Pharmacological Mechanisms Underlying the Hepatoprotective Effects of

backgroundThe number of hepatocellular carcinoma (HCC) cases worldwide has increased significantly. As a traditional Chinese medicine (TCM) with a long history,

methodsIn this study, we applied a network pharmacology-based strategy and experimental verification to systematically unravel the underlying mechanisms of EH against HCC. First, six active ingredients of EH were screened from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) by the ADME method. Subsequently, 52 potential targets of 6 active ingredients acting on HCC were screened from various databases, including TCMSP, DGIdb, SwissTargetPrediction, CTD, and GeneCards. Then, by constructing protein-protein interaction (PPI) network from STRING, we displayed the intricate connections among these 52 targets through Cytoscape software. We also applied enrichment analysis, including Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses, to provide an outline and set of concepts for describing gene functions and the advanced functions of biological systems of these 52 targets from genomic and molecular level information. Finally, molecular docking and biological experiments were used to reconfirm these results.

resultsWe hypothesized that EH might exert anti-HCC activity by acting on hub genes, including RELA, MMP9, PTGS2, ESR1, EGFR, AR, AKT1, HIF1A, AHR, CYP3A4, ABCG2, and MMP2. Moreover, based on GO and KEGG analysis, we speculated that EH may exert hepatoprotective effects on HCC through the following mechanisms: regulation of the PI3K-AKT signaling pathway to promote apoptosis and inhibit the abnormal proliferation of HCC, downregulation of HIF-1A expression by activating the HIF-1 signaling pathway, prevention of HCC by regulating lipid metabolism, and inhibition of nonalcoholic fatty liver disease (NAFLD) by the cytochrome P450 subfamily. Subsequent biological experiments verified that EH inhibits the PI3K-AKT signaling pathway through its active ingredients, quercetin, and wedelolactone, thereby inhibiting the proliferation of HCC cells and promoting the apoptosis of HCC cells.

conclusionsThe network pharmacological strategy provides an efficient method to systematically explore the pharmacological mechanism of EH in HCC. Our study demonstrated that the anti-HCC proliferation activity of EH is mainly exerted by two active ingredients (quercetin and wedelolactone), which inhibit the proliferation of HCC cells (HepG2 and Huh-7) by inhibiting PI3K-AKT signaling.