Rebamipide inhibits tumor necrosis factor-α-induced interleukin-8 expression by suppressing the NF-κB signal pathway in human umbilical vein endothelial cells

This study was designed to identify the inhibitory effect of rebamipide on tumor necrosis factor-alpha (TNF-alpha)-induced interleukin-8 (IL-8) production and nuclear factor-kappa B (NF-kappa B) activation in human umbilical vein endothelial cells (HUVECs). After stimulation with TNF-alpha, HUVECs were treated with rebamipide in a dose-dependent manner. The viability of HUVECs was assessed using methylthiazol tetrazolium assay after 24 h incubation with rebamipide. TNF-alpha-induced IL-8 expression was determined by enzyme-linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (RT-PCR). TNF-alpha-induced I kappa B-alpha phosphorylation and translocation of NF-kappa B p65 subunit into nucleus in endothelial cells were assessed using immunoblot analysis. We found that rebamipide decreased the expression of IL-8 in the dose-dependent manner under the treatment with TNF-alpha 10 ng/ml. TNF-alpha-induced degradation of I kappa B-alpha at 15 min was maximally observed and rebamipide (2 mM) inhibited TNF-alpha-induced phosphorylation of I kappa B-alpha in the cytoplasm of endothelial cells by western blot analysis. Rebamipide also suppressed TNF-alpha-stimulated NF-kappa B p65 nuclear translocation. Rebamipide suppresses TNF-alpha-induced IL-8 production through (1) inhibition of I kappa B-alpha phosphorylation in the cytoplasm and (2) blockage of NF-kappa B p65 protein transport into the nucleus. We suggest that the anti-inflammatory effect of rebamipide is related to the down-regulation of IL-8 expression that is important in endothelial inflammation.