TNF-induced apoptosis occurs at a higher dose of TNF than that required to activate NF-κB. (a) Comparison by EMSA of NF-κB activation after TNF injection or PH. Nuclear protein was prepared from livers of mifepristone-treated single-transgenic mice killed at the indicated times after injection of 25 μg/kg TNF (left lanes) or PH (right lanes). (b) NF-κB EMSA. Nuclear extracts were prepared from livers of C57BL6 mice killed 30 minutes after injection with the indicated dose of TNF. (c) IκBα Western blot. Liver protein lysates were prepared from mice described in b and immunoblotted for IκBα. (d) Caspase-3 activity. Mice were killed 4 hours after treatment with the indicated dose of TNF in combination with either mifepristone (mif) or D-galactosamine (GalN). For mifepristone, double-transgenic mice were used. For D-galactosamine, C57BL6 mice were used. One hundred micrograms of liver protein lysates were incubated with fluorogenic DEVD-AMC as described in Methods. *P < 0.05 compared with vehicle controls. (e) AST/ALT activity. Serum was collected from mice killed 7 hours after treatment with TNF in combination with mifepristone or D-galactosamine, as described in d.