Neutrophils stimulated by apolipoprotein(a) generate fragments that are stronger inhibitors of plasmin formation than apo(a)

Leila B. Lamanuzzi; El Mostafa Mtairag; Gabriella Pepe; Eduardo Anglés-Cano

doi:10.1160/TH04-04-0241

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2004; 92(05): 1066-1075
DOI: 10.1160/TH04-04-0241

Wound Healing and Inflammation/Infection

Schattauer GmbH

Neutrophils stimulated by apolipoprotein(a) generate fragments that are stronger inhibitors of plasmin formation than apo(a)

Authors

Leila B. Lamanuzzi

¹Department of Biochemistry & Molecular Biology & IBBE-CNR, University of Bari, Italy

³INSERM U460, CHU Bichat-Claude Bernard, Paris, France
El Mostafa Mtairag

²Hassan II University, School of Sciences I, Department of Biology, Casablanca, Morocco

³INSERM U460, CHU Bichat-Claude Bernard, Paris, France
Gabriella Pepe

¹Department of Biochemistry & Molecular Biology & IBBE-CNR, University of Bari, Italy
Eduardo Anglés-Cano

³INSERM U460, CHU Bichat-Claude Bernard, Paris, France

Abstract

Summary

Apolipoprotein(a), the plasminogen-like component of lipoprotein(a), is transformed into fragments by polymorphonuclear neutrophils (PMNs) elastase. Since stimulated PMNs express urokinase-type plasminogen activator (uPA), we sought to investigate the relevance of apo(a) fragmentation on plasminogen activation by neutrophils. Freshly isolated human PMNs stimulated by a 10 kringle recombinant apo(a), r-apo(a), activate plasminogen in a specific and saturable manner (Km = 476 ± 42 nM, Vmax = 896 ± 18 pmol min-1). This activation is prevented by amiloride, an inhibitor of u-PA, and ɛ-aminocaproic acid, ɛ-ACA, a lysine analogue that blocks plasminogen binding to PMNs. Stimulation of PMNs by apo(a) results in the formation of elastase-derived apo(a) fragments. These fragments produce a concentration-dependent decrease in the formation of plasmin. Addition of elastase inhibitors to PMNs prevented degradation of apo(a) and partially restored the formation of plasmin. In a similar manner, isolated r-apo(a) fragments were able to produce a 100% decrease in plasmin generation as compared to intact r-apo(a). These data indicate that apo(a) fragments produce a more pronounced inhibition in the generation of cellbound plasmin by uPA than the parent apo(a). These effects of apo(a) and its fragments were neutralised by a monoclonal antibody directed against the lysine-binding site of apo(a). This mechanism may be of biological relevance to the effects of Lp(a) in conditions where PMNs accumulate and release elastase, i.e. thrombus lysis and inflammatory lesions.

