Platelet-derived growth factor-BB (PDGF-BB) regulation of migration and focal adhesion kinase phosphorylation in rabbit aortic vascular smooth musclecells: roles of phosphatidylinositol 3-kinase and mitogen-activated protein kinases

Objective: Phosphatidylinositol 3'-kinase (PI3-kinase) is implicated in cel l migration and focal adhesion kinase (FAK) phosphorylation. In contrast, i t has been proposed that mitogen-activated protein (MAP) kinases are essent ial for proliferation but may be dissociated from chemotactic signalling. W e investigated the roles of PI3-kinase and p42/p44 MAP kinases in cell migr ation and FAK tyrosine phosphorylation induced by platelet-derived growth f actor-BB (PDGF-BB)in rabbit aortic vascular smooth muscle cells (VSMCs). Th e roles of PI3-kinase and MAP kinase pathways in the chemotactic response t o insulin-like growth factor-I (IGF-I) were also examined. Methods: The rol es of PI3-kinase and p42/p44 MAP kinases were assessed using the PIS-kinase inhibitors, wortmannin and LY294002, and an inhibitor of MAP kinase kinase , PD98059. PI3-kinase activity was measured by phosphatidylinositol phospho rylation in anti-phosphotyrosine immunoprecipitates and by thin layer chrom atography of phosphorylated products. Phosphorylation was assessed by immun oprecipitation with anti-phosphotyrosine antibodies and Western blotting wi th FAK-specific antibody. Migration was evaluated in a chemotaxis chamber u sing polycarbonate filters with an 8-mm pore size. Results: Neither wortman nin nor LY294002 significantly reduced PDGF-BB stimulation of FAK tyrosine phosphorylation, chemotaxis or immunofluorescent staining of focal adhesion s in VSMCs. PD98059, a specific inhibitor of MAP kinase activation, did not inhibit FAK tyrosine phosphorylation but markedly inhibited the migratory response of VSMCs to PDGF-EB. IGF-I also stimulated migration of VSMCs, and , relative to the effect of PDGF-BE, induced smaller increases in PIS-kinas e and MAP kinase activities. Both wortmannin and PD98059 partially inhibite d the migratory response to IGF-I. Conclusions: PDGF-BB stimulation of both FAK tyrosine phosphorylation and migration in VSMCs are not dependent on a ctivation of PI3-kinase. While PDGF-BB stimulation of FAK tyrosine phosphor ylation is not dependent on p42/p44 MAP kinase activation, PDGF-BB and IGF- I bath stimulate p42/p44 MAP kinase activity and the chemotactic response t o these factors is partially dependent on MAP kinase activation. (C) 1999 E lsevier Science B.V. All rights reserved.