Print Email Facebook Twitter Tissue factor pathway inhibitor in endothelial cells colocalizes with glycolipid microdomains/caveolae: Regulatory mechanism(s) of the anticoagulant properties of the endothelium Title Tissue factor pathway inhibitor in endothelial cells colocalizes with glycolipid microdomains/caveolae: Regulatory mechanism(s) of the anticoagulant properties of the endothelium Author Lupu, C. Goodwin, C.A. Westmuckett, A.D. Emeis, J.J. Scully, M.F. Kakkar, V.V. Lupu, F. Gaubius Instituut TNO Publication year 1997 Abstract Tissue factor pathway inhibitor (TFPI), the main downregulator of the procoagulant activity of tissue factor·factor VIIa complex, locates in human endothelial cells (EC) in culture as well-defined clusters uniformly distributed both on the cell surface and intracellularly. We here demonstrate by immunofluorescence that TFPI colocalizes in EC with caveolin, urokinase- type plasminogen activator receptor, and glycosphingolipids. The localization of TFPI in caveolae in resting endothelium is proved by double immunogold electron microscopy for TFPI and caveolin. After ultracentrifugation of rat lung or EC homogenates through density gradients of Nycodenz, TFPI was highly enriched at densities of 1.05 to 1.08 g/mL, together with caveolin and alkaline phosphatase. By ELISA, more than half of the cellular TFPI was detected in Triton X-100-insoluble extracts of EC. TFPI incorporates [1- 3H]ethanolamine and is cleaved from the cell surface by phosphatidylinositol-phospholipase C, indicating a specific glycosylphosphatidylinositol-anchorage mechanism for TFPI in the plasma membrane. Clustering of TFPI and its localization in caveolae are dependent on the presence of cholesterol in the membrane. Agonist-induced stimulation of EC caused marked changes of distribution for both TFPI and caveolin at subcellular level, with subsequent increase of the cell surface-associated inhibitory activity toward tissue factor·factor VIIa. Our findings suggest that, beside their function in transcytosis, potocytosis, cell surface proteolysis, and regulation of signal transduction, caveolae also play a direct role in the regulation of EC anticoagulant properties. Subject AnimalsCalcimycinCalciumCaveolin 1CaveolinsCell CompartmentationCell FractionationCell Line, TransformedCell MembraneCells, CulturedCentrifugation, Density GradientChloratesCholesterolEndocytosisEndothelium, VascularFactor VIIaFluorescent Antibody Technique, IndirectGlycosylphosphatidylinositolsHexosaminesHumansImmunohistochemistryIonophoresLipoproteinsMembrane LipidsMembrane ProteinsMicroscopy, ImmunoelectronPhosphatidylinositol Diacylglycerol-LyasePhospholipase CPolysaccharide-LyasesRatsSolubilityThrombinUmbilical VeinsUrinary Plasminogen Activator To reference this document use: http://resolver.tudelft.nl/uuid:d17083b6-d855-4d9d-a4b7-5a24b2427d74 TNO identifier 234184 ISSN 1079-5642 Source Arteriosclerosis, Thrombosis, and Vascular Biology, 17 (11), 2964-2974 Document type article Files To receive the publication files, please send an e-mail request to TNO Library.