Download PDF
Thromb Haemost 1985; 53(03): 337-342
DOI: 10.1055/s-0038-1661310
Original Article
Schattauer GmbH Stuttgart

Studies on the Effect of Platelet Inhibitors on Platelet Adhesion to Collagen and Collagen-Induced Human Platelet Activation

S Krishnamurthi
The Thrombosis Research Unit, King’s College School of Medicine and Dentistry, London, UK
,
V V Kakkar
The Thrombosis Research Unit, King’s College School of Medicine and Dentistry, London, UK
› Author Affiliations
Further Information

Publication History

Received 12 June 1984

Accepted 01 March 1985

Publication Date:
18 July 2018 (online)

Also available at
  PDF Download  Permissions and Reprints

Summary

The effect of pyridoxal 5’-phosphate (PALP) and trifluoperazine (TFPZ), the calmodulin antagonist, on in vitro platelet adhesion to collagen and collagen-induced platelet activation was studied using platelet-rich-plasma (PRP) or washed platelets (WPL). Platelet aggregation and [14C]-5HT release induced by “threshold” or low concentrations of collagen (0.6 μg/ ml) in PRP were completely abolished by PALP (24 mM), TFPZ (250 μM) as well as indomethacin (10 μM). At higher concentrations of collagen (10–15 μg/ml) in PRP and WPL, the use of stirred and unstirred platelets treated with collagen enabled a distinction to be made between aggregation and adhesion- mediated release reaction. Platelet aggregation and the aggregation-mediated release reaction induced by these concentrations of collagen in stirred platelets were completely abolished by PALP, TFPZ and indomethacin although neither adhesion to collagen nor the adhesion-mediated release reaction of unstirred platelets was significantly affected by these inhibitors. Interestingly, both adhesion and the adhesion-mediated release reaction were abolished by concentrations of PALP 10–40 fold higher than those required to abolish aggregation. Collagen-induced platelet aggregation, but not platelet adhesion, was inhibited in resuspended platelets pretreated with PALP and NaBH4 indicating a separation in the membrane sites involved in aggregation and adhesion. The results further emphasize the distinction between adhesion and aggregation-mediated events with regards to collagen with the latter being more susceptible to inhibition by antiplatelet agents such as PALP and TFPZ.

Keywords

Platelet adhesion - Pyridoxal 5’-phosphate - Trifluoperazine
 

  • References

  • 1 Baumgartner HR, Muggli R, Tschopp TB, Turitto VT. Platelet adhesion, release and aggregation in flowing blood: effects of surface properties and platelet function. Thromb Haemostas 1976; 35: 124-138
    PubMedSearch in Google Scholar
  • 2 Wilner GD, Nossel HL, Procupez TL. Aggregation of platelets by collagen: polar active sites of insoluble human collagen. Am J Physiol 1971; 220: 1074-1079
    PubMedSearch in Google Scholar
  • 3 Jamieson GA, Smith DF, Kosow DP. Possible role of collagen: glucosyltransferase in platelet adhesion. Mechanistic considerations Thrombos Diathes Haemorrh 1975; 33: 668-671
    PubMedSearch in Google Scholar
  • 4 Wang C, Miyata T, Weksler B, Rubin AL, Stenzel KH. Collagen- induced platelet aggregation and release. I. Effects of side-chain modifications and role of arginyl residues. Biochim Biophys Acta 1978; 544: 555-567
    CrossrefPubMedSearch in Google Scholar
  • 5 Meyer FA, Frojmovic MM. Characteristics of the major platelet membrane site used in binding to collagen. Thromb Res 1979; 15: 755-767
    CrossrefPubMedSearch in Google Scholar
  • 6 Legrand YJ, Karniguian A, Le Francier P, Fauvel F, Caen JP. Evidence that a collagen-derived nanopeptide is a specific inhibitor of platelet-collagen interaction. Biochem Biophys Res Commun 1980; 96: 1579-1585
    CrossrefPubMedSearch in Google Scholar
  • 7 Chesney CM, Pifer DD, Crofford LJ, Huch KM. Reevaluation of the role of the polar groups of collagen in the platelet-collagen interaction. Am J Pathol 1983; 112: 200-206
    PubMedSearch in Google Scholar
  • 8 Zucker MB, Peterson J. Effect of acetylsalicyclic acid, other nonsteroidal anti-inflammatory agents, and dipyridamole on human blood platelets. J Lab Clin Med 1970; 76: 66-75
    PubMedSearch in Google Scholar
  • 9 Cazenave JP, Kinlough-Rathbone RL, Packham MA, Mustard JF. The effect of acetylsalicylic acid and indomethacin on rabbit platelet adherence to collagen and the subendothelium in the presence of a low or high hematocrit. Thromb Res 1978; 13: 971-981
    CrossrefPubMedSearch in Google Scholar
  • 10 Kinlough-Rathbone RL, Cazenave JP, Packham MA, Mustard JF. Effect of inhibitors of the arachidonate pathway on the release of granule contents from rabbit platelets adherent to collagen. Lab Invest 1980; 42: 28-34
    PubMedSearch in Google Scholar
  • 11 Cazenave JP, Dejana E, Kinlough-Rathbone RL, Richardson M, Packham MA, Mustard JF. Prostaglandins I2 and E1 reduce rabbit and human platelet adherence without inhibiting serotonin release from adherent platelets. Thromb Res 1979; 15: 273-279
    CrossrefPubMedSearch in Google Scholar
  • 12 Weiss HJ, Turitto VT. Prostacyclin inhibits platelet adhesion and thrombus formation on subendothelium. Blood 1979; 53: 244-250
    PubMedSearch in Google Scholar
  • 13 Krishnamurthi S, Westwick J, Kakkar VV. Regulation of human platelet activation - analysis of cyclooxygenase and cyclic AMP- dependent pathways. Biochem Pharmacol 1984; 33: 3025-3035
    CrossrefPubMedSearch in Google Scholar
  • 14 Fasella P. Pyridoxal phosphate. Ann Rev Biochem 1967; 36: 185-203
    CrossrefPubMedSearch in Google Scholar
  • 15 Krishnamurthi S, Westwick J, Kakkar VV. Effect of pyridoxal 5’- phosphate (PALP) on human platelet aggregation, dense granule release and thromboxane B2 generation - role of Schiff base formation. Thromb Haemostas 1982; 48: 136-141
    PubMedSearch in Google Scholar
  • 16 Krishnamurthi S, Westwick J, Kakkar VV. Studies on the mechanism of pyridoxal 5’-phosphate-induced inhibition of human platelet activation. Thromb Haemostas 1983; 50: 280 (Abstr.)
    PubMedSearch in Google Scholar
  • 17 Subbarao K, Kuchibhotla J, Kakkar VV. Pyridoxal 5’-phosphate - a new physiological inhibitor of blood coagulation and platelet function. Biochem Pharmacol 1979; 28: 531-534
    CrossrefPubMedSearch in Google Scholar
  • 18 Kornecki E, Feinberg H. Pyridoxal phosphate inhibition of platelet function. Am J Physiol 1980; 238: H54-H60
    PubMedSearch in Google Scholar
  • 19 Lam SC-T, Harfenist EJ, Packham MA, Mustard JF. Investigation of possible mechanism of pyridoxal 5’-phosphate inhibition of platelet reactions. Thromb Res 1980; 20: 633-645
    CrossrefPubMedSearch in Google Scholar
  • 20 Rinehart JF, Greenberg LD. Arteriosclerotic lesions in pyridoxine deficient monkeys. Am J Pathol 1949; 25: 481-484
    PubMedSearch in Google Scholar
  • 21 Rinehart JF, Greenberg LD. Pathogenesis of experimental arteriosclerosis in pyridoxine deficiency with notes on similarities to human arteriosclerosis. Arch Pathol 1951; 51: 12-18
    PubMedSearch in Google Scholar
  • 22 McCully KS, Wilson RB. Homocystine theory of arteriosclerosis. Atherosclerosis 1975; 22: 215-227
    CrossrefPubMedSearch in Google Scholar
  • 23 Levin RM, Weiss B. Binding of trifluoperazine to the calcium- dependent activator of cyclic nucleotide phosphodiesterase. Mol Pharmacol 1977; 13: 690-697
    PubMedSearch in Google Scholar
  • 24 Kao KJ, Sommers JR, Pizzo SV. Modulation of platelet shape and membrane receptor binding by Ca2+-calmodulin complex. Nature 1981; 292: 82-84
    CrossrefPubMedSearch in Google Scholar
  • 25 Packham MA, Warrior ES, Glynn MF, Senyi AS, Mustard JF. Alteration of the response of platelets to surface stimuli by pyrazole compounds. J Exp Med 1967; 126: 171-188
    CrossrefPubMedSearch in Google Scholar
  • 26 Scheffel U, McIntyre PA, Evatt B, Dvornicky JA, Natarajan TK, Bolling DR, Murphy EA. Evaluation of 111Indium as a new photon yield gamma emitting physiological platelet label. John Hopkins Med J 1977; 140: 285-293
    PubMedSearch in Google Scholar
  • 27 Legrand YJ, Fauvel F, Kartalis G, Wautier JL, Caen JP. Specific and quantitative method for estimation of platelet adhesion to fibrillar collagen. J Lab Clin Med 1979; 94: 438-446
    PubMedSearch in Google Scholar
  • 28 Lowry OH, Roberts NR, Kapphahn JI. The fluorometric measurement of pyridine nucleotides. J Biol Chem 1957; 224: 1047-1064
    PubMedSearch in Google Scholar
  • 29 Cabantchik IZ, Balshin M, Breuer W, Rothstein A. Pyridoxal phosphate. An anionic probe for protein amino groups exposed on the outer and inner surfaces of intact human red blood cells J Biol Chem 1975; 250: 5130-5136
    PubMedSearch in Google Scholar
  • 30 Eger R, Rifkin DB. The preparation and use of pyridoxal [32P]- phosphate as a labelling reagent for proteins on the outer surface of membranes. Biochim Biophys Acta 1977; 470: 70-83
    CrossrefPubMedSearch in Google Scholar
  • 31 Lahav J, Meyer FA. On the role of the major platelet membrane glycoproteins in platelet adhesion to collagen. Thromb Res 1981; 22: 457-468
    CrossrefPubMedSearch in Google Scholar
  • 32 Pollard HB, Tack-Goldman K, Pazoles CJ, Creutz CE, Shulman NR. Evidence for control of serotonin secretion from human platelets by hydroxyl ion transport and osmotic lysis. Proc Natl Acad Sci USA 1977; 74: 5295-5299
    CrossrefPubMedSearch in Google Scholar
  • 33 Feinstein MB, Hadjian RA. Effects of the calmodulin antagonist trifluoperazine on stimulus-induced calcium mobilization, aggregation, secretion and protein phosphorylation in platelets. Mol Pharmacol 1982; 21: 422-431
    PubMedSearch in Google Scholar
  • 34 Brass LF, Bensusan HB. The role of collagen quaternary structure in the platelet: collagen interaction. J Clin Invest 1974; 54: 1480-1487
    CrossrefPubMedSearch in Google Scholar
  • 35 Nossel HL, Wilner GD, LeRoy EC. Importance of polar groups for initiating blood coagulation and aggregating platelets. Nature 1969; 221: 75-76
    CrossrefPubMedSearch in Google Scholar
  • 36 Wilner GD, Nossel HL, LeRoy EC. Aggregation of platelets by collagen. J Clin Invest 1968; 47: 2616-2621
    CrossrefPubMedSearch in Google Scholar