Redox regulation of ERK1/2 activation induced by sphingosine 1-phosphate in fibroblasts: involvement of NADPH oxidase and platelet-derived growth factor receptor.

Sphingosine 1-phosphate (S1P) is a sphingolipid metabolite synthesized after stimulation with growth factors or cytokines. S1P extracellular effects are mediated through specific Gi-protein coupled receptors (GPCRs). Recently, we demonstrated in NIH3T3 fibroblasts stimulated by platelet-derived growth factor (PDGF) or S1P the NADPH oxidase activation and the H(2)O(2) intracellular level increase trough the Gi protein involvement. NIH3T3 fibroblast cell cultures were used. Western blot and quantitative analyses by Chemidoc-Quantity-One software were performed. H(2)O(2) level was assayed by fluorescence spectrophotometric analysis, and cell proliferation by counted manually or ELISA kit. This study demonstrates, in NIH 3T3 fibroblasts, a novel redox regulated mechanism of S1P-induced activation of ERK 1/2 related to NADPH oxidase activity and intracellular H(2)O(2) level increase with PDGF receptor tyrosine kinase involvement through a transactivation mechanism. This event is mediated by S1P(1) and S1P(3) receptors by Gi proteins and can contribute to S1P mitogenic signaling. These results can be related to mechanisms of cross-talk previously identified between receptor tyrosine kinase, including PDGFreceptor, and several GPCR ligands. The redox-sensitive ERK1/2 and PDGFr tyrosine kinase activity could be targets for therapies in diseases in which deregulation of intracellular oxidative status and the consequent alteration of S1P and/or PDGF signaling pathway are involved.