Expression of exogenous tissue factor pathway inhibitor in vivo suppresses thrombus formation in injured rabbit carotid arteries

AbstractActivation of inflammatory and procoagulant mechanisms is thought to contribute significantly to the initiation of restenosis, a common complication after balloon angioplasty of obstructed arteries. During this process, expression of tissue factor (TF) represents one of the major physiologic triggers of coagulation that results in thrombus formation and the generation of additional signals leading to vascular smooth muscle cell (VSMC) proliferation and migration. In this study, we have investigated the mechanisms by which inhibition of coagulation at an early stage through overexpression of tissue factor pathway inhibitor (TFPI), an endogenous inhibitor of TF, might reduce restenosis. In a rabbit femoral artery model, percutaneous delivery of TFPI using a recombinant adenoviral vector resulted in a significant reduction of the intimamedia ratio 21 days after injury. Investigating several markers of inflammation and coagulation, we found reduced neointimal expression of monocyte chemoattractant protein-1 (MCP-1), lesional monocyte infiltration, and expression of vascular TF, matrix metalloproteinase-2 (MMP-2), and MMP-9. Moreover, overexpression of TFPI suppressed the autocrine release of platelet-derived growth factor BB (PDGF-BB), MCP-1, and MMP-2 in response to factors VIIa and Xa from VSMCs in vitro and inhibited monocyte TF activity. These results suggest that TFPI exerts its action in vivo through not only thrombotic, but also nonthrombotic mechanisms.

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Endogenous Tissue Factor Pathway Inhibitor Modulates Thrombus Formation in an In Vivo Model of Rabbit Carotid Artery Stenosis and Endothelial Injury

Circulation ◽

10.1161/01.cir.102.1.113 ◽

2000 ◽

Vol 102 (1) ◽

pp. 113-117 ◽

Cited By ~ 25

Author(s):

Massimo Ragni ◽

Paolo Golino ◽

Plinio Cirillo ◽

Annalisa Scognamiglio ◽

Orlando Piro ◽

...

Keyword(s):

Carotid Artery ◽

Tissue Factor ◽

Carotid Artery Stenosis ◽

Tissue Factor Pathway Inhibitor ◽

Thrombus Formation ◽

Endothelial Injury ◽

In Vivo Model ◽

Rabbit Carotid Artery ◽

Tissue Factor Pathway

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Adenovirus-mediated Gene Transfer of Recombinant Tissue Factor Pathway Inhibitor in Vitro and in Vivo

Journal of the American College of Cardiology ◽

10.1016/s0735-1097(97)84333-1 ◽

1998 ◽

Vol 31 (2) ◽

pp. 145A

Author(s):

P Zoldhelyi

Keyword(s):

Gene Transfer ◽

Tissue Factor ◽

Tissue Factor Pathway Inhibitor ◽

Tissue Factor Pathway

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Effect of Depolymerized Holothurian Glycosaminoglycan (DHG) on Tissue Factor Pathway Inhibitor: In Vitro and In Vivo Studies

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657643 ◽

1997 ◽

Vol 78 (02) ◽

pp. 864-870 ◽

Cited By ~ 9

Author(s):

Hideki Nagase ◽

Kei-ichi Enjyoji ◽

Yu-ichi Kamikubo ◽

Keiko T Kitazato ◽

Kenji Kitazato ◽

...

Keyword(s):

Tissue Factor ◽

Tissue Factor Pathway Inhibitor ◽

Factor Xa ◽

Free Form ◽

Initial Reaction ◽

Extrinsic Pathway ◽

Factor Xa Inhibition ◽

Tissue Factor Pathway

SummaryDepolymerized holothurian glycosaminoglycan (DHG) is a glycosaminoglycan extracted from the sea cucumber Stichopus japonicusSelenka. In previous studies, we demonstrated that DHG has antithrombotic and anticoagulant activities that are distinguishable from those of heparin and dermatan sulfate. In the present study, we examined the effect of DHG on the tissue factor pathway inhibitor (TFPI), which inhibits the initial reaction of the tissue factor (TF)-mediated coagulation pathway. We first examined the effect of DHG on factor Xa inhibition by TFPI and the inhibition of TF-factor Vila by TFPI-factor Xa in in vitro experiments using human purified proteins. DHG increased the rate of factor Xa inhibition by TFPI, which was abolished either with a synthetic C-terminal peptide or with a synthetic K3 domain peptide of TFPI. In contrast, DHG reduced the rate of TF-factor Vila inhibition by TFPI-factor Xa. Therefore, the effect of DHG on in vitroactivity of TFPI appears to be contradictory. We then examined the effect of DHG on TFPI in cynomolgus monkeys and compared it with that of unfractionated heparin. DHG induced an increase in the circulating level of free-form TFPI in plasma about 20-fold when administered i.v. at 1 mg/kg. The prothrombin time (PT) in monkey plasma after DHG administration was longer than that estimated from the plasma concentrations of DHG. Therefore, free-form TFPI released by DHG seems to play an additive role in the anticoagulant mechanisms of DHG through the extrinsic pathway in vivo. From the results shown in the present work and in previous studies, we conclude that DHG shows anticoagulant activity at various stages of coagulation reactions, i.e., by inhibiting the initial reaction of the extrinsic pathway, by inhibiting the intrinsic Xase, and by inhibiting thrombin.

