BC down-regulates Bfl-1 expression by inhibiting NF-κB activity. A , A549 cells infected by BC or control (Con) adenovirus were treated with 40 ng/ml gemcitabine for 72 h. Total RNA was isolated and subjected to RT-PCR for Bfl-1 and β-actin. B , A549 cells were infected with BC or control adenovirus and treated with 40 ng/ml gemcitabine or not. 50 μg of total cellular proteins were subjected to SDS-PAGE and then immunoblotted using anti-Bfl-1, -Bax, -Bcl-xL, -Bcl-2, and -α-tubulin antibodies. Data are representative of three independent experiments. C , A549 cells, transfected with the pNF-κB-Luc reporter gene and β-gal-lacZ plasmid, were infected with BC or control adenovirus and then treated with 40 or 400 ng/ml of gemcitabine for 48 h. Luciferase and β-galactosidase activities were determined using a luminometer. The values shown represent luciferase activities relative to β-gal, and the means ± SDs from three independent experiments are plotted. *p < 0.001 by the Student's t test for difference in tumor volumes between experimental groups. D , A549 cells infected with BC or control adenovirus were treated with 0, 40, or 400 ng/ml gemcitabine for 48 h. Proteins from nuclear and cytosolic fractions were separately isolated, and 50 μg aliquots were subjected to SDS-PAGE and then immunoblotted using anti-p65/relA, -lamin B, and -α-tubulin antibodies. Data are representative of three independent experiments.