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Immunocytochemical evidence for the translocation of α-granule membrane glycoprotein IIb/IIIa (integrin αIIbβ3) of human platelets to the surface membrane during the release reaction

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The localization of glycoprotein (GP) IIb/IIIa (integrin αIIbβ3) in both resting and thrombin-activated platelets was studied immunocytochemically. By the pre-embedding method where only the GP IIb/IIIa molecules on the surface of platelets were immunostained, the distribution of protein A-colloidal gold label was randomly distributed along the surface membrane of resting platelets at a density of 18.0±2.7 gold particles/μm of membrane. At 15 s after stimulation by 0.1 U/ml of thrombin in an unstirred platelet suspension, the spheroid-shaped platelets with pseudopodia still had normal numbers of α-granules, and the density of gold particles was 19.7±3.6 particles/μm. At 5 min, the α-granules were no longer present because of the release reaction, and the density of gold particles significantly increased (27.0±3.7 particles/μm; p<0.01). In immunostained ultra-thin frozen sections, the gold particles were detected not only on the surface membrane, including the open canalicular system (OCS), but also on the α-granule membranes of resting platelets. At 30 s after thrombin stimulation the α-granules fused with the OCS, resulting in the formation of a swollen OCS, which still had gold particles on its membrane. At 5 min, the gold particles were detected on the membrane of the swollen OCS located near the surface membrane, while very few gold particles were present on the membrane of the OCS in the central part of the platelets. These results demonstrate that α-granule membrane GPIIb/IIIa translocates to the surface membrane through the membrane of the OCS. Also the translocation of α-granule membrane GPIIb/IIIa gives rise to an actual increase in GPIIb/IIIa on the surface membrane during the release reaction induced by thrombin.

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  1. Beckerle MC, Miller DE, Bertagnolli ME, Locke SJ (1989) Activation-dependent redistribution of the adhesion plaque protein, talin, in intact human platelets. J Cell Biol 109:3333–3346

  2. Bennett JS, Vilaire G (1979) Exposure of platelet receptors by ADP and epinephrine. J Clin Invest 64:1393–1401

  3. Carlsson L, Markey F, Blikstad I, Persson T, Lindberg U (1979) Reorganization of actin in platelets stimulated by thrombin as measured by the DNase I inhibition assay. Proc Natl Acad Sci USA 76:6376–6380

  4. Coller BS (1985) 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 76:101–108

  5. Coller BS (1989) Activation-specific platelet antigens. In: Kunicki TJ, George JN (eds) Platelet immunobiology. Lippincott, Philadelphia, pp 166–189

  6. Cramer EM, Savidge GF, Vainchenker W, Berndt MC, Pidard D, Caen JP, Masse J-M, Breton-Gorius J (1990) Alpha-granule pool of glycoprotein IIb-IIIa in normal and pathologic platelets and megakaryocytes. Blood 75:1220–1227

  7. Fox JEB, Phillips DR (1983) Polymerization and organization of actin filaments within platelets. Semin Hematol 20:243–259

  8. Gerrard JM, Lint D, Sims PJ, Wiedmer T, Fugate RD, McMillan E, Robertson C, Israels SJ (1991) Identification of a platelet dense granule membrane protein that is deficient in a patient with the Hermansky-Pudlak syndrome. Blood 77:101–112

  9. Gogstad GO, Hagen I, Korsmo R, Solum NO (1981) Characterization of the proteins of isolated human platelet α-granules: evidence for a separate α-granule-pool of the glycoproteins IIb and IIIa. Biochim Biophys Acta 670:150–162

  10. Hartwig JH, Chambers KA, Stossel TP (1989) Association of gelsolin with actin filaments and cell membranes of macrophages and platelets. J Cell Biol 108:467–479

  11. Hynes RO (1987) Integrins: a family of cell surface receptors. Cell 48:549–554

  12. Isenberg WM, McEver RP, Phillips DR, Shuman MA, Bainton DF (1987) The platelet fibrinogen receptor: an immunogold-surface replica study of agonist-induced ligand binding and receptor clustering. J Cell Biol 104:1655–1663

  13. Jennings LK, Phillips DR (1982) Purification of glycoproteins IIb and III from human platelet membranes and characterization of a calcium-dependent glycoprotein IIb-IIIa complex. J Biol Chem 257:10458–10466

  14. Jennings LK, Fox JEB, Wdwards HH, Phillips DR (1981) Changes in the cytoskeletal structure of human platelets following thrombin activation. J Biol Chem 256:6927–6932

  15. Lombardo VT, Hodson E, Roberts JR, Kuniki TJ, Zimmerman TS, Ruggeri ZM (1985) Independent modulation of vWF and fibrinogen binding to the platelet membrane GPIIb/IIIa complex as demonstrated by monoclonal antibody. J Clin Invest 76:1950–1958

  16. McEver RP, Martin MN (1984) A monoclonal antibody to a membrane glycoprotein binds only to activated platelets. J Biol Chem 259:9799–9804