Keywords

Neutrophils - apolipoprotein(a) - elastase - plasminogen - proteolysis

Full Text

References

References
1 Borregaard N, Cowland JB. Granules of the human neutrophilic polymorphonuclear leukocyte. Blood 1997; 89: 3503-21.
2 Heiple JM, Ossowski L. Human neutrophil plasminogen activator is localized in specific granules and is translocated to the cell surface by exocytosis. J Exp Med 1986; 164: 826-40.
3 Pedersen TE, Plesner T. Subcellular distribution of urokinase and urokinase receptor in human neutrophils determined by immunoelectron microscopy. Ultrastruct Pathol 2000; 24: 175-82.
4 Herren T, Burke TA. Regulation of plasminogen binding to neutrophils. Blood 2001; 97: 1070-8.
5 Miles LA, Dahlberg CM. Role of cell-surface lysines in plasminogen binding to cells: identification of α-enolase as a candidate plasminogen receptor. Biochemistry 1991; 30: 1682-91.
6 McLean JW, Tomlinson JE. cDNA sequence of human apolipoprotein(a) is homologous to plasminogen. Nature 1987; 300: 132-7.
7 Danesh J, Collins R. Lipoprotein(a) and coronary heart disease. Meta-analysis of prospective studies. Circulation 2000; 102: 1082-5.
8 Kang C, Durlach V. Lipoprotein(a) isoforms display differences in affinity for plasminogen binding to human mononuclear cells. Arterioscler Thromb Vasc Biol 1997; 17: 2036-43.
9 Miles LA, Plow EF. Plasminogen receptors: ubiquitous sites for cellular regulation of fibrinolysis. Fibrinolysis 1988; 02: 61-71.
10 Hall SW, Humphries JE. Inhibition of cell surface receptor-bound plasmin by alpha2-antiplasmin and alpha2-macroglobulin. J Biol Chem 1991; 266: 12329-36.
11 Eberhard T, Kronvall G. Surface bound plasmin promotes migration of Streptococcus pneumoniae through reconstituted basement membranes. Microb Pathol 1999; 26: 175-81.
12 Salonen EV, Saksela O. Plasminogen and tissue-type plasminogen activator bind to immobilized fibronectin. J Biol Chem 1985; 260: 12302-7.
13 Lijnen HR. Plasmin and matrix metalloproteinases in vascular remodeling. Thromb Haemost 2001; 86: 324-33.
14 Annes JP, Munger JS. Making sense of latent TGFb activation. J Cell Sci 2003; 116: 217-24.
15 Moir E, Booth NA. Polymorphonuclear leucocytes mediate endogenous thrombus lysis via a u-PA-dependent mechanism. Brit J Haem 2001; 113: 72-80.
16 Scanu AM, Edelstein C. Learning about the structure and biology of human lipoprotein through dissection by enzymes of the elastase family: facts and speculations. J Lipid Res 1997; 38: 2193-206.
17 Edelstein C, Italia JA. Polymorphonuclear cells isolated from human peripheral blood cleave lipoprotein(a) and apolipoprotein(a) at multiple interkringle sites via the enzyme elastase. J Biol Chem 1997; 272: 11079-87.
18 Scapini P, Nesi L. Generation of biologically active angiostatin kringle 1-3 by activated human neutrophils. J Immunol 2002; 168: 5798-804.
19 Schmitt M, Kanayama N. Elastase released from human granulocytes stimulated with Nformyl-chemotactic peptide prevents activation of tumor cell prourokinase (pro-uPA). FEBS Lett 1989; 255: 83-8.
20 Moir E, Robbie LA. Polymorphonuclear leucocytes have two opposing roles in fibrinolysis. Thomb Haemost 2001; 87: 1006-10.
21 Owen CA, Campbell MA. Cell surface-bound elastase and cathepsin G on human neutrophils: a novel, non-oxidative mechanism by which neutrophils focus and preserve catalytic activity of serine proteinases. J Cell Biol 1995; 131: 775-89.
22 Maeda S, Abe A. Transient changes of serum lipoprotein(a) as an acute phase protein. Atherosclerosis 1989; 78: 145-50.
23 Dominguez M, Rojas G. Kringles of the plasminogen-prothrombin gene family share conformational epitopes with recombinant apo-lipoprotein(a): specificity of the fibrinbinding site. Biochim Biophys Acta 2001; 1548: 72-80.
