Transforming growth factor-beta (TGF-beta) reportedly induces vascular endothelial growth factor (VEGF) synthesis in osteoblast-like MC3T3-E1 cells. We have recently shown that TGF-beta activates p44/p42 mitogen-activated protein (MAP) kinase and p38 MAP kinase in these cells. In the present study, we investigated the exact mechanism of TGF-beta behind the synthesis of VEGF in MC3T3-E1 cells. PD98059 and U-0126, specific inhibitors of MEK, suppressed the VEGF synthesis induced by TGF-beta. U-0126 inhibited the TGF-beta-induced p44/p42 MAP kinase phosphorylation. SB203580 and PD169316, inhibitors of p38 MAP kinase, reduced the TGF-beta-stimulated VEGF synthesis. SB202474, a negative control for p38 MAP kinase inhibitor, did not affect the VEGF synthesis. A combination with PD98059 and SB203580 almost completely suppressed the TGF-beta-induced VEGF synthesis. Retinoic acid, which alone failed to affect VEGF synthesis, markedly enhanced the VEGF synthesis stimulated by TGF-beta. Retinoic acid enhanced the TGF-beta-increased levels of VEGF mRNA. The amplifications by retinoic acid of TGF-beta-increased VEGF synthesis and levels of VEGF mRNA were reduced by PD98059 or SB203580. The combination of PD98059 and SB203580 almost completely suppressed the enhancement by retinoic acid of VEGF synthesis induced by TGF-beta. Taken together, our results strongly suggest that both p44/p42 MAP kinase and p38 MAP kinase take part in TGF-beta-stimulated VEGF synthesis in osteoblasts, and that retinoic acid upregulates the VEGF synthesis.