Pomolic Acid from the Dong Botanical Drug Madeng’ai Suppresses TNF-α-Induced Inflammatory Response in RA-HFLS by Inhibiting NF-κB Signaling Pathway Activation

Objective: This study aimed to investigate whether pomolic acid (PA), a predicted bioactive metabolite of the Dong botanical drug Madeng’ai (MDA), suppresses inflammatory cytokine expression by inhibiting nuclear factor-κB (NF-κB) pathway activation in a tumor necrosis factor-α (TNF-α)-induced human rheumatoid arthritis fibroblast-like synoviocyte (RA-HFLS) model. Methods: PA content in MDA from different regions and harvest years was quantified via High-Performance Liquid Chromatography (HPLC). Network analysis was employed as a hypothesis-generating tool to predict potential targets and pathways, followed by molecular docking to validate the binding affinity of PA to core targets of the NF-κB pathway, and ADMET prediction to evaluate its pharmacokinetic properties and safety profile. The RA-HFLS inflammatory model was induced by TNF-α. Cell viability and inflammatory cytokine secretion were assessed using Cell Counting Kit-8 (CCK-8) and enzyme-linked immunosorbent assay (ELISA). NF-κB signaling pathway activation and downstream gene expression were examined by Western blot and reverse transcription quantitative polymerase chain reaction (RT-qPCR), respectively. Results: HPLC analysis revealed that MDA samples from Guizhou harvested in 2019 contained the highest PA content (0.1117 mg/g). Network analysis predicted the NF-κB signaling pathway as a candidate mechanism underlying PA’s potential anti-inflammatory effects. Molecular docking showed that PA stably bound to IKKβ, p65, and IκBα, while ADMET prediction indicated favorable intestinal absorption, low drug–drug interaction risk, and good genetic safety, albeit with potential hepatotoxicity and reproductive toxicity risks. In the TNF-α-induced RA-HFLS model, PA dose-dependently inhibited abnormal cell proliferation and significantly reduced the secretion of pro-inflammatory cytokines TNF-α and interleukin-6 (IL-6). Mechanistic studies indicated that PA suppressed the activation of the NF-κB signaling pathway, thereby downregulating the mRNA expression of inflammatory genes such as IL-6, TNF-α, and interleukin-1β (IL-1β). Conclusions: PA, a bioactive metabolite of the Dong botanical drug MDA, may inhibit NF-κB signaling pathway activation, thereby downregulating TNF-α-induced inflammatory cytokine expression in RA-HFLS, demonstrating its in vitro anti-inflammatory potential.