PDF herunterladen
Thromb Haemost 1996; 76(06): 1038-1046
DOI: 10.1055/s-0038-1650704
Original Article
Schattauer GmbH Stuttgart

A Novel Regulatory Epitope Defined by a Murine Monoclonal Antibody to the Platelet GPIIb-IIIa Complex (αIIbβ3 Integrin)

Michihide Tokuhira
1   The Department of Internal Medicine, School of Medicine, Keio University, Tokyo, Japan
,
Makoto Handa
2   The Department of Blood Center, School of Medicine, Keio University, Tokyo, Japan
,
Tetsuji Kamata
1   The Department of Internal Medicine, School of Medicine, Keio University, Tokyo, Japan
,
Atsushi Oda
1   The Department of Internal Medicine, School of Medicine, Keio University, Tokyo, Japan
,
Masahiko Katayama
3   The Department of Biotechnology Research Laboratories, Takara Shuzo Co., Ltd., Otsu, Japan
,
Yoshiaki Tomiyama
4   The Second Department of Internal Medicine, Osaka University Medical School, Osaka, Japan
,
Mitsuru Murata
1   The Department of Internal Medicine, School of Medicine, Keio University, Tokyo, Japan
,
Yohko Kawai
5   The Department of Laboratory Medicine, School of Medicine, Keio University, Tokyo, Japan
,
Kiyoaki Watanabe
5   The Department of Laboratory Medicine, School of Medicine, Keio University, Tokyo, Japan
,
Yasuo Ikeda
1   The Department of Internal Medicine, School of Medicine, Keio University, Tokyo, Japan
› Institutsangaben
Weitere Informationen

Publikationsverlauf

Received 18. März 1996

Accepted after resubmission 05. August 1996

Publikationsdatum:
11. Juli 2018 (online)

Auch verfügbar auf
  als PDF herunterladen  Lizenzen und Reprints

Summary

We characterized a murine monoclonal antibody, PT25-2 (IgG1), raised against washed human platelets. The antibody and its Fab fragments were both capable of inducing platelet aggregation in a fibrinogen-dependent manner and induced 125I-fibrinogen binding to unstimulated platelets (120,000 molecules/platelet at a 100 nM IgG concentration). The antibody immunoprecipitated the αIIbβ3 complex from lysates of iodinated platelets but did not react with the respective subunits when complex formation was disrupted by treatment with 5 mM EDTA at 37°C for 30 min. However, simply removing the extracellular divalent cation with EDTA had no effect on antibody binding indicating that the antibody’s epitope depends upon a conformational structure maintained by αβ subunit association. Antibody binding to unstimulated, washed platelets yielded binding parameters (Kd = 40 nM, Bmax = 100,000 molecules/platelet), which were found to be virtually unchanged when binding was performed using thrombin or RGDS-peptide-stimulated platelets. Thus, the PT25-2 antibody defines a novel regulatory epitope expressed by the αIIbβ3 integrin on unstimulated, quiescent platelets.

 

