Figure 4

Sensitization of melanoma cells to TRAIL-induced apoptosis by 2-DG is largely due to up-regulation of TRAIL-R2. A, Mel-RM and MM200 cells were treated with a TRAIL-R2/Fc chimera (10 μg/ml) before the addition of 2-DG (10 μM) and TRAIL (200 ng/ml) for a further 24 hours. Apoptosis was measured by the propidium iodide method using flow cytometry. B, Mel-RM and M200 cells were treated the caspase-8 specific inhibitor z-IETD-fmk (30 μM) for 1 hour before the addition of 2-DG (10 μM) and TRAIL (200 ng/ml) for a further 24 hours. Apoptosis was measured by the propidium iodide method using flow cytometry. C, Mel-RM and MM200 cells were treated with a TRAIL-R1/Fc chimera (10 μg/ml) before the addition of 2-DG (10 μM) and TRAIL (200 ng/ml) for a further 24 hours. Apoptosis was measured by the propidium iodide method using flow cytometry. D, Mel-RM and MM200 cells were transfected with the control or TRAIL-R2 siRNA. Left panel: Twenty-four hours later, whole cell lysates were subjected to Western blot analysis. Right panel: Twenty-four hours later, the cells were treated with 2-DG (10 μM) and TRAIL (200 ng/ml) for a further 24 hours. Apoptosis was measured by the propidium iodide method using flow cytometry. The data shown are either the mean ± SE (A, B, C, & the right panel of D), or representative (the left panel of D), of three individual experiments.