Abstract
BACKGROUND: Acute lung injury (ALI) poses significant clinical challenges due to its irreversible alveolar damage and the limitation of available regenerative therapies. Emerging evidence suggests that macrophage-epithelial crosstalk plays a pivotal role in lung repair; however, the specific molecular mediators underlying this process remain largely undefined. METHODS: To address this gap, we isolated and characterized macrophage-derived exosomes (MD-Exos) using dynamic light scattering, transmission electron microscopy (TEM), and immunoblotting. Proteomic analysis and molecular docking were employed to reveal interactions between BMPR2 on exosomes and BMPR1B on epithelial cells. Single-cell RNA sequencing (scRNA-seq) was utilized to map alveolar cell dynamics. Biochemical assays and confocal colocalization were performed to validate SMAD1 signaling activation. The biodistribution of exosomes was tracked via near-infrared imaging, and AT2-to-AT1 transdifferentiation was assessed through multiplex immunofluorescence and pseudotime trajectory analysis. RESULTS: Proteomic profiling of MD-Exos identified BMPR2 as the predominant component. Molecular docking studies confirmed a strong binding affinity between exosomal BMPR2 and epithelial BMPR1B. Single-cell RNA sequencing and biochemical analyses revealed significant alterations in alveolar macrophage (34% vs 27%) and epithelial cell populations during injury, accompanied by enhanced cellular communication. The characterized macrophage-derived exosomes (163.6 ± 70.2 nm) demonstrated efficient pulmonary targeting, with peak accumulation occurring at 4 hours post-administration. Mechanistically, the formation of the BMPR2-BMPR1B complex activated SMAD1-dependent signaling pathways, as evidenced by strong BMPR1B-SMAD1 colocalization (correlation coefficient 0.94 ± 0.02) and enhanced ID1 expression. CONCLUSION: The BMPR2-BMPR1B interaction was demonstrated to accelerate type II to type I alveolar epithelial cell transdifferentiation, thereby facilitating tissue repair in ALI. Comprehensive toxicological assessment confirmed the safety profile of exosome administration across major organ systems. These findings establish exosomal BMPR2 as a crucial mediator of pulmonary repair through specific molecular recognition and signaling activation, providing new therapeutic strategies for treating acute lung injury.