24 Kang C, Dominguez M. Lp(a) particles mold fibrin-binding properties of apo(a) in sizedependant manner. Arterioscler Thromb Vasc Biol 2002; 22: 1232-8.
25 Anglés-Cano E, Loyau S. A novel kringle-4 number based recombinant apo(a) standard for human apo(a) phenotyping. J Lipid Res 1999; 40: 354-9.
26 Fleury V, Anglés-Cano E. Characterization of the binding of plasminogen to fibrin surfaces: the role of carboxy-terminal lysines. Biochemistry 1991; 30: 7630-8.
27 Mtairag EM, Abdelghaffar H. Role of extracellular calcium in in vitro uptake and intraphagocytic location of macrolides. Antimicrob Agents Chemother 1995; 39: 1676-82.
28 Gaussem P, Grailhe P. Sodium dodecyl sulfate-induced dissociation of complexes between human tissue plasminogen activator and its specific inhibitor. J Biol Chem 1993; 268: 12150-5.
29 Fleury V, Loyau S. Molecular assembly of plasminogen and tissue-type plasminogen activator on an evolving fibrin surface. Eur J Biochem 1993; 216: 549-56.
30 Meilhac O, Ho-Tin-Noe B. Pericellular plasmin induces smooth muscle cell anoikis. FASEB J 2003; 17: 1301-3.
31 Haslett C, Guthrie LA. Modulation of multiple neutropihil functions by preparative methods or trace concentrations of bacterial lipopolysaccharide. Am J Pathol 1985; 119: 101-10.
32 Morrison DC, Jacobs DM. Binding of polymixin B to the lipid A portion of bacterial lipopolysaccharides. Immunochemistry 1976; 13: 813-8.
33 Bannatyne RM, Harnett NM. Inhibition of biological effects of endotoxin on neutrophils by polymixin B sulfate. J Infect Dis 1977; 136: 469-73.
34 Plesner T, Ploug M. The receptor for urokinase-type plasminogen activator and urokinase is translocated from two distinct intracellular compartments to the plasma membrane on stimulation of human neutrophils. Blood 1994; 83: 808-15.
35 Edelstein C, Italia JA. Functional and metabolic differences between elastase-generated fragments of human lipoprotein(a) and apolipoprotein(a). J Lipid Res 1996; 37: 1786-801.
36 Dollery CM, Owen CA. Neutrophil elastase in human atherosclerotic plaques: production by macrophages. Circulation 2003; 107: 2829-36.
37 Rabinovitch M. EVE and beyond, retro and perspective insights. Am J Physiol 1999; 277: L5-L12.
38 Plow EF, Herren T. The cell biology of the plasminogen system. FASEB J 1995; 09: 939-45.
39 Yurewicz EC, Zimmerman M. Cytochalasin B-dependent release of azurophil granule enzymes from human polymorphonuclear leukocytes. Inflammation 1977; 02: 259-64.
40 Hallet MB, Lloyds D. Neutrophil priming: the cellular signals that say ‘amber’ but not ‘green’. Immunol Today 1995; 16: 264-8.
41 Rubel C, Fernandez GC. Fibrinogen promotes neutrophil activation and delays apoptosis. J Immunol 2001; 166: 2002-10.
42 Condliffe AM, Kitchen E. Neutrophil priming: pathophysiiological consequences and underlying mechanisms. Clin Sc 1998; 94: 461-71.
43 Wiedow O, Luademann J. Elafin is a potent inhibitor of proteinase-3. Biochim Biophys Res Comm 1991; 174: 6-10.
44 Miles LA, Plow EF. Receptor mediated binding of the fibrinolytic components, plasminogen and urokinase, to peripheral blood cells. Thromb Haemost 1987; 58: 936-42.
45 Felez J, Miles LA. Characterization of cellular binding sites and interactive regions within reactants required for enhancement of plasminogen activation by tPA on the surface of leukocytic cells. Thromb Haemost 1996; 76: 577-84.
46 Sinniger V, Merton RE. Regulation of tissue plasminogen activator activity by cells. Domains responsible for binding and mechanism of stimulation. J Biol Chem 1999; 274: 12414-22.
47 Hoff HF, O’Neil J. Immunochemically detectable lipid-free apo(a) in plasma and in human atherosclerotic lesions. Chem Phys Lipids 1994; 67-68: 271-80.
48 Miles LA, Dahlberg CM. The cell-binding domains of plasminogen and their function in plasma. J Biol Chem 1988; 263: 11928-34.