  • References

  • 1 Ginsberg MH, Du X, O’Toole TE, Loftus JC, Plow EF. Platelet integrins. Thromb Haemost 1993; 70: 87-93
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 2 Ruggeri ZM. von Willebrand factor and fibrinogen. Curr Opin Cell Biol 1993; 5: 898-906
    CrossrefPubMedGoogle Scholar
  • 3 Phillips DR, Charo IF, Parise LV, Fitzgerald LA. The platelet membrane glycoprotein IIb-IIIa complex. Blood 1988; 71: 831-843
    PubMedGoogle Scholar
  • 4 Shattil SJ, Hoxie JA, Cunningham M, Brass LF. Changes in the platelet membrane glycoprotein IIb-IIIa complex during platelet activation. J Biol Chem 1985; 260: 11107-11114
    PubMedGoogle Scholar
  • 5 Hynes R. Integrins; Versatility, modulation, and signaling in cell adhesion. Cell 1992; 69: 11-25
    CrossrefPubMedGoogle Scholar
  • 6 Ginsberg MH, Du X, Plow EF. Inside-out integrin signaling. Curr Opin Cell Biol 1992; 4: 766-771
    CrossrefPubMedGoogle Scholar
  • 7 Cohen I, DeVries A. Platelet contractile regulation in an isometric system. Nature 1973; 246: 36-37
    CrossrefPubMedGoogle Scholar
  • 8 Phillips DR, Jennings LK, Edwards HH. Identification of membrane proteins mediating the interaction of human platelets. J Cell Biol 1980; 86: 77-86
    CrossrefPubMedGoogle Scholar
  • 9 Calvete JJ, Mann K, Schafer W, Femandez-Lafuente R, Guisan JM. Proteolytic degradation of the RGD-binding and non-RGD-binding conformers of human platelet integrin glycoprotein IIb/IIIa; clues for identification of regions involved in the receptor’'s activation. Biochem J 1994; 298: 1-7
    CrossrefPubMedGoogle Scholar
  • 10 Chen YP, O’Toole TE, Ylanne J, Rosa JP, Ginsberg MH. A point mutation in the integrin β3 cytoplasmic domain (S752-->P) impairs bidirectional signaling through αIIbβ3 (platelet glycoprotein IIb-IIIa). Blood 1994; 841: 857-865
    PubMedGoogle Scholar
  • 11 Vassallo Jr R, Kieber-Emmons T, Cichowski K, Brass LF. Structure-function relationships in the activation of platelet thrombin receptors by receptor-derived peptides. J Biol Chem 1992; 267: 6081-6085
    PubMedGoogle Scholar
  • 12 Huang MM, Indik Z, Brass LF, Hoxie JA, Schreiber AD, Brugge JS. Activation of FcyγII induces tyrosine phosphorylation of multiple proteins including FcγRII. J Biol Chem 1992; 267: 5467-5473
    PubMedGoogle Scholar
  • 13 Tomiyama Y, Kunicki TJ, Zipf TF, Ford SB, Aster RH. Response of human platelets to activating monoclonal antibodies; Importance of FcγRII (CD32) phenotype and level of expression. Blood 1992; 80: 2261-2268
    PubMedGoogle Scholar
  • 14 Ikeda Y, Handa M, Kamata T, Kawano K, Kawai Y, Watanabe K, Kawakami K, Sakai K, Fukuyama M, Itagaki I, Yoshioka A, Ruggeri ZM. Transmembrane calcium influx associated with von Willebrand factor binding to GPIb in the initiation of shear-induced platelet aggregation. Thromb Haemost 1993; 69: 496-502
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 15 Gulino D, Ryckewaert JJ, Andrieuxx A, Rabiet MJ, Marguerie G. Identification of a monoclonal antibody against platelet GPIIb that interacts with a calcium-binding site and induces aggregation. J Biol Chem 1990; 265: 9575-9581
    PubMedGoogle Scholar
  • 16 Kouns WC, Wall CD, White MM, Fox CF, Jennings LK. A conformation-dependent epitope of human platelet glycoprotein IIIa. J Biol Chem 1990; 265: 20594-20601
    PubMedGoogle Scholar
