Ibuprofen impairs fracture healing by disrupting neutrophil-mediated tissue repair

Abstract

Neutrophils are among the first immune cells recruited to fractures, yet their contribution to bone repair remains poorly understood. Anti-inflammatory use has been linked to increased risk of non-union fractures, while prolonged neutrophil activity or failed clearance of neutrophil-derived DNA in the form of neutrophil extracellular traps (NETs), can also impair fracture healing. Using dynamic live imaging and cellular perturbation in adult zebrafish, we show that neutrophils play a key role in shaping the early fracture microenvironment. Neutrophils rapidly accumulated at fractures where they released NETs and exhibited the activation of gene programmes associated with matrix remodelling. Compromising neutrophil recruitment, via ibuprofen treatment or orthogonal genetic and pharmacological perturbations, dramatically increased incidence of fracture non-union and lead to altered callus architecture through delay of osteoblast differentiation and impaired mineralisation. Mechanistically, neutrophils actively interacted with key structural proteins, including laminin, fibronectin, and collagen I, through uptake, recycling and active de novo secretion pathways. Together, our findings reveal that neutrophils establish a provisional emergency fracture matrix that templates subsequent bone repair, providing a mechanistic link between early non-steroidal anti-inflammatory drug (NSAID) exposure and impaired skeletal healing.