Epidermal growth factor (EGF) does not alter NF-κB activation by tumor necrosis factor alpha (TNF-α). (a) Chondrocytes transfected with a κB reporter were treated with TNF-α (30 ng/ml), EGF (10 ng/ml) or TNF-α + EGF. Reporter activity was assessed after 24 hours. Data were corrected for a constitutively expressed reporter (pSV40-RL), presented as mean relative luciferase units (RLU) ± standard error of the mean, and analyzed using one-way analysis of variance followed by a Tukey–Kramer post-test. a Significant difference from basal κB activity at P < 0.001 and no significant difference between TNF-α and TNF-α + EGF. (b) The effect of EGF on sustained NF-κB activity induced by TNF-α was determined by treating confluent chondrocytes with TNF-α (30 ng/ml) alone for 24 hours, EGF (10 ng/ml) alone for 20 hours, or TNF-α for 4 hours followed by the addition of EGF (10 ng/ml) for co-treatment for a further 20 hours. Nuclear extracts were prepared and analyzed by electrophoretic mobility shift assay for activation of NF-κB. The reporter assay and band shift shown are representative of three independent experiments.