Mechanisms of Xiaoyao Huaxian Formula in the treatment of liver fibrosis based on network pharmacology combined with molecular docking and in vivo experiments

Abstract

Abstract Purpose Xiaoyao Huaxian Formula (XYHXF) shows promise in treating liver fibrosis (LF), but the mechanism is unknown. We sought to elucidate this using network pharmacology, docking and animal experiments.Methods The overlapping targets were determined between those of the main active ingredients of XYHXF using the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform and the Traditional Chinese Medicine Integrated Database and LF-related targets from the Therapeutic Target Database, Online Mendelian Inheritance in Man, and DisGeNET. Protein-protein interaction (PPI) network was generated with Cytoscape 3.7.2. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed using R language. Docking was conducted with AutoDockTools for the key targets and active ingredients of XYHXF. An LF rat model was established with subcutaneously injected carbon tetrachloride followed by ethanol gavage and feeding a high-fat, low-protein diet, and an oral dose of 2.72 g/kg of XYHXF was tested in this model. Liver function was assessed with serum ALT and AST measurements, HE and Masson staining, and western blot of TGF-β, α-SMA, PI3K, p-PI3K, AKT, and p-AKT proteins in liver.Results Overall, 143 active ingredients of XYHXF with 520 potential targets and 2,070 targets of LF were retrieved. The 220 overlapping targets included STAT3, MAPK3, MAPK1, AKT1, and SRC. GO and KEGG analysis revealed overlapping targets that included the response to lipopolysaccharide, response to molecule of bacterial origin, reactive oxygen species metabolic process, and PI3K-Akt signaling pathway. Docking demonstrated potential binding between quercetin/luteolin and AKT1. In rats, compared to the LF group, the XYHXF group had significantly attenuated liver injury and fibrosis, reductions in ALT, AST, α-SMA, and TGF-β; and down-regulation of p-PI3K/PI3K and p-AKT/AKT ratios.Conclusion XYHXF may act on inflammatory processes via multiple active ingredients and treat LF by regulating PI3K-Akt signaling.