In Fas-deficient Ipr/Ipr mice, endotoxin induces a severe septic shock syndrome associated with a high mortality rate due to overproduction of tumor necrosis factor-a (TNF-α) by macrophages. In order to determine if excessive TNF-α production might be due to defective activation-induced cell death (AICD), we investigated the roles of Fas and tumor necrosis factor receptor I (TNFRI) in AICD of macrophages. AICD of macrophages was analyzed in C57BL/6-+/+ mice, CSlBL/6-lpr/lpr mice, C57BL/6-TNFRI knockout mice (Tnfrf) and in C57BL/6-Tnfrr°-Ipr/lpr double mutant mice. Stimulation of purified peritoneal macrophages from C57BL/6-+/+ mice with lipopolysaccharide (LPS) (250 ng/ml) led to AICD, which was partially inhibited by addition of either Fas fusion protein or anti-TNF-α, and completely inhibited by addition of both. Surprisingly, there was increased AICD after LPS-stimulation of macrophages obtained from CSTBL/6-lprllpr mice compared to C57BL/6-+/+ mice. In CSlBL/6-lpr/lpr mice, high LPS-induced AICD was not affected by the addition of Fas fusion protein, but was completely blocked by the addition of anti-TNF-α antibody. There was also high AICD after LPS-stimulation of macrophages obtained from C57BL/6-rfr/' mice, which was completely blocked by the addition of Fas fusion protein, but unaffected by the addition of anti-TNF-α antibody. LPS-stimulation of macrophages obtained from double mutant C51BU6-Tnfrf0/0-lpr/lpr mice did not result in AICD. These results indicate that TNFRI and Fas-mediated signals are the major pathways involved in AICD of macrophages, that either pathway can compensate for the loss of the other, and that Fas-mediated apoptosis in LPS-activated macrophages plays a crucial role in down-modulation of TNF-ot production.