Vitamin C inhibits GM-CSF–induced MAP kinase phosphorylation.

(A) Accumulation of vitamin C by U937 cells. Cells were incubated for the time indicated with 500 μM DHA (closed circle) or 500 μM AA (open circle). The insert shows the accumulation of vitamin C incubation with increasing concentrations of DHA for 30 minutes. The uptake of DHA and AA was determined as described in “Materials and methods.” (B) U937 cells treated with either buffer (–), 500 μM AA, or 500 μM DHA for 30 minutes were incubated with 1 nM GM-CSF for 1 minute. Phosphorylated (p-MAPK) and nonphosphorylated MAP kinase (MAPK) were detected by immunoblotting as shown in the upper and lower panels, respectively. (C) DHA did not affect ligand or GM-CSF receptor functions. DHA and GM-CSF were incubated together for 5 minutes before addition (GM-CSF/DHA) or added simultaneously (GM-CSF+DHA). Concentrations of DHA are indicated by 10 μM and 100 μM. The phosphorylated and nonphosphorylated MAP kinase are shown in the upper and lower panels, respectively. A schematic representation of the experiments is shown below the figure.