9.2.27PE and ABT-737 in Melanoma Figure 4. ABT-737 induces ER stress and cell death in melanoma cells. (A) Increased GRP78 and peIF2a protein levels in FEMX, Melmet-5 and MelRM melanoma cells treated with ABT-737 (10 mM) up to 48 h, indicating ER stress. CHOP protein was only increased with 9.2.27PE treatment. Both ABT-737 and 9.2.27PE caused inactivation of PARP, indicating apoptosis. GAPDH and amidoblack staining was used as a loading control. Tunicamycin (3 mM), was used as a positive control for ER stress. The blot is representative of at least three independent experiments, except for the 32 and 48 h time point where the experiments were repeated twice. Lane 1: Vehicle (DMSO), 2: ABT-737 1 h, 3: ABT-737 2 h, 4: ABT-737 4 h, 5: ABT-737 8 h, 6: ABT-737 16 h, 7: ABT-737 24 h, 8: ABT-737 32 h 9: ABT-737 48 h, 10: Vehicle (PBS 0.1% human serum albumin), 11: 9.2.27PE 8 h, 12: 9.2.27PE 16 h, 13: 9.2.27PE 24 h, 14: Tunicamycin 8 h. (B) Mitochondrial membrane potential was measured using the JC-1 dye. A decrease in red/green ratio is indicative of depolarization of the mitochondrial membrane. The data represents the mean 6 SD. (C) FEMX, Melmet-5 and MelRM cells were treated with ABT-737 (10 mM) for 24–48 h before stained with propidium iodide for quantitation of dead cells. An increase in the percentage sub-G1 represents increased cell death compared to control cells. The data represents the mean 6 SD. doi:10.1371/journal.pone.0024012.g004 PLoS ONE | www.plosone.org 8 September 2011 | Volume 6 | Issue 9 | e24012