Neutrophil-promoted macrophage state transition coordinates endothelial migration and barrier resealing in zebrafish...

Microvessels are frequently damaged, yet the in vivo temporal logic that couples endothelial gap closure to barrier resealing, and the immune-cell interactions that coordinate these steps, remains poorly defined. Using live imaging of laser-injured zebrafish intersegmental vessels, we define a staged repair program: endothelial cells migrate to bridge the gap, trapped erythrocytes are cleared, and vascular permeability rises transiently before resealing. Neutrophils arrive first, whereas macrophages persist and undergo a neutrophil-dependent functional transition. Early tnfa-associated macrophages promote endothelial migration and erythrocyte debris removal, while later ccl34a.4⁺ pro-remodeling macrophages support barrier restoration. Neutrophil ablation delays this transition and selectively prolongs permeability defects. Neutrophils release CD63⁺ extracellular vesicles that are taken up by macrophages, and inhibiting EV/exosome secretion phenocopies key features of neutrophil loss. These findings provide a temporal framework to dissect immune-endothelial coordination during microvascular repair.