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Semin Thromb Hemost 2001; 27(5): 495-502
DOI: 10.1055/s-2001-17960
Copyright © 2001 by Thieme Medical Publishers, Inc., 333 Seventh Avenue, New York, NY 10001, USA. Tel.: +1(212) 584-4662

Effects of a Heparin-Binding Protein on Blood Coagulation and Platelet Function

Phoebe Kaiser1 , Job  Harenberg1 , Jeanine M. Walenga2 , Günther Huhle1 , Christina Giese1 , Margaret  Prechel2 , Debra Hoppensteadt2 , Jawed Fareed2
  • 1First Department of Medicine, University Hospital, Mannheim, Germany
  • 2Departments of Pathology and Pharmacology, Cardiovascular Institute, Loyola University Medical Center, Maywood, Illinois
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Publication History

Publication Date:
22 October 2001 (online)

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ABSTRACT

The objective of this study was to characterize the heparin-binding properties of a protein secreted by mouse myeloma cells. The characterization was performed using clinical assays, such as heparin activity assays and heparin-induced thrombocytopenia (HIT) platelet activation assays. The tests were performed in the presence of heparin, low-molecular-weight heparins (LMWH), or heparinoids and either heparin-binding protein (HBP) or saline to determine whether the HBP affects the activity of heparins. The characterization of the HBP using heparin activity assays showed that the HBP shortened the prolonged clotting times of the activated partial thromboplastin time (aPTT) and thrombin clotting time induced by high concentrations of unfractionated heparin. The chromogenic assays for antithrombin (AT), thrombin inhibition, and factor Xa inhibition demonstrated that this effect is related to heparin concentrations below 0.5 IU/ml. The Heptest assay did not detect these differences. The HBP did not modify the anticoagulant effect of any LMWH or low- or high-sulfated glycosaminoglycans in the aPTT assay. Activation of donor platelets in the presence of unfractionated heparin, platelet factor 4 (PF4), and HIT-serum was not counteracted by the HBP in any of the assays. The characterization of the HBP using a PF4-enzyme-linked immunosorbent assay (ELISA) confirmed the lack of structural identity with PF4. However, the optical density data indicated that the protein structure may be similar to PF4 by binding to a PF4 antibody. These data suggest that the HBP isolated from mouse myeloma cells has a low affinity to heparin and interacts with the secondary binding site to AT and also perhaps to PF4.

KEYWORD

Heparin-binding protein - heparin - low-molecular-weight heparin (LMWH) - heparin-induced thrombocytopenia (HIT) - platelet factor 4

REFERENCES

  • 1 Kure S, Yoshie O. A syngeneic monoclonal antibody to murine meth-A sarcoma (HepSS-l) recognizes heparan sulfate glycosaminoglycan (HS-GAG): cell density and transformation dependent alteration in cell surface HS-GAG defined by HepSS-1.  J Immunol . 1986;  137 3900-3908
    PubMedGoogle Scholar
  • 2 van-den Born J, van-den Heuvel P L, Bakker M A. A monoclonal antibody against GBM heparan sulfate induces acute selective proteinuria in rats.  Kidney Int . 1992;  41 115-123
    PubMedGoogle Scholar
  • 3 Yagamata M, Kimata K, Oike Y. A monoclonal antibody that specifically recognizes a glucuronic acid 2-sulfate-containing determinant in intact chondroitin sulfate chain.  J Biol Chem . 1987;  262 4146-4152
    PubMedGoogle Scholar
  • 4 Caterson B, Christner J E, Baker J R. Identification of a monoclonal antibody that specifically recognizes corneal and skeletal keratan sulfate.  J Biol Chem . 1983;  258 8848-8854
    PubMedGoogle Scholar
  • 5 Mehmet H, Scudder P, Tang P W. The antigenic determinants recognized by three monoclonal antibodies to keratan sulphate involve sulphated hepta- or larger oligosaccharides of the poly(N-acetyllactosamine) series.  Eur J Biochem . 1986;  157 385-391
    CrossrefPubMedGoogle Scholar
  • 6 Pejler G, Lindahl U, Larm O. Monoclonal antibodies specific for oligosaccharides prepared by partial nitrous acid deamination of heparin.  J Biol Chem . 1988;  263 5197-5201
    PubMedGoogle Scholar
  • 7 Gitel S N, Medina V M, Wessler S. Preparation and identification of a population of antibodies that recognize carbodiimide-modified heparin.  Blood . 1985;  655 902-911
    PubMedGoogle Scholar
  • 8 Strauss A H, Travassos L R, Takahashi H K. A monoclonal antibody (ST-I) directed to the native heparin chain.  Anal Biochem . 1992;  201 1-8
    CrossrefPubMedGoogle Scholar
  • 9 Huhle G, Harenberg J, Malsch R, Heene D L. Comparison of three heparin bovine-serum-albumin binding-methods for production of anti-heparin antibodies.  Semin Thromb Hemost . 1994;  20 193-204
    PubMedGoogle Scholar
  • 10 Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.  Anal Biochem . 1976;  72 248-254
    CrossrefPubMedGoogle Scholar
  • 11 Lämmli U K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4.  Nature . 1970;  227 680-685
    PubMedGoogle Scholar
  • 12 Switzer R C, Merrill C R, Shifrin S. A highly sensitive silver stain for detecting proteins and peptides in polyacrylamid gels.  Anal Biochem . 1979;  98 231-237
    CrossrefPubMedGoogle Scholar
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