  • 17 O’Toole TE, Loftus JC, Du X, Glass AA, Ruggeri ZM, Shattil SJ, Plow EF, Ginsberg MH. Affinity modulation of the αIIbβ3 integrin (platelet GPIIb-IIIa) is an intrinsic property of the receptor. Cell Regul 1990; 1: 883-893
    CrossrefPubMedGoogle Scholar
  • 18 Frelinger ALIII, Cohen I, Plow EF, Smith MA, Roberts J, Lam SC, Ginsberg MH. Selective inhibition of integrin function by antibodies specific for ligand-occupied receptor conformers. J Biol Chem 1990; 265: 6346-6352
    PubMedGoogle Scholar
  • 19 Honda S, Tomiyama Y, Pelletier A, Annis D, Honda Y, Orchekowski R, Ruggeri ZM, Kunicki TJ. Topography of ligand-induced binding sites, including a novel cation-sensitive epitope (AP5) at the amino terminus, of the human integrin β3 Subunit. J Biol Chem 1995; 20: 11947-11954
    PubMedGoogle Scholar
  • 20 Du X, Plow EF, Frelinger ALIII, O’Toole TE, Loftus JC, Ginsberg MH. Ligands “activate” integrin αIIbβ3 (platelet GPIIb-IIIa). Cell 1991; 65: 409-416
    CrossrefPubMedGoogle Scholar
  • 21 Niewiarowski S, Kornecki E, Budzynski AZ, Morinelli TA, Tuszynski GP. Fibrinogen interaction with platelet receptors. Ann N Y Acad Sci 1983; 408: 536-555
    CrossrefPubMedGoogle Scholar
  • 22 Pidard D, Frelinger ALIII, Bouillot C, Nurden AT. Activation of the fibrinogen receptor on human platelets exposed to alpha chymotrypsin. Relationship with a major proteolytic cleavage at the carboxyterminus of the membrane glycoprotein IIb heavy chain. Eur J Biochem 1991; 200: 437-447
    CrossrefPubMedGoogle Scholar
  • 23 Zucker MB, Masiello NC. Platelet aggregation caused by dithiothreitol. Thromb Haemost 1984; 51: 119-124
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 24 Kouns WC, Steiner B, Kunicki TJ, Moog S, Jutzi J, Jennings LK, Cazenave JP, Lanza F. Activation of the fibrinogen binding site on platelets isolated from a patient with the Strasbourg I variant of Glanzmann’s thrombasthenia. Blood 1994; 84: 1108-1115
    PubMedGoogle Scholar
  • 25 Du X, Gu M, Weisel JW, Nagaswami C, Bennett JS, Bowditch R, Ginsberg MH. Long range propagation of conformational changes in integrin αIIbβ3. J Biol Chem 1993; 268: 23087-23092
    PubMedGoogle Scholar
  • 26 Frelinger ALIII, Du X, Plow EF, Ginsberg MH. Monoclonal antibodies to ligand-occupied conformers of integrin αIIbβ3 (glycoprotein IIb-IIIa) alter receptor affinity, specificity, and function. J Biol Chem 1991; 266: 17106-17111
    PubMedGoogle Scholar
  • 27 Walsh PN, Biggs R, Gagnatelli G. Platelet antiheparin activity. Assay based on factor-Xa inactivation by heparin and antifactor Xa. Br J Haematol 1974; 26: 405-419
    PubMedGoogle Scholar
  • 28 Handa M, Titani K, Holland LZ, Roberts JR, Ruggeri ZM. The von Willebrand factor-binding domain of platelet membrane glycoprotein lb. Characterization by monoclonal antibodies and partial amino acid sequence analysis of proteolytic fragments. J Biol Chem 1986; 261: 12579-12585
    PubMedGoogle Scholar
  • 29 Newman PJ, Allen RW, Kahn RA, Kunicki TJ. Quantitation of membrane glycoprotein IIIa on intact human platelets using the monoclonal antibody, AP-3. Blood 1985; 65: 227-232
    PubMedGoogle Scholar
  • 30 Lombardo VT, Hodson E, Roberts JR, Kunicki TJ, Zimmerman TS, Ruggeri ZM. Independent modulation of von Willebrand factor and fibrinogen binding to the platelet membrane glycoprotein IIb/IIIa complex as demonstrated by monoclonal antibody. J Clin Invest 1985; 76: 1950-1958
    CrossrefPubMedGoogle Scholar
