Download PDF
Thromb Haemost 1991; 65(01): 096-101
DOI: 10.1055/s-0038-1647461
Original Article
Schattauer GmbH Stuttgart

Binding of Platelet Agglutinating Protein p 37 from the Plasma of a Patient with Thrombotic Thrombocytopenic Purpura to Human Platelets

Eric C-Y Lian
The Hemophilia and Thrombosis Center, W. J. Harrington Center for Blood Diseases, University of Miami School of Medicine and the Research and Medical Services, Veterans Administration Medical Center, Miami, Florida, USA
,
Farooq A Siddiqui
The Hemophilia and Thrombosis Center, W. J. Harrington Center for Blood Diseases, University of Miami School of Medicine and the Research and Medical Services, Veterans Administration Medical Center, Miami, Florida, USA
› Author Affiliations
Further Information

Publication History

Received 29 August 1989

Accepted after revision 08 August 1990

Publication Date:
02 July 2018 (online)

  PDF Download  Permissions and Reprints

Summary

We have previously reported the purification of a 37 kDa platelet agglutinating protein (PAP p37) from the plasma of a patient with thrombotic thrombocytopenic purpura (TTP) that was shown to be present in a subset of TTP patients. To gain further insight into the interaction between PAP p37 and platelets, we have studied the properties of PAP p37 binding to platelets. Washed human platelets from two normal donors and two TTP patients after recovery were used for the experiments. The PAP p 37 binding to platelets was specific, concentration dependent and saturable. Scatchard analysis demonstrated about 20,564–27,090 PAP p37 binding sites per platelet. Stimulation of platelets with thrombin or ADP did not have any significant effect on its binding.

Thiol- and serine-specific protease inhibitors did not inhibit PAP p37 binding to the platelets. Sugars such as glucose, fructose, mannose, and sialic acid, at 40 mM, inhibited its binding to platelets by 44%, 73%, 79%, and 91% respectively, but galactose and amino sugars did not have any significant effect. At 250 μg/ml, Concanavalin-A inhibited 42% of binding, but other lectins, such as phytohemagglutinin-P, potato lectin and helix pomatia lectin (snail), did not. Pre-incubation of 125I-PAP p37 with the adult human IgG, decreased its binding to the platelets. The monoclonal antibodies to GP Ib (6D1) and GP IIb-IIIa complex (10E5) did not inhibit the binding of 125I-PAP p37 to platelets. Fibrinogen and von Willebrand factor did not affect the binding either. These results suggest that PAP p37 binds to platelets on the sites other than GP lb or GP IIb-IIIa complex.

 

  • References

  • 1 Moschcowitz E. An acute febrile pleiochromic anemia with hyaline thrombosis of the terminal arterioles and capillaries: An undescribed disease. Arch Int Med 1925; 36: 89-93
    CrossrefPubMedGoogle Scholar
  • 2 Amorosi EL, Ultman JE. Thrombotic thrombocytopenic purpura: Report of 16 cases and review of literature. Medicine (Baltimore) 1966; 45: 139-159
    CrossrefPubMedGoogle Scholar
  • 3 Kwaan HC. The pathogenesis of thrombotic thrombocytopenic purpura. Semin Thromb Hemostas 1979; 3: 184-198
    Article in Thieme ConnectPubMedGoogle Scholar
  • 4 Myers TJ, Wakem CJ, Ball ED, Tremont SJ. Thrombotic thrombocytopenic purpura: Combined treatment with plasmapheresis and anti-platelet agents. Ann Intern Med 1980; 92: 149-155
    CrossrefPubMedGoogle Scholar
  • 5 Lian EC-Y, Harkness DR, Byrnes JJ, Wallach H, Nunez R. The presence of a platelet aggregating factor in the plasma of patients with thrombotic thrombocytopenic purpura and its inhibition by normal plasma. Blood 1979; 53: 333-338
    CrossrefPubMedGoogle Scholar
  • 6 Ansell J, Slepchuck NI, Pechet L. Thrombotic thrombocytopenic purpura. Blood 1979; 54: 959-960
    PubMedGoogle Scholar
  • 7 Brandt JT, Kennedy MS, Senhauser DA. Platelet aggregating factor in thrombotic thrombocytopenic purpura. Lancet 1979; 2: 463-464
    PubMedGoogle Scholar
  • 8 Burnes ER, Zucker-Franklin D. Pathologic effects of plasma from patients with thrombotic thrombocytopenic purpura on platelets and cultured vascular endothelial cells. Blood 1982; 60: 1030-1037
    PubMedGoogle Scholar
  • 9 Kelton JG, Moore J, Santos A, Sheridan D. Detection of a platelet-agglutinating factor in thrombotic thrombocytopenic purpura. Ann Intern Med 1984; 101: 589-593
    CrossrefPubMedGoogle Scholar
  • 10 Lian EC-Y, Savaraj N. Effects of platelet inhibitors on the platelet aggregation induced by plasma from patients with thrombotic thrombocytopenic purpura. Blood 1981; 58: 354-359
    PubMedGoogle Scholar
  • 11 Lian EC-Y, Mui P, Siddiqui FA, Chiu AY, Chiu LL. Inhibition of platelet aggregating activity in thrombotic thrombocytopenic purpura plasma by normal adult immunoglobulin G. J Clin Invest 1984; 73: 548-555
    CrossrefPubMedGoogle Scholar
  • 12 Siddiqui FA, Lian EC-Y. Novel platelet-agglutinating protein from a thrombotic thrombocytopenic purpura plasma. J Clin Invest 1985; 76: 1330-1337
    CrossrefPubMedGoogle Scholar
  • 13 Fowler WE, Fretto LJ, Hamilton KK, Erickson HP, McKee PA. Substructure of human von Willebrand factor. J Clin Invest 1985; 76: 1491-1500
    CrossrefPubMedGoogle Scholar
  • 14 Walsh PN, Mills D CB, White JG. Metabolism and function of human platelets washed by albumin density gradient separation. Br J Haematol 1977; 36: 287-296
    PubMedGoogle Scholar
  • 15 Sinha D, Seaman FS, Koshy A, Knight LC, Walsh PN. Blood coagulation factor XIa binds specifically to a site on activated human platelets distinct from that for factor XI. J Clin Invest 1984; 73: 1550-1556
    CrossrefPubMedGoogle Scholar
  • 16 Bradford 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
  • 17 Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature (London) 1970; 227: 680-685
    CrossrefPubMedGoogle Scholar
  • 18 Lian EC-Y. Pathogenesis of thrombotic thrombocytopenic purpura. Semin Hematol 1987; 24: 82-100
    PubMedGoogle Scholar
  • 19 Siddiqui FA, Lian EC-Y. Platelet-agglutinating protein p37 from a thrombotic thrombocytopenic purpura plasma forms a complex with human immunoglobulin G. Blood 1988; 71: 299-304
    PubMedGoogle Scholar
  • 20 Phillips DR, Jakabova M. Calcium dependent protease in human platelets. Specific cleavage of platelet polypeptides in the presence of added Ca++ . J Biol Chem 1977; 252: 5602-5605
    PubMedGoogle Scholar
  • 21 White GC. Calcium-dependent proteins in platelets: Response of calcium-activated protease in normal and thrombasthenic platelets to aggregating agents. Biochim Biophys Acta 1980; 631: 130-138
    CrossrefPubMedGoogle Scholar
  • 22 Truglia JA, Stracher A. Purification and characterization of a calcium dependent sulfhydryl protease from human platelets. Biochem Biophys Res Commun 1981; 100: 814-822
    CrossrefPubMedGoogle Scholar
  • 23 Yoshida N, Weskler B, Nachman R. Purification of human platelet calcium-activated protease. Effect on platelet and endothelial function. J Biol Chem 1983; 258: 7168-7174
    PubMedGoogle Scholar
  • 24 Fox JE, Reynolds CC, Phillips DR. Calcium-dependent proteolysis occurs during platelet aggregation. J Biol Chem 1983; 258: 9973-9981
    PubMedGoogle Scholar
  • 25 Vassalli P, Simon G, Rouiller C. Ultrastructural study of platelet changes initiated in vivo by thrombin. J Ultrastruct Res 1964; 11: 374-387
    CrossrefPubMedGoogle Scholar
  • 26 Packham MA, Kinglough-Rathbone RL, Reimers HJ. et al Mechanisms of platelet aggregation independent of adenosine diphosphate. In: Prostaglandins in Hematology Silver MJ. et al. (eds) Spectrum; New York: 1977: 247-276
  • 27 Nachman RL, Leung LL. Complex formation of platelet membrane glycoproteins lib and Ilia with fibrinogen. J Clin Invest 1982; 69: 263-269
    CrossrefPubMedGoogle Scholar
  • 28 Kinglough-Rathbone RL, Packham MA, Mustard JF. Effect of amino sugars on platelet aggregation and on fibrinogen binding. Thromb Haemostas 1984; 52: 75-80
    PubMedGoogle Scholar
  • 29 Scatchard G. The attractions of proteins for small molecules and ions. Ann NY Acid Sci 1949; 51: 660-672
    PubMedGoogle Scholar
  • 30 Lian EC-Y, Siddiqui FA, Jamieson GA, Tandon NN. Platelet agglutinating protein p37 causes platelet agglutination through its binding to membrane glycoprotein IV. Thromb Haemostas 1991; 65: 102-106
    PubMedGoogle Scholar