Fig. 3

Activation of NF-κB pathway by dTCTP in BEAS-2B cells. a NF-κB-driven luciferase activity was measured in dTCTP-treated BEAS-2B cells that were transiently transfected with pNF-κB-Luc. To compensate for transfection efficiency, cells were cotransfected with pRL-TK vector, and the results are presented as a ratio of firefly luficerase to Renilla luciferase. Values represent mean ± SEM, n = 3. ^†p < 0.05; control vs 1 μg/ml, *p < 0.05, **p < 0.01, ***p < 0.001; control vs 10 μg/ml. b Nuclear extracts of BEAS-2B cells were subjected to supershift assays with either no antibody or the indicated antibodies. They were incubated with nuclear extracts for 20 min prior to binding reactions. Arrows, DNA-binding specific complexes. Results are representative of three experiments. c BEAS-2B cells were treated with 10 μg/ml of dTCTP for the times indicated. Cells were harvested and nuclear and cytosol fractions were isolated. Each compartment was analyzed by immunoblotting. The experiments were repeated three times. d NF-κB nuclear translocation was assessed by immunofluorescence staining for NF-κB p65 (green) in BEAS-2B cells exposed to 10 μg/ml of dTCTP for indicated times. Nucleus was detected by DAPI (blue). Similar results are obtained from three independent experiments and the scale bars were 50 μm