The suppressive effect of the glucocorticoid dexamethasone (DEX) on purified CD4+ T cells was found to depend on the activation pathway. In contrast to anti-CD3- or PHA-induced T cell proliferation, the alternative pathway of T cell activation, i.e., through anti-CD2 and anti-CD28, appeared largely resistant to DEX. By titrating anti-CD28 or the protein kinase C (PKC) activator PMA in the DEX-sensitive systems, it was demonstrated that inhibition by DEX could be abrogated by enhancing the CD28 signal or by stimulation of the PKC-dependent pathway. Supraoptimal concentrations of PMA were inhibitory for proliferation and this effect was partly prevented by DEX. These data suggest that the outcome of the effect of DEX on CD4+ T cells is dependent on the activation pathway, in particular the role and composition of the transcription factor AP-1.