Lipopolysaccharide (LPS), a part of the Gram-negative bacteria cell wall, is a potent inducer of tumor necrosis factor (TNF). TNF is an important mediator in Gram-negative infections such as meningococcal septic shock, but its harmful action can be prevented by the natural occurring soluble (s) TNF receptors (sTNFR) sp55 and sp75. In this study, the effect of LPS on release of sTNFR was investigated. First, we found a selective increase in human whole-blood sp75 TNFR levels following LPS stimulation, accompanied by no increase in sp55. Separating the different blood cell populations, mononuclear cells (PBMC) selectively released sp75 upon LPS stimulation, while LPS induced a minor increase in sp75 release from polymorphonuclear granulocytes. Interestingly, in co-cultures of PBMC and granulocytes, the release of LPS-induced sp75 TNFR was enhanced. Second, adherent monocytes were also found to selectively release sp75 TNFR upon LPS stimulation, where Neisseria meningitidis LPS was found to be 100-1000 times more potent in inducing sp75 release than Escherichia coli LPS. Using flow cytometry, the monocyte membrane distribution of both TNFR were found to be increased after LPS stimulation. Third, human umbilical vein endothelial cells selectively released sp55 TNFR after stimulation with LPS. We conclude that mononuclear and endothelial cells might be the main sources of soluble p75 and p55 TNFR, respectively, observed in Gram-negative sepsis, although these receptors are released in vivo more rapidly than they are in vitro.