Leukocyte recruitment into tissue compartments is a complex process central to all inflammatory responses. This process involves multiple discrete sequential steps that are orchestrated by a diverse array of chemoattractant molecules, including lipid metabolites (e.g., leukotrienes), proteolytic fragments of serum proteins (e.g., complement) or secreted chemotactic cytokines (e.g., chemokines). Though chemically diverse, these chemoattractants all induce cell migration by activating a family of related chemoattractant G protein-coupled receptors (GPCRs). Although these chemoattractants appear to have overlapping functions in vitro, in vivo studies have uncovered a complexity of leukocyte migration control not appreciated by simple in vitro chemotaxis assays. In the prior funding period, we have contributed to this evolving concept by defining the sequential chemoattractant control of effector T cell migration into the asthmatic airway, initially directed by the leukotriene B4 (LTB4) receptor BLT1 and subsequently by chemokine receptors. In this period, we have also discovered that neutrophil synthesis of LTB4 and BLT1-induced neutrophil responses to LTB4 are absolutely required for the development of immune complex-induced inflammatory arthritis. Surprisingly, we found that the primary pathogenic role for this autocrine LTB4-BLT1 loop is to enable neutrophils to enter the joint via other chemoattractant receptors, indicating that different chemoattractants collaborate to mediate different phases of neutrophil recruitment. We have also found that BLT1 signaling in the neutrophil is upstream of IL-1 and chemokine generation, and that the neutrophil is the critical cellular source of IL-1 in immune-complex induced arthritis. Based on these novel findings, the central hypothesis of this renewal application is that multiple chemoattractant receptors cooperate sequentially to direct neutrophil trafficking into the inflamed joint. We describe four phases of neutrophil recruitment into the joint as: Initiation, Amplification, Facilitation, and Progression. We propose that these phases are sequentially controlled by different chemoattractant receptors, including the complement receptor C5aR, the LTB4 receptor BLT1, and the chemokine receptors CCR1 and CXCR2. Specifically, we propose to: (1) determine the mechanism of initial neutrophil recruitment into the joint and activation upstream of BLT1 signaling;(2) determine the role of the inflammasome in BLT1-induced neutrophil IL-1 production in the amplification of neutrophil recruitment and correlate this to human neutrophils isolated from the blood and joints of patients with active rheumatoid arthritis;(3) determine the mechanism by which BLT1 on facilitates leukocyte transendothelial migration;and (4) determine the respective and differential roles of the neutrophil chemokine receptors CCR1, CXCR1, CXCR2, and their chemokine ligands, in the progression of neutrophil accumulation into the inflamed joint. These studies will define the unique and sequential roles of chemoattractant receptors in the control of neutrophil trafficking in inflammatory arthritis.
Many chronic inflammatory diseases arise from the abnormal accumulation of white blood cells in affected tissue, such as the joint in rheumatoid arthritis. White blood cells are drawn into inflamed tissue from the blood by chemical messengers that are released at sites of inflammation. We have identified a protein called BLT1 that is present upon the surface of white blood cells that is a receptor for one of these important messengers, LTB4. We have found that LTB4 signaling through BLT1 is absolutely required for white blood cells to enter inflamed tissue in certain mouse models of disease, most strikingly in a model of inflammatory arthritis. Further understanding of how BLT1 collaborates with other related receptors, such as the chemokine receptors, to mediate these effects is the focus of this proposal, and will allow for the development of new therapies aimed at inhibiting white blood cell recruitment into inflamed tissue. We expect such therapies will be relevant for arthritis as well as many other chronic inflammatory diseases.
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