It is known that 1,25(OH)2D3 inhibits IL-2, IFN-gamma, and GM-CSF mRNA accumulation in activated T cells. While 1,25(OH)2D3 enhances macrophage competence, its effect on cytokine gene expression in monocytes is less well defined. Using Northern blot analysis, we examined the effect of 1,25(OH)2D3 pretreatment on IL-1 beta and TNF gene expression in LPS- and PHA-stimulated human PBMNC and several human myeloid cell lines (U937 and THP1). In PBMNC, preincubation had no effect on the steady-state level of LPS-induced TNF or IL-1 beta mRNA. When PHA was used to stimulate pretreated PBMNC, a 60-80% inhibition of TNF mRNA levels was observed. There was no effect on IL-1 beta mRNA. In U937 cells, 1,25(OH)2D3 preincubation resulted in a 4-fold increase in the level of TNF and IL-1 beta mRNA levels. Pretreatment had no effect on TNF or IL-1 beta gene expression in THP1 cells. The observation that PHA-induced TNF gene expression is modulated by 1,25(OH)2D3 is novel. Possible mechanisms by which 1,25(OH)2D3 preincubation may influence mitogen-specific inducible cytokine gene expression in different cell types are discussed.