The Significant Enhancement of Fibrinolysis by Calcium Ion in a Cell Free System: The Shortening of Euglobulin Clot Lysis Time by Calcium Ion

 

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Summary

We investigated the roles of calcium ion (Ca²⁺) on euglobulin clot lysis time (ECLT) and found that the physiological concentration of Ca²⁺ significantly (4-5 times) shortened ECLT. The shortening was observed at the concentration of Ca²⁺ higher than 1.5-2.0 mM. Other divalent cations such as Mg²⁺, Zn²⁺ or Mn²⁺ didn’t change ECLT. Anti-tPA antibody or plasminogen activator inhibitor-1 prolonged ECLT in the absence of Ca²⁺, whereas they had no effect on ECLT shortened by Ca²⁺. Cl inactivator also had no effect. When barium absorbed plasma was employed, the shortening of ECLT by Ca²⁺ wasn’t observed, whereas it was recovered by the readdition of barium absorbed fraction. When factor X deficient plasma was employed, the shortening of ECLT by Ca²⁺ was also not observed. Thus, Ca²⁺ enhances fibrinolysis in a cell free system by a novel pathway in which the presence of factor X is prerequisite.

• References

• 1 Marsh N. Measurement of fibrinolysis and fibrinolytic components. New York: Johp Wiley & Sons; 1981: 206-241
  Google Scholar
• 2 Davidson JF, Walker ID. Assessment of die fibrinolytic system. In: Haenaostasis and Thrombosis Bloom AL, Ahomas DP. (eds) New York: Churchill Livingstone; 1987: 953-966
• 3 Rijken DC, Wijngaards G, Welbergen J. Relationship between tissue plasminogpp activator and the activators in blood and vascular wall. Thromb Res 1980; 18: 815-830
  CrossrefPubMedGoogle Scholar
• 4 Urano T, Sakakibara K, Rydzewski A, Urano S, Takada Y, Takada A. Relationships between euglohulin clot lysis time and the plasma levels of tissue plasminogen activator gnd plasminogen activator inhibitor 1. Thromb Haemost 1990; 63: 82-86
  Article in Thieme ConnectPubMedGoogle Scholar
• 5 Urano T, Sumiyoshi K, Pietraszek MH, Takada Y, Takada A. PAI-1 plays an important role in the expression of f-PA activity in the euglobulin clot lysis by controlling the concentration pf free t-PA. Thromb Haemost 1991; 66: 474-478
  Article in Thieme ConnectPubMedGoogle Scholar
• 6 Ratnoff OD. Studies on a proteolytic enzyme in human plasma. J Exp Med 1952; 96: 319-329
  CrossrefPubMedGoogle Scholar
• 7 Carlson RH, Gamick RL, Jones AJ, Meunier AM. The determination of recombinant human tissue-type plasminogen activator activity by turbidi-metry using a microcentrifpgal analyzer. Anal Biochem 1988; 168: 428-435
  CrossrefPubMedGoogle Scholar
• 8 Urano S, Metzger AR, Castellino FJ. Plasmin-mediated fibrinolysis by variant recombinant tissue plasminogen activators. Proc Natl Acad Sci USA 1989; 86: 2568-2571
  CrossrefPubMedGoogle Scholar
• 9 Deutsch DG, Mertz ET. Plasminogen: purification from human plasma by affinity chromatography. Science 1970; 170: 1095-1096
  CrossrefPubMedGoogle Scholar
• 10 Takada A, Takada Y. Interaction of Cls and Cl inactivator in the presence of heparin, dextran sulfate and protamine sulfate. Thrombosis Research 1980; 18: 847-859
  CrossrefPubMedGoogle Scholar
• 11 Mullertz S, Clemmensen I. The primary inhibitor of plasmin in human plasma. Biochem J 1976; 159: 545-553
  CrossrefPubMedGoogle Scholar
• 12 Wiman B. Affinity-chromatographic purification of human alpha2-antiplas-min. Biochem J 1980; 191: 229-232
  CrossrefPubMedGoogle Scholar
• 13 Lawrence D, Strandberg L, Grundstrom T, Ny T. Purification of active human plasminogen activator inhibitor 1 from Escherichia coll Comparison with natural and recombinant forms purified from eucaryotic cells. Eur J Biochem 1989; 186: 523-533
  CrossrefPubMedGoogle Scholar
• 14 Biggs R, Macfarlane RG. Human blood coagulation and its disorders (3rd ed.). Blackwell scientific publications; Oxford: 1962: 371
  Google Scholar
• 15 Laemmli KU. Cleavage of structual proteins during the assembly of the head of bacteriophage T4. Nature 1970; 227: 680-685
  CrossrefPubMedGoogle Scholar
• 16 Granelli-Pipemo A, Reich M. A study of proteases and protease-inhibitor complexes in biological fluids. J Exp Med 1978; 148: 223-234
