We investigated the effect of blood mononuclear cell-conditioned medium on prostacyclin (PGI2) production by human umbilical vein endothelial cells in culture (HUVEC), and compared the potency of the conditioned medium in PGI2 production with that of various cytokines and lipopolysaccharide (LPS). HUVEC which had been preincubated with LPS, interleukin-1 alpha (IL-1 alpha), IL-1 beta, tumor necrosis factor (TNF alpha), or interferon-gamma (IFN-gamma) produced more PGI2 than control cells in response to thrombin. However, the HUVEC preincubated with the conditioned medium made with mononuclear cells with or without LPS (LPS-Mo-CM, Mo-CM) produced more PGI2 than those preincubated with LPS, IL-1 alpha, IL-1 beta, TNF alpha, or IFN-gamma. Although the concentrations or IL-1 beta and TNF alpha in the post-culture medium of HUVEC treated with LPS-Mo-CM were much higher than those with Mo-CM, LPS-Mo-CM which was made with 13,000/ml of mononuclear cells and 1 microgram/ml of LPS did not significantly augment the subsequent PGI2 production by HUVEC as compared with Mo-CM made with the same numbers of mononuclear cells. PGI2 production by Mo-CM-treated HUVEC still exceeded that of control cells, even when an excess amount of antibody to TNF alpha and/or IL-1 alpha was added to the Mo-CM. It is possible that Mo-CM contains unknown cytokines besides IL-1 and TNF which stimulate the HUVEC to produce PGI2.