Fig. 5.

MnTBAP blocks DR5 expression, caspase-8 activation, and DNA damage, whereas dominant-negative caspase-8 prevents cell death induced by the resveratrol/HDACI regimen. A and B, U937 cells were treated with 50 μM resveratrol (RESV) with or without 15 nM LBH-589 (LBH) or 1.5 μM vorinostat (Vor) in the absence or presence of 400 μM MnTBAP for 24 h, after which cells were lysed and subjected to Western blot analysis to monitor the expression of DR5, γH2A.X, and caspase-8. Each lane was loaded with 30 μg of protein; blots were stripped and reprobed for β-actin to ensure equivalent loading and transfer. CF, cleaved fragment; Casp, caspase. C, U937 cells stably transfected with dominant-negative (DN) caspaspe-8 or the empty-vector control (pcDNA3.1) were treated with 50 μM resveratrol with or without 15 nM LBH-589 or 1.5 μM vorinostat for 24 h, after which Western blot analysis was performed to monitor the cleavage/activation of caspase-8 and caspase-3. Blots were stripped and reprobed with antibodies to β-actin, to ensure equivalent loading and transfer. D, flow cytometry was performed to assess cell death after 7-aminoactinomycin D staining. C, control; R, resveratrol; L, LBH-589; V, vorinostat. Values represent the mean ± S.D. of triplicate determinations performed on three separate occasions.