Figure 3. The Ras and Raf pathways mediate S1P-promoted PDGF-A expression and angiogenesis. (A) Cells were pretreated for 30 min with manumycin A (10 μM) and GW5074 (10 μM), or transfected with Ras and Raf siRNAs then stimulated with S1P (10 μM). PDGF-A expression was examined by qPCR assays (n=5). (B, C) The CM was applied to EPCs and analyses assessed migratory and tube formation activity (n=4). (D) JJ012 cells were incubated with S1P; Ras and Raf activity was examined by Western blot assay (n=3). (E) JJ012 cells were pretreated with manumycin A for 30 min, then stimulated with S1P and Raf phosphorylation was examined (n=3). Results are expressed as the mean ± SEM. *p < 0.05 as compared with the control group; #p < 0.05 as compared with the S1P-treated group.