Modulation of the erythropoietin-induced proliferative pathway by cAMP in vascular smooth muscle cells.

We previously reported that erythropoietin (Epo) has a mitogenic effect on rat vascular smooth muscle cells (VSMC) and that activation of the mitogen-activated protein kinase (MAPK) cascade is an important mediator for Epo-induced mitogenesis. An increase in intracellular cAMP has an antiproliferative effect on VSMC. We therefore hypothesized that cAMP effectors inhibit Epo-induced MAPK activation in rat VSMC. When we exposed VSMC to recombinant human Epo (rHuEpo), DNA synthesis was increased. Forskolin (Fsk) or cilostazol (Cil) decreased the DNA synthesis stimulated by rHuEpo. Coincubation with Rp-cAMPS triethylamine canceled the suppression of DNA synthesis and MAPK activity by Fsk. Both rHuEpo and phorbol 12-myristate 13-acetate upregulated phosphorylations of MEK and MAPK. Pretreatment with Fsk inhibited these phosphorylations. Protein kinase C inhibitors also suppressed MEK and MAPK phosphorylations. Moreover, Fsk induced phosphorylation of Raf-1 at serine-259. These results indicated that cAMP inhibited Epo-induced MAPK activation and that this suppression might be regulated upstream or at Raf-1. The results also suggested that these agents, which could accumulate cAMP, might be protective for Epo-stimulated direct action.