Targeting the proline-glycine-proline-protease feed-forward loop attenuates primary graft dysfunction after lung transplantation.

Introduction: Primary graft dysfunction (PGD) is the leading cause of early mortality after lung transplantation, yet no targeted therapy exists. We investigated whether the collagen-derived matrikine proline-glycine-proline (PGP) drives neutrophil-predominant injury in PGD and whether its neutralization confers protection. Methods: Human mini-bronchoalveolar lavage (BAL) fluid was collected 72 hours post-transplantation from recipients with grade 3 PGD and non-PGD controls. In parallel, a murine orthotopic lung transplantation model incorporating 18 hours of cold ischemia was used to reproduce PGD; mice received vehicle (PBS) or the PGP-sequestering tripeptide L-arginine-threonine-arginine (RTR) immediately before reperfusion. Histology, immunofluorescence, LC-MS/MS quantification of acetyl-PGP (acPGP), gelatin zymography for active MMP-9, and ELISA for MMP-9 and prolyl endopeptidase (PE) were performed four hours later. Results: Human PGD BAL contained approximately fourfold higher acPGP, along with significantly elevated MMP-9 and PE, compared with PGD 0 controls. Murine PGD allografts similarly demonstrated dense neutrophilic infiltrates and increased acPGP, MMP-9, and PE expression. RTR treatment markedly reduced histologic injury, neutrophil accumulation, and composite PGD scores while improving oxygenation and allograft lung function. RTR also restored acPGP, MMP-9, PE, and active MMP-9 levels to near-baseline compared with vehicle-treated PGD allografts. Conclusion: These findings delineate a feed-forward PGP-protease circuit linking extracellular matrix degradation to neutrophil recruitment and vascular leak. Neutralizing PGP effectively disrupts this circuit, attenuating graft injury. By connecting extracellular matrix-derived signals to innate immune activation, this work broadens the immunopathologic framework of PGD.