The transforming growth factor-β (TGF-β) signaling pathway is a central driver in the pathogenesis of pulmonary fibrosis (PF), and strategies targeting this pathway demonstrate therapeutic potential. However, the ubiquitous blockade of TGF-β signaling is associated with detrimental effects due to its pleiotropic involvement in physiological processes. Here, we demonstrate that blocking neuropilin-1 (NRP-1), a high-affinity TGF-β co-receptor, with host defense peptide melittin (MLT) attenuates PF progression. Molecular docking and surface plasmon resonance (SPR) demonstrated direct binding between MLT and NRP-1. In vitro studies revealed that MLT selectively targeted endothelial cells with high NRP-1 receptor expression, suppressing endothelial-to-mesenchymal transition (EndMT) via inhibition of TGF-β/Smad and MAPK pathways activation. In a bleomycin-induced PF model, elevated NRP-1 expression enhanced fluorescence-labeled MLT targeting in fibrotic lungs, enabling MLT to exert dose-dependent anti-fibrotic effects through EndMT suppression and significantly improve survival in PF mice. Furthermore, in vivo imaging showed that MLT-loaded peptide-lipid nanoparticles (M-pLNPs) formed a depot sustaining lung fluorescence for over 24 h, starkly contrasting with free MLT clearance within 6 h, which may contribute to comparable anti-fibrotic efficacy at a lower dose. Therefore, our results suggest a novel mechanism involving co-receptor blockade for the anti-fibrotic effect of MLT, highlighting its potential as a therapeutic candidate for PF.

The transforming growth factor-β (TGF-β) signaling pathway is a central driver in the pathogenesis of pulmonary fibrosis (PF), and strategies targeting this pathway demonstrate therapeutic potential. However, the ubiquitous blockade of TGF-β signaling is associated with detrimental effects due to its pleiotropic involvement in physiological processes. Here, we demonstrate that blocking neuropilin-1 (NRP-1), a high-affinity TGF-β co-receptor, with host defense peptide melittin (MLT) attenuates PF progression. Molecular docking and surface plasmon resonance (SPR) demonstrated direct binding between MLT and NRP-1. In vitro studies revealed that MLT selectively targeted endothelial cells with high NRP-1 receptor expression, suppressing endothelial-to-mesenchymal transition (EndMT) via inhibition of TGF-β/Smad and MAPK pathways activation. In a bleomycin-induced PF model, elevated NRP-1 expression enhanced fluorescence-labeled MLT targeting in fibrotic lungs, enabling MLT to exert dose-dependent anti-fibrotic effects through EndMT suppression and significantly improve survival in PF mice. Furthermore, in vivo imaging showed that MLT-loaded peptide-lipid nanoparticles (M-pLNPs) formed a depot sustaining lung fluorescence for over 24 h, starkly contrasting with free MLT clearance within 6 h, which may contribute to comparable anti-fibrotic efficacy at a lower dose. Therefore, our results suggest a novel mechanism involving co-receptor blockade for the anti-fibrotic effect of MLT, highlighting its potential as a therapeutic candidate for PF.