Abstract 282: Endothelial-specific Deletion of Inhibitor of Kappa-B Kinase-β (IKK-β)
The serine-threonine kinase, IKK-β, is required for classical activation of the NF-κB family of transcription factors which are central regulators of inflammation and cell survival that are activated in a variety of vascular diseases.
[Methods and Results]: To examine the role of IKK-β and NF-κB in endothelial cells (EC), we generated mice with EC-specific deletion of IKK-β (IKK-βΔEC) on an ApoE−/− background using Tie2-Cre transgenic and IKK-βflox/flox mice. IKK-βΔEC were born at less than the expected Mendelian frequency (7/48 vs 24/48 expected, p<0.0001) and were ~25% smaller than Cre-negative littermates (p<0.003). Vascular permeability assessed by in vivo microscopy of ear capillaries after intravenous injection of Evans blue dye was substantially increased (p<0.04). Several IKK-βΔEC phenotypes were similar to Akt1−/− mice, prompting us to examine Akt1 in the IKK-βΔEC mice. Interestingly, phospho-Akt was markedly reduced in vascular endothelium from IKK-βΔEC mice. In vitro studies of primary cultured EC demonstrated that deletion of IKK-β inhibited phosphorylation of Akt and increased permeability, while overexpression of either wild-type (WT) or kinase-deficient (KD) IKK-β enhanced Akt phosphorylation. Immunoprecipitation experiments demonstrated IGF-I-enhanced physical interaction of IKK-β with Akt. Moreover, IKK-β deletion inhibited Akt trafficking to lipid rafts rich in caveolae, an essential step in its activation. Expression of either WT or KD IKK-β in IKK-β-deleted EC in vitro restored normal Akt trafficking and reduced endothelial permeability. These results are in contrast to those reported with deletion of IKK-β in hepatocytes which have relatively few caveolae.
[Conclusion] These data demonstrate that IKK-β modulates Akt subcellular trafficking and activation in EC through a kinase-independent mechanism. Understanding interactions between these signaling pathways may provide important insights in conditions marked by disregulated inflammation and/or cell survival including atherosclerosis and ischemic-injury.