Activation of the coagulation mechanism on tumor necrosis factor-stimulated cultured endothelial cells and their extracellular matrix

The role of flow and factor IX/IXa

Pim N M Tijburg, Jane Ryan, David Stern, Barry Wollitzky, Sara Rimon, Abraham Rimon, Dean Handley, Peter Nawroth, Jan J. Sixma, Philip G. De Groot

Research output: Contribution to journalArticle

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Abstract

Infusion of tumor necrosis factor (TNF) into tumor-bearing mice led to intravascular clot formation with fibrin deposition in microvessels in the tumor bed in close association with the vessel wall, which could be prevented by active site-blocked factor IXa (IXai). This observation prompted us to examine the role of the intrinsic system in activation of the coagulation mechanism on TNF-stimulated human endothelial cell monolayers and endothelial-derived matrix during exposure to purified coagulation factors or flowing blood. Treatment of endothelial cells in intact monolayers with TNF induced expression of the procoagulant cofactor tissue factor (TF) in a dose-dependent manner, and after removal of the cells, TF was present in the matrix. TNF-treated endothelial cell monolayers exposed to blood anticoagulated with low molecular weight heparin induced activation of coagulation. Addition of IXai blocked the procoagulant response on TNF-treated endothelial cells, and consistent with this, the presence of factor IX/VIIIa enhanced endothelial TF/factor VII(a) factor X activation over a wide range of cytokine concentrations (0-600 PM). When TF-dependent factor X activation on endothelial cells was compared with preparations of subendothelium, the extracellular matrix was 10-20 times more effective. IXai blocked TF/factor VII(a) mediated activated coagulation on matrix, but only at lower concentration of TNF (<50 pM). Similarly, enhancement of factor Xa formation on matrix by factors IX/VIIIa was most evident at lower TNF concentrations. When anticoagulated whole blood flowing with a shear of 300 s-1 was exposed to matrices from TNF-treated endothelial cells, but not matrices from control cells, fibrinopeptide A (FPA) generation, fibrin deposition, and platelet aggregate formation were observed. FPA generation could be prevented by a blocking antibody to TF and by active site-blocked factor Xa (Xai) over a wide range of TNF concentrations (0-600 pM), whereas IXai only blocked FPA generation at lower TNF concentrations (<50 pM). Activation of coagulation on matrix from TNF-stimulated endothelial cells was dependent on the presence of platelets, indicating the important role of platelets in propagating the reactions leading to fibrin formation. These observations demonstrate the potential of cytokine-stimulateed endothelium and their matrix to activate coagulation and suggest the importance of the intrinsic system in factor Xa formation on cellular surfaces.

Original languageEnglish (US)
Pages (from-to)12067-12074
Number of pages8
JournalJournal of Biological Chemistry
Volume266
Issue number18
StatePublished - 1991
Externally publishedYes

Fingerprint

Factor IXa
Factor IX
Endothelial cells
Coagulation
Extracellular Matrix
Cultured Cells
Endothelial Cells
Tumor Necrosis Factor-alpha
Chemical activation
Thromboplastin
Fibrinopeptide A
Factor Xa
Factor VIIIa
Fibrin
Platelets
Factor X
Factor VII
Blood Platelets
Monolayers
Blood

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Activation of the coagulation mechanism on tumor necrosis factor-stimulated cultured endothelial cells and their extracellular matrix : The role of flow and factor IX/IXa. / Tijburg, Pim N M; Ryan, Jane; Stern, David; Wollitzky, Barry; Rimon, Sara; Rimon, Abraham; Handley, Dean; Nawroth, Peter; Sixma, Jan J.; De Groot, Philip G.

In: Journal of Biological Chemistry, Vol. 266, No. 18, 1991, p. 12067-12074.

