MAPK p38 regulates the stability of IFN-γ mRNA. (A) Freshly isolated human PBLs were stimulated with IL-12 and IL-18 (20 ng/mL each) for 4 hours, and then actinomycin D was added in the presence of 1 μM SB203580 or vehicle (0.1% DMSO). Total RNA was purified at the intervals shown and Northern blotted for IFN-γ mRNA. 28S rRNA was quantified as a loading control. (B) The experiment described in panel A was performed 4 times and IFN-γ mRNA was normalized against 28S rRNA and plotted against time on a semilogarithmic graph. Error bars indicate SEM. (C) NK cells were purified by negative selection and stimulated with IL-12 and IL-18 for 4 hours, then actinomycin D was added in the presence of 1 μM SB203580 or vehicle (0.1% DMSO). Total RNA was purified at the intervals shown and IFN-γ mRNA was quantified by ribonuclease protection assay. This experiment was performed twice with identical results.