Protein S levels modulate the activated protein C resistance phenotype induced by elevated prothrombin levels

Summary

Elevated plasma prothrombin levels, due to the prothrombin 20210 G/A mutation or to acquired causes, area risk factor for venous thrombosis,partly because of prothrombin-mediated inhibition of the protein C anticoagulant pathway and consequent activated proteinC (APC) resistance. We determined the effect of plasma prothrombin concentration on the APC resistance phenotype and evaluated the role of protein S levels asa modulating variable. The effect of prothrombin and protein S levels on APC resistance was investigated in reconstituted plasma systems and in a population of healthy individuals using both the aPTT-based and the thrombin generation-based APC resistance tests. In reconstituted plasma, APC resistance increased at increasing prothrombin concentration in both assays. Enhanced APC resistance was caused by the effect of prothrombin on the clotting time in the absence of APC in the aPTT-based test, and on thrombin formation in the presence of APC in the thrombin generation-based test. In plasma from healthy individuals prothrombin levels were highly correlated to protein S levels. Since prothrombin and proteinS had opposite effects on the APC resistance phenotype, the prothrombin/protein S ratio was a better predictor of APC resistance than the levels of either protein alone. Prothrombin titrations in plasmas containing different amounts of proteinS confirmed that proteinS levels modulate the ability of prothrombin to induce APC resistance. These findings suggest that carriers of the prothrombin 20210 G/A mutation, who have a high prothrombin/protein S ratio, may experience a higher thrombosis risk than non-carriers with comparable prothrombin levels.

