A novel mechanism for the inhibition of NF-kappaB activation in vascular endothelial cells by natural antioxidants.

The activation of Nuclear Factor kappa B (NF-kappaB) in vascular endothelial cells, in response to biochemical or biomechanical stimuli, is associated with vascular pathologies such as atherosclerosis. The present manuscript studies the ability of the natural antioxidant-pomegranate wine (PW), to inhibit tumor necrosis factor alpha (TNF-alpha) or shear stress-mediated-NF-kappaB activation in vascular endothelial cells and compares it to that of red wine (RW) and N-acetyl cysteine (NAC). PW and RW act as potent antioxidants in vascular endothelial cells, inhibiting the oxidation of 2',7'-dichloroflurescin diacetate in TNF-alpha treated cells. PW (as well as RW and NAC) acted as potent inhibitors of NF-kappaB activation (migration into the nucleus and DNA binding activity) in vascular endothelial cells. Nevertheless, PW and NAC failed to inhibit TNF-a induced serine 32/36 phosphorylation and IkappaBalpha degradation. Surprisingly, these antioxidants alone induced enhanced IkappaB serine phosphorylation, which was not accompanied by IkappaBalpha degradation, or NF-kappaB nuclear translocation. This phosphorylation did not involve serine 32/36. Furthermore, we show for the first time that NAC inhibited TNF-alpha mediated phosphorylation of p65 (ser536), whereas PW had no effect on this phosphorylation. Thus, natural antioxidants may serve as potent NF-kappaB inhibitors in vascular endothelial cells, yet act through unique and divergent pathways.