Apoptosis induction, caspase-8 cleavage, and the decrease in c-FLIP levels due to DX1 treatment in NB4, HP100-1 and K562 cells.

<p>(<b>A</b>) <b>Apoptotic cells.</b> HL-60, HP100-1, NB4 and K562 cells were treated with 10 µM DX1 for 9 h. The percentage of apoptotic cells was determined by morphological observation after AO/EB staining as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0015843#s2" target="_blank">Materials and Methods</a>. (<b>B</b>) <b>The protein levels and cleavage of PARP, caspase-8 and c-FLIP.</b> NB4, HP100-1 and K562 cells were treated with or without 10 µM DX1 for 10 h. The levels of each protein were detected using specific antibodies as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0015843#s2" target="_blank">Materials and Methods</a>. (<b>C</b>) <b>MG-132 enhancement of DX1-induced apoptosis.</b> Cells were treated with DMSO (control), MG-132 0.5 µM, DX1 8 µM or MG132 plus DX1 for 10 h (NB4 and HP100-1 cells) or for 36 h (K562 cells). The percentage of apoptotic cells was determined morphologically after AO/EB staining as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0015843#s2" target="_blank">Materials and Methods</a>. * <i>P</i><0.05 compared to cells treated with DX1 alone. (<b>D</b>) <b>MG-132 enhancement of DX1-induced cleavage of PARP and caspase-8 in K562 cells.</b> K562 cells were treated with DMSO (control), MG-132 0.5 µM, DX1 8 µM or MG-132 plus DX1 for 36 h. The levels of each protein were detected using specific antibodies as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0015843#s2" target="_blank">Materials and Methods</a>. (<b>E</b>) <b>The influence of c-FLIP siRNA on DX1-induced cleavage of PARP and caspase-8 in K562 cells.</b> K562 cells were incubated with control siRNA or c-FLIP siRNA for 15 h and then treated with or without 10 µM DX1 for 24 h. The levels of each protein were deteced using specific antibodies as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0015843#s2" target="_blank">Materials and Methods</a>.