TNF is a highly pleiotropic cytokine. The recent identification of two distinct cellular receptors for TNF may provide explanations for the many different TNF activities. We have investigated the expression of the two receptor types, TNFR alpha (75 kDa) and TNFR beta (55 kDa), in human PBMC. Both receptors were found simultaneously expressed by cytofluorimetric, radioligand binding and Northern analysis of naive as well as PHA-activated PBMC. The expression levels in the CD14+ and CD14- subsets were different. Both receptors were strongly expressed in the CD14+ subset. The expression of the receptors in the CD14-, CD3+, CD4+, and CD8+ subsets was lower and similar among these subsets, but TNFR alpha was expressed at higher level than TNFR beta. To dissect the functional roles of the two receptors, we studied the growth factor activity of TNF in the late proliferative responses of PBMC to PHA. In the first approach, the activity of either receptor was blocked by neutralizing, receptor type specific antibodies. In a second approach, the ligand, TNF, was inhibited by a neutralizing antiserum, and the cells were restimulated using type-specific anti-TNFR antibodies with agonistic activity. It was found that both receptor types mediated signals required for proliferative responses of PBMC to PHA from day 4 to day 8 in culture. The cell responses to the activation of either receptor type appeared to be independent, because one receptor could not compensate for the reduction in cell activation caused by blocking the other receptor type.