Caspase inhibition reveals functional cooperation between p55-and p75-TNF receptors in cell necrosis

TNF-induced caspase activation is critically involved in both apoptosis and protection from cell necrosis, We have investigated the roles of the p55- and p75-TNF receptors (TNFR1 and TNFR2) in the induction of mouse L-M cell death in the presence of a caspase inhibitor (zVAD-fmk) and a transcription inhibitor (actinomycin D), i.e. under conditions in which protective pathw ays requiring caspase activation and protein synthesis were blocked, Cytome tric analysis after TNF treatment showed that apoptosis was inhibited, whil e necrosis was highly activated. In contrast, apoptosis was observed in cel ls treated with TNF and actinomycin D alone. Stimulation of TNFR1 was suffi cient to induce either cell necrosis or apoptosis, even when me blocked end ogenous TNF with an anti-murine TNF antibody, Experiments based on the use of receptor-agonist and antagonist antibodies also showed that TNFR2 contri butes to cell necrosis and apoptosis, Simultaneous stimulation of TNFR2 and TNFR1 with specific agonists indicated that TNFR2 functionally cooperates with TNFR1 to potentiate the response indirectly, by inducing endogenous TN F cytotoxicity. Caspase inhibitors enhanced the cytotoxic effect of endogen ous TNF, suggesting that TNFR2 modulation can regulate the global necrotic response to TNF, TNFR2, modulation could play an important role in determin ing the response to TNF in pathophysiological conditions characterized by c aspase down-regulation and local TNF production.