芒柄花素通过HSP90/AKT调控扩张型心肌病心力衰竭大鼠心肌细胞凋亡及其机制

Formononetin regulates dilated cardiomyopathy-mediated heart failure in rats via HSP90/AKT cardiomyocyte apoptosis and mechanism

目的探讨芒柄花素(formononetin,FMN)对扩张型心肌病心力衰竭大鼠心肌细胞凋亡及HSP90/AKT的影响。方法通过超声心动图、ELISA法、组织学染色、TUNEL染色,观察不同剂量FMN对扩张型心肌病心力衰竭大鼠的保护作用及心肌细胞凋亡情况。从TCMSP、DisGeNet、GeneCards等数据库获取FMN作用于扩张型心肌病心力衰竭的潜在靶点及其交集,再根据蛋白质互作网络(PPI)获得两者交集的关键靶点,并通过分子对接和Western blotting对关键靶点进行验证。结果FMN可降低扩张型心肌病心力衰竭大鼠左心室收缩末期内径(LVIDS)、左心室舒张末期内径(LVIDD)水平及NT-pro BNP、cTn-T、CK、CK-MB和LDH含量,提高射血分数(EF)和短轴缩短率(FS)水平,减少心肌细胞凋亡;FMN与网络药理学筛出的10个关键靶点(AKT1、HSP90AA1、CASP3、MAPK1、MMP9、SRC、ALB、HRAS、IGF1、EGFR)均具有较强的结合作用,以HSP90AA1、AKT1结合作用最强,同时FMN可降低心肌组织HSP90、AKT及下游CASP3蛋白表达,提高p-AKT的表达。结论FMN可能通过抑制HSP90的表达,促进AKT磷酸化,降低CASP3的表达,减少心肌细胞凋亡,改善心肌组织损伤,达到治疗扩张型心肌病心力衰竭的目的。

Objective To investigate the effects of formononetin(FMN)on cardiomyocyte apoptosis and HSP90/AKT in rats with dilated cardiomyopathy-mediated heart failure.Methods Echocardiography,ELISA,histological staining,and TUNEL staining were used to observe the protective effect of different doses of FMN on dilated cardiomyopathy-mediated heart failure in rats and the apoptosis of cardiomyocytes.The potential targets of formononetin on dilated cardiomyopathy-mediated heart failure were obtained from TCMSP,DisGeNet,GeneCards,and other databases,the key targets were obtained according to the protein-protein interaction(PPI)network,and the key targets were verified by molecular docking.Western blotting was used to further verify the regulatory role of key targets in the treatment of dilated cardiomyopathy-mediated heart failure with formononetin.Results Formononetin could reduce the levels of LVIDS,LVIDD,NT-pro BNP,cTn-T,CK,CK-MB,and LDH in rats with dilated cardiomyopathy-mediated heart failure,increase the levels of EF and FS,and reduce the apoptosis of cardiomyocytes.FMN had a strong binding effect on 10 key targets(AKT1,HSP90AA1,CASP3,MAPK1,MMP9,SRC,ALB,HRAS,IGF1,and EGFR)screened by network pharmacology,with HSP90AA1 and AKT1 having the strongest binding effect.Formononetin decreased the expression of HSP90,AKT and downstream CASP3 protein,but increased the expression of p-AKT in myocardial tissue.Conclusion Formononetin may inhibit the expression of HSP90,promote phosphorylation of AKT to p-AKT,and inhibit the expression of CASP3,thereby reducing the apoptosis of cardiomyocytes and improving myocardial tissue damage,so as to achieve the purpose of treating dilated cardiomyopathy-mediated heart failure.