Figure 5

PEE-evoked human T24 cell apoptosis via the mitochondrial pathway and death receptor pathway. Human T24 or J82 cells were treated with 50 or 100 μg/ml of PEE for different time durations. Various effector proteins involved in apoptosis were detected by western blotting with the antibodies against procaspase-3 (A), procaspase-9 (C), procaspase-8 (D), Bax/BcL-2 (E), DR4 and DR5 (F) respectively as described in section 2. T24 cells were investigated in all panels except panel A as indicated. The blot in each diagram was the typical result of three independent experiments except pro-caspase-3 examination in J82 cell (one experiment) and DR4 as well as DR5 detection (two independent experiments) in T24 cells. The densitometer-intensity data (mean ± S.D.) were shown under each blot. (B) For the counteract study, human T24 cells were incubated with 2 μM of caspase-3 inhibitor Z-DEVD-FMK for 4 hours and then 100 μg/ml of PEE was incorporated with the inhibitor-pretreated cells for 48 hours. After incubation, the cells were collected for annexin V/PI analyses.