(105) - The Real Time Visualization of Neutrophil Extracellular Trap Formation in Lung Transplants Reveals a Critical Role in Tolerance

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Methods: The pericardiophrenic artery was procured from donor lungs transplanted in our clinical program and was implanted into the abdominal aorta of immune deficient mice. Of 17 lung recipients, 10 (43.47%) developed severe PGD (grade 2-3) and 13 (56.52%) did not develop PGD (grade 0-1) at 24 h. Negative control mice received no human leukocyte reconstitution (neg. co). PBMC PGD+ group mice received 5x106 allogeneic human peripheral blood mononuclear cells (PBMC) from recipients with severe PGD, whereas PBMC PGDgroup mice received PBMC from patients without PGD. Two groups of animals were reconstituted with the respective PBMC additionally enriched with autologous CD4+CD25 high cells (putative regulatory T cells, Treg) from recipients with severe PGD (PBMC+Treg PGD+ group) or without PGD (PBMC+Treg PGDgroup). Transplant arteriosclerosis was histologically assessed 28 days after PBMC reconstitution. Results: The neg. co group showed only mild thickening of the intima (13.86±14.22%). In the PBMC PGD+ group, intimal thickening obliterating the vessel lumen was significantly more severe than in the PBMC PGDgroup (41.71 ± 15.01% vs. 10.52 ± 10.22%, p= 0.001). Then, intimal thickening was significantly inhibited in the PBMC+Treg PGD+ group as compared to the PBMC PGD+ group (3,42 ± 5,41% vs. 41.71 ± 15.01%, p= 0.016). In the experiments using PBMC from lung recipients without PGD, enriching Treg also further suppressed the development of transplant arteriosclerosis (3.88 ± 5.54% PBMC PGDvs. 10.52 ± 10.22%, PBMC+Treg PGD-, p= 0.010). Conclusion: Leukocytes from lung transplant recipients with PGD transfer proinflammatory properties eliciting transplant arteriosclerosis into a humanized mouse model. This indicates a link between early graft dysfunction and the later development of chronic rejection. Importantly, however, transplant arteriosclerosis remains sensitive to inhibition by autologous Treg, suggesting a novel cell therapy-based approach for the treatment of patients with severe PGD.