Network pharmacology exploration reveals the bioactive compounds and molecular mechanisms by which Gui-Qi-Yang-Xin Decoction suppresses myocardial fibrosis

Background: Gui-Qi-Yang-Xin Decoction (GQYXD), a Chinese medicine formula, has been shown to exert obvious therapeutic effects on myocardial fibrosis (MF). However, the bioactive compounds and underlying pharmacological mechanisms remain to be elucidated.Methods:The present study aimed to determine the bioactive compounds and potential action mechanisms of GQYXD in the treatment of MF using an integrated network pharmacology approach. All compounds were identified from the corresponding database and active compounds were selected according to their oral bioavailability and drug-likeness index. The components of GQYXD and the corresponding drug targets were retrieved through drug-target databases and the potential MF genes were obtained from the GeneCard database. The abovementioned ingredients and related targets were used to construct ‘compound–target–disease’, protein-protein interaction (PPI) and target-pathway networks. Functional and pathway enrichment analyses were performed to determine the key biological processes and important pathways using the clusterProfiler package in R programming language. Experimental validation was performed using haematoxylin-eosin staining, Masson’s trichrome staining and immunohistochemistry analysis in isoproterenol-induced MF rats, and western blot analysis, phalloidin staining and immunofluorescence staining were used to verify the proposed mechanism of protection on H9C2 cells.Results: Fifty-five bioactive components and 59 putative targets that were directly related to GQYXD and MF were collected. The results of functional enrichment analysis revealed that responses to lipopolysaccharides, oxidative stress, hypoxia, nutrient levels and mechanical stimuli were the key biological processes and were regulated simultaneously by six direct targets, including prostaglandin-endoperoxide synthase 2 (PTGS2), mitogen-activated protein kinase 14 (MAPK14), protein kinase 1 (AKT1) mitogen-activated protein kinase 8 (MAPK8), interleukin-6 (IL-6) and interleukin-1β (IL-1β). KEGG pathway enrichment analysis found that five signalling pathways were closely related to MF treatment by GQYXD. Animal experiments showed that GQYXD could significantly alleviate isoproterenol-induced MF. In vitro cell experiment results showed that GQYXD treatment could ameliorate organisational damage to the cell cytoskeleton in H9C2 induced by isoproterenol. Down-regulated expression of PTGS2, MAPK14 and MAPK8 and up-regulated expression of AKT1 was demonstrated both in animal and cell experiments.Conclusion: GQYXD could alleviate MF based on multiple components, targets and pathways. This study also demonstrated the network pharmacology approach as an effective tool to reveal the mechanisms of traditional Chinese medicine (TCM) from a holistic perspective.