Techniques to Examine Platelet Adhesive Interactions Under Flow

  • Suhasini Kulkarni
  • Warwick S. Nesbitt
  • Sacha M. Dopheide
  • Sascha C. Hughan
  • Ian S. Harper
  • Shaun P. Jackson
Part of the Methods In Molecular Biology™ book series (MIMB, volume 272)

Abstract

Platelet adhesion and aggregation at sites of vessel-wall injury are critical for the arrest of bleeding and for the development of vaso-occlusive thrombi at sites of atherosclerotic-plaque rupture. These adhesive interactions are critically dependent on multiple receptors on the platelet surface (GPIb/V/IX, GPVI, integrins αIIbβ3 and α2β1) and their specific ligands in the subendothelium (von Willebrand Factor, collagen) and plasma (von Willebrand Factor, fibrinogen) (1,2). In vivo, these receptor-ligand interactions are exposed to a broad range of shear stresses generated by blood flow, ranging from 20–200/s in veins to 800–10,000/s in arteries (3). In stenotic vessels, shear rates can approach 40,000/s. The development of in vitro methodologies mimicking physiological and pathophysiological flow conditions has significantly improved our understanding of the role of shear in regulating platelet functional responses. In general, the effects of shear stress have been studied with platelets in suspension using rotational devices such as the Couette or cone-plate viscometer. Alternatively, the effects of shear on platelets have been evaluated in a laminar-flow device such as the tubular, annular, or parallel-plate flow chamber. Rotational viscometers are ideal for the examination of shear effects on platelet adhesive interactions in the absence of platelet-surface interactions (i.e., platelets in suspension). Such studies are important in determining the mechanisms of platelet activation occurring in areas of vascular stenosis where shear rates are elevated well above physiological levels. Thrombus formation, however, does not generally occur with platelets in suspension but rather involves the progressive accrual of platelets onto vascular subendothelium and subsequently onto immobilized platelets. As such, the in vitro investigation of platelet function under conditions of physiological and pathological shear has been greatly facilitated by laminar flow devices.

