Figure 4

Treatment with POMx inhibited the activation and DNA binding activity of NF-κB in PMACI-stimulated KU812 cells. KU812 cells (3 × 10⁶ cells ml^-1) were pretreated with POMx (20, 40 μg/ml) for 1 h and stimulated by PMA (40 nM) plus A23187 (1 μM) for 2 h. (A) IκBα degradation and NF-κB translocation was analyzed by Western immunoblotting using antibodies specific for the p65 subunit of NF-κB (C-NF-κB, cytoplasmic NF-κB; N-NF-κB, nuclear NF-κB). (B) Band intensities were obtained as described above Data shown is cumulative of two experiments and the O.D. values (pixels/band) are mean ± SD. *P < 0.001 Vs PMACI alone (N-NF-κB), ^#P < 0.05 Vs PMACI alone (C-NF-κB), ^##P < 0.05 Vs PMACI alone (IκBα). (C) NF-κB luciferase assay: Cells were transiently transfected with the NF-κB luciferase reporter construct or empty vector and the NF-κB-dependent transcriptional activity was determined by luciferase activity using a commercially available kit (Promega). Each bar represents the mean ± SD of three independent experiments. *P < 0.001 Vs PMACI alone.