The natural coumarin 7-methoxy-8-isopentenyloxycoumarin alleviates LPS-induced inflammation by suppressing NF-κB and the MAPK/AP-1 pathways and accelerating COX-2 proteasomal degradation.
Inflammatory diseases impose a significant global healthcare burden, necessitating the development of effective anti-inflammatory agents. Many clinically used drugs are isolated from plants or derived from natural products via semi-synthetic procedures. Among natural products, coumarins have demonstrated broad-spectrum pharmacological activities. This study investigates the anti-inflammatory mechanisms of an uncommon natural coumarin, 7-methoxy-8-isopentenyloxycoumarin (7MI), isolated from the seed of the higher Mediterranean plant Magydaris pastinacea. 7MI's effects were assessed in LPS-stimulated Raw264.7 macrophages using immunoblotting, RT-qPCR, ChIP assay, chase assay, reporter assay, ELISA, phosphatase activity assay, and transfection. 7MI significantly reduced the release of pro-inflammatory cytokines, including IL-6 and MCP-1, and suppressed cyclooxygenase-2 (COX-2) expression. Mechanistically, 7MI attenuated canonical LPS-induced TLR4 signaling by inhibiting the parallel activation of the NF-κB and MAPK/AP-1 pathways. 7MI blocked NF-κB p65 nuclear translocation and its transcriptional activity. Concurrently, 7MI enhanced MAPK phosphatase-1 (MKP-1) activity, which deactivated p38MAPK, thereby dampening activation of AP-1 and the synergistic transcription factor CCAAT/enhancer-binding protein (C/EBP)β. Beyond transcriptional suppression, 7MI exerted a distinct post-translational effect by accelerating COX-2 proteasomal degradation through inhibition of the deubiquitinases USP7 and USP22, leading to enhanced K48-linked polyubiquitination of the COX-2 protein. Crucially, in vivo administration of 7MI dramatically improved survival rate and mitigated multi-organ damage in a murine model of fatal sepsis. These findings suggest that 7MI exerts broad-spectrum anti-inflammatory effects by simultaneously suppressing canonical TLR4-driven transcriptional cascades to halt de novo inflammatory mediator synthesis and by promoting pro-inflammatory protein clearance, highlighting its potential as a lead compound for LPS-associated inflammatory disorders.