Fig. 5

Effects of KC-53 on TRAF2 and IκΒα phosphorylation, and p65 translocation to the nucleus. a Cells were incubated with vehicle, KC-53 or PS-341 (for the times indicated) followed by incubation with TNFα for 20 min. Whole cell extracts were analyzed by immunoblotting. b Cells were incubated to either vehicle, KC-53 or PS-34 as indicated followed by the addition of TNFα for 20 min. The nuclear localization of p65 was examined by immunoblotting. Histone H3 levels were determined as a loading control and the α-Tubulin levels as an indicator of cytosolic contamination. Numbers signifies intensity values and are expressed as fold change compared to the control. c Cells were subjected to treatments either with the vehicle or TNFα (1 ng/ml) for 4 h, or KC-53 (5 μΜ) for 6 h plus TNFα for 4 h. The PCR products were normalized to those obtained from GAPDH mRNA amplification. The results in panels (a) and (b) are representative of three repetitions. The results in (c) panels represent the mean ± SEM of two replicates and are representative of three independent experiments. (*p value <0.05, **p value <0.01, ***p value <0.001)