Pectolinarigenin Induces Mitochondrion-dependent Apoptosis, Causes G2/M Arrest, and Inhibits Migration and Invasion in Rheumatoid Arthritis–Fibroblast-like Synoviocytes

Export Pectolinarigenin (PEC), a bioactive flavonoid extracted from Cirsium japonicum, demonstrates anti-inflammatory and anticancer properties with potential to inhibit the tumor-like invasion of rheumatoid arthritis–fibroblast-like synoviocytes (RA-FLSs). This study aimed to evaluate PEC’s effects on apoptosis, cell cycle arrest, migration, and invasion in RA-FLSs and elucidate the underlying mechanisms. In vitro experiments used the human RA-FLS cell line MH7A, primary rheumatoid arthritis–human FLSs, and normal cellular controls. Interventions included PEC alone or combined with the epidermal growth factor receptor (EGFR) inhibitor gefitinib (GEF) or agonist NSC228155, with methotrexate as a positive control. Potential targets of PEC were prioritized using a computational framework and then experimentally validated. Cell viability was detected by cell counting kit-8; apoptosis was analyzed by flow cytometry with Annexin V/propidium iodide and JC-1 staining; cell cycle by DNA quantitation; migration by scratch assay; invasion by transwell assay; and protein expression by western blotting. Computational analysis identified EGFR, protein kinase B (AKT), and matrix metalloproteinase-9 (MMP-9) as PEC’s core targets, with molecular dynamics confirming stable PEC-EGFR binding. PEC selectively inhibited RA-FLS viability with minimal toxicity to normal cells. Mechanistically, PEC induced mitochondrial-dependent apoptosis by upregulating Bcl2-associated X, downregulating B-cell lymphoma-extra large, and activating caspase-3 and poly ADP-ribose polymerase cleavage. Furthermore, it caused G2/M phase arrest by suppressing cyclin-dependent kinases 1/2 and cyclin B, while upregulating cyclin-dependent kinase inhibitors p21/p27. PEC also potently inhibited cell migration and invasion by reversing the epithelial–mesenchymal transition (modulating E-cadherin and N-cadherin) and downregulating MMP-9. Importantly, these coordinated effects were primarily mediated through the EGFR/AKT pathway. The critical role of the EGFR/AKT pathway was demonstrated by the observations that GEF synergistically enhanced PEC’s efficacy, whereas the EGFR agonist NSC228155, through restoring phospho-EGFR and phospho-AKT levels, significantly attenuated PEC-induced apoptosis, cell cycle arrest, and inhibition of invasion. In conclusion, our findings demonstrate that PEC suppresses multiple pathogenic hallmarks of RA-FLSs by inhibiting the EGFR/AKT axis, highlighting its promise as a novel targeted therapeutic candidate for RA.