Regulation of NF-κB activation and nuclear translocation by exogenous nitric oxide (NO) donors in TNF-α activated vascular endothelial cells

Abstract

Nitric oxide (NO) is a unique mediator which may promote or suppress inflammation. In this study, we examine the effect of exogenous NO on nuclear translocation of nuclear factor-kappa B (NF-κB) in quiescent human umbilical vein endothelial cells (HUVECs) subsequently activated by tumor necrosis factor-α (TNF-α), and in HUVECs previously activated by TNF-α, a model of vascular inflammation. Quiescent and activated HUVECs are exposed to exogenous NO donors of varying half-lives and the degree of NF-κB translocation into the nucleus determined by unique application of immunofluorescence image analysis in whole cells and correlative biochemical analysis of activated NF-κB proteins in the nucleus. NO donors with shorter half-lives are more effective in blocking the activation and translocation of NF-κB, when added to quiescent HUVECs prior to cellular activation by TNF-α. However, in previously activated HUVECs where NF-κB had relocated into the cytoplasm, addition of short half-life NO donors, but not TNF-α, induced re-translocation of NF-κB back into the nucleus sustaining the inflamed cell phenotype. These data suggest that NO as an inhibitor or activator of NF-κB may depend on the state of activation of vascular endothelial cells in which it contacts. Additionally, in activated cells, NO may modulate expression of NF-κB-dependent gene products, when cytokines are ineffective.