IL-16, in a CD4-dependent manner, induces high affinity IL-2R (CD25) selectively on CD4+ T cells. Based on this observation, we determined the relative effects of IL-16 on IL-2Rα, β, and γ expression on CD4+ T cells and of IL-16/IL-2 cotreatment of resting human PBMC obtained from normal individuals on CD4+ T cell proliferation and cytokine production, in vitro. IL-16 increased CD4+ T cell IL-2Rα and β expression, but had no effect on expression of IL-2Rγ. There was marked synergy of thymidine uptake and expansion of CD4+ T cell numbers in the presence of IL-16 and IL-2 or IL-16 and IL-15 compared with the responses to any of the cytokines alone. By 4 wk, IL-16/IL-2-cotreated PBMC cultures were predominantly CD4+, CD25+ CD45RO T cells. Of the cytokines measured, IL-16 treatment alone was sufficient to induce synthesis of granulocyte-macrophage CSF by 2 wk. IL-16/IL-2 cotreatment did not appear to induce selective proliferation of any Th subset, as cytokines of both Th1 (eg, IFN-γ) and Th2 (eg, IL-5) types were synthesized by the expanded cell populations at 2 and 4 wk. These results suggest that IL-16 can prime CD4+ T cells for IL-2 responsiveness, and therefore may be a useful adjunct to IL-2 therapy for immune reconstitution in disease or therapeutic conditions resulting in CD4+ T cell depletion.