Inhibitors that blocked ceramide-induced DNA fragmentation did not prevent Fas-induced DNA fragmentation. (A) Jurkat cells were treated with ceramide and various inhibitors for 4 hours. The sub-G₁ DNA content was quantitated as described in Fig1. Additional inhibitors used were as follows: dbcAMP, 1 mmol/L; C₆₀ (C3/100), 100 μmol/L; C₆₀ (D3/100), 100 μmol/L; C₆₀ (D3/10), 10 μmol/L. (B) Jurkat cells were stimulated with anti-Fas antibody CH11 (125 ng/mL) in the absence or presence of the inhibitors for 14 hours, and the sub-G₁ DNA content was determined. The inhibitors used were as follows: forskolin, 50 μmol/L; dbcAMP, 2.5 mmol/L; C₆₀ (C3), 100 μmol/L; C₆₀ (D3), 100 μmol/L; PDTC, 200 μmol/L; Z-VAD-FK (ICEi), 300 μmol/L. None of the inhibitors alone induced DNA fragmentation at the concentrations used, except PDTC, in which an 1% to 2% increase over control was observed. The result is the average of duplicates, with standard deviation shown as an error bar. Those not shown are too small in scale. Experiments were repeated three times with the same results.