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The Journal of Immunology, 2002, 168: 4559-4566.
Copyright © 2002 by The American Association of Immunologists

Signaling Pathways Involved in IL-8-Dependent Activation of Adhesion Through Mac-11

Mimi Takami*, Valeri Terry{dagger} and Lilli Petruzzelli2,{dagger}

Divisions of * Gastroenterology and {dagger} Hematology/Oncology, Department of Internal Medicine, University of Michigan Medical Center and Department of Veterans Affairs Medical Center, Ann Arbor, MI 48109

In human neutrophils, IL-8 induces chemotaxis, the respiratory burst, and granule release, and enhances cellular adhesion, a {beta}2 integrin-dependent event. IL-8 stimulates neutrophil adhesion to purified fibrinogen in a Mac-1-dependent manner. Mitogen-activated protein kinase (MAPK) activation was detected in human neutrophil lysates after treatment with IL-8 and PMA, but not the activating mAb CBR LFA 1/2. IL-8-stimulated neutrophil adhesion to fibrinogen was blocked 50% by the MAPK/extracellular signal-related kinase-activating enzyme inhibitor PD098059. Adhesion was blocked ~75% by inhibition of the phosphatidylinositol-3 kinase (PI3K) pathway with LY294002, supporting that activation of both MAPK and PI3K may play a role in IL-8-dependent inside-out signals that activate Mac-1. Activation of MAPK was inhibited in IL-8-stimulated cells in the presence of PI3K inhibitors LY294002 or wortmannin, supporting a model in which PI3K is upstream of MAPK. IL-8-stimulated neutrophil adhesion was inhibited 50% by bisindolylmaleimide-I, implicating protein kinase C (PKC) in the intracellular signaling from the IL-8R to Mac-1. A 74-kDa molecular mass species was detected by an activation-specific Ab to PKC when cells were stimulated with PMA or IL-8, but not a {beta}2-activating Ab. Inhibition of either MAPK or PKC resulted in partial inhibition of IL-8-stimulated polymorphonuclear neutrophil adhesion, and treatment with both inhibitors simultaneously completely abolished IL-8-stimulated adhesion to ligand. Inhibition of PI3K blocked MAPK activation, but not PKC activation, suggesting a branch point that precedes PI3K activation. These data suggest that both MAPK and PKC are activated in response to IL-8 stimulation, and that these may represent independent pathways for {beta}2 integrin activation in neutrophils.




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