The long-term goal of this project is to define the receptors and the molecular mechanisms by which neutrophils interact with immune complexes (ICs) and activate signaling pathways that contribute to cellular functions. These events are fundamental to physiologic inflammation, but their dysregulation is a defining feature in IC-mediated disorders. Work on this project demonstrated a direct role for neutrophil FcgammaRIII (FcgRIII) in IC-induced neutrophil recruitment in vitro under physiological flow and following in situ IC deposition in a model of anti-glomerular basement membrane (GBM) nephritis. A murine model of IC deposition that was amenable to intravital microscopy was developed to provide mechanistic details of the recruitment process. Fc'yRs promoted slow leukocyte rolling; a step that favors adhesion and FcgRIII in particular was essential for firm adhesion and transmigration. Previous studies have shown that the leukocyte CD18 integrin Mac-1 (CD11b/CD18) is required for IC-mediated adhesion. FcgRIII physically associates with Mac-1 on the cell surface through lectin-carbohydrate interactions and FcgRIIl-ICs interactions leads to Mac-1 activation. Studies in Mac-1 deficient (-/-) mice revealed an essential role for Mac-1 in FcgRIII dependent adhesion and antibody dependent cytotoxicity (ADCC) in vivo. Mac-1 on murine neutrophils was also needed for ADCC mediated by transgenically expressed human FcaR. Together our studies place murine neutrophil FcRs and Mac-1 at critical junctures in acute inflammation and suggest that their cooperation in IC-mediated functions is biologically important. The molecular basis of FcgRIII-Mac-1 cooperation remains unclear. Preliminary data suggest that a residue in the extracellular I-like domain of the CD18 subunit and activation of Mac-1 ligand binding promotes neutrophil interactions with ICs.
The aims i n the current application builds on our ongoing work based on the central hypothesis that human FcgRIIIB and IIA and Mac-1 are required for neutrophil recruitment and cytotoxicity in vivo, and FcyR-Mac-1 cooperate, throuqh interactions of their extracellular domains, to promote IC-mediated neutrophil functions.
The aims are: I) To identify the in vivo role of human neutrophil FcgRIIA and the GPl-linked FcgRIIIB (for which there are no genetic equivalents in mice) in ICinduced neutrophil recruitment and adhesion, and examine the biological importance of Mac-1 in the context of these human FcgRs; and II) To elucidate the molecular basis for FcgR-Mac-1 cross-talk focusing on the role of the extracellular CD1 lb ligand binding I and lectin-like domains, and the I-like domain of CD18 in this cross-talk. These studies should provide valuable data on human FcgR functions and their cooperation with Mac-1 in vivo, which could have direct clinical relevance, and will provide new concepts on how Mac-1 and FcgR cross-talk to promote neutrophil adhesion to ICs.
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