Tissue factor pathway inhibitor (TFPI) produces factor Xa (FXa)-dependent feedback inhibition of factor VIIa/tissue factor (FVIIa/TF)-induced coagulation. In humans, TFPI is expressed as two isoforms. TFPI1 contains three tandem Kunitz-type protease inhibitor domains, Kunitz-1 binds FVIIa in the FVIIa/TF complex and the second binds FXa. Kunitz-3 does not possess protease inhibitor activity. TFPI2 lacks the Kunitz-3 and the C-terminus of TFPI1. The latter is required for TFPI1's optimal inhibition of FXa and its anticoagulant activity in one-stage coagulation assays. TFPI1 interacts with protein S (PS), which enhances its anti-FXa activity, and with factor V (FV), which may prolong its clearance from plasma. Both forms of TFPI associate with the surface of cells in a glycosylphosphatidylinositol (GPI)-anchor dependent fashion, but through different mechanisms. TFPI1, present in plasma and secreted by activated platelets, appears to bind to another GPI-anchored protein(s), whereas TFPI2 contains an intrinsic GPI-anchor. We plan to determine the structures within TFPI1 responsible for its binding to PS and FV and further explore its interactions with cell surfaces. The form(s) of TFPI, which circulate bound to lipoproteins, will be identified and the relevance of a potential additional isoform of TFPI, TFPI4, will be investigated. TFPI gene-disrupted mice (TFPI KO) die intrautero of widespread thrombosis and a consumptive coagulopathy, but are rescued by a low level of either of two transgenes with FVIIa/TF inhibitory activity. One, Tie2-hTFPI1, directs expression in endothelial cells, the other, Tf-mXK1-hAlb directs expression into plasma. Transgene-rescued TFPI KO mice and appropriate bone marrow transplantation studies will be used to assess the role of platelet- associated TFPI1 and to determine the effect of low levels of endogenous TFPI on thrombosis, inflammation, and atherosclerosis using mouse models.

Public Health Relevance

TFPI is a plasma and cell-associated protein, which exists as two isoforms (TFPI1 and TFPI2) and plays an important role in regulating the initiation of coagulation. The goals of this proposal are to determine the mechanism and relevance of its interactions with circulation lipoproteins, cell surfaces, and two other coagulation-related proteins, Protein S and Factor V, and to determine the effects of TFPI deficiency on thrombosis, inflammation, and atherosclerosis using mouse models.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
4R37HL077193-10
Application #
8889772
Study Section
Hemostasis and Thrombosis Study Section (HT)
Program Officer
Link, Rebecca P
Project Start
2004-07-15
Project End
2019-07-31
Budget Start
2014-08-01
Budget End
2015-07-31
Support Year
10
Fiscal Year
2014
Total Cost
$415,181
Indirect Cost
$142,931
Name
Washington University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
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Kanse, Sandip M; Declerck, Paul J; Ruf, Wolfram et al. (2012) Factor VII-activating protease promotes the proteolysis and inhibition of tissue factor pathway inhibitor. Arterioscler Thromb Vasc Biol 32:427-33
Broze Jr, George J; Girard, Thomas J (2012) Tissue factor pathway inhibitor: structure-function. Front Biosci (Landmark Ed) 17:262-80
Girard, Thomas J; Tuley, Elodee; Broze Jr, George J (2012) TFPI? is the GPI-anchored TFPI isoform on human endothelial cells and placental microsomes. Blood 119:1256-62
Ndonwi, Matthew; Tuley, Elodee A; Broze Jr, George J (2010) The Kunitz-3 domain of TFPI-alpha is required for protein S-dependent enhancement of factor Xa inhibition. Blood 116:1344-51