E-SELECTIN EXPRESSION IN HUMAN ENDOTHELIAL-CELLS BY TNF-ALPHA-INDUCEDOXIDANT GENERATION AND NF-KAPPA-B ACTIVATION

Because reactive oxygen species (ROS) can function as second messenger s and regulate the activation of the transcription factor nuclear fact or (NF)-kappa B, we investigated the possible role of tumor necrosis f actor-alpha (TNF-alpha)-induced ROS generation in endothelial cells in signaling E-selectin gene transcription. We demonstrated that stimula tion of human pulmonary artery endothelial cells with TNF-alpha (100 U /ml) resulted in ROS production using the oxidant-sensitive dye 5 (and 6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate bis(acetoxymeth yl)ester. Pretreatment with N-acetyl-L-cysteine (NAC) or pyrrolidine d ithiocarbamate (PDTC) for 0.5 h inhibited TNF-alpha-induced generation of ROS as well as activation of NF-kappa B and E-selectin mRNA and th e cell surface protein expression. These findings indicate that TNF-al pha induces NF-kappa B activation and the resultant E-selectin gene ex pression by a pathway that involves formation of ROS and that E-select in expression can be inhibited by the antioxidant action of NAC or PDT C. The results support the hypothesis that generation of ROS in endoth elial cells induced by proinflammatory cytokines such as TNF-alpha, is a critical signal mediating E-selectin expression.