  • 31 Niiya K, Hodson E, Bader R, Byers-Ward V, Koziol JA, Plow EF, Ruggeri ZM. Increased surface expression of the membrane glycoprotein IIb/IIIa complex induced by platelet activation. Relationship to the binding of fibrinogen and platelet aggregation. Blood 1987; 70: 475-483
    PubMedGoogle Scholar
  • 32 Pidard D, Montgomery RR, Bennett JS, Kunicki TJ. Interaction of AP-2, a monoclonal antibody specific for the human platelet glycoprotein IIb-IIIa complex, with intact platelets. J Biol Chem 1983; 258: 12582-12586
    PubMedGoogle Scholar
  • 33 McEver RP, Baenzinger NL, Majerus PW. Isolation and quantitation of the platelet membrane glycoprotein deficient in thrombasthenia using a monoclonal hybridoma antibody. J Clin Invest 1980; 66: 1311-1318
    CrossrefPubMedGoogle Scholar
  • 34 Tomiyama Y, Tsubakio T, Piotrowicz RS, Kurata Y, Loftus JC, Kunicki TJ. The Arg-Gly-Asp (RGD) recognition site of platelet glycoprotein IIb-IIIa on nonactivated platelets is accessible to high-affmity macromolecules. Blood 1992; 79: 2303-2312
    PubMedGoogle Scholar
  • 35 Holmsen H, Holmsen I, Bemhardsen A. Microdetermination of adenosine diphosphate and adenosine triphosphate in plasma with firefly luciferase system. Anal Biochem 1966; 17: 456-473
    CrossrefPubMedGoogle Scholar
  • 36 Oda A, Daley JF, Kang J, Smith M, Ware JA, Salzman EW. Quasi-simulta-neous measurement of ionized calcium and a-granule release in individual platelets. Am J Physiol 1991; 260 (Cell Physiol, 29) 242-258
    CrossrefPubMedGoogle Scholar
  • 37 Grynkiewicz G, Poenie M, Tsien RY. A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem 1985; 260: 3440-3450
    PubMedGoogle Scholar
  • 38 Tomiyama Y, Take H, Ikeda H, Titani T, Furubayasi T, Mizutani H, Yamamoto N, Tandon NN, Sekiguchi S, Jamieson GA, Kurata Y, Yonezawa T, Tarui S. Identification of the platelet-specific alloantigen, NaKa, on platelet membrane glycoprotein IV. Blood 1990; 75: 684-687
    PubMedGoogle Scholar
  • 39 Kunicki TJ, Pidard D, Rosa JP, Nurden AT. The formation of Ca++-dependent complexes of platelet membrane glycoproteins IIb and IIIa in solution as determined by crossed immunoelectrophoresis. Blood 1981; 58: 268-278
    PubMedGoogle Scholar
  • 40 Brass LF, Shattil SJ, Kunicki TJ, Bennett JS. Effect of calcium on the stability of the platelet membrane glycoprotein IIb-IIIa complex. J Biol Chem 1985; 260: 7875-7881
    PubMedGoogle Scholar
  • 41 Fitzgerald LA, Phillips DR. Calcium regulation of the platelet membrane glycoprotein IIb-IIIa complex. J Biol Chem 1985; 260: 11366-11374
    PubMedGoogle Scholar
  • 42 Shattil SJ, Brass LF, Bennett JS, Pandhi P. Biochemical and functional consequences of dissociation of the platelet membrane glycoprotein IIb-IIIa complex. Blood 1985; 66: 92-98
    PubMedGoogle Scholar
  • 43 Pidard D, Didry D, Kunicki TJ, Nurden AT. Temperature-dependent effects of EDTA on the membrane glycoprotein IIb-IIIa complex and platelet aggregability. Blood 1986; 67: 604-611
    PubMedGoogle Scholar
  • 44 McEver RP, Bennett EM, Martin MN. Identification of two structurally and functionally distinct sites on human platelet membrane glycoprotein IIb-IIIa using monoclonal antibodies. J Biol Chem 1983; 258: 5269-5275
    PubMedGoogle Scholar
  • 45 Bennett JS, Hoxie JA, Leitman SF, Vilaire G, Cines DB. Inhibition of fibrinogen binding to stimulated human platelets by a monoclonal antibody. Proc Natl Acad Sci USA 1983; 80: 2417-2421
