High glucose-induced NF-κB activation occurs via tyrosine phosphorylation of IκBα in human glomerular endothelial cells:: involvement of Syk tyrosine kinase

Activation of nuclear factor-kappa B (NF-kappa B) occurs by dissociation from I kappa B after serine or tyrosine phosphorylation of I kappa B alpha, but the way of NF-kappa B activation by high glucose has not been defined. High glucose is known to activate NF-kappa B via protein kinase C and reactive oxygen species (ROS). In this study, we investigated how high glucose activates NF-kappa B for CC chemokine ligand 2 production in cultured human glomerular endothelial cells. High glucose increased nuclear translocation of p65 and also increased NF-kappa B DNA binding activity. High glucose-induced NF-kappa B activation occurred without degradation of I kappa B alpha. In agreement with this, there was no increase in serine phosphorylation of I kappa B alpha, while tyrosine phosphorylation of I kappa B alpha was increased by high glucose. High glucose increased the generation of ROS, whereas both alpha-lipoic acid and N-acetylcysteine scavenged the ROS and decreased high glucose-induced tyrosine phosphorylation of I kappa B alpha, nuclear translocation of p65, and NF-kappa B DNA binding activity. Protein kinase C pseudosubstrate inhibited high glucose-induced ROS production, tyrosine phosphorylation of I kappa B alpha, and nuclear translocation of p65. Both BAY 61-3606, a specific inhibitor of Syk protein-tyrosine kinase, and small interfering RNA directed against Syk inhibited high glucose-induced tyrosine phosphorylation of I kappa B alpha as well as p65 nuclear translocation. High glucose increased tyrosine phosphorylation of Syk, while it was inhibited by alpha-lipoic acid and protein kinase C pseudosubstrate. In summary, high glucose-induced NF-kappa B activation occurred not by serine phosphorylation of I kappa B alpha. Our data suggest that ROS-mediated tyrosine phosphorylation of I kappa B alpha is the mechanism for high glucose-induced NF-kappa B activation, and Syk may play a role in tyrosine phosphorylation of I kappa B alpha.