Research output: Contribution to journalArticle

Tijburg, PNM, Ryan, J, Stern, D, Wollitzky, B, Rimon, S, Rimon, A, Handley, D, Nawroth, P, Sixma, JJ & De Groot, PG 1991, 'Activation of the coagulation mechanism on tumor necrosis factor-stimulated cultured endothelial cells and their extracellular matrix: The role of flow and factor IX/IXa', Journal of Biological Chemistry, vol. 266, no. 18, pp. 12067-12074.
Tijburg, Pim N M ; Ryan, Jane ; Stern, David ; Wollitzky, Barry ; Rimon, Sara ; Rimon, Abraham ; Handley, Dean ; Nawroth, Peter ; Sixma, Jan J. ; De Groot, Philip G. / Activation of the coagulation mechanism on tumor necrosis factor-stimulated cultured endothelial cells and their extracellular matrix : The role of flow and factor IX/IXa. In: Journal of Biological Chemistry. 1991 ; Vol. 266, No. 18. pp. 12067-12074.
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abstract = "Infusion of tumor necrosis factor (TNF) into tumor-bearing mice led to intravascular clot formation with fibrin deposition in microvessels in the tumor bed in close association with the vessel wall, which could be prevented by active site-blocked factor IXa (IXai). This observation prompted us to examine the role of the intrinsic system in activation of the coagulation mechanism on TNF-stimulated human endothelial cell monolayers and endothelial-derived matrix during exposure to purified coagulation factors or flowing blood. Treatment of endothelial cells in intact monolayers with TNF induced expression of the procoagulant cofactor tissue factor (TF) in a dose-dependent manner, and after removal of the cells, TF was present in the matrix. TNF-treated endothelial cell monolayers exposed to blood anticoagulated with low molecular weight heparin induced activation of coagulation. Addition of IXai blocked the procoagulant response on TNF-treated endothelial cells, and consistent with this, the presence of factor IX/VIIIa enhanced endothelial TF/factor VII(a) factor X activation over a wide range of cytokine concentrations (0-600 PM). When TF-dependent factor X activation on endothelial cells was compared with preparations of subendothelium, the extracellular matrix was 10-20 times more effective. IXai blocked TF/factor VII(a) mediated activated coagulation on matrix, but only at lower concentration of TNF (<50 pM). Similarly, enhancement of factor Xa formation on matrix by factors IX/VIIIa was most evident at lower TNF concentrations. When anticoagulated whole blood flowing with a shear of 300 s-1 was exposed to matrices from TNF-treated endothelial cells, but not matrices from control cells, fibrinopeptide A (FPA) generation, fibrin deposition, and platelet aggregate formation were observed. FPA generation could be prevented by a blocking antibody to TF and by active site-blocked factor Xa (Xai) over a wide range of TNF concentrations (0-600 pM), whereas IXai only blocked FPA generation at lower TNF concentrations (<50 pM). Activation of coagulation on matrix from TNF-stimulated endothelial cells was dependent on the presence of platelets, indicating the important role of platelets in propagating the reactions leading to fibrin formation. These observations demonstrate the potential of cytokine-stimulateed endothelium and their matrix to activate coagulation and suggest the importance of the intrinsic system in factor Xa formation on cellular surfaces.",
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T1 - Activation of the coagulation mechanism on tumor necrosis factor-stimulated cultured endothelial cells and their extracellular matrix

T2 - The role of flow and factor IX/IXa

AU - Tijburg, Pim N M

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AU - Stern, David

AU - Wollitzky, Barry

AU - Rimon, Sara

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AU - Handley, Dean

AU - Nawroth, Peter

AU - Sixma, Jan J.

AU - De Groot, Philip G.

