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J Biol Chem. 2014 Nov 7;289(45):31647-61. doi: 10.1074/jbc.M114.569665. Epub 2014 Sep 28.

Platelets contain tissue factor pathway inhibitor-2 derived from megakaryocytes and inhibits fibrinolysis.

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From the UCLA/Orthopaedic Hospital Department of Orthopaedic Surgery.
the Department of Molecular and Medical Pharmacology.
the Department of Pediatrics, Division of Hematology, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
the Department of Molecular and Medical Pharmacology, the Molecular Biology Institute, UCLA, Los Angeles, California 90095 and.
the Department of Medicine, Division of Pulmonology and Critical Care, and.
From the UCLA/Orthopaedic Hospital Department of Orthopaedic Surgery, the Molecular Biology Institute, UCLA, Los Angeles, California 90095 and


Tissue factor pathway inhibitor-2 (TFPI-2) is a homologue of TFPI-1 and contains three Kunitz-type domains and a basic C terminus region. The N-terminal domain of TFPI-2 is the only inhibitory domain, and it inhibits plasma kallikrein, factor XIa, and plasmin. However, plasma TFPI-2 levels are negligible (≤20 pM) in the context of influencing clotting or fibrinolysis. Here, we report that platelets contain significant amounts of TFPI-2 derived from megakaryocytes. We employed RT-PCR, Western blotting, immunohistochemistry, and confocal microscopy to determine that platelets, MEG-01 megakaryoblastic cells, and bone marrow megakaryocytes contain TFPI-2. ELISA data reveal that TFPI-2 binds factor V (FV) and partially B-domain-deleted FV (FV-1033) with K(d) ~9 nM and binds FVa with K(d) ~100 nM. Steady state analysis of surface plasmon resonance data reveal that TFPI-2 and TFPI-1 bind FV-1033 with K(d) ~36-48 nM and bind FVa with K(d) ~252-456 nM. Further, TFPI-1 (but not TFPI-1161) competes with TFPI-2 in binding to FV. These data indicate that the C-terminal basic region of TFPI-2 is similar to that of TFPI-1 and plays a role in binding to the FV B-domain acidic region. Using pull-down assays and Western blots, we show that TFPI-2 is associated with platelet FV/FVa. TFPI-2 (~7 nM) in plasma of women at the onset of labor is also, in part, associated with FV. Importantly, TFPI-2 in platelets and in plasma of pregnant women inhibits FXIa and tissue-type plasminogen activator-induced clot fibrinolysis. In conclusion, TFPI-2 in platelets from normal or pregnant subjects and in plasma from pregnant women binds FV/Va and regulates intrinsic coagulation and fibrinolysis.


Fibrinolysis; Hemostasis; Plasmin; Platelet; Protease Inhibitor

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