Corticosteroids are the most common therapeutic approach for control of tissue inflammation. Combination IL-2/IL-4 is known to induce T-cell steroid resistance. This can be reversed with IFN-gamma; however, the mechanism by which this occurs is unknown. In the current study, we found that treatment of peripheral blood mononuclear cells with combination IL-2/IL-4 for 48 hours, but not with IL-2 or IL-4 alone, abrogated dexamethasone (DEX)-induced glucocorticoid receptor (GCR)-alpha nuclear translocation in both CD4(+) and CD8(+) T cells. The presence of IL-4 significantly down-regulated IFN-gamma production by IL-2-stimulated cells. Importantly, addition of IFN-gamma to the IL-2/IL-4 combination restored GCRalpha nuclear translocation in response to DEX. Furthermore, DEX-induced mitogen-activated protein kinase (MAPK) phosphatase-1 induction, used as a readout for corticosteroid-induced transactivation, was significantly greater (P < 0.05) in media and IL-2/IL-4/IFN-gamma-treated conditions compared with IL-2/IL-4-treated cells. The combination of IL-2/IL-4 induced p38 MAPK activation in CD3(+) cells (30.5 +/- 5.7% cells expressed phospho-p38 MAPK versus no phospho-p38 MAPK expression after media treatment). The presence of the p38 MAPK inhibitor, SB203580, or IFN-gamma inhibited p38 MAPK phosphorylation and enhanced GCRalpha nuclear translocation in response to DEX. These data indicate that combination IL-2/IL-4 inhibits GCRalpha nuclear translocation in human T cells, and this effect is reversed by IFN-gamma via inhibition of p38 MAPK activation.