PY - 1991

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N2 - Infusion of tumor necrosis factor (TNF) into tumor-bearing mice led to intravascular clot formation with fibrin deposition in microvessels in the tumor bed in close association with the vessel wall, which could be prevented by active site-blocked factor IXa (IXai). This observation prompted us to examine the role of the intrinsic system in activation of the coagulation mechanism on TNF-stimulated human endothelial cell monolayers and endothelial-derived matrix during exposure to purified coagulation factors or flowing blood. Treatment of endothelial cells in intact monolayers with TNF induced expression of the procoagulant cofactor tissue factor (TF) in a dose-dependent manner, and after removal of the cells, TF was present in the matrix. TNF-treated endothelial cell monolayers exposed to blood anticoagulated with low molecular weight heparin induced activation of coagulation. Addition of IXai blocked the procoagulant response on TNF-treated endothelial cells, and consistent with this, the presence of factor IX/VIIIa enhanced endothelial TF/factor VII(a) factor X activation over a wide range of cytokine concentrations (0-600 PM). When TF-dependent factor X activation on endothelial cells was compared with preparations of subendothelium, the extracellular matrix was 10-20 times more effective. IXai blocked TF/factor VII(a) mediated activated coagulation on matrix, but only at lower concentration of TNF (<50 pM). Similarly, enhancement of factor Xa formation on matrix by factors IX/VIIIa was most evident at lower TNF concentrations. When anticoagulated whole blood flowing with a shear of 300 s-1 was exposed to matrices from TNF-treated endothelial cells, but not matrices from control cells, fibrinopeptide A (FPA) generation, fibrin deposition, and platelet aggregate formation were observed. FPA generation could be prevented by a blocking antibody to TF and by active site-blocked factor Xa (Xai) over a wide range of TNF concentrations (0-600 pM), whereas IXai only blocked FPA generation at lower TNF concentrations (<50 pM). Activation of coagulation on matrix from TNF-stimulated endothelial cells was dependent on the presence of platelets, indicating the important role of platelets in propagating the reactions leading to fibrin formation. These observations demonstrate the potential of cytokine-stimulateed endothelium and their matrix to activate coagulation and suggest the importance of the intrinsic system in factor Xa formation on cellular surfaces.

AB - Infusion of tumor necrosis factor (TNF) into tumor-bearing mice led to intravascular clot formation with fibrin deposition in microvessels in the tumor bed in close association with the vessel wall, which could be prevented by active site-blocked factor IXa (IXai). This observation prompted us to examine the role of the intrinsic system in activation of the coagulation mechanism on TNF-stimulated human endothelial cell monolayers and endothelial-derived matrix during exposure to purified coagulation factors or flowing blood. Treatment of endothelial cells in intact monolayers with TNF induced expression of the procoagulant cofactor tissue factor (TF) in a dose-dependent manner, and after removal of the cells, TF was present in the matrix. TNF-treated endothelial cell monolayers exposed to blood anticoagulated with low molecular weight heparin induced activation of coagulation. Addition of IXai blocked the procoagulant response on TNF-treated endothelial cells, and consistent with this, the presence of factor IX/VIIIa enhanced endothelial TF/factor VII(a) factor X activation over a wide range of cytokine concentrations (0-600 PM). When TF-dependent factor X activation on endothelial cells was compared with preparations of subendothelium, the extracellular matrix was 10-20 times more effective. IXai blocked TF/factor VII(a) mediated activated coagulation on matrix, but only at lower concentration of TNF (<50 pM). Similarly, enhancement of factor Xa formation on matrix by factors IX/VIIIa was most evident at lower TNF concentrations. When anticoagulated whole blood flowing with a shear of 300 s-1 was exposed to matrices from TNF-treated endothelial cells, but not matrices from control cells, fibrinopeptide A (FPA) generation, fibrin deposition, and platelet aggregate formation were observed. FPA generation could be prevented by a blocking antibody to TF and by active site-blocked factor Xa (Xai) over a wide range of TNF concentrations (0-600 pM), whereas IXai only blocked FPA generation at lower TNF concentrations (<50 pM). Activation of coagulation on matrix from TNF-stimulated endothelial cells was dependent on the presence of platelets, indicating the important role of platelets in propagating the reactions leading to fibrin formation. These observations demonstrate the potential of cytokine-stimulateed endothelium and their matrix to activate coagulation and suggest the importance of the intrinsic system in factor Xa formation on cellular surfaces.

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