The effect of fibrin(ogen) on thrombin generation and decay

Summary

Defibrination causes a ~30% decrease of thrombin generation (TG) which can be restored by adding native fibrinogen in its original concentration (3 mg/ml). The fibrinogen variant γA/γ′, which binds thrombin with high affinity, is over four times more efficient in this respect than the more common γA/γA form. By using high tissue factor concentrations we accelerated prothrombin conversion so as to obtain a descending part of the TG curve that was governed by thrombin decay only. From that part we calculated the antithrombin (AT)- and α2-macroglobulin- dependent decay constants at a series of concentrations of native, γA/γA and γA/γ′ fibrinogen. We found that the increase of TG in the presence of fibrinogen is primarily due to a dose-dependent decrease of thrombin inactivation by α2-macroglobulin, where the γA/γ′ form is much more active than the γA/γA form. AT-dependent decay is somewhat decreased by γA/γ′ fibrinogen but hardly by the γA/γA form. We assume that binding of thrombin to fibrin(ogen) interferes with its binding to inhibitors. Attenuation of decay only in part explains the stimulating effect of fibrinogen on TG, as fibrinogen stimulates prothrombin conversion, regardless of the fibrinogen variant.

Note: Part of this work was presented at the ISTH meeting in 2013.

Keywords

Fibrinogen - thrombin generation - gamma chain - thrombin decay - prothrombin conversion

