Thrombin Stimulates Expression of Tissue-Type Plasminogen Activator and Plasminogen Activator lnhibitor Type 1 in Cultured Human Vascular Smooth Muscle Cells

Johann Wojta; Marisa Gallicchio; Hans Zoellner; Peter Hufnagl; Karena Last; Enrico L Filonzi; Bernd R Binder; John A Hamilton; Katherine McGrath

doi:10.1055/s-0038-1649607

Thrombosis and Haemostasis, Inhaltsverzeichnis

Thromb Haemost 1993; 70(03): 469-474
DOI: 10.1055/s-0038-1649607

Original Article

Fibrinolysis

Schattauer GmbH Stuttgart

Thrombin Stimulates Expression of Tissue-Type Plasminogen Activator and Plasminogen Activator lnhibitor Type 1 in Cultured Human Vascular Smooth Muscle Cells

Johann Wojta

¹The Dept. Diagnostic Haematology, Royal Melbourne Hospital, Parkville, Victoria, Australia

²The Dept. Medical Physiology, Laboratory for Clinical and Experimental Physiology, University of Vienna, Austria

,

Marisa Gallicchio

¹The Dept. Diagnostic Haematology, Royal Melbourne Hospital, Parkville, Victoria, Australia

,

Hans Zoellner

³The Dept. Medicine, University of Melbourne, Parkville, Victoria, Australia

,

Peter Hufnagl

²The Dept. Medical Physiology, Laboratory for Clinical and Experimental Physiology, University of Vienna, Austria

,

Karena Last

³The Dept. Medicine, University of Melbourne, Parkville, Victoria, Australia

,

Enrico L Filonzi

³The Dept. Medicine, University of Melbourne, Parkville, Victoria, Australia

,

Bernd R Binder

²The Dept. Medical Physiology, Laboratory for Clinical and Experimental Physiology, University of Vienna, Austria

,

John A Hamilton

³The Dept. Medicine, University of Melbourne, Parkville, Victoria, Australia

,

Katherine McGrath

¹The Dept. Diagnostic Haematology, Royal Melbourne Hospital, Parkville, Victoria, Australia

› Institutsangaben

Abstract

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Summary

The effect of thrombin on the fibrinolytic potential of human vascular smooth muscle cells (SMC) in culture was studied. SMC of different origin responded to thrombin treatment with a dose and time dependent increase in tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type-1 (PAI-1) levels in both cell lysates and conditioned media with maximum effects achieved at 10-20 IU/ml thrombin. PAI-1 antigen levels also increased in the extracellular matrix of thrombin treated SMC. PAI-2 levels in cell lysates of such SMC were not affected by thrombin. The effect was restricted to active thrombin, since DFP-thrombin and thrombin treated with hirudin showed no increasing effect on t-PA and PAI-1 levels in SMC.

Enzymatically active thrombin also caused a four-fold increase in specific PAI-1 mRNA and a three-fold increase in t-PA mRNA.

Furthermore we demonstrated the presence of high and low affinity binding sites for thrombin on the surface of SMC with a K _D = 4.3 × 10⁻¹⁰ M and 9.0 × 10⁴ sites per cell and a KD = 0.6 × 10⁻⁸ M and 5.8 × 10⁵ sites per cell respectively.

Thrombin could come in contact with SMC in case of vascular injury or following gap formation between endothelial cells. Our data support the idea that besides its known proliferative effect for SMC, thrombin could also modulate their fibrinolytic system.

