Synthesis of Formononetin Derivatives and Cardioprotective Effects

This study aims to design and synthesize a series of novel formononetin (FMN) derivatives and explore their protective effects on oxygen glucose deprivation/relapse (OGD/R) damage to H9C2 cells, along with their molecular regulatory mechanisms. The OGD/R model was established to simulate myocardial ischemia–reperfusion injury. The protective effects of these novel compounds on H9C2 cells were evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method, while the apoptosis rate, myocardial enzyme activity, and autophagy reaction post-compound treatment were assessed using kit-based methods. The formation of autophagosomes in H9C2 cells was observed via transmission electron microscopy, and the expression levels of autophagy-related proteins phosphatidylinositol 3-kinase (PI3K), Akt, Beclin-1, and P62 were determined using Western blotting. The experimental findings demonstrated that compounds 1–6 (C1–6) exhibited varying degrees of protective effects on damaged H9C2 cells, generally outperforming the parent compound FMN. Among these compounds, C4 demonstrated the most significant activity, even surpassing the positive control drug diltiazem. Further mechanistic investigations revealed that C4 could mitigate apoptosis rates, reduce the activity of myocardial enzyme (such as aspartate aminotransferase (AST), lactate dehydrogenase (LDH), and CK), diminish the number of autophagic vesicles, and restore excessive autophagy. Additionally, C4 exerted its protective effects by downregulating the expression of autophagic proteins PI3K, Akt, Beclin-1, P62, LC3 and ATG12. These results indicated that C4 regulates autophagy through the PI3K/Akt/Beclin-1 signaling pathway, thereby exerting a protective effect on cardiomyocytes. Therefore, C4 emerges as a potential myocardial protective drug, offering a new research direction and strategy for the treatment of myocardial ischemia–reperfusion injury.