• References

• 1 Jackson CM, Nemerson Y. Blood coagulation. Ann Rev Biochem 1980; 49: 765-811.
• 2 Hemker HC, Beguin S. Phenotyping the clotting system. Thromb Haemost 2000; 84: 747-751.
• 3 Hemker HC, Kessels H. Feedback mechanisms in coagulation. Haemostasis 1991; 21: 189-196.
• 4 Tanaka KA, Key NS, Levy JH. Blood coagulation: hemostasis and thrombin regulation. Anesth Analg 2009; 108: 1433-1446.
• 5 Hoffman M, Monroe DM. Coagulation 2006: a modern view of hemostasis. Hematol Oncol Clin North Am 2007; 21: 1-11.
• 6 Hemker HC, Beguin S. Thrombin generation in plasma: its assessment via the endogenous thrombin potential. Thromb Haemost 1995; 74: 134-138.
• 7 Hemker HC, Giesen P, Al Dieri R. et al. Calibrated automated thrombin generation measurement in clotting plasma. Pathophysiol Haemost Thromb 2003; 33: 4-15.
• 8 Kumar R, Beguin S, Hemker HC. The influence of fibrinogen and fibrin on thrombin generation--evidence for feedback activation of the clotting system by clot bound thrombin. Thromb Haemost 1994; 72: 713-721.
• 9 de Bosch NB, Mosesson MW, Ruiz-Saez A. et al. Inhibition of thrombin generation in plasma by fibrin formation (Antithrombin I). Thromb Haemost 2002; 88: 253-258.
• 10 Seegers WH, Nieft M, Loomis EC. Note on the Adsorption of Thrombin on Fibrin. Science 1945; 101: 520-521.
• 11 Seegers WH, Johnson JF, Fell C. An antithrombin reaction to prothrombin activation. Am J Physiol 1954; 176: 97-103.
• 12 Hemker HC, Kremers R. Data management in Thrombin Generation. Thromb Res 2013; 131: 3-11.
• 13 Mosesson MW. Antithrombin I Inhibition of thrombin generation in plasma by fibrin formation. Thromb Haemost 2003; 89: 9-12.
• 14 Doolittle RF, Hong S, Wilcox D. Evolution of the fibrinogen gamma’ chain: implications for the binding of factor XIII, thrombin and platelets. J Thromb Haemost 2009; 07: 1431-1433.
• 15 Higgins DL, Lewis SD, Shafer JA. Steady state kinetic parameters for the thrombin-catalyzed conversion of human fibrinogen to fibrin. J Biol Chem 1983; 258: 9276-9282.
• 16 Mosesson MW. Fibrinogen and fibrin structure and functions. J Thromb Haemost 2005; 03: 1894-1904.
• 17 Alexander KS, Fried MG, Farrell DH. Role of Electrostatic Interactions in Binding of Thrombin to the Fibrinogen gamma’ Chain. Biochemistry 2012; 51: 3445-3450.
• 18 Pospisil CH, Stafford AR, Fredenburgh JC. et al. Evidence that both exosites on thrombin participate in its high affinity interaction with fibrin. J Biol Chem 2003; 278: 21584-21591.
• 19 Rosing J, Tans G, Govers-Riemslag JW. et al. The role of phospholipids and factor Va in the prothrombinase complex. J Biol Chem 1980; 255: 274-283.
• 20 Hemker HC, Giesen P, AlDieri R. et al. The calibrated automated thrombogram (CAT): a universal routine test for hyper- and hypocoagulability. Pathophysiol Haemost Thromb 2002; 32: 249-253.
• 21 Hendrix H, Lindhout T, Mertens K. et al. Activation of human prothrombin by stoichiometric levels of staphylocoagulase. J Biol Chem 1983; 258: 3637-3644.
• 22 Mosesson MW, Sherry S. The preparation and properties of human fibrinogen of relatively high solubility. Biochemistry 1966; 05: 2829-2835.
• 23 Siebenlist KR, Meh DA, Mosesson MW. Plasma factor XIII binds specifically to fibrinogen molecules containing gamma chains. Biochemistry 1996; 35: 10448-10453.
• 24 Okwuosa TM, Klein O, Chan C. et al. 13-year long-term associations between changes in traditional cardiovascular risk factors and changes in fibrinogen levels: The Coronary Artery Risk Development in Young Adults (CARDIA) study. Atherosclerosis 2013; 226: 214-219.
• 25 van den Herik EG, Cheung EY, de Lau LM. et al. Fibrinogen gamma’ levels in patients with intracerebral hemorrhage. Thromb Res 2012; 129: 807-809.
• 26 Meh DA, Siebenlist KR, Mosesson MW. Identification and characterization of the thrombin binding sites on fibrin. J Biol Chem 1996; 271: 23121-23125.
• 27 Hogg PJ, Jackson CM, Winzor DJ. Effects of thermodynamic nonideality in kinetic studies: evidence of an expanded intermediate complex in the inactivation of thrombin by antithrombin III. Biochim Biophys Acta 1991; 1073: 609-613.
• 28 Barrett AJ, Starkey PM. The interaction of alpha 2-macroglobulin with proteinases. Characteristics and specificity of the reaction, and a hypothesis concerning its molecular mechanism. Biochem J 1973; 133: 709-724.
• 29 Seegers WH. Antithrombin III: a backward glance o’er travel’d roads. Adv Exp Med Biol 1975; 52: 195-215.
• 30 Bouton MC, Boulaftali Y, Richard B. et al. Emerging role of serpinE2/protease nexin-1 in hemostasis and vascular biology. Blood 2012; 119: 2452-2457.
• 31 Carrell RW. alpha 1-Antitrypsin: molecular pathology, leukocytes, and tissue damage. J Clin Invest 1986; 78: 1427-1431.
• 32 Rau JC, Mitchell JW, Fortenberry YM, Church FC. Heparin cofactor II: discovery, properties, and role in controlling vascular homeostasis. Semin Thromb Hemost 2011; 37: 339-348.
• 33 Lovely RS, Moaddel M, Farrell DH. Fibrinogen gamma’ chain binds thrombin exosite II. J Thromb Haemost 2003; 01: 124-131.
• 34 Fredenburgh JC, Stafford AR, Leslie BA. et al. Bivalent binding to gammaA/ gamma’-fibrin engages both exosites of thrombin and protects it from inhibition by the antithrombin-heparin complex. J Biol Chem 2008; 283: 2470-2477.
• 35 Kumar R, Beguin S, Hemker HC. The effect of fibrin clots and clot-bound thrombin on the development of platelet procoagulant activity. Thromb Haemost 1995; 74: 962-968.
• 36 Beguin S, Kumar R. Thrombin, fibrin and platelets: a resonance loop in which von Willebrand factor is a necessary link. Thromb Haemost 1997; 78: 590-594.
• 37 von dem Borne PA, Meijers JC, Bouma BN. Effect of heparin on the activation of factor XI by fibrin-bound thrombin. Thromb Haemost 1996; 76: 347-353.
• 38 Harpel PC, Brower MS. Alpha 2-macroglobulin: an introduction. Ann NY Acad Sci 1983; 421: 1-9.
• 39 Travis J, Salvesen GS. Human plasma proteinase inhibitors. Ann Rev Biochem 1983; 52: 655-709.
• 40 Hogg PJ, Jackson CM. Fibrin monomer protects thrombin from inactivation by heparin-antithrombin III: implications for heparin efficacy. Proc Natl Acad Sci USA 1989; 86: 3619-3623.
• 41 Hemker HC, de Smedt E, Hemker PW. During coagulation, thrombin generation shifts from chemical to diffusional control. J Thromb Haemost 2005; 03: 2399-2400.
• 42 Wilhelmsen L, Svardsudd K, Korsan-Bengtsen K. et al. Fibrinogen as a risk factor for stroke and myocardial infarction. N Engl J Med 1984; 311: 501-505.
• 43 Farrell DH. gamma’ Fibrinogen as a novel marker of thrombotic disease. Clin Chem Lab Med 2012; 50: 1903-1909.
• 44 Uitte de Willige S, de Visser MC, Houwing-Duistermaat JJ. et al. Genetic variation in the fibrinogen gamma gene increases the risk for deep venous thrombosis by reducing plasma fibrinogen gamma’ levels. Blood 2005; 106: 4176-4183.
• 45 Cheung EY, Vos HL, Kruip MJ. et al. Elevated fibrinogen gamma’ ratio is associated with cardiovascular diseases and acute phase reaction but not with clinical outcome. Blood 2009; 114: 4603-4604 author reply 4604-4605.
• 46 Omarova F, Uitte De Willige S, Ariens RA. et al. Inhibition of thrombin-mediated factor V activation contributes to the anticoagulant activity of fibrinogen gamma’. J Thromb Haemost 2013; 11: 1669-1678.