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Referenzen

References
1 Clowes AW, Clowes MM, Au YPT, Reidy MA, Belin D. Smooth muscle cells express urokinase during mitogenesis and tissue-type plasminogen activator during migration in injured rat carotid artery. Circ Res 1990; 67: 61-67
2 Booyse FM, Scheinbuks J, Radek J, Osikowicz G, Feder S, Quarfoot AJ. Immunological identification and comparison of plasminogen activator forms in cultured normal human endothelial cells and smooth muscle cells. Thromb Res 1981; 24: 495-504
3 Levin EG, Loskutoff DJ. Comparative studies of the fibrinolytic activity of cultured vascular cells. Thromb Res 1979; 15: 869-878
4 Laug WE, Aebersold R, Jong A, Rideout W, Bergman BL, Baker J. Isolation of multiple types of plasminogen activator inhibitors from vascular smooth muscle cells. Thromb Haemostas 1989; 61: 517-521
5 Bell L, Madri JA. Influence of the angiotensin system on endothelial and smooth muscle cell migration. Am J Path 1990; 137: 7-12
6 Reily CF, McFall RC. Platelet-derived growth factor and transforming growth factor-Beta regulate plasminogen activator inhibitor-1 synthesis in vascular smooth muscle cells. J Biol Chem 1991; 266: 9419-9427
7 Nelken NA, Soifer SJ, O'Keefe J, Vu TKH, Charo IF, Coughlin SR. Thrombin receptor expression in normal and atherosclerotic human arteries. J Clin Invest 1992; 90: 1614-1621
8 Schneiderman J, Sawdey MS, Keeton MR, Bordin GM, Bernstein EF, Dilley RB, Loskutoff DJ. Increased type 1 plasminogen activator inhibitor gene expression in atherosclerotic human arteries. Proc Nat Acad Sci USA 1992; 89: 6998-7002
9 Loskutoff DJ. Effect of thrombin on the fibrinolytic activity of cultured bovine endothelial cells. J Clin Invest 1979; 64: 329-332
10 Levin EG, Marzec U, Anderson J, Harker LA. Thrombin stimulates tissue plasminogen activator release from cultured human endothelial cells. J Clin Invest 1984; 74: 1988-1995
11 Gelehrter TD, Sznycer-Laszuk R. Thrombin inducation of plasminogen activator inhibitor in cultured human endothelial cells. J Clin Invest 1986; 77: 165-169
12 Van Hinsbergh VWM, Sprengers ED, Kooistra T. Effect of thrombin on the production of plasminogen activators and PA inhibitor-1 by human foreskin microvascular endothelial cells. Thromb Haemostas 1987; 57: 148-153
13 Dichek D, Quertermous T. Thrombin regulation of mRNA levels of tissue plasminogen activator and plasminogen activator inhibitor-1 in cultured human umbilical vein endothelial cells. Blood 1989; 74: 222-228
14 Chamley-Campbell J, Campbell GR, Ross R. The smooth muscle cell in culture. Physiol Rev 1979; 59: 1-61
15 Gospodarowicz D. Purification of a fibroblast growth factor from bovine pituitary. J Biol Chem 1975; 250: 2515-2520
16 Wojta J, Holzer M, Hufnagl P, Christ G, Hoover RL, Binder BR. Hyperthermia stimulates plasminogen activator inhibitor type 1 (PAI-1) expression in human umbilical vein endothelial cells (HUVEC) in vitro. Am J Pathol 1991; 39: 911-920
17 Wojta J, Turcu L, Wagner OF, Korninger C, Binder BR. Evaluation of fibrinolytic capacity by a combined assay system for tissue-type plasminogen activator antigen and function using monoclonal anti-tissue type plasminogen activator antibodies. J Lab Clin Med 1987; 109: 665-671
18 Wojta J, Binder BR, Huber K, Hoover RL. Evaluation of fibrinolytic capacity in plasma during thrombolytic therapy with single (scu-PA) or two-chain urokinase type plasminogen activator (tcu-PA) by a combined assay system for urokinase type plasminogen activator antigen and function. Thromb Haemostas 1989; 61: 289-293