  17. Metzelaar MJ, Wijngaard PJ, Peters PJ, Sixma JJ, Nieuwenhuis HK, Clevers HC (1991) CD63: a novel lysosomal membrane glycoprotein cloned by a screening procedure for intracellular antigens in eukaryotic cells. J Biol Chem 266:3239–3245

  18. Michelson AD, Ellis PA, Barnard MR, Matic GB, Viles AF, Kestin AS (1991) Downregulation of the platelet surface glycoprotein Ib-IX complex in whole blood stimulated by thrombin, adenosine diphosphate, or an in vivo wound. Blood 77:770–779

  19. Mustard JF, Kinlough-Rathbone RL, Packham MA, Perry DW, Harfenist EJ, Pai KRM (1979) Comparison of fibrinogen association with normal and thrombasthenic platelets on exposure to ADP or chymotrypsin. Blood 54:987–993

  20. Nieuwenhuis HK, Oosterhout JJG van, Rosemuller E, Iwaarden F van, Sixma JJ (1987) Studies with a monoclonal antibody against activated platelets: evidence that a secreted 53000-molecular weight lysosome-like granule protein is exposed on the surface of activated platelets in the circulation. Blood 70:838–845

  21. Niiya K, Hodson E, Bader R, Byers-Ward V, Koziol JA, Plow EF, Ruggeri ZM (1987) Increased surface expression of the membrane glycoprotein IIb/IIIa complex induced by platelet activation. Relationship to the binding of fibrinogen and platelet aggregation. Blood 70:475–483

  22. Painter RG, Prodouz KN, Gaarde W (1985) Isolation of a subpopulation of glycoprotein IIb-III from platelet membranes that is bound to membrane actin. J Cell Biol 100:652–657

  23. Phillips DR, Charo IF, Parise LV, Fitzgerald LA (1988) The platelet membrane glycoprotein IIb-IIIa complex. Blood 71:831–843

  24. Polley MJ, Leung LLK, Clark FY, Nachman RL (1981) Thrombin-induced platelet membrane glycoprotein IIb and IIIa complex formation. An electron microscope study. J Exp Med 154:1058–1068

  25. Stenberg PE, McEver RP, Shuman MA, Jacques YV, Bainton DF (1985) A platelet alpha-granule membrane protein (GMP-140) is expressed on the plasma membrane after activation. J Cell Biol 101:880–886

  26. Suzuki H, Yamamoto N, Tanoue K, Yamazaki H (1987) Glycoprotein Ib distribution on the surface of platelets in resting and activation states: an electron microscope study. Histochem J 19:125–136

  27. Suzuki H, Katagiri Y, Tsukita S, Tanoue K, Yamazaki H (1990) Localization of adhesive proteins in two newly subdivided zones in electron-lucent matrix of human platelet α-granules. Histochemistry 94:337–344

  28. Suzuki H, Tanoue K, Yamazaki H (1991) Morphological evidence for the association of plasma membrane glycoprotein IIb/IIIa with the membrane skeleton in human platelets. Histochemistry 96:31–39

  29. Tanoue K, Hasegawa S, Yamaguchi A, Yamamoto N, Yamazaki H (1987) A new variant of thrombasthenia with abnormality glycosylated GPIIb/IIIa. Thromb Res 47:323–333

  30. Tokuyasu KT (1989) Use of poly(vinylpyrrolidone) and poly(vinylalcohol) for cryoultramicrotomy. Histochem J 21:163–171

  31. Van der Meulen J, Furuya W, Grinstein S (1983) Isolation and partial characterization of platelet α-granule membranes. J Membr Biol 71:47–59

  32. Wencel-Drake JD (1990) Plasma membrane GPIIb/IIIa. Evidence for a cycling receptor pool. Am J Pathol 136:61–70

  33. Wencel-Drake JD, Plow EF, Kunicki TJ, Woods VL, Keller DM, Ginsberg MH (1986) Localization of internal pools of membrane glycoproteins involved in platelet adhesive responses. Am J Pathol 124:324–334

  34. White JG (1974) Electron microscopic studies of platelet secretion. In: Spaet TH (ed) Progress in hemostasis and thrombosis, vol 2. Grune and Stratton, New York London, pp 49–98

  35. Yamamoto N, Kitagawa H, Yamamoto K, Tanoue K, Yamazaki H (1989) Calcium ions and the conformation of glycoprotein IIIa that is essential for binding to platelets: analysis by a new monoclonal anti-GPIIIa antibody, TM83. Blood 73:1552–1560

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Correspondence to H. Suzuki.

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Suzuki, H., Nakamura, S., Itoh, Y. et al. Immunocytochemical evidence for the translocation of α-granule membrane glycoprotein IIb/IIIa (integrin αIIbβ3) of human platelets to the surface membrane during the release reaction. Histochemistry 97, 381–388 (1992). https://doi.org/10.1007/BF00270384

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  • Integrin
  • Thrombin
  • Freeze Section
  • Surface Membrane
  • Gold Particle