<0.05),LVEDD,LVESD,myocardial cell diameter,the expression levels of CVF,COL-Ⅰ,COL-Ⅲ,BNP,ANP and P62 proteins,and the level of LDH increased(P<0.05).Conclusion SY can improve the antioxidant capacity of AMI myocardium,reduce the degree of myocardial hypertrophy,alleviate the process of myocardial fibrosis,and reverse myocardial remodeling,which may be related to the activation of AMPK/mTOR signal pathway and the up regulation of myocardial autophagy.