Objective <p>This study aimed to elucidate the molecular mechanism of Er Miao San (EMS) in treating rheumatoid arthritis (RA) by integrating network pharmacology, molecular docking, and experimental validation.</p> Methods <p>Using public databases, potential targets of EMS, RA-related targets, and NF-κB pathway targets were cross-referenced to identify common genes. These were analyzed via protein-protein interaction (PPI)network, GO/KEGG enrichment analyses (via DAVID), and hub gene identification (using Cytoscape and cytoHubba). Core predictions were validated by molecular docking, with therapeutic efficacy confirmed in adjuvant-induced arthritis (AA)rats and NF-κB pathway inhibition confirmed in TNF-α-induced fibroblast-like synoviocytes (FLSs).</p> Results <p>Integrated analysis identified 34 common targets of Er Miao San (EMS) for rheumatoid arthritis (RA) treatment, with NFKBIA, RELA, and TNF recognized as the top hub genes. Molecular docking revealed stable binding between these targets and the core active components (fumarine, berberine, and wogonin). Experimentally, EMS alleviated joint pathology in AA rats and concentration-dependently suppressed TNF-α-induced proliferation of FLSs and the secretion of IL-1β and IL-6. Mechanistically, EMS inhibited NF-κB signaling by reducing the phosphorylation of IκBα and p65 and blocking the nuclear translocation of p65.</p> Conclusion <p>The therapeutic effects of EMS on RA are mediated through the inhibition of the NF-κB signaling pathway, thereby elucidating its mechanism of action.</p>