Leukotriene B4 (LTB4) is a potent lipid chemoattractant that classically has been associated with myeloid cell chemotaxis. We isolated a novel murine seven transmembrane spanning G protein-coupled receptor, which we identified as BLTR1, an LTB4 receptor, and generated BLTRl-deficient (BLTR1 -/) mice by targeted gene disruption. Characterization of this mouse revealed that BLTR1 is responsible for LTB4- mediated leukocyte calcium flux, chemotaxis, and firm adhesion to endothelium. Further, despite functional redundancy with other chemoattractant-receptor pairs in vitro, loss of BLTR1 function significantly reduced eosinophil recruitment into the peritoneum in a model of peritonitis. Follow-up studies of BLTR1 -/- mice have allowed us to make several novel observations about the roles of LTB4 and BLTR1 in leukocyte recruitment that will be the focus of this grant proposal. We have found that loss of BLTR1 function significantly and specifically impairs neutrophil transendothelial migration (TEM) in vitro and in vivo. Secondly, we have found that BLTR1 is highly expressed and functional on certain subsets of activated T cells and important for their trafficking in vivo. Additionally, we have found that recruitment of BLTR1 /- T cells into the airways early in allergic pulmonary inflammation is profoundly impaired, despite preserved trafficking of these cells into the lung parenchyma. Based on these novel preliminary findings, the central hypothesis of this proposal is that LTB4 and BLTR1 play a critical role in facilitating the ability of leukocytes to traverse endothelial, interstitial and epithelial barriers. We hypothesize that interactions between leukocytes and barriers to migration they encounter, e.g. endothelial cells, tissue interstitium, and epithelial cells, induces the production of LTB4. We hypothesize that the LTB4 produced by these interactions in turn activates leukocyte BLTR1, which facilitates penetration of these barriers by the migrating leukocytes. We believe that the LTB4 biosynthetic pathway uniquely positions this chemoattractant-receptor pair for this role. LTB4 is produced by serial enzymatically catalyzed reactions, and hence can be produced much more rapidly than peptide chemokines, which require transcription and translation. The rapid kinetics of LTB4 biosynthesis enables leukocytes, endothelial cells and epithelial cells to produce LTB4 contemporaneously with leukocyte migration across cellular and extracellular barriers. In this grant, we propose to investigate this hypothesis as it relates to neutrophil and lymphocyte recruitment. Specifically we propose to: (1) determine to cellular source of the LTB4 that facilitates neutrophil transendothelial migration; (2) define the roles of BLTR1 and CXCR2 in neutrophil recruitment into the airways; (3) determine the role of LTB4 and BLTR1 in antigen specific T cell trafficking; (4) determine the role of LTB4 and BLTR1 in T cell recruitment into the airways of the lung.
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