    CrossrefPubMedGoogle Scholar
  • 46 Coller BS. A new murine monoclonal antibody reports an activation-dependent change in the conformation and/or microenvironment of the platelet glycoprotein IIb/IIIa complex. J Clin Invest 1985; 76: 101-108
    CrossrefPubMedGoogle Scholar
  • 47 Tomiyama Y, Brojer E, Ruggeri ZM, Shattil SJ, Smiltneck J, Gorski J, Kumar A, Kieber-Emmons T, Kunicki TJ. A molecular model of RGD ligands. J Biol Chem 1992; 267: 18085-18092
    PubMedGoogle Scholar
  • 48 Kovach NL, Carlos TM, Yee E, Harlan JM. A monoclonal antibody to β1 integrin (CD29) stimulates VLA-dependent adherence of leukocytes to human umbilical vein endothelial cells and matrix components. J Cell Biol 1992; 116: 499-509
    CrossrefPubMedGoogle Scholar
  • 49 Arroyo AG, Sánchez-Mateos P, Campanero MR, Martín-Padura I, Dejana E, Sánchez-Madrid F. Regulation of the VLA integrin-ligand interactions through the β1 subunit. J Cell Biol 1992; 117: 659-670
    CrossrefPubMedGoogle Scholar
  • 50 Takada Y, Puzon W. Identification of a regulatory region of integrin β1 subunit using activating and inhibiting antibodies. J Biol Chem 1993; 268: 17597-17601
    PubMedGoogle Scholar
  • 51 van Kooyk Y, Weder P, Hogervorst F, Verhoeven Aj, van Seventer G, te Velde AA, Borst J, Keizer GD, Figdor CG. Activation of LFA-1 though a Ca2+-dependent epitope stimulates lymphocyte adhesion. J Cell Biol 1991; 112: 345-354
    CrossrefPubMedGoogle Scholar
  • 52 Andrew D, Shock A, Ball E, Ortlepp S, Bell J, Robinson M. KIM 185, a monoclonal antibody to CD 18 which induces a change in the conformation of CD 18 and promotes both LFA-1- and CR3-dependent adhesion. Eur J Immunol 1993; 23: 2217-2222
    CrossrefPubMedGoogle Scholar
  • 53 Bednarczyk JL, Szabo MC, Wygant JN, Lazarovits AI, McIntyre BW. Identification of a combinatorial epitope expressed by the integrin α4β1 heterodimer involved in the regulation of cell adhesion. J Bio Chem 1994; 269: 8348-8354
    PubMedGoogle Scholar
  • 54 Hato T, Watanabe A, Nakatani S, Minamoto Y, Fujita S. A novel mechanism for exposure of fibrinogen binding sites of GPIIb-IIIa by a monoclonal antibody. Thromb Haemost 1995; 73: 138-143
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 55 Yamamoto N, Kitagawa H, Yamamoto K, Tanoue K, Yamazaki H. Calcium ions and the conformation of glycoprotein IIIa that is essential for fibrinogen binding to platelets; Analysis by a new monoclonal anti-GPIIIa antibody, TM83. Blood 1989; 73: 1552-1560
    PubMedGoogle Scholar
  • 56 D’Souza SE, Haas TA, Piotrowicz RS, Byers-Ward V, McGrath DE, Soule HR, Ciemiewski C, Plow EF, Smith JW. Ligand and cation binding are dual functions of a discrete segment of the integrin β3 subunit: Cation displacement is involved in ligand binding. Cell 1994; 79: 659-667
    CrossrefPubMedGoogle Scholar
  • 57 Peerschke EL. Regulation of platelet aggregation by post-fibrinogen binding events. Thromb Haemost 1995; 73: 862-867
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 58 Huang MM, Lipfert L, Cunningham M, Brugge JS, Ginsberg MH, Shattil SJ. Adhesive ligand binding to integrin αIIbβ3 stimulates tyrosine phosphorylation of novel protein substrates before phosphorylation of ppl25FAK. J Cell Biol 1994; 122: 473-483
    PubMedGoogle Scholar
  • 59 Kouns WC, Jennings LK. Activation-independent exposure of the GPIIb-IIIa fibrinogen receptor. Thromb Res 1991; 63: 343-354
    CrossrefPubMedGoogle Scholar