Keywords

Burner Formaldehyde Mercury Aspirin Syringe 

References

  1. 1.
    Savage, B., Almus-Jacobs, F., and Ruggeri, Z. M. (1998) Specific synergy of multiple substrate-receptor interactions in platelet thrombus formation under flow. Cell 94, 657–666.PubMedCrossRefGoogle Scholar
  2. 2.
    Dopheide, S. M., Yap, C. L., and Jackson, S. P. (2001) Dynamic aspects of platelet adhesion under flow. J. Exp. Pharm. Phys. 28, 355–363.CrossRefGoogle Scholar
  3. 3.
    Kroll, M. H., Hellums, D., McIntire, L. V., Schafer, A. I., and Moake, J. L. (1996) Platelets and shear stress. Blood 88, 1525–1541.PubMedGoogle Scholar
  4. 4.
    Savage, B., Saldivar, E., and Ruggeri, Z. M. (1996) Initiation of platelet adhesion by arrest onto fibrinogen or translocation on von Willebrand factor. Cell 84, 289–297.PubMedCrossRefGoogle Scholar
  5. 5.
    Alevriadou, B. R., Moake, J. L., Turner, N. A., Ruggeri, Z. M., Folie, B. J., Phillips, M. D., et al. (1993) Real-time analysis of shear-dependent thrombus formation and its blockade by inhibitors of von Willebrand factor binding to platelets. Blood 81, 1263–1276.PubMedGoogle Scholar
  6. 6.
    Ruggeri, Z. M., Dent, J. A., and Saldivar, E. (1999) Contribution of distinct adhesive interactions to platelet aggregation in flowing blood. Blood 94, 172–178.PubMedGoogle Scholar
  7. 7.
    Kulkarni, S., Dopheide, S. M., Yap, C. L., Heel, K. A., Harper, I. S., and Jackson, S. P. (2000) A revised model of platelet aggregation. J. Clin. Invest. 105, 783–791.PubMedCrossRefGoogle Scholar
  8. 8.
    Baumgartner, H. R., Stemerman, M. B., and Spaet, T. H. (1971) Adhesion of blood platelets to subendothelial surface: distinct from adhesion to collagen. Experientia 27, 283–285.PubMedCrossRefGoogle Scholar
  9. 9.
    Baumgartner, H. R. and Haudenschild, C. (1972) Adhesion of platelets to subendothelium. Ann NY Acad. Sci. 201, 22–36.PubMedCrossRefGoogle Scholar
  10. 10.
    Montgomery, R. R. and Zimmerman, T. S. (1978) von Willebrand’s disease antigen II. A new plasma antigen deficient in severe von Willebrand’s disease. J. Clin. Invest. 61, 1498–1507.PubMedCrossRefGoogle Scholar
  11. 11.
    Moake, J. L., Turner, N. A., Stathopoulos, N. A., Nolasco, L. H., and Hellums, J. D. (1986) Involvement of large plasma von Willebrand factor (vWF) multimers and unusually large vWF forms derived from endothelial cells in shear stress-induced platelet aggregation. J. Clin. Invest. 78, 1456–1461.PubMedCrossRefGoogle Scholar
  12. 12.
    Moake, J. L., Turner, N. A., Stathopoulos, N. A., Nolasco, L. H., and Hellums, J. D. (1988) Shear-induced platelet aggregation can be mediated by vWF released from platelets, as well as by exogenous large or unusually large vWF multimers, requires adenosine diphosphate, and is resistant to aspirin. Blood 71, 1366–1374.PubMedGoogle Scholar
  13. 13.
    Jakobsen, E. and Kierulf, P. (1970) A modified β-alanine precipitation procedure to prepare fibrinogen free of anti-thrombin III and plasminogen. Thromb. Res. 3, 145–149.CrossRefGoogle Scholar
  14. 14.
    Cazanave, J. P., Hemmendinger, S., Beretz, A., Sutter-Bay, A., and Launay, J. (1983) L’agrégation plaquettaire: outil d’investigation clinique et d’étude pharmacologique méthodologie. Ann. Biol. Clin. 41, 167–179.Google Scholar
  15. 15.
    Denis, C., Methia, N., Frenette, P. S., Rayburn, H., Ullmann-Cullere, M., Hynes, R. O., et al. (1998) A mouse model of severe von Willebrand disease: defects in haemostasis and thrombosis. Proc. Natl. Acad. Sci. USA 95, 9524–9529.PubMedCrossRefGoogle Scholar
  16. 16.
    Baumgartner, H. R., Tschopp, T. B., and Weiss, H. J. (1977) Platelet interaction with collagen fibrils in flowing blood. II. Impaired adhesion-aggregation in bleeding disorders. A comparison with subendothelium. Thromb Haemost. 37, 17–28.PubMedGoogle Scholar
  17. 17.
    Yap, C. L., Hughan, S. C., Cranmer, S. L., Nesbitt, W. S., Rooney, M. M., Giuliano, S., et al. (2000) Synergistic adhesive interactions and signaling mechanisms operating between platelet glycoprotein Ib/IX and integrin αIIbβ3. Studies in human platelets and transfected chinese hamster ovary cells. J. Biol. Chem. 275, 41,377–41,388.PubMedCrossRefGoogle Scholar
  18. 18.
    Nesbitt, W. S., Kulkarni, S., Giuliano, S., Goncalves, I., Nesbitt, W. S., Kulkarni, S., et al. (2002) Distinct glycoprotein Ib/V/IX and integrin αIIbβ3-dependent calcium signals cooperatively regulate platelet adhesion under flow. J. Biol. Chem. 277, 2965–2972.PubMedCrossRefGoogle Scholar
  19. 19.
    Dopheide, S. M., Maxwell, M. J., and Jackson, S. P. (2002) Shear-dependent tether formation during platelet translocation on von Willebrand factor. Blood 99, 159–167.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc. 2004

Authors and Affiliations

  • Suhasini Kulkarni
    • 1
  • Warwick S. Nesbitt
    • 1
  • Sacha M. Dopheide
    • 1
  • Sascha C. Hughan
    • 1
  • Ian S. Harper
    • 2
  • Shaun P. Jackson
    • 1
  1. 1.Australian Centre for Blood Diseases, Department of MedicineMonash UniversityClaytonAustralia
  2. 2.Australian Centre for Blood Diseases, The Microscopy & Imaging Research FacilityMonash UniversityClaytonAustralia

Personalised recommendations