Rapid Detection Of Factor V Leiden (fvq506) By Non-radioactive Single Strand Conformation Polymorphism (sscp)

The transition G→A at position 506 of the factor V gene is responsible for resistance to the anticoagulant effect of activated protein C (APC-resistance), and represents the most common hereditary risk factor for venous thrombosis. A comparison of the ability of non-radioactive single-strand conformation polymorphism (SSCP) and of a standard PCR procedure followed by further digestion with MnlI to detect this factor V gene (FVQ 506) transition indicates that these is a good agreement between the two methods. Non-radioactive SSCP analysis therefore represents a rapid and sensitive alternative for the diagnosis of this important point mutation.415379381Anderson, F.A., Wheeler, B., Goldberg, R.J., Hosmer, D.W., Forcier, A., Patwardhan, N.A., Physician practices in the prevention of venous thromboembolism (1993) Ann Int Med, 115, pp. 591-595Colman, R.W., Hirsh, J., Marder, V.J., Salzman, E.W., Hemostasis and Thrombosis (1994) Basic Principles and Clinical Practice, 3rd Edn, , Lippincott, PhiladelphiaBauker, K.A., Rosenberg, R.D., The hypercoagulable state (1991) Disorders of Hemostasis 2nd Edn., pp. 267-291. , Ratnoff OD & Forbes CD ed. Saunders, PhiladelphiaDahlback, B., Carlsson, M., Svensson, P.J., 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 (1993) Proc Natl Acad Sci USA, 90, pp. 1004-1008Koster, T., Rosendaal, F.R., Ronde, H., Briét, E., Vandenbroucke, J., Bertina, R.M., Venous thrombosis due to poor anticoagulant response to activated protein C: Leiden Thrombophilia Study (1993) Lancet, 342, pp. 1503-1506Griffin, J.H., Evatt, B., Wideman, C., Fernández, J.A., Anticoagulant protein C pathway defective in majority of thrombophilic patients (1993) Blood, 82, pp. 1989-1993Svensson, P.J., Dahlbäck, B., Resistance to activated protein C as a basis for venous thrombosis (1994) N Engl J Med, 330, pp. 517-522Bertina, R.M., Koelemann, B.P.C., Koster, T., Rosendaal, F.R., Dirven, R.J., Ronde, H., Mutation in blood coagulation factor V associated with resistance to activated protein C (1994) Nature, 369, pp. 64-67Orita, M., Iwahana, H., Kanazawa, H., Hayashi, K., Sekiya, T., Detection of polymorphisms of human DNA by gel electrophoresis as single-strand conformation polymorphisms (1989) Proc Natl Acad Sci USA, 86, pp. 2766-2770Grompe, M., The rapid detection of unknown mutation in nucleic acids (1993) Nature Genetics, 5, pp. 111-117Sugano, K., Kyogoku, A., Fukayama, N., Ohkura, H., Shimosato, Y., Sekiya, T., Hayashi, K., Rapid and simple detection of c-Ki-ras2 gene codon 12 mutations by nonradioisotopic single-strand conformation polymorphism analysis (1993) Lab Invest, 68, pp. 361-366Arruda, V.R., Annichini-Bizzacchi, J.M., Costa, F.F., Reitsma, P.H., Factor, V., Leiden (FVQ506) is common in Brazilian population (1995) Am J Hematol, 39, pp. 242-243Millar, S.A., Dykes, D.D., Polesky, H.F., A simple salting out procedure for extracting DNA from nucleated cell (1988) Nucleic Acids Res, 16, p. 1215Saiki, R.K., Gelfand, D.H., Stoffel, S., Scharf, S.J., Higushi, R., Horn, G.T., Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase (1983) Science, 239, pp. 487-491Lunghi, B., Iacoviello, L., Gemmati, D., Dilasio, M.G., Castoldi, E., Pinotti, M., Detection of new polymorphic markers in the factor V gene: Association with factor V levels in plasma (1996) Thromb Haemostas, 75, pp. 45-48Engel, H., Zwang, L., VanHiet, H.H.D.M., Michiels, J.J., Stibbe, J., Lindemans, J., Phenotyping and genotyping of coagulation factor V Leiden (1996) Thromb Haemostas, 75, pp. 267-26

The Mutation Ala677 → Val In The Methylene Tetrahydrofolate Reductase Gene: A Risk Factor For Arterial Disease And Venous Thrombosis

Mild hyperhomocysteinemia has been identified as a risk factor for arterial disease and for venous thrombosis. Individuals homozygous for the thermolabile variant of the methylene tetrahydrofolate reductase gene (MTHFR) which results from a common mutation Ala677 → Val and is found in 5-15% of the general population, have significantly elevated plasma homocysteine levels and may account for one of the genetic risk factors in vascular disease. We have analyzed the prevalence of MTHFR-T homozygotes in patients with arterial disease or venous thrombosis. We studied 191 patients with arterial disease and 127 individuals with venous thrombosis and compared with 296 unmatched controls. The results showed that there was a high prevalence of homozygotes for the mutated MTHFR-T allele among a group of patients with arterial disease (19%) in the absence of hyperlipoproteinemia, hypertension, and diabetes mellitus when compared to controls (4%), odds ratio of 5.52 (95% C.I., 2.27 to 13.51). The prevalence of homozygotes among patients with venous thrombosis was 11%, odds ratio of 2.93 (95% C.I., 1.23 to 7.01). The risk of venous thrombosis remained high, odds ratio of 2.63, even after we excluded 27 patients with hereditary thrombophilia (e.g. factor V Leiden, dysfibrinogenemia, deficiency of protein C, protein S, antithrombin III, or factor XII) from the 127 overall cases with venous thrombosis. These data support the hypothesis that being a homozygote for the MTHFR-T is a risk factor for the development of arterial diasease and also for venous thrombosis.775818821Ueland, P.M., Refsum, H., Plasma homocysteine, a risk factor for vascular disease: Plasma levels in health, disease, and drug therapy (1989) J Lab Clin Med, 114, pp. 473-501Kang, S.S., Wong, P.W.K., Malinow, M.R., Hyperhomocysteinemia as a risk factor for occlusive vascular disease (1992) Annu Rev Nutr, 12, pp. 279-298Brattström, L., Lindgren, A., Hyperhomocysteinemia as a risk factor for stroke (1992) Neurol Res, 14, pp. 81-84Verhoef, P., Hennekens, C.H., Malinow, R., Kok, F.J., Willet, W.C., Stampfer, M.J., A prospective study of plasma homocysteine and risk of ischemic stroke (1994) Stroke, 25, pp. 1924-1930Boers, G.H.J., Smals, A.G.H., Trijbels, F.J.M., Fowler, B., Bakkeren, J.A.J.M., Schoonderwaldt, H.C., Kleijer, W.J., Kloppenborg, P.W.C., Heterozygosity for homocystinuria in peripheral and cerebral occlusive arterial disease (1985) N Engl J Med, 313, pp. 709-715Boushey, C.J., Beresford, S.S.A., Omenn, G.S., Motulsky, A.G., A quantitative assessment of plasma homocysteine as a risk factor for vascular disease: Probable benefits of increasing folic acid intakes (1995) JAMA, 274, pp. 1049-1057Clarke, R., Daly, L., Robinson, K., Naughten, E., Cahalane, S., Fowler, B., Graham, I., Hyperhomocysteinemia: An independent risk factor for vascular disease (1991) N Engl J Med, 324, pp. 1149-1155Falcon, C.R., Cattaneo, M., Panzeri, D., Martinelli, I., Mannucci, P.M., High prevalence of hyