Mechanism of berberine in the treatment of polycystic ovary syndrome revealed by network pharmacology analysis: Focusing on ferroptosis, ROS, immunity and inflammation.

Berberine, a bioactive component isolated from traditional Chinese medicine, functions as a multitarget regulator and has been proven to ameliorate reproductive endocrine disorders and metabolic abnormalities in patients with polycystic ovary syndrome (PCOS). Nevertheless, the exact molecular mechanisms underlying its therapeutic effects remain to be fully elucidated. In this study, we integrated multi-omics data analysis with in vitro experimental validation. Differentially expressed genes associated with PCOS were screened from the Gene Expression Omnibus (GEO) database, and berberine-targeted genes were retrieved from public databases to construct a protein-protein interaction (PPI) network. KGN and Ishikawa cell models were employed to verify the regulatory effects of berberine on cell proliferation, oxidative stress and nuclear receptor signaling. Finally, molecular docking and molecular dynamics simulations were performed to analyze the binding patterns, so as to comprehensively decipher the molecular mechanism of berberine against PCOS. A total of 32 common targets were identified via network pharmacology analysis, which were mainly enriched in metabolism-, reproduction- and inflammation-related signaling pathways. Further screening identified 8 hub genes that constituted a nuclear receptor-kinase interaction module. Enrichment analysis indicated that these hub genes were cell-specifically correlated with N6-methyladenosine (m6A) modification, ferroptosis and immune inflammation. In vitro cellular assays demonstrated that berberine suppressed cell proliferation, elevated reactive oxygen species (ROS) levels and reduced intracellular iron concentrations. Molecular simulation results revealed the favorable binding energy of berberine to target receptors and stable RMSD values of the ligand-receptor complexes, uncovering a multi-axis network regulatory mechanism of berberine. Collectively, this study adopts an integrated multi-method strategy and confirms that berberine targets the nuclear receptor-kinase interaction module. It exerts multi-axis regulatory effects on PCOS by modulating m6A modification, ferroptosis, ROS accumulation and immune inflammatory responses.