Previous studies have demonstrated that 1,25-dihydroxyvitamin D3 (calcitriol) is a potent inhibitor of human T lymphocyte proliferation. It has been reported that only CD4+ cells are sensitive to the anti-proliferative action of calcitriol. To further evaluate this observation, we first performed cell cycle analysis of unfractionated phytohemagglutinin-stimulated peripheral blood mononuclear cells (PBMC) in the absence or presence of calcitriol. The CD4/CD8 ratio was similar between control and treated cells for each phase of cycle (G0----G1, S, G2 + M), suggesting that calcitriol did not selectively block proliferation of either T cell subpopulation. Secondly, the growth-inhibitory activity of calcitriol on PBMC selectively depleted of either CD4+ or CD8+ cells was comparable to that observed with unfractionated PBMC. Furthermore, the induction of transferrin receptors was inhibited by calcitriol to a comparable degree in each T cell subset, suggesting that equivalent inhibition of transition into late G1 was observed. Finally, calcitriol inhibited the proliferation of highly purified T cell subsets (greater than 99% pure) to equivalent degrees. These data suggest that T cell subsets defined by either the CD4 or by the CD8 antigen are both sensitive to the growth inhibitory effects of calcitriol.