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Logo of jcinvestThe Journal of Clinical Investigation
J Clin Invest. 1984 November; 74(5): 1625–1633.
PMCID: PMC425339

Complex formation of platelet thrombospondin with plasminogen. Modulation of activation by tissue activator.


Thrombospondin (TSP), a multifunctional alpha-granule glycoprotein of platelets, binds fibrinogen, fibronectin, heparin, and histidine-rich glycoprotein and thus may play an important role in regulating thrombotic influences at vessel surfaces. In this study we have demonstrated that purified human platelet TSP formed a complex with purified human plasminogen (Plg). Complex formation was detected by rocket immunoelectrophoresis of mixtures of the purified radiolabeled proteins. Significant complex formation of fluid-phase Plg with adsorbed TSP was also demonstrated by enzyme-linked immunosorbent assay (ELISA). The complex formation was specific, saturable, and inhibited by excess fluid-phase TSP, with an apparent KD of approximately 35 nM. In both ELISA and rocket immunoelectrophoresis systems, complex formation was inhibited by 10 mM epsilon-amino-n-caproic acid, implying that there is a role for the lysine binding sites of Plg in mediating the interaction. TSP also formed a complex with plasmin as detected by ELISA but did not directly inhibit plasmin activity measured with a synthetic fluorometric substrate or with a 125I-fibrin plate assay. TSP, when incubated with Plg before addition to 125I-fibrin plates significantly inhibited the generation of plasmin activity by tissue plasminogen activator (TPA) in a manner that was calcium dependent. A kinetic study of Plg activation by TPA in the presence of TSP demonstrated that Michaelis-Menten kinetics were followed and that TSP acted as a noncompetitive inhibitor. These studies support the hypothesis that TSP, acting as a multifunctional regulator in focal areas of active hemostasis, could serve as a prothrombotic influence, leading to increased deposition of fibrin.

