Macrophage cell death induced by heme is dependent on autocrine TNF production and TNFR1 signaling. (A) Peritoneal macrophages (2 × 10⁵/well) from C57Bl/6 mice were treated for 6 hours with heme (30μM) in the absence or in the presence of 10% FCS. Bone marrow-derived macrophages (2 × 10⁵/well) from WT and Tnfr1^−/− (B), WT and Tlr4^−/− (C), WT, Trif^−/− and Myd88^−/− (D) were left untreated (−) or stimulated with heme at 30μM and/or TNF at 0.5 ng/mL (+) for 6 hours and supernatants were collected for LDH determination. Data are representative of 3 independent experiments performed in triplicates and represent mean ± SEM (*P ≤ .05; **P ≤ .001; ***P ≤ .0001).