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The 536C→T Transition in the Human Tissue Factor Pathway Inhibitor (TFPI) Gene Is Statistically Associated with a Higher Risk for Venous Thrombosis

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614619 ◽

1999 ◽

Vol 82 (07) ◽

pp. 1-5 ◽

Cited By ~ 14

Author(s):

Michael Schmidt ◽

Christian Götting ◽

Britt Schwenz ◽

Stefan Lange ◽

Gert Müller-Berghaus ◽

...

Keyword(s):

Venous Thrombosis ◽

Tissue Factor ◽

Tissue Factor Pathway Inhibitor ◽

Blood Donors ◽

Amino Acid Position ◽

Immunological Methods ◽

Venous Thromboembolic ◽

Coagulation Inhibitors ◽

Tissue Factor Pathway

SummaryTissue factor pathway inhibitor (TFPI) is an important regulator in the extrinsic blood coagulation pathway. Although the regulatory biochemical role of TFPI is evident, the clinical significance of this proteinase inhibitor remains to be elucidated. The definition of a clinical TFPI deficiency seems to be more complex than that of other coagulation inhibitors because the activity and concentration of circulating TFPI can not be considered a true measure of in vivo levels. Its determination in plasma samples by immunological methods or functional assays has been shown to be inadequate in the detection of a clinical deficiency.Therefore, we screened genomic DNA samples of blood donors and thrombotic patients for alterations in the TFPI gene to assess the influence of a modified TFPI in venous thromboembolic diseases. We detected a single nucleotide substitution in exon 7 (536C→T) leading to a proline to leucine exchange at amino acid position 151 of the protein ([P151L]TFPI) and found the prevalence of heterozygous carriers in German unrelated blood donors to be 0.2% (n = 5120).Four unrelated persons out of 14 probands carrying the genetic variation could be linked to venous thrombosis. For calculation of a potential risk for venous thrombosis for carriers of the mutation we investigated healthy blood donors about thrombotic events. 7 out of 308 blood donors were found to have a history of venous thrombosis, one of them carried the TFPI mutation. Statistical calculation showed a significant relative risk for venous thrombosis for individuals with the trait (odds ratio, 9.3; confidence interval, 1.8-48.6; p <0.01).

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Caveolin-1 Enhances Tissue Factor Pathway Inhibitor Exposure and Function on the Cell Surface

Journal of Biological Chemistry ◽

10.1074/jbc.m503333200 ◽

2005 ◽

Vol 280 (23) ◽

pp. 22308-22317 ◽

Cited By ~ 18

Author(s):

Cristina Lupu ◽

Xiaohong Hu ◽

Florea Lupu

Keyword(s):

Tissue Factor ◽

Tissue Factor Pathway Inhibitor ◽

Factor Viia ◽

Protease Production ◽

Extrinsic Pathway ◽

Caveolin 1 ◽

Tissue Factor Pathway ◽

Quaternary Complex ◽

And Function

Tissue factor pathway inhibitor (TFPI) blocks tissue factor-factor VIIa (TF-FVIIa) activation of factors X and IX through the formation of the TF-FVIIa-FXa-TFPI complex. Most TFPI in vivo associates with caveolae in endothelial cells (EC). The mechanism of this association and the anticoagulant role of caveolar TFPI are not yet known. Here we show that expression of caveolin-1 (Cav-1) in 293 cells keeps TFPI exposed on the plasmalemma surface, decreases the membrane lateral mobility of TFPI, and increases the TFPI-dependent inhibition of TF-FVIIa. Caveolae-associated TFPI supports the co-localization of the quaternary complex with caveolae. To investigate the significance of these observations for EC we used RNA interference to deplete the cells of Cav-1. Functional assays and fluorescence microscopy revealed that the inhibitory properties of TFPI were diminished in EC lacking Cav-1, apparently through deficient assembly of the quaternary complex. These findings demonstrate that caveolae regulate the inhibition by cell-bound TFPI of the active protease production by the extrinsic pathway of coagulation.

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Recombinant full-length tissue factor pathway inhibitor fails to bind to the cell surface: implications for catabolism in vitro and in vivo

Blood ◽

10.1182/blood.v95.6.1973 ◽

2000 ◽

Vol 95 (6) ◽

pp. 1973-1978 ◽

Cited By ~ 7

Author(s):

Guyu Ho ◽

Masaaki Narita ◽

George J. Broze ◽

Alan L. Schwartz

Keyword(s):

Tissue Factor ◽

Tissue Factor Pathway Inhibitor ◽

Feedback Inhibition ◽

Factor Xa ◽

Full Length ◽

Pharmacokinetic Studies ◽

Dependent Manner ◽

Carboxy Terminus ◽

Tissue Factor Pathway

Abstract Tissue factor pathway inhibitor (TFPI) plays a key role in the regulation of tissue factor-initiated blood coagulation secondary to loss of the integrity of the blood vessel wall. TFPI is a naturally occurring Kunitz-type protease inhibitor that inhibits coagulation factor Xa and, in a factor Xa-dependent manner, mediates feedback inhibition of the factor VIIa/tissuefactor catalytic complex. In vivo full-length TFPI is thought to be primarily bound to the vascular endothelium and the high affinity binding requires an intact carboxy terminus. Here we describe a full-length TFPI molecule, expressed in mouse C127 cells (TFPIC127), which exhibits virtually no cellular binding yet contains the intact carboxy terminus. This TFPI (TFPIC127) is neither internalized nor degraded via the TFPI endocytic receptor, LDL-receptor–related protein. Pharmacokinetic studies of TFPIC127 in vivo demonstrate a 10-fold prolongation in the plasma half-life, compared with that of bacterial recombinant TFPI.

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