Download PDF
Thromb Haemost 1995; 74(02): 736-742
DOI: 10.1055/s-0038-1649806
Original Article
Platelets
Schattauer GmbH Stuttgart

Protein Tyrosine Phosphorylation in Human Platelets during Shear Stress-Induced Platelet Aggregation (SIPA) Is Regulated by Glycoprotein (GP) Ib/IX as well as GP IIb/IIIa and Requires Intact Cytoskeleton and Endogenous ADP

Atsushi Oda
1   The Division of Hematology, Department of Internal Medicine, Keio University, Tokyo, Japan
,
Kenji Yokoyama
1   The Division of Hematology, Department of Internal Medicine, Keio University, Tokyo, Japan
,
Mitsuru Murata
1   The Division of Hematology, Department of Internal Medicine, Keio University, Tokyo, Japan
,
Michihide Tokuhira
1   The Division of Hematology, Department of Internal Medicine, Keio University, Tokyo, Japan
,
Kosei Nakamura
1   The Division of Hematology, Department of Internal Medicine, Keio University, Tokyo, Japan
,
Makoto Handa
2   The Blood Center, School of Medicine, Keio University, Tokyo, Japan
,
Kiyoaki Watanabe
3   The Department of Laboratory Medicine, School of Medicine, Keio University, Tokyo, Japan
,
Yasuo Ikeda
1   The Division of Hematology, Department of Internal Medicine, Keio University, Tokyo, Japan
› Author Affiliations
Further Information

Publication History

Received 25 February 1994

Accepted after revision 22 February 1995

Publication Date:
06 July 2018 (online)

  PDF Download  Permissions and Reprints

Summary

Shear stress-induced platelet aggregation (SIPA) may be essential in thrombus formation in pathologically stenotic arteries. Intracellular events during SIPA are, however, poorly understood. Washed platelets were exposed to shear stress (108 dyne/cm2) in the presence of von Willebrand factor (vWf, 10 μg/ml) and 1 mM CaCl2 for various time intervals, and then lyzed in SDS. Platelet proteins were separated by 10% SDS-PAGE and tyrosine phosphorylated proteins were detected by immunoblotting with an anti-phosphotyrosine monoclonal antibody. Increased tyrosine phosphorylation of proteins of 130, 100, 85, 74, 70, 64, 58, and 40 kDa was observed within 30 s after the beginning of exposure of platelets to high shear force and the degree of tyrosine phosphorylation continued to increase up to approximately 2 min after the exposure. A monoclonal antibody (MoAb) against vWf-binding domain of glycoprotein (GP) Ibα (GUR83-35), anti-vWf MoAb that inhibits binding of vWf to GPIbα (NMC-4), or a MoAb against GP IIb/IIIa complex (AP-2) inhibited SIPA as well as tyrosine phosphorylation of these proteins. Apyrase (an ADP scavenger, 2 U/ml), EDTA (5 mM), or RGDS peptide (200 μg/ml) also had inhibitory effects on both SIPA and tyrosine phosphorylation. However, Cytochalasin D (2 μM) or staurosporin (1 μM) did not affect SIPA, while they inhibited SIPA-associated tyrosine phosphorylation of those proteins. SIPA-associated tyrosine phosphorylation is a novel post-aggregatory pathway in signal transduction, which is dependent on the binding of vWf to GP Ib/IX and GP IIb/IIIa, endogenous ADP, and intact cytoskeleton.

 

