Abstract 1,25-Dihydroxyvitamin-D 3 (1,25-D 3) is known to inhibit DNA synthesis, immunoglobulin and lymphokine production [interleukin-2 (IL-2), gamma interferon (G-IFN), and granulocyte-monocyte colony-stimulating factor (GM-CSF)] by mitogen-stimulated human peripheral blood mononuclear cells (PBMCs). Recent data suggest these inhibitory effects are mediated at the gene level through inhibition of mRNA accumulation of specific lymphokines in the activated cells. In previous studies, we have demonstrated the CD8 + T cell population was less sensitive to the anti-proliferative actions of 1,25-D 3 than CD4 + T cells. The purpose of this investigation was to further assess ability of 1,25-D 3 to regulate CD4 + and CD8 + T cell functions. Initial experiments showed that 1,25-D 3 inhibited both IL-2 production and mRNA accumulation in mitogen-stimulated PBMC. However, IL-2 receptor (IL-2R) expression and mRNA accumulation in stimulated PBMC was not affected by 1,25-D 3. Both FACS sorted CD4 + and CD8 + T cells expressed IL-2R equally upon stimulation and neither showed an inhibitory effect on this expression by 1,25-D 3. Human CD4 + and CD8 + T cells showed a stimulus-specific production of IL-2. CD4 + cells stimulated with mitogen and HLA-DR positive accessory cells produced measurable levels of IL-2 that were completely inhibited by 1,25-D 3. CD8 + T cells did not generate measurable amounts of IL-2 in this system. However, CD4 + and CD8 + T cells produced large amounts of IL-2 when stimulated with mitogen and a protein kinase C activator, phorbol mystirate acetate (PMA). Under these circumstances, both CD4 + and CD8 + T cell IL-2 production was inhibited completely by 1,25-D 3. These data suggest that IL-2R expression in PBMCs and T cell subsets is equal and unaffected by 1,25-D 3 while IL-2 production in T cell subsets is stimulus-specific and completely inhibited by 1,25-D 3.