Figure 7.

5-FU-induced necrosis in C3KO cells is ROS-dependent. HCT116 WT and C3KO cells were treated with DMSO or 5-FU (50 μg/ml). (a, b) Mitochondrial outer membrane potential measured by MitoTracker Red CMXRos at 48 h. (c) Production of mitochondrial reactive oxygen species (Mt. ROS) measured by mitoSox at 24 h with or without glutathione (GSH, 10 μM). (d) Fractions of PI+/Annexin- cells treated as in (c). (e) Levels of HMGB1 in the medium from cells treated as in (c). (f) The effects of GSH on the caspase-8/RIP1/FADD complex at 24 h in the presence of z-VAD (20 μM). (g) 5-FU induced production of mitochondrial reactive oxygen species (Mt. ROS) at 24 h in control (GPF) or RIP1 knockdown (shRIP1) WT and C3KO cells. (b–d and g) Data are the mean+S.E.M. of triplicate wells. *P <0.05, **P<0.01