  • References

  • 1 O’Brien JR. Shear-induced platelet aggregation. Lancet 1990; 335: 711-713
    CrossrefPubMedGoogle Scholar
  • 2 Ruggeri ZM, Ware J. von Willebrand factor. FASEB J 1993; 7: 308-316
    CrossrefPubMedGoogle Scholar
  • 3 Wurzinger U. Histophysiology of the circulating platelet. In: Advance in Anatomy, Embryology and Cell Biolog. Beck L, Hild W, Kriz W, Pauly JL, Schielbler TH. (eds) Berlin, Heidelberg: Springer; 1990. 120 I 97
    Google Scholar
  • 4 Moake JL, Turner NA, Stehopolous NA, Nolasco LH, Heliums JD. Shear- induced platelet aggregation can be mediated by vWb released from platelets, as well as by exogenous large or unusually large vWF multimeres, requires adenosine diphosphate, and is resistant to aspirin. Blood 1981; 58: 678-682
    PubMedGoogle Scholar
  • 5 Moake JL, Turner NA, Stehopolous NA, Nolasco LH, Heliums JD. Involvement of large plasma von Willebrand factor (vWL) and unusually large vWL forms derived from endothelial cells in shear-induced platelet aggregation. J Clin Invest 1986; 78: 1456-1461
    CrossrefPubMedGoogle Scholar
  • 6 Ikeda Y, Handa M, Kawano K, Kamata T, Murata M, Araki Y, Anbo H, Kawai Y, Walanabe K, Itagaki I, Sakai K, Ruggeri ZM. The role of von Willebrand factor and fibrinogen in platelet aggregation under varying shear stress. J Clin Invest 1991; 87: 1234-1240
    CrossrefPubMedGoogle Scholar
  • 7 Goto S, Ikeda Y, Murata M, Handa M, Takahashi E, Yoshioka A, Fujimura Y, Fukuyama M, Handa S, Ogawa S. Epinephrine augments von Willebrand factor-dependent shear-induced platelet aggregation. Circulation 1992; 86: 1859-1863
    CrossrefPubMedGoogle Scholar
  • 8 Murata M, Fukuyama M, Satoh K, Fujimura Y, Yoshioka A, Takahashi H, Handa M, Kawai Y, Walanabe K, Ikeda Y. Low shear stress can initiate von Willebrand factor-dependent platelet aggregation in patients with type MB and platelet-type von Willebrand disease. J Clin Invest 1993; 92: 1555-1558
    CrossrefPubMedGoogle Scholar
  • 9 Chow TW, Heliums JD, Moake JL, Kroll MH. Shear stress-induced von Willebrand factor binding to platelet glycoprotein lb initiates calcium influx associated with aggregation. Blood 1992; 80: 113-120
    PubMedGoogle Scholar
  • 10 Ikeda Y, Handa M, Kamata T, Kawano K, Kawai Y, Watanabe K, Kawakami K, Sakai K, Fukuyama M, Itagaki I, Yoshioka A, Ruggeri ZM. Transmembrane calcium influx associated with von Willebrand factor binding to GP lb in the initiation of shear-induced platelet aggregation. Thromb Haemost 1993; 69: 496-502
    Article in Thieme ConnectPubMedGoogle Scholar
  • 11 Kroll MH, Heliums JD, Guo Z, Durante W, Razdan K, Hrbolich JK, Schafer AI. Protein kinase C is activated in platelets subjected to pathological shear stress. J Biol Chem 1993; 268: 3520-3524
    PubMedGoogle Scholar
  • 12 Siess W. Molecular mechanism of platelet activation. Physiol Rev 1989; 12: 71-95
    PubMedGoogle Scholar
  • 13 Oda A, Daley JF, Kang J, Smith M, Ware JA, Salzman EW. Quasisimultaneous measurement of ionized calcium and a-granule release in individual platelets. Am J Physiol 1991; 260: C242-C248
    CrossrefPubMedGoogle Scholar
  • 14 Oda A, Daley JF, Claudia Cabral, Kang J, Smith M, Salzman EW. Heterogeneity in filamentous actin content among individual human blood platelets. Blood 1992; 79: 920-927
    PubMedGoogle Scholar
  • 15 Oda A, Druker BJ, Smith M, Salzman EW. Inhibition of sodium nitro- prusside or PGE, of tyrosine phosphorylation induced in platelets by thrombin and ADP. Am J Physiol 1992; 262: C701-C707
    CrossrefPubMedGoogle Scholar
  • 16 Oda A, Druker BJ, Smith M, Salzman EW. Association of pp60src with Triton X-100-insoluble residue in human blood platelets requires platelet aggregation and actin polymerization. J Biol Chem 1992; 267: 20075-20081
