OBJECTIVE To explore the mechanism of action of Qihuang Zhuyu formula (QHZYF) in improving depression after myocardial infarction (MI), with a focus on revealing its regulatory effect on the inflammatory response of the heart and brain.

METHODS The active ingredients of QHZYF and the action targets for intervening in depression after MI were analyzed by using ultra-performance liquid chromatography-high-resolution mass spectrometry (UPLC-Q-TOF/MS) combined with network pharmacology and molecular docking. A rat model of depression after MI was established by ligation of the left anterior descending coronary artery combined with chronic restraint stress. Echocardiography was used to evaluate cardiac function, hematoxylin-eosin (HE) and Masson staining were used to evaluate myocardial injury, behavioral tests were used to detect melancholic behaviors, Nissl staining was used to evaluate hippocampal neuron injury. Western blot detection of tumor necrosis factor receptor 2 (TNFR2), phosphatidylinositol-3-kinase (PI3K), phosphorylated seronine protein kinase (p-AKT), seronine protein kinase (AKT), tumor necrosis factor receptor 1 (TNFR1), phosphorylated nuclear factor κB (p-NF-κB), and nuclear factor κB (NF-κB) in cardiac and hippocampal tissues was conducted. The levels of serum IL-6 and IL-10 were detected by enzyme-linked immunosorbent assay (ELISA), and the expression of TNFR1 and TNFR2 was detected by immunohistochemical technique (IHC). In vitro experiments, co-culture of rat cardiomyocyte line H9C2 cells and rat adrenal pheochromocytoma cell line with high differentiation PC12 cells was conducted, TNFR1 inhibitor (H398) and TNFR2 agonist (C-6His) were administered for intervention, and the expression of TNFR2, PI3K, p-AKT, AKT, TNFR1, NF-κB, p-NF-κB was detected by Western blot. Observe the apoptosis of cells by TUNEL staining, ELISA was used to detect the levels of IL-6 and IL-10 in the cell supernatant.

RESULTS Network pharmacological analysis indicates that the TNF signaling pathway was a key target for the treatment of depression after MI with the QHZYF. In vivo experiments have confirmed that the intervention of QHZYF could significantly improve the cardiac function, myocardial tissue and hippocampal neuron structure damage of depressed rats after MI, and improve their depression-like behaviors. At the molecular level, the high-dose group of QHZYF significantly upregulated TNFR2, p-AKT/AKT, and IL-10 in cardiac and hippocampal tissues (P < 0.01), and downregulated TNFR1, p-NF-κB/NF-κB and IL-6 (P < 0.01). In vitro experiments showed that the drug-containing serum of QHZYF significantly upregulated the expression of TNFR2, p-AKT/AKT and IL-10 in H9C2 and PC12 cells (P < 0.01), downregulated the expression of TNFR1, p-NF-κB/NF-κB and IL-6 (P < 0.01), and significantly inhibited cell apoptosis (P < 0.01). Furthermore, experiments on the combined application of H398 or C-6His further confirmed that its protective and anti-inflammatory effects were mediated by regulating the TNFR2/PI3K/AKT and TNFR1/NF-κB pathways.

CONCLUSION QHZYF improves the homeostasis of heart and brain inflammation by regulating the TNF pathway, and ameliorates myocardial injury and depressive state in depressed rats after MI.