In response to inflammatory stimuli, neutrophils become activated resulting in increased adhesiveness, chemotaxis, generation of toxic oxygen metabolites and upregulation of IgG Fc receptor (FcR) and complement receptor function. Although these neutrophil responses are critical for host defense, they also lead to significant tissue damage contributing to many disease processes. In vivo studies have demonstrated that monoclonal antibody (mAb) blockage of a family of leukocyte surface receptors, the beta2 integrins, can significantly decrease this tissue destruction at inflammatory sites. In vitro studies by the applicant have shown that CR3 is involved in neutrophil activation via the IgG FcRs, including generation of the chemotaxin and proinflammatory LTB4, and augmentation of FcR dependent adhesion and phagocytosis. In order to investigate the molecular mechanism of this FcR/CR3 interaction, a transfection system has been developed that mimics the functional interaction of FcRII and CR3 in neutrophils. In this system, FcR-mediated phagocytosis only occurs in cells transfected with CR3. Recent data demonstrate that a specific cytoskeletal protein, paxillin, becomes tyrosine phosphorylated in a CR3 dependent manner in the transfected K562 and in PMN. Furthermore, CR3 transfectants show a general decrease in other phosphotyrosine containing proteins compared with controls. In addition, phagocytosis of IgG-opsonized particles is maximal when CR3 is cotransfected with the leukocyte tyrosine phosphatase, SHP. Together these data suggest the hypothesis that CR3 is involved in the regulation of tyrosine phosphorylation associated with PMN activation. The applicant proposes to localize the domains in CR3 necessary for cooperation with FcRII, and to test the hypothesis that the CR3 effect on FcRII function is through modulation of tyrosine phosphorylation. To accomplish these goals, the applicant proposes to (1) determine the effect of mutations of CR3 on complement- and IgG-dependent phagocytosis, adhesion, and cytoskeletal associations; (2) investigate the role of CR3 in regulation of tyrosine phosphorylation, and (3) characterize the phagocytic function of tyrosine phosphatase (SHP) transfected cells. Understanding the mechanism of interaction between FcRII and CR3 may be useful in designing strategies to control inflammatory tissue damage. Pharmacologic inhibition of the interaction or its consequences has the possibility of being antiinflammatory and at the same time not affecting CR3-dependent adhesion of PMN to endothelium, which is critical for normal host defense. Potential applications are quite broad including treatment of autoimmune diseases, as well as preservation of myocardium following infarcts.
| Gresham, H D; Graham, I L; Griffin, G L et al. (1996) Domain-specific interactions between entactin and neutrophil integrins. G2 domain ligation of integrin alpha3beta1 and E domain ligation of the leukocyte response integrin signal for different responses. J Biol Chem 271:30587-94 |