Keywords

Prothrombin - protein S - thrombin generation and APC resistance

• References

• 1 Dahlbäck B. Blood coagulation. Lancet 2000; 355: 1627-32.
• 2 Walker FJ, Fay PJ. Regulation of blood coagulation by the proteinC system. FASEB J 1992; 06: 2561-7.
• 3 Heeb MJ, Mesters RM, Tans G. et al. Binding of protein S to factor Va associated with inhibition of prothrombinase that is independent of activated protein C. J Biol Chem 1993; 268: 2872-7.
• 4 Hackeng TM, van’t Veer C, Meijers JC. et al. Human proteinS inhibits prothrombinase complex activity on endothelial cells and platelets via direct interactions with factors Va and Xa. J Biol Chem 1994; 269: 21051-8.
• 5 Bertina RM, Koeleman BP, Koster T. et al. Mutation in blood coagulation factor V associated with resistance to activated protein C. Nature 1994; 369: 64-7.
• 6 Dahlbäck B, Carlsson M, Svensson PJ. Familial thrombophilia due to a previously unrecognized mechanism characterized by poor anticoagulant response to activated protein C: prediction of a cofactor to activated protein C. Proc Natl Acad Sci U S A 1993; 90: 1004-8.
• 7 Poort SR, Rosendaal FR, Reitsma PH. et al. A common genetic variation in the 3’-untranslated region of the prothrombin gene is associated with elevated plasma prothrombin levels and an increase in venous thrombosis. Blood 1996; 88: 3698-703.
• 8 Kyrle PA, Mannhalter C, Béguin S. et al. Clinical studies and thrombin generation in patients homozygous or heterozygous for the G20210A mutation in the prothrombin gene. Arterioscler Thromb Vasc Biol 1998; 18: 1287-91.
• 9 Wolberg AS, Monroe DM, Roberts HR. et al. Elevated prothrombin results in clots with an altered fiber structure: a possible mechanism of the increased thrombotic risk. Blood 2003; 101: 3008-13.
• 10 Koenen RR, Tans G, van Oerle R. et al. The APC-independent anticoagulant activity of proteinS in plasma is decreased by elevated prothrombin levels due to the prothrombin G20210A mutation. Blood 2003; 102: 1686-92.
• 11 Seré KM, Rosing J, Hackeng TM. Inhibition of thrombin generation by protein S at low procoagulant stimuli: implications for maintenance of the hemostatic balance. Blood 2004; 104: 3624-30.
• 12 Colucci M, Binetti BM, Tripodi A. et al. Hyperprothrombinemia associated with prothrombin G20210A mutation inhibits plasma fibrinolysis through a TAF-Imediated mechanism. Blood 2004; 103: 2157-61.
• 13 Smirnov MD, Safa O, Esmon NL. et al. Inhibition of activated proteinC anticoagulant activity by prothrombin. Blood 1999; 94: 3839-46.
• 14 Mitchell CA, Jane SM, Salem HH. Inhibition of the anticoagulant activity of protein S by prothrombin. J Clin Invest 1988; 82: 2142-7.
• 15 Norstrøm EA, Tran S, Dahlbäck B. Prothrombin inhibition of the individual activated protein C cleavage sites in factor Va. PhD thesis University of Lund. Sweden: 2004
• 16 Tripodi A, Chantarangkul V, Mannucci PM. Hyperprothrombinemia may result in acquired activated protein C resistance. Blood 2000; 96: 3295-6.
• 17 Castaman G, Tosetto A, Simioni M. et al. Phenotypic APC resistance in carriers of the A20210 prothrombin mutation is associated with an increased risk of venous thrombosis. Thromb Haemost 2001; 86: 804-8.
• 18 Seré KM, Janssen MP, Willems GM. et al. Purified protein S contains multimeric forms with increased APC-independent anticoagulant activity. Biochemistry 2001; 40: 8852-60.
• 19 Di Scipio RG, Hermodson MA, Yates SG. et al. A comparison of human prothrombin, factor IX (Christmas factor), factor X (Stuart factor), and protein S. Biochemistry 1977; 16: 698-706.
• 20 Govers-Riemslag JW, Speijer H, Zwaal RF. et al. The effects of bovine prothrombin fragment 1 and fragment 1. 2 on prothrombin activation. Thromb Res 1985; 38: 375-88.
• 21 Castoldi E, Brugge JM, Nicolaes GA. et al. Impaired APC cofactor activity of factor V plays a major role in the APC resistance associated with the factor V Leiden (R506Q) and R2 (H1299R) mutations. Blood 2004; 103: 4173-9.
• 22 Hemker HC, Wielders S, Kessels H. et al. Continuous registration of thrombin generation in plasma, its use for the determination of the thrombin potential. Thromb Haemost 1993; 70: 617-24.
• 23 Koenen RR, Thomassen MC, Tans G. et al. Effect of oral contraceptives on the anticoagulant activity of protein S in plasma. Thromb Haemost 2005; 93: 853-9.
• 24 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: 274-83.
• 25 Taralunga C, Gueguen R, Visvikis S. et al. Phenotypic sensitivity to activated protein C in healthy families: importance of genetic components and environmental factors. Br J Haematol 2004; 126: 392-7.
• 26 de Visser MC, van Hylckama Vlieg A, Tans G. et al. Determinants of the APTT-based and ETP-based APC sensitivity tests. J Thromb Haemost 2005; 03: 1488-94.
• 27 Curvers J, Thomassen MC, Rimmer J. et al. Effects of hereditary and acquired risk factors of venous thrombosis ona thrombin generation-based APC resistance test. Thromb Haemost 2002; 88: 5-11.
• 28 De Ronde H, Bertina RM. Laboratory diagnosis of APC-resistance: A critical evaluation of the test and the development of diagnostic criteria. Thromb Haemost 1994; 72: 880-6.
• 29 Grand’Maison A, Bates SM, Johnston M. et al. “ProC Global”: a functional screening test that predicts recurrent venous thromboembolism. Thromb Haemost 2005; 93: 600-4.
• 30 Souto JC, Almasy L, Blangero J. et al. Genetic regulation of plasma levels of vitamin K-dependent proteins involved in hematostatis: results from the GAIT Project. Genetic Analysis of Idiopathic Thrombophilia. Thromb Haemost 2001; 85: 88-92.
• 31 van Hylckama Vlieg A, Callas PW, Cushman M. et al. Inter-relation of coagulation factors and D-dimer levels in healthy individuals. J Thromb Haemost 2003; 01: 516-22.