  CrossrefPubMedGoogle Scholar
• 17 Sakata Y, Aoki N. Cross-linking of alpha2-plasmin inhibitor to fibrin by fibrin-stabilizing factor. J Clin Invest 1980; 65: 290-297
  CrossrefPubMedGoogle Scholar
• 18 Sakata Y, Aoki N. Significance of cross-linking of alpha2-plasmin inhibitor to fibrin in inhibition of fibrinolysis and in hemostasis. J Clin Invest 1982; 69: 536-542
  CrossrefPubMedGoogle Scholar
• 19 Furie B, Furie BC. The molecular basis of blood coagulation. Cell 1988; 53: 505-518
  CrossrefPubMedGoogle Scholar
• 20 Esnouf MP. The action of vitamin K. In: Human Blood Coagulation, Haemostasis and Thrombosis Rosemary B, Rizza CR. (eds) Oxford: Blackwell Scientific Publications; 1984: 37-38
• 21 Curtis CG. Plasma factor XIII. In: Haemostasis and Thrombosis Bloom AL, Thomas DP. (eds) New York: Churchill Livingstone; 1981: 216-220
• 22 Ogston D. The Physiology of Hemostasis. Cambridge, Massachusetts: Harvard University Press; 1983: 106-107
  Google Scholar
• 23 Esnouf MP. Isolation of Vitamin K-dependent proteins. In: Human Blood Coagulation, Haemostasis and Thrombosis Biggs R, Rizza CR. (eds) London: Blackwell Scientific Publications; 1984: 34-35
• 24 Walker FJ, Fay PJ. Regulation of blood coagulation by the protein C system. FASEB J 1992; 6: 2561-2567
  CrossrefPubMedGoogle Scholar
• 25 Sakata Y, Loskutoff DJ, Gladson CL, Hekman CM, Griffin JH. Mechanism of protein C-dependent clot lysis: role of plasminogen activator inhibitor. Blood 1986; 68: 1218-1223
  PubMedGoogle Scholar
• 26 de Fouw N, Haverkate F, Bertina RM, Koopman J, van Wijngaarden A, van Hinsbergh V. The cofactor role of protein S in the acceleration of whole blood clot lysis by activated protein C in vitro. Blood 1986; 67: 1189-1192
  PubMedGoogle Scholar
• 27 de Fouw N, de Jong Y, Haverkate F, Bertina RM. Activated protein C increases fibrin clot lysis by neutralization of plasminogen activator inhibitor - no evidence for a cofactor role of protein S. Thromb Haemost 1988; 60: 328-333
  Article in Thieme ConnectPubMedGoogle Scholar
• 28 Stenflo J. Structure and Function of Protein C. Seminars in Thrombosis and Hemostasis 1984; 10: 109-121
  Article in Thieme ConnectPubMedGoogle Scholar
• 29 Esmon CT. Protein C. In: Progress in Hemostasis and Thrombosis Spaet TH. (ed) Orlando, Florida: Grune & Stratton; 1984: 25-54
• 30 Oslen PH, Esmon NL, Esmon CT, Laue TM. Ca²⁺ Dependence of the Interactions between Protein C, Thrombin, and the Esterase Fragment of Thrombomodulin. Analysis by Ultracentrifugation. Biochem 1992; 31: 746-754
  PubMedGoogle Scholar
• 31 Bajzar L, Fredenburgh JC, Nesheim M. The activated protein C-mediated enhancement of tissue-type plasminogen activator-induced fibrinolysis ip a cell-free system. J Biol Chem 1990; 265: 16948-16954
  PubMedGoogle Scholar
• 32 Jackson CM, Nemerson Y. Blood coagulation. Annu Rey Biochepi 1980; 49: 765-811
  PubMedGoogle Scholar
• 33 Sugo T, Bjork J, Holmgren A, Stenflo J. Calcippi-bipdipg properfips of bovine factor X lacking the gamma-carboxyglutamic acid-containing region. J Bipl Clpp 1984; 259: 5705-5710
  PubMedGoogle Scholar
• 34 Skogen WF, Bushong DS, Johnson AE, Cox AC. The role of the Gla domain in the activation of bovine coagulation factor X by the snake venom protein XCP. Biochemical and Biophysical Research Communications 1983; 111: 14-20
  CrossrefPubMedGoogle Scholar
• 35 Ellis V, Scully M, Kakkar V. The effect of divalent metal cations on the inhibition of human coagulation factor Xa by plasma proteinase inhibitors. Biochem Biophys Acta 1983; 747: 123-129
  CrossrefPubMedGoogle Scholar
• 36 Heeb MJ, Gruber A, Griffin JH. Identification of divalent metal ion-dependent inhibition of activated protein C by alpha₂-macroglobulin and alpha₂-antiplasmin in blood and comparisons to inhibition of factor Xa, thrombin, and plasmin. J Biol Chem 1991; 266: 17606-17612
  PubMedGoogle Scholar
• 37 von Kaulla KN, Shultz RL. Methods for the evaluation of human fibrinolysis. Am J Clin Path 1958; 29: 104-112
  CrossrefPubMedGoogle Scholar