Inhibition of the endothelial cell activation by antithrombin in vitro

• Published in: Thrombosis and haemostasis Vol. 92; no. 6; p. 1420
• Main Authors: Uchiba, Mitsuhiro, Okajima, Kenji, Kaun, Christoph, Wojta, Johann, Binder, Bernd R
• Format: Journal Article
• Language: English
• Published: Germany 01.12.2004
• Subjects: Anti-Inflammatory Agents - pharmacology; Antithrombins - metabolism; Blotting, Western; Carbazoles - pharmacology; Cells, Cultured; Colforsin - pharmacology; Cyclic AMP - metabolism; Cyclic AMP-Dependent Protein Kinases - antagonists & inhibitors; DNA - metabolism; Dose-Response Relationship, Drug; E-Selectin - biosynthesis; Endothelial Cells - cytology; Endothelial Cells - metabolism; Endothelium, Vascular - cytology; Endothelium, Vascular - metabolism; Enzyme-Linked Immunosorbent Assay; Heparin - metabolism; Humans; Immunohistochemistry; Immunoprecipitation; In Vitro Techniques; Indoles - pharmacology; Interleukin-1 - metabolism; Lipopolysaccharides - metabolism; NF-kappa B - metabolism; Nuclear Proteins - metabolism; p38 Mitogen-Activated Protein Kinases - metabolism; Phosphorylation; Protein Binding; Pyrroles - pharmacology; RNA, Messenger - metabolism; Time Factors; Trans-Activators - metabolism; Tumor Necrosis Factor-alpha - metabolism; Umbilical Veins - cytology
• ISSN: 0340-6245
• DOI: 10.1160/TH04-03-0139

We examined whether antithrombin (AT) inhibits tumor necrosis factor (TNF)-alpha-induced endothelial cell activation to elucidate molecular mechanism(s) of the anti-inflammatory activity of AT. AT inhibited the increase in E-selectin expression in cultured human umbilical vein endothelial cells (HUVECs) stimulated with TNF-alpha. In contrast, chemically modified AT that lacks affinity for heparin did not. AT inhibited the TNF-alpha-induced interaction of NF-kappaB p65 with p300, a homologue of cAMP-responsive element binding protein (CREB)-binding protein (CBP). AT increased both intracellular levels of cAMP and binding of phosphorylated-CREB to DNA in HUVECs. Forskolin showed the inhibitory effect similar to that of AT and pretreatment of HUVECs with KT-5720, an inhibitor of protein kinase A, reversed the inhibitory effect of AT. These observations suggested that AT inhibited the TNF-alpha-induced increase in E-selectin expression in HUVECs by inhibiting the interaction of NF-kappaB with CBP/p300 through cAMP-dependent protein kinase A-induced CREB activation. This inhibitory activity of AT might depend on its binding to heparin-like substances on the endothelial cell. Such an inhibitory effect of AT on TNF-alpha-induced endothelial cell activation might at least partly contribute to its anti-inflammatory activity.