    PubMedGoogle Scholar
  • 17 Oda A, Druker BJ, Ariyoshi H, Smith M, Salzman EW. pp60src is an endogenous substrate for calpain in human blood platelets. J Biol Chem 1993; 268: 12603-12606
    PubMedGoogle Scholar
  • 18 Kugelmass AD, Oda A, Monahan MD, Cabral C, Ware JA. Human platelet activation by cocaine. Circulation 1993; 88: 876-883
    CrossrefPubMedGoogle Scholar
  • 19 Frangioni JV, Oda A, Smith M, Salzman EW, Neel BG. Calpain catalyzed cleavage and subcellular relocation of protein phosphotyrosine phosphatase IB (PTB-1B) in human platelets. EMBO J 1993; 12: 4843-4856
    PubMedGoogle Scholar
  • 20 Ferrel JE Jr, Martin S. Tyrosine specific protein tyrosine phosphorylation is regulated by thrombin. Mol Cell Biol 1988; 8: 3603-3610
    CrossrefPubMedGoogle Scholar
  • 21 Golden A, Brugge JS. Thrombin treatment induces rapid changes in tyrosine phosphorylation in platelets. Proc Natl Acad Sci USA 1989; 86: 901-905
    CrossrefPubMedGoogle Scholar
  • 22 Golden A, Brugge JS, Shattil SJ. Blood platelets express membrane glycoproteins IIb-IIIa in agonist-induced tyrosine phosphorylation of platelet proteins. J Cell Biol 1990; 111: 3117-3127
    CrossrefPubMedGoogle Scholar
  • 23 Haimovich B, Lipfert L, Brugge JS, Shattil SJ. Tyrosine phosphorylation and cytoskeletal reorganization in platelets are triggered by interaction of integrin receptors with immobilized ligands. J Biol Chem 1993; 268: 15868-15877
    PubMedGoogle Scholar
  • 24 Clark E, Brugge JS. Redistribution of activated pp60c_srcto integrin dependent cytoskeletal complexes in thrombin-stimulated platelets. Mol Cell Biol 1993; 13: 1863-1871
    CrossrefPubMedGoogle Scholar
  • 25 Huang M-M, Lipfert L, Cunningham M, Brugge JS, Ginsberg MH, Shattil SJ. Adhesive ligand binding to integrin aIIbP3stimulates tyrosine phosphorylation of novel protein substrates prior to phosphorylation of ppl25FAK . J Cell Biol 1993; 122: 473-483
    CrossrefPubMedGoogle Scholar
  • 26 Lipfert L, Heimovich B, Schaller MD, Cobbs BS, Parsons JT. Integrin- dependent phosphorylation and activation of the protein tyrosine kinase ppl25FAKin platelets. J Cell Biol 1992; 119: 905-912
    CrossrefPubMedGoogle Scholar
  • 27 Wong S, Reynolds AB, Papkoff J. Platelet activation leads to increased c-src kinase activity and association of c-src with an 85-kDa tyrosine phosphoprotein. Oncogenes 1992; 7: 2407-2415
    PubMedGoogle Scholar
  • 28 Clark EA, Brugge JS. Tyrosine phosphorylation of platelets. Trends Cardiovasc Med 1993; 3: 218-227
    CrossrefPubMedGoogle Scholar
  • 29 Fujimura Y, Miyata S, Nishida S, Miura M, Kaneda A, Yoshioka A, Fukui H, Katayama M, Tudenham EG D, Usami Y, Titani K. The interaction of botrocetin with normal or variant von Willebrand factor (type IIA and IIB) and its inhibition by monoclonal antibodies that block receptor binding. Thromb Haemost 1992; 68: 464-469
    Article in Thieme ConnectPubMedGoogle Scholar
  • 30 Pidard D, Montgomery RR, Bennet JS, Kunicki TJ. Interaction of Ap-2, a monoclonal antibody specific for the human platelet glycoprotein IIb-IIIa complex, with intact platelets. J Biol Chem 1983; 258: 12582-12586
    PubMedGoogle Scholar
  • 31 Lamemli UK. Cleavage of structual proteins during assembly of the head of bacteriophage T4. Nature (London) 1970; 227: 680-685
    CrossrefPubMedGoogle Scholar
  • 32 King WG, Kucera GL, Sorisky A, Zhang J, Rittenhouse SE. Protein kinase C regulates the stimulated accumulation of 3-phosphorylated phosphoinositides in platelets. Biochem J 1991; 278: 475-480
    CrossrefPubMedGoogle Scholar
  • 33 Razdan K, Hellumus JD, Kroll MH. Shear-stress induced von Willebrand factor binding to platelets causes the activation of tyrosine kinase(s). Biochem J 1993; 302: 661-666
    PubMedGoogle Scholar