Figure 1

Sulindac sulfide induces activation of the NF-κB pathway and increases cell death. (A) Quantification of apoptosis and necrosis of sulindac sulfide (SS) treated cells (4 hours), assessed by trypan blue exclusion assay or FACS analysis of Annexin-V-Fluos/PI stained cells. Bars represent mean percentage of cells in each population ± SEM (n = 3-4). (B) Photomicrographs of HCT-15 cells treated with sulindac sulfide for 8 hours (phase-contrast, scale bars 60 μm and insert 24 μm). (C) HCT-15 cells were treated with the indicated doses of sulindac sulfide or the control DMSO (equivalent to the highest dose SS) for 2 and 4 hours, after which lysates were collected and western blot analysis was performed for IκBα and the loading control β-actin. Densitometry of IκBα levels normalized to β-actin. Error bars represent SEM (n = 3-4). (D) qPCR analysis for A20, ICAM1 and IL-8 mRNA expression. HCT-15 cells were treated with the indicated concentrations of SS or the control. Gene expression was normalized to the house-keeping gene GAPDH. The data are presented as fold change ± SEM (from 3 independent experiments). (E) Western blot for IκBα levels in cells treated with SS or the control for the indicated time points. Densitometry of IκBα levels normalized to β-actin. Error bars represent SEM.