19 Wagner OF, Binder BR. Purification of an active plasminogen activator inhibitor immunologically related to the endothelial type plasminogen activator inhibitor from the conditioned media of a human melanoma cell line. J Biol Chem 1986; 261: 14474-14478
20 Resch I, Krutisch G, Geiger M, Binder BR. ELISA systems for active and total plasminogen activator inhibitor 1 (PAI-1) antigen. Thromb Haemostas 1989; 62: 229 (abstr)
21 Knudsen BS, Harpel PC, Nachman RL. Plasminogen activator inhibitor is associated with the extracellular matrix of cultured bovine smooth muscle cells. J Clin Invest 1987; 80: 1082-1089
22 Kruithof EKO, Vassalli JD, Schleuning WD, Mattaliano RJ, Bachmann F. Purification and characterization of a plasminogen activator inhibitor from the histiocytic lymphoma cell line U-937. J Biol Chem 1986; 261: 11207-11213
23 Antalis TM, Clark MA, Barnes T, Lehrbach PR, Devine PL, Schevzov G, Goss NH, Stephens RW, Tolstoshev P. Cloning and expression of a cDNA coding for a human monocyte-derived plasminogen activator inhibitor. Proc Natl Acad Sci USA 1985; 85: 985-989
24 Chirgwin JM, Przybyla AE, McDonald RJ, Rutter WJ. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry 1979; 18: 5294-5299
25 Glenn KC, Carney DH, Fenton JW, Cunningham DD. Thrombin active site regions required for fibroblast receptor binding and initiation of cell division. J Biol Chem 1980; 252: 6609-6616
26 Fenton JW. Thrombin. Ann NY Acad Sci 1986; 485: 5-11
27 Fenton JW, Bing DH. Thrombin active site regions. Semin Thromb Hemostas 1986; 12: 200-208
28 White JG. Platelet morphology and function. In: Hematology. Williams WJ, Butler E, Erslve AJ, Lichtman MJ. (eds) New York, NY: McGraw-Hill; 1983: 1128-1135
29 Bizios R, Lai L, Fenton JW, Malik AB. Thrombin induced chemotaxis and aggregation of neutrophils. J Cell Physiol 1986; 128: 485-490
30 Wilner GD, Danitz MP, Mudd MS, Hsieh KH, Fenton JW. Selective immobilization of alpha thrombin by surface bound fibrin. J Lab Clin Med 1981; 97: 402-411
31 Laposata M, Dovnarsky DK, Solkin HS. Thrombin induced gap formation in confluent endothelial cell monolayers in vitro. Blood 1983; 62: 549-556
32 Garcia JGN, Siflinger-Birnboim A, Bizios R, Del Vecchio, Fenton JW, Malik AB. Thrombin induced increase in albumin permeability across the endothelium. J Cell Physiol 1986; 128: 96-104
33 Bar-Shavit R, Benezra M, Eldor A, Hy-Am E, Fenton JW, Wilner GD, Vlodavsky I. Thrombin immobilized to extracellular matrix is a potent mitogen for vascular smooth muscle cells: nonenzymatic mode of action. Cell Reg 1990; 1: 453-464
34 de Fouw NJ, van Hinsbergh VWM, de Jong YF, Haverkate F, Bertina RM. The interaction of activated protein C and thrombin with plasminogen activator inhibitor released from human endothelial cells. Thromb Haemostas 1987; 57: 176-182
35 He CJ, Rondeau E, Medcalf R, Lacave R, Schleuning WD, Sraer JD. Thrombin increases proliferation and decreases fibrinolytic activity of kidney glomerular epithelial cells. J Cell Physiol 1991; 146: 131-140
36 Awbrey BJ, Hoak JC, Owen EG. Binding of human thrombin to cultured human endothelial cells. J Biol Chem 1979; 254: 4092-4095
37 Villamediana LM, Rondeau E, He CH, Medcalf RL, Peraldi MAN, Lacave R, Delarue F, Sraer JD. Thrombin regulates components of the fibrinolytic system in human mesangial cells. Kidney Int 1990; 38: 956-961
38 Au YPT, Kenagy RD, Clowes AW. Heparin selectively inhibits the transcription of tissue-type plasminogen activator in primate arterial smooth muscle cell during mitogenesis. J Biol Chem 1992; 267: 3438-3444