BackgroundDepression is a common complication after myocardial infarction (MI) that can seriously affect the prognosis of MI. PurposeTo investigate whether formononetin could ameliorate MI injury and depressive behaviours in a mouse model of MI with depression and elucidate its underlying molecular mechanisms. MethodsHaemodynamic measurements (systolic blood pressure (SYS), the maximum rate of rise of LV pressure (± dp/dtmax)) and behavior tests (tail suspension test, sucrose preference test, forced swimming test) were used to evaluate the effects of formononetin on male C57BL/6N mice after left anterior descending (LAD) coronary artery ligation and chronic unpredictable stress. RT-qPCR, immunohistochemistry, immunofluorescence analysis, western blotting, molecular docking technology, surface plasmon resonance and gene-directed mutagenesis were used to clarify the underlying mechanism. ResultsFormononetin significantly suppressed the depressive behaviours and improved cardiac dysfunction in MI with depression mice model. Formononetin inhibited M1 polarization in macrophages/microglia, while promoting M2 polarization. Importantly, elevated serum IL-6 and IL-17A levels were found in patient with MI, and the patient serum induced M1 microglial polarization; however, formononetin reversed the polarization. Further mechanistic studies showed that formononetin inhibited GSK-3β activity and downstream Notch1 and C/EBPα signaling pathways. Covalent molecular docking showed that formononetin bound to Cys199 of GSK-3β and it has a high affinity for GSK-3β. When Cys199 was mutation, the inhibitory effect of formononetin on GSK-3β activity and M1 polarization in macrophages/microglia were also partly blocked. ConclusionsOur results firstly uncovered that formononetin improved cardiac function and suppressed depressive behaviours in mice after MI with depression by targeting GSK-3β to regulate macrophage/microglial polarization. More importantly, IL-6 and IL-17A produced after MI may cause neuroinflammation, which might be the key factors for depression. Formononetin may be a potential drug for treating MI with depression.