Both the p42/p44 MAP kinase and the PI3 kinase pathways are implicated in VEGF production by K562 cells. (A) K562 cells (100 000 cells/mL) were maintained in 10% serum containing medium for 24 hours in the absence (control) or in the presence of 2 μM imatinib mesylate (STI571), 5 μM UO126, and/or 5 μM LY204002. VEGF-A production in conditioned medium following 24 hours of treatment was tested by ELISA. These results are representative of 2 independent experiments performed in triplicate ± SE. (B) K562 cells were incubated for different times with 1 μM imatinib. Cells were lysed and proteins were separated by electrophoresis on 10% polyacrylamide gels. Proteins were then blotted onto a PVDF membrane and tested for the presence of either antiphospho p42/p44 MAP kinases, antiphospho AKT, or anti-Hsp-60. A positive control for AKT activation by a PMA treatment is also shown. Arrows indicate the position of phosphorylated proteins and the loading control (Hsp-60).