(A) p65 recruitment to IL-8 promoter was analyzed by ChIP in PC3 cells pre-incubated 12 hours with control DMSO or 5 μM Bay-117082, and then treated 24 hours with 0, 0.1 or 1 μM BZ. The results were quantified by real time PCR and presented as the change in occupancy over the human IGX1A (SA Biosciences) sequence control. The data represent the mean +/−SE of four experiments; asterisks denote a statistically significant (p<0.05) change compared to control cells pre-treated with DMSO. (B) Western analysis of CE and NE prepared from PC3 cells treated with increasing concentrations of BZ for 24 hours, and analyzed by using p65, S536P-p65 and actin antibodies. Each lane corresponds to approximately 5x104 cells. (C) Densitometric evaluation of p65 and S536P-p65 levels in nuclear extracts of BZ-treated PC3 cells. The nuclear p65 and S536P-p65 bands were scanned and their densities were normalized to actin. The values for NE of untreated cells were arbitrarily set to 1, and the other values are presented relative to these values. The data represent the means of three experiments +/−SE.