Effect of NiCl2 on IKKβ and IκBα.
Exposure of HUVECs to NiCl2 leads to activation of IKKβ and degradation of IκBα. (A) Endothelial cells were stimulated for 30 minutes with different concentrations of NiCl2 as indicated. Thereafter, cell extracts were prepared and IKKβ was assayed in immune-complex kinase assays using GST-IκBα as a substrate. (B) HUVECs were exposed to 1.5 mM NiCl2 for 0, 5, 10, 15, 30, 45, or 60 minutes. IKKβ activity was determined by immune-complex kinase assays as described above. Protein loads were controlled by Western blot with the use of an antiserum to IKKβ. (C) HUVECs stimulated with 1.5 mM NiCl2 or 2 ng/mL TNFα for 0, 5, 15, 45, or 90 minutes were processed for Western blot analysis.IκBα immunoreactivity is visualized as a 40-kd protein.
