The indications for lung transplantation have expanded significantly, resulting in a clinical growth of over 50% in the last decade. However, the survival following lung transplant is the worst compared to other solid organs with only 80% and 50% of patients alive at 1 and 5 years, respectively (www.unos.org). Primary graft dysfunction (PGD), resulting from ischemia-reperfusion injury, affects over 50% of recipients within 24 hours of transplantation and has emerged as the most important risk factor for both short-term mortality as well as long-term graft loss from chronic rejection. As such, therapies directed to ameliorate PGD have the highest potential for improving lung transplant outcomes. Prior reports have demonstrated that recruitment and extravasation of neutrophils into the allograft leads to the formation of neutrophil extracellular traps which play a pivotal role in the development of PGD. Depleting neutrophils can ameliorate PGD, but is not clinically feasible given their importance in pathogen clearance. Accordingly, we have focused on understanding the mechanisms that drive neutrophil recruitment to the lung following transplantation. We discovered that orphan nuclear receptor (NR4A1)-dependent non-classical monocytes (NCM), characterized by CD14dimCD16++ in humans and Ly6ClowCX3CR1highCCR2- in mice, are bound to the pulmonary endothelium via a leukocyte integrin (lymphocyte factor associated antigen-1) and retained in the donor lungs despite the current techniques for organ perfusion and preservation. Following murine and human lung transplantation, activated donor-derived NCM mediate neutrophil recruitment and extravasation into the allograft through mechanisms that remain unknown. Our published and preliminary data suggest that activation of toll-like receptors in donor-derived NCM leads to the production of neutrophil chemoattractant, macrophage inflammatory protein 2 (MIP-2). The donor NCM also release IL-1 and contribute to enhanced vascular permeability in the allograft. Importantly, using a genetic lineage tracing system we found that donor-derived NCM also migrate to distant organs after transplant where they recruit neutrophils. Collectively, these data support our hypothesis that donor-derived NCM are activated following lung ischemia-reperfusion, drive neutrophil influx into the allograft, and promote secondary injury to distant organs. We have developed a model of murine vascularized orthotopic lung transplantation where lungs from NR4A1-deficient donor mice, which selectively lack NCM, can be reconstituted with fluorescent wild-type or genetically mutated NCM to study the signaling pathways in donor-derived NCM and their contribution to lung IRI. Accordingly, we will dissect the molecular mechanisms underlying NCM-mediated injury in the allograft and bystander tissues. Specifically, we will determine: 1) The toll-like receptors responsible for the activation of donor NCM, 2) The mechanisms through which donor NCM migrate to distant organs and mediate bystander injury, 3) The role of NLRP3 inflammasome in the production of IL-1 and extravasation of neutrophils into the extravascular space. Through these experiments, we will identify pathways that could be clinically targeted, prior to transplantation, to ameliorate PGD without causing recipient toxicity.
|Effective start/end date||1/1/19 → 12/31/23|
- National Heart, Lung, and Blood Institute (5R01HL145478-02)