Analysis of the F8 Gene in Individuals with High Plasma Factor VIII:C Levels and Associated Venous Thrombosis

Summary

High FVIII:C levels have previously been shown to be an independent risk factor for thrombosis with 4.8 times higher potential risk of thrombosis in individuals with FVIII:C levels greater than 1.5 u/ml. Recently, we found that raised FVIII:C levels are largely attributable to elevated FVIII:Ag levels. The determinants of FVIII:Ag levels are unclear and might be partly genetic. The promoter of the F8 gene has recently been characterised we therefore investigated the promoter and the 3’ terminus of the F8 gene for possible polymorphisms associated with raised FVIII:Ag levels in 62 selected individuals with a thrombotic tendency. We confirm previous reports that raised FVIII:C levels are largely attributable to an elevation in FVIII:Ag and this is also associated with elevation of vWF; non-O blood group; relatively short APTT and relatively low APC ratio. We screened 1140bp of the proximal promoter including the protein binding sites identified by DNase I footprint analysis by SSCP, however no polymorphisms were identified. Direct DNA sequence analysis of the region –542 to +165 failed to identify any sequence polymorphisms. The recently described polymorphism in the polyadenylation cleavage site in the prothrombin gene associated with increased prothrombin activity prompted us to screen the region surrounding the 3’ terminus of the F8 gene for polymorphisms but we found none.

^* Present address: Dr. E. P. G. Mansvelt, Department of Haematological Pathology, Medical Faculty, University of Stellenbosch, PO Box 19063, Tygerberg, 7505, South Africa

• References

• 1 Bertina RM, Koeleman BPC, Koster T, Rosendaal FR, Dirven RJ, De Ronde H, Van der Velden PA, Reitsma PH. Mutation in blood coagulation factor V associated with resistance to activated protein C.. Nature 1994; 369: 64-7.
• 2 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 USA 1993; 90: 1004-8.
• 3 Koster T, Blann AD, Briët E, Vandenbroucke JP, Rosendaal FR. Role of clotting factor VIII in effect of von Willebrand factor on occurrence of deep-vein thrombosis.. Lancet 1995; 345: 152-5.
• 4 O’Donnell J, Tuddenham EGD, Manning R, Kemball-Cook G, Johnson D, Laffan M. High prevalence of elevated factor VIII levels in patients referred for thrombophilia screening: Role of increased synthesis and relationship to the acute phase reaction.. Thromb Haemost 1997; 77: 825-8.
• 5 Scopes D, Berg LP, Krawczak M, Kakkar VV, Cooper DN. Polymorphic variation in the human protein C (PROC) gene promoter can influence transcriptional efficiency in vitro.. Blood Coagul Fibrinolysis 1995; 6: 317-21.
• 6 Spek CA, Koster T, Rosendaal FR, Bertina RM, Reitsma PH. Genotypic variation in the promoter region of the protein C gene is associated with plasma protein C levels and thrombotic risk.. Arterioscler Thromb Vasc Biol 1995; 15: 214-8.
• 7 Dawson SJ, Wiman B, Hamsten A, Green F, Humphries S, Henney AM. The two allele sequences of a common polymorphism in the promoter of the plasminogen activator inhibitor-1 (PAI-1) gene respond differently to interleukin-1 in HepG2 cells.. J Biol Chem 1993; 268: 10739-45.
• 8 Thomas AE, Green FR, Kelleher CH, Wilkes HC, Brennan PJ, Meade TW, Humphries SE. Variation in the promoter region of the beta fibrinogen gene is associated with plasma fibrinogen levels in smokers and non-smokers.. Thromb Haemost 1991; 65: 487-90.
• 9 Sacchi E, Tagliabue L, Scoglio R, Baroncini C, Coppola R, Bernardi F, Mannucci PM. Plasma factor VII levels are influenced by a polymorphism in the promoter region of the FVII gene.. Blood Coagul Fibrinolysis 1996; 7: 114-7.
• 10 Poort SR, Rosendaal FR, Reitsma PH, Bertina RM. 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.
• 11 Rotblat F, Tuddenham EG. Immunologic studies of factor VIII coagulant activity (VIII:C) 1.. Assays based on a haemophilic and an acquired antibody to VIII: Thromb Res 1981; 21: 431-45.
• 12 Kunkel LM, Smith KD, Boyer SH, Borgaonkar DS, Wachtel DS, Miller OJ, Berg WR, Jones HW, Rary JM. Analysis of human Y chromosome-specific reiterated DNA in chromosome variants.. Proceedings Of The National Academy Of Sciences Of The United States Of America 1977; 74: 1245-9.
• 13 Rabes JP, Trossaert M, Conard J, Samama M, Giraudet P, Boileau C. Single point mutation at Arg506 of factor V associated with APC resistance and venous thromboembolism: improved detection by PCR-mediated site-directed mutagenesis [letter].. Thromb Haemost 1995; 74: 1379-80.
• 14 Michaelides K, Schwaab R, Lalloz MRA, Schmidt W, Tuddenham EGD. Mutational analysis: new mutations.. In: PCR2. A practical approach.. McPherson MJ, Hames BD, Talyor GR. (eds). Oxford, IRL Press, Oxford University Press,; 1995. pp 255-88.
• 15 McKenna R, Bachmann F, Miro Quesada M. Thrombo-embolism in patients with abnormally short activated partial thromboplastin time.. Thromb Haemost 1977; 38: 893-9.
• 16 Henkens CM, Bom VJ, Van der Meer J. Lowered APC-sensitivity ratio related to increased factor VIII-clotting activity [letter].. Thromb Haemost 1995; 74: 1198-9.
• 17 Simioni P, Prandoni P, Lensing AW, Scudeller A, Sardella C, Prins MH, Villalta S, Dazzi F, Girolami A. The risk of recurrent venous thromboembolism in patients with an Arg506→Gln mutation in the gene for factor V (factor V Leiden).. N Engl J Med 1997; 336: 399-403.
• 18 Laffan MA, Manning R. The influence of factor VIII on measurement of activated protein C resistance.. Blood Coagul Fibrinolysis 1996; 7: 761-5.
• 19 Noe DA, Murphy PA, Bell WR, Siegel JN. Acute-phase behavior of factor VIII procoagulant and other acute-phase reactants in rabbits.. Am J Physiol 1989; 257: R49-56.
• 20 Palareti G, Legnani C, Guazzaloca G, Frascaro M, Grauso F, De Rosa F, Fortunato G, Coccheri S. Activation of blood coagulation after abrupt or stepwise withdrawal of oral anticoagulants – A prospective study.. Thromb Haemost 1994; 72: 222-6.
• 21 Figueiredo MS, Brownlee GG. cis-acting elements and transcription factors involved in the promoter activity of the human factor VIII gene.. J Biol Chem 1995; 270: 11828-38.
• 22 McGlynn LK, Mueller CR, Begbie M, Notley CR, Lillicrap D. Role of the liver-enriched transcription factor hepatocyte nuclear factor 1 in transcriptional regulation of the factor VIII gene.. Molecular And Cellular Biology 1996; 16: 1936-45.
• 23 Kamphuisen PW, Houwing-Duistermaat JJ, van Houwelingen HC, Eiken-boom JCJ, Bertina RM, Rosendaal FR. Familial clustering of factor VIII and von Willebrand factor levels.. Thromb Haemost 1998; 79: 323-7.