Expression of TRAIL receptors in HMCLs treated with PD184352 and ATO. (A) HMCLs were seeded at 2.5 × 105 cells/mL in the presence of DMSO (vehicle) or PD (1 μM) for 3 hours and then were incubated for 24 hours with ATO (2 μM), after which cells were lysed and subjected to Western blot analysis to monitor the expression of TRAIL receptors using goat polyclonal anti-DR4 (Santa Cruz Biotechnology), goat polyclonal anti-DR5 (Santa Cruz Biotechnology), rabbit polyclonal anti-DcR1 (Cell Signaling Technology), and rabbit polyclonal anti-DcR2 (Imgenex). β-Actin levels are shown for confirmation of equal protein loading. (B) TRAIL receptors and β-actin bands were subjected to densitometric scanning using the TINA 2 software, and the ratio DR4/β-actin, DR5/β-actin, DcR1/β-actin, or DcR2/β-actin was calculated. (C) DR4, DR5, DcR1, DcR2, and β-actin bands were subjected to densitometric scanning using the TINA 2 software, and the (DR4 + DR5)/(DcR1 + DcR2) ratio was calculated. (D) The same blots were subsequently stripped and reprobed for procaspase-8 and Bid. CTR indicates control; PD, PD184352 (1 μM); ATO, arsenic trioxide (2 μM).
