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Plasmin induces apoptosis of aortic valvular myofibroblasts

Authors

  • Najlah Kochtebane,

    1. Inserm, UMR698, Hemostasis, Bio-Engineering and Cardiovascular Remodelling, Paris 7 Denis Diderot University, Hôpital Bichat-Claude Bernard, 46 Rue Henri Huchard, 75877 Paris Cedex 18, France
    2. Laboratoire de Biochimie, Unité de Recherche 02/UR/09-01, Institut Supérieur de Biotechnologie, Université de Monastir, 5000 Monastir, Tunisia
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  • Christine Choqueux,

    1. Inserm, UMR698, Hemostasis, Bio-Engineering and Cardiovascular Remodelling, Paris 7 Denis Diderot University, Hôpital Bichat-Claude Bernard, 46 Rue Henri Huchard, 75877 Paris Cedex 18, France
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  • Stéphanie Passefort,

    1. Inserm, UMR698, Hemostasis, Bio-Engineering and Cardiovascular Remodelling, Paris 7 Denis Diderot University, Hôpital Bichat-Claude Bernard, 46 Rue Henri Huchard, 75877 Paris Cedex 18, France
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  • Patrick Nataf,

    1. Cardiovascular Surgery Unit, Paris 7 Denis Diderot University, Hôpital Bichat-Claude Bernard, 46 Rue Henri Huchard, 75877 Paris Cedex 18, France
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  • David Messika-Zeitoun,

    1. Cardiology Unit, AP-HP, Paris 7 Denis Diderot University, Hôpital Bichat-Claude Bernard, 46 Rue Henri Huchard, 75877 Paris Cedex 18, France
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  • Aghleb Bartagi,

    1. Laboratoire de Biochimie, Unité de Recherche 02/UR/09-01, Institut Supérieur de Biotechnologie, Université de Monastir, 5000 Monastir, Tunisia
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  • Jean-Baptiste Michel,

    1. Inserm, UMR698, Hemostasis, Bio-Engineering and Cardiovascular Remodelling, Paris 7 Denis Diderot University, Hôpital Bichat-Claude Bernard, 46 Rue Henri Huchard, 75877 Paris Cedex 18, France
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  • Eduardo Anglés-Cano,

    1. Inserm, U919, Serine Proteases in Neurovascular Pathology, GIP Cyceron/Bd Henri Becquerel, 14074, Caen, France
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  • Marie-Paule Jacob

    Corresponding author
    1. Inserm, UMR698, Hemostasis, Bio-Engineering and Cardiovascular Remodelling, Paris 7 Denis Diderot University, Hôpital Bichat-Claude Bernard, 46 Rue Henri Huchard, 75877 Paris Cedex 18, France
    • Inserm UMR698, Hôpital Bichat-Claude Bernard, 46 Rue Henri Huchard, 75877 Paris Cedex 18, France.
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  • No conflicts of interest were declared.

Abstract

Previous studies have described remodelling of the extracellular substratum by matrix metalloproteinases (MMPs) in aortic valves. However, involvement of the fibrinolytic system has not yet been analysed. We hypothesized that plasminogen and plasminogen activator(s) are present in aortic valves and that plasminogen activation could induce the degradation of adhesive proteins and apoptosis of the valvular myofibroblasts. We employed ELISA, western blotting, fibrin-agar zymography, and immunochemistry to detect components of the plasminogen activation system, in samples of aortic valves and valvular myofibroblasts in primary culture. Using myofibroblast cultures, real-time measurement of plasminogen activation was performed in the absence and presence of inhibitors (amiloride, ε-aminocaproic acid, and an MMP inhibitor); the degradation of fibronectin was visualized on western blots; and the apoptotic process was assessed by detection of phosphatidylserine exposure (binding of FITC-annexin V) and DNA fragmentation (TUNEL and ELISA). We demonstrate that a time- and plasminogen concentration-dependent generation of plasmin occurs on the surface of cultured valvular myofibroblasts expressing both u-PA and t-PA. Only u-PA appears to activate plasminogen as t-PA is essentially found in complex with PAI-1. Plasmin-dependent degradation of pericellular proteins, such as fibronectin, leads to cell detachment and apoptosis. In conclusion, various proteins of the fibrinolytic system are synthesized in vitro by cultured myofibroblasts from aortic valves, leading to plasmin-dependent cell detachment-induced apoptosis, a biological process named anoikis. The presence of plasminogen in aortic valves suggests that this process may be operating in vivo and may participate in valvular tissue remodelling, as also suggested by the finding of apoptotic cells in valvular tissue. This is the first demonstration of the presence and potential role of enzymes of the fibrinolytic system in aortic valves. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

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