title: Two independent binding sites on monolayers of human endothelial cells are responsible for interaction with coagulation factor VII and factor VIIa.
abstract: The interaction of radiolabeled factor VII (FVII) and factor VIIa (FVIIa) with endotoxin-stimulated endothelial cells (EC), known to express tissue factor (TF), and unstimulated EC was studied. FVII/FVIIa binding to EC-monolayers was saturable within 4.5-6 h, reversible, temperature and calcium dependent on both, endotoxin-stimulated and on unstimulated EC. Upon 2 h of incubation on EC, FVII was partially converted to FVIIa in the absence of protease inhibitors. The affinity of this binding was Kd = 45.4 +/- 18.7 nM with a calculated number of binding sites Bmax = 3.75 +/- 0.31 x 10(6) molecules/cell. In addition to unlabeled FVII and FVIIa, other vitamin K-dependent proteins reduced binding of [125I]-FVII/FVIIa to about 60-70%, and this type of common binding site for vitamin K-dependent proteins revealed a Kd = 32.2 +/- 5.6 nM and a Bmax = 3.03 +/- 0.14 x 10(6) molecules/cell. Moreover, in the presence of 1 microM prothrombin to suppress common binding sites, only on endotoxin-stimulated EC additional inhibition of FVII/FVIIa binding was achieved by anti-TF antibodies. The characteristics of the FVII/FVIIa-TF interaction with a Kd = 17.2 +/- 5.2 nM and a Bmax = 342,000 +/- 1,100 binding sites/cell revealed a similar saturation kinetics in radioligand binding and in functional factor X activation within 90-120 min. These data indicate the presence of at least two independent binding sites for FVII/FVIIa on stimulated EC of which about 10% are TF specific.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Haemostasis Research Unit, Kerckhoff-Klinik, Max-Planck-Institut, Bad Nauheim, Germany.
authors.forename: U
authors.initials: U
authors.lastname: Reuning
authors.forename: K T
authors.initials: KT
authors.lastname: Preissner
authors.forename: G
authors.initials: G
authors.lastname: Müller-Berghaus
mesh_terms: D001665:Binding Sites; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D004731:Endotoxins / Q000494:pharmacology; D005167:Factor VII / Q000378:metabolism*; D015942:Factor VIIa / Q000378:metabolism*; D015951:Factor Xa / Q000096:biosynthesis; D006801:Humans; D007700:Kinetics; D008050:Lipid A / Q000494:pharmacology; D013925:Thromboplastin / Q000378:metabolism
chemical_list: D004731:Endotoxins; D008050:Lipid A; D005167:Factor VII; D013925:Thromboplastin; D015942:Factor VIIa; D015951:Factor Xa