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Selected References

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  • Lawler JW, Slayter HS, Coligan JE. Isolation and characterization of a high molecular weight glycoprotein from human blood platelets. J Biol Chem. 1978 Dec 10;253(23):8609–8616. [PubMed]
  • Baenziger NL, Brodie GN, Majerus PW. A thrombin-sensitive protein of human platelet membranes. Proc Natl Acad Sci U S A. 1971 Jan;68(1):240–243. [PubMed]
  • Margossian SS, Lawler JW, Slayter HS. Physical characterization of platelet thrombospondin. J Biol Chem. 1981 Jul 25;256(14):7495–7500. [PubMed]
  • Hagen I. Effects of thrombin on washed, human platelets: changes in the subcellular fractions. Biochim Biophys Acta. 1975 Jun 12;392(2):242–254. [PubMed]
  • Gerrard JM, Phillips DR, Rao GH, Plow EF, Walz DA, Ross R, Harker LA, White JG. Biochemical studies of two patients with the gray platelet syndrome. Selective deficiency of platelet alpha granules. J Clin Invest. 1980 Jul;66(1):102–109. [PMC free article] [PubMed]
  • Phillips DR, Jennings LK, Prasanna HR. Ca2+-mediated association of glycoprotein G (thrombinsensitive protein, thrombospondin) with human platelets. J Biol Chem. 1980 Dec 25;255(24):11629–11632. [PubMed]
  • Mosher DF, Doyle MJ, Jaffe EA. Synthesis and secretion of thrombospondin by cultured human endothelial cells. J Cell Biol. 1982 May;93(2):343–348. [PMC free article] [PubMed]
  • Saglio SD, Slayter HS. Use of a radioimmunoassay to quantify thrombospondin. Blood. 1982 Jan;59(1):162–166. [PubMed]
  • McPherson J, Sage H, Bornstein P. Isolation and characterization of a glycoprotein secreted by aortic endothelial cells in culture. Apparent identity with platelet thrombospondin. J Biol Chem. 1981 Nov 10;256(21):11330–11336. [PubMed]
  • Jaffe EA, Ruggiero JT, Leung LK, Doyle MJ, McKeown-Longo PJ, Mosher DF. Cultured human fibroblasts synthesize and secrete thrombospondin and incorporate it into extracellular matrix. Proc Natl Acad Sci U S A. 1983 Feb;80(4):998–1002. [PubMed]
  • Jaffe EA, Leung LL, Nachman RL, Levin RI, Mosher DF. Thrombospondin is the endogenous lectin of human platelets. Nature. 1982 Jan 21;295(5846):246–248. [PubMed]
  • Leung LL, Nachman RL. Complex formation of platelet thrombospondin with fibrinogen. J Clin Invest. 1982 Sep;70(3):542–549. [PMC free article] [PubMed]
  • Leung LL. Role of thrombospondin in platelet aggregation. J Clin Invest. 1984 Nov;74(5):1764–1772. [PMC free article] [PubMed]
  • Lahav J, Schwartz MA, Hynes RO. Analysis of platelet adhesion with a radioactive chemical crosslinking reagent: interaction of thrombospondin with fibronectin and collagen. Cell. 1982 Nov;31(1):253–262. [PubMed]
  • Leung LL, Nachman RL, Harpel PC. Complex formation of platelet thrombospondin with histidine-rich glycoprotein. J Clin Invest. 1984 Jan;73(1):5–12. [PMC free article] [PubMed]
  • Lijnen HR, Hoylaerts M, Collen D. Heparin binding properties of human histidine-rich glycoprotein. Mechanism and role in the neutralization of heparin in plasma. J Biol Chem. 1983 Mar 25;258(6):3803–3808. [PubMed]
  • Lijnen HR, Hoylaerts M, Collen D. Isolation and characterization of a human plasma protein with affinity for the lysine binding sites in plasminogen. Role in the regulation of fibrinolysis and identification as histidine-rich glycoprotein. J Biol Chem. 1980 Nov 10;255(21):10214–10222. [PubMed]
  • Deutsch DG, Mertz ET. Plasminogen: purification from human plasma by affinity chromatography. Science. 1970 Dec 4;170(3962):1095–1096. [PubMed]
  • Harpel PC. Alpha2-plasmin inhibitor and alpha2-macroglobulin-plasmin complexes in plasma. Quantitation by an enzyme-linked differential antibody immunosorbent assay. J Clin Invest. 1981 Jul;68(1):46–55. [PMC free article] [PubMed]
  • McConahey PJ, Dixon FJ. A method of trace iodination of proteins for immunologic studies. Int Arch Allergy Appl Immunol. 1966;29(2):185–189. [PubMed]
  • Unkeless JC, Tobia A, Ossowski L, Quigley JP, Rifkin DB, Reich E. An enzymatic function associated with transformation of fibroblasts by oncogenic viruses. I. Chick embryo fibroblast cultures transformed by avian RNA tumor viruses. J Exp Med. 1973 Jan 1;137(1):85–111. [PMC free article] [PubMed]
  • Hoylaerts M, Rijken DC, Lijnen HR, Collen D. Kinetics of the activation of plasminogen by human tissue plasminogen activator. Role of fibrin. J Biol Chem. 1982 Mar 25;257(6):2912–2919. [PubMed]
  • Lawler JW, Slayter HS. The release of heparin binding peptides from platelet thrombospondin by proteolytic action of thrombin, plasmin and trypsin. Thromb Res. 1981 May 1;22(3):267–279. [PubMed]
  • Wiman B, Wallén P. The specific interaction between plasminogen and fibrin. A physiological role of the lysine binding site in plasminogen. Thromb Res. 1977 Feb;10(2):213–222. [PubMed]
  • Lijnen HR, Collen D. Interaction of plasminogen activators and inhibitors with plasminogen and fibrin. Semin Thromb Hemost. 1982 Jan;8(1):2–10. [PubMed]
  • Unkeless JC, Gordon S, Reich E. Secretion of plasminogen activator by stimulated macrophages. J Exp Med. 1974 Apr 1;139(4):834–850. [PMC free article] [PubMed]
  • Wilson EL, Dowdle E. Secretion of plasminogen activator by normal, reactive and neoplastic human tissues cultured in vitro. Int J Cancer. 1978 Oct 15;22(4):390–399. [PubMed]
  • Strickland S, Beers WH. Studies on the role of plasminogen activator in ovulation. In vitro response of granulosa cells to gonadotropins, cyclic nucleotides, and prostaglandins. J Biol Chem. 1976 Sep 25;251(18):5694–5702. [PubMed]
  • Strickland S, Reich E, Sherman MI. Plasminogen activator in early embryogenesis: enzyme production by trophoblast and parietal endoderm. Cell. 1976 Oct;9(2):231–240. [PubMed]
  • Moonen G, Grau-Wagemans MP, Selak I. Plasminogen activator-plasmin system and neuronal migration. Nature. 1982 Aug 19;298(5876):753–755. [PubMed]
  • Hoal EG, Wilson EL, Dowdle EB. The regulation of tissue plasminogen activator activity by human fibroblasts. Cell. 1983 Aug;34(1):273–279. [PubMed]

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