Tumor Necrosis Factor α-Induced Endothelial Tissue Factor Is Located on the Cell Surface Rather Than in the Subendothelial Matrix

• Published in: Blood Vol. 84; no. 5; pp. 1559 - 1566
• Main Authors: Mulder, André B., Hegge-Paping, Karin S.M., Magielse, Christel P.E., Blom, Nel R., Smit, Jan W., van der Meer, Jan, Halie, M.Ruud, Bom, Victor J.J.
• Format: Journal Article
• Language: English
• Published: Washington, DC Elsevier Inc 01.09.1994; The Americain Society of Hematology
• Subjects: Antibodies, Monoclonal; Biological and medical sciences; Blood coagulation. Blood cells; Cell Membrane - drug effects; Cell Membrane - metabolism; Cell Membrane - ultrastructure; Cells, Cultured; Coagulation factors; Endothelium, Vascular - drug effects; Endothelium, Vascular - metabolism; Endothelium, Vascular - ultrastructure; Extracellular Matrix - metabolism; Extracellular Matrix - ultrastructure; Factor Xa - analysis; Factor Xa - metabolism; Flow Cytometry; Fundamental and applied biological sciences. Psychology; Humans; Immunoassay; Immunoglobulin G; Microscopy, Electron; Molecular and cellular biology; Sensitivity and Specificity; Thromboplastin - biosynthesis; Thromboplastin - metabolism; Tumor Necrosis Factor-alpha - pharmacology; Umbilical Veins; Human; Cell culture; Endothelial cell; Coagulation; Tissue factor; Cell surface; Endothelium
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V84.5.1559.1559

Because there is no consensus regarding the precise distribution of induced endothelial tissue factor (TF), we studied TF activity in and on tumor necrosis factor α-stimulated cultured human umbilical vein endothelial cells (ECs) and their underlying matrix. TF was mainly expressed on the cell surface. Only small traces were found on the apical surface suggesting that TF is predominantly located on the basolat-eral side of the cell membrane. The presence of TF on the cell surface was confirmed by flow cytometry. Subendothelial TF activity appeared to be dependent upon the procedure used to remove the stimulated EC monolayer. Whereas ammonium hydroxide or hypotonic lysis resulted in relatively high levels of matrix-associated TF, virtually no TF was found on the matrix after mild enzymatic detachment of stimulated ECs. Cell removal with EDTA resulted in intermediate levels of matrix-associated TF. Neither the enzymatic treatment nor EDTA degraded or removed this TF activity. Similar patterns were observed for matrix-associated TF antigen and EC surface markers. Electron microscopic analysis showed cell fragments on the matrix after monolayer lysis. The findings strongly suggest that induced endothelial TF associated with the subendothelial matrix actually represents TF on EC remnants.