INTERLEUKIN-8 REGULATION OF THE RAS RAF/MITOGEN-ACTIVATED PROTEIN-KINASE PATHWAY IN HUMAN NEUTROPHILS/

Interleukin-8 (IL-8), the prototypic member of the CXC subfamily of ch emokines, induces in neutrophils chemotaxis, the respiratory burst, gr anule release, and increased cell adhesion, The IL-8 receptor is a sev en-transmembrane spanning receptor coupled to specific heterotrimeric G proteins including G(i) and G(16). IL-8 stimulation of its receptor on neutrophils activates Ras GTP loading and the mitogen-activated pro tein kinase (MAPK) pathway including Raf-1 and B-Raf. The properties o f IL-8 stimulation of the MAPK pathway differ from those observed for chemoattractants such as C5a. Even though Ras GTP loading is similar f or IL-8 and C5a, the maximal activation of Raf-1 and B-Raf is approxim ately 2-fold and 3-7-fold, respectively, less for IL-8 than that obser ved for C5a. Raf-1 activation is rapid but transient, returning to nea r basal levels by 10 min. B-Raf activation is slower in onset and does not return to basal levels for nearly 30 min, IL-8 activation of MAPK follows a time course suggesting an involvement of both Raf-1 and B-R af. Surprisingly, wortmannin, at low concentrations, inhibits Raf-1, B -Raf, and MAPK activation in response to IL-8 and C5a demonstrating a role for phosphatidylinositol 3-kinase in the activation of Raf kinase s in G protein-coupled receptor systems in human neutrophils. Furtherm ore, wortmannin inhibits IL-8 stimulated granule release and neutrophi l adherence, These findings demonstrate the control of Raf kinases, th e MAPK pathway and specific neutrophil functions by phosphatidylinosit ol 3-kinase enzymes.