Brain-death (BD) induces a systemic inflammatory response known to impair tissue perfusion and promote graft immunogenicity. Monocytes play a significant role in the regulation of inflammatory responses in experimental models of BD. However, the mechanics of activation, differentiation and tissue recruitment of monocyte populations after BD remains a matter of investigation. In this study we used a non-human primate model of BD to investigate monocyte activation and differentiation kinetics in peripheral blood and in transplantable organs. BD was induced by inflation of a subdurally placed catheter and animals (n=5) were monitored for a 20-hour period. Sham operated animals (n=2) and living donors (LD, n=7) served as controls. Peripheral blood and bone marrow (BM) cells were analyzed by FACS to determine cell composition and activation state. Our results indicate that 12 hours after BD induction there is a 56% increase in total leukocyte count. This increase was mostly driven by neutrophils which increased by 32% and we observed a 3-fold reduction in the monocyte fraction over the course of the 20-hour experimental period. Furthermore, we documented a progressive shift in intermediate monocytes (CD14+, CD16+) at baseline to a classical phenotype (CD14+, CD16dim) at 20 hours post-BD. Analysis of cell populations in the BM at 20 hours showed a predominance of immature and classical monocytes. We also observed increased CD68+ immunohistochemical staining in the liver, heart, pancreas and lung of BD donors when compared to LD controls. Furthermore, renal biopsy specimens were analyzed by microarray and we documented 240 genes that were significantly up-regulated in the context of BD with significant enrichment of biological processes related to acute inflammatory response (p<0.001), macrophage activation (p<0.05), mononuclear cell migration (p<0.05) and cytokine production (p<0.001). There were few significantly regulated markers of dendritic cell, neutrophil, T-cells, and B-cell function. These data indicate that BD induces mobilization of naive BM and circulating monocytes into transplantable organs. Moreover, naive monocytes undergo transcriptional and phenotypical changes indicative of activation and maturation at the tissue level. The impact of monocyte differentiation patterns in the modulation of adaptive immunity after renal transplantation is currently being explored.

CITATION INFORMATION: Chlebeck P, Danobeitia J, Zitur L, Zens T, Xu X, Odorico S, D'Alessandro A, Fernandez L. Patterns of Monocyte Differentiation and Activation in a Non-Human Primate Model of Brain Death. Am J Transplant. 2017;17 (suppl 3).

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