Thromb Haemost 1988; 59(01): 054-061
DOI: 10.1055/s-0038-1642565
Review Article
Schattauer GmbH Stuttgart

Identity of Saturable Calcium-Binding Sites on Blood Platelets and Their Involvement in Platelet Aggregation

Geoffrey I Johnston
The Department of Medicine, University Hospital, Queen’s Medical Centre, Nottingham, UK
*   Current address:, Dept. of Medicine/Cardiovascular Blology Research Program, Oklahoma University Health Science Center-line 825 Northeast 13th Street Oklahoma City, Oklahoma 73104, USA
,
Stanley Heptinstall
The Department of Medicine, University Hospital, Queen’s Medical Centre, Nottingham, UK
› Institutsangaben
Weitere Informationen

Publikationsverlauf

Received 07. April 1987

Accepted after revision 02. Oktober 1987

Publikationsdatum:
18. April 2018 (online)

Summary

Extracellular Ca2+ ions are required for platelet aggregation and we show that they enter two platelet pools. One pool is rapidly filled and easily displaced by EGTA. The second is filled more slowly and is not displaced by EGTA. The EGTA-displaceable pool is believed to be surface-located and was found to contain at least one class of saturable binding sites as well as a class of non-saturable binding sites. The saturable sites were found to be highly selective for Ca2+ (dissociation constant, 3.5 × IO−7 M) even in the presence of 1 mM Mg2+ ions, and they took up between 261,000 and 307,000 Ca2+ ions/platelet. Full occupancy of the saturable binding sites appeared to be necessary for platelet aggregation to proceed. We also studied platelets that were unable to aggregate normally, either due to the congenital bleeding disorder Glanzmann’s thrombastenia or due to experimental manipulation. In both cases wc found decreased Ca2+uptake specifically by the saturable Ca2+ binding sites, and that this was associated with decreased number of GP IIb/IIIa molecules expressed on these platelets. We suggest that the Ca2+binding sites involved in platelet aggregation are located on the GP Ilb/IIIa complexes and may be involved in holding the glycoproteins in the complex together, and that the binding sites need to be fully occupied before aggregation can proceed.

 
  • References

  • 1 Born GV R, Cross MJ. Effects of inorganic ions and of plasma proteins on the aggregation of blood platelets by ADR. J Physiol 1964; 70: 397-414
  • 2 Hovig T. The effect of calcium and magnesium on rabbit blood platelet aggregation in vitro. Thrombos Diathes Hemorrh 1964; 12: 179-200
  • 3 Mitchell JR A, Sharp AA. Platelet clumping in vitro. Br J Haematol 1964; 10: 78-93
  • 4 Heptinstall S. The use of a chelating ion-exchange resin to evaluate the effects of the extracellular calcium concentration on adenosine diphosphate induced aggregation of human blood platelets. Thromb Haemostas 1976; 36: 208-220
  • 5 Robblee LS, Shepro D. The effect of external calcium and lanthenum on platelet calcium content and on the release reaction. Biochim Biophys Acta 1976; 436: 448-459
  • 6 Peerschke EI, Grant RA, Zucker MB. Decreased association of 45calcium with platelets unable to aggregate due to thrombasthenia or prolonged calcium deprivation. Br J Haematol 1980; 46: 247-256
  • 7 Brass LF, Shattil SJ. Changes in surface-bound and exchangeable calcium during platelet activation. J Biol Chem 1982; 257: 14000-14005
  • 8 Kunicki TJ, Pidard D, Rosa JR, Nurden AT. The formation of Ca++-dependant complexes of platelet membrane glycoproteins IIb and Ilia in solution as determined by crossed immunoelectrophoresis.. Blood 1981 58: 268-278
  • 9 George JN, Nurden AT, Phillips DR. Molecular defects in interactions of platelets with the vessel wall. N Engl I Med 1984; 311: 1084-1098
  • 10 Brass LF, Shattil SJ. Identification and function of the high affinity binding sites for Ca2+ on the surface of platelets. J Clin Invest 1984; 73: 626-632
  • 11 Peerschke EI. pH and magnesium after 45calcium binding to platelets at sites other than glycoprotein I or Ilb/IIIa. Proc Soc Exp Biol Med 1985; 179: 232-239
  • 12 Zucker MB, Grant RA. Nonreversible loss of platelet aggregability induced by calcium deprivation. Blood 1978; 52: 505-514
  • 13 Taylor PM, Heptinstall S. The abilities of human blood platelets to bind extracellular calcium and to be aggregated by adenosine diphosphate are related. Br J Haematol 1980; 46: 115-122
  • 14 Fitzgerald LA, Phillips DR. Calcium regulation of the platelet membrane glycoprotein Ilb-IIIa complex. J Biol Chem 1985; 260: 11366-11374
  • 15 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
  • 16 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
  • 17 Tangen O, Berman HJ. Gel filtration of blood platelets: a methodological report. Adv Exp Med Biol 1972; 34: 235-243
  • 18 Burgess-Wilson ME, Cockbill SR, Johnston GI, Heptinstall S. Platelet aggregation in whole blood patients with Glanzmann’s thrombasthenia. Blood 1987; 69: 38-42
  • 19 Portzehl H, Caldwell PC, Ruegg JC. The dependence of contraction and relaxation of muscle fibres from the crab MAIA SQUINADO on the internal concentration of free calcium ions. Biochim Biophys Acta 1964; 79: 581-591
  • 20 Scatchard G. The attraction of protein for small molecules and ions. Ann NY Acad Sci 1949; 51: 660-672
  • 21 Adams J, Heptinstall S, Mitchell JR A. A six-channel automated platelet aggregometer. Thrombos Diathes Haemorrh 1975; 34: 821-824
  • 22 Johnston GI, Heptinstall S, Robins RA, Price MR. The expression of glycoproteins on single blood platelets from healthy individuals and from patients with congenital bleeding disorders. Biochem Biophys Res Comm 1984; 123: 1091-1098
  • 23 Jones D, Fritschy J, Garson J, Nokes TJ C, Kemshead JT, Hardisty RM. A monoclonal antibody binding to human medulloblastoma cells and to the platelet glycoprotein IIb-IIIa complex. Br J Haematol 1984; 57: 621-631
  • 24 Johnston GI, Heptinstall S. Examination of glycoproteins on intact blood platelets by using monoclonal antibodies and the fluorescence activated cell sorter. Biochem Soc Trans 1985; 13: 109 (Abstr.)
  • 25 Steiner M, Tateishi T. Distribution and transport of calcium in human platelets. Biochim Biophys Acta 1974; 367: 232-246
  • 26 Brass LF. Ca2+ homeostasis in unstimulated platelets. J Biol Chem 1985; 259: 12563-12570
  • 27 Brass LF, Shattil SJ. Identification of the saturable binding sites for Ca2+ on the surface of human platelets. Thromb Haemostas 1983; 50: 327 (Abstr.)
  • 28 Phillips DR, Baughan AK. Fibrinogen binding to human platelet plasma membranes. Identification of two steps requiring divalent cations. J Biol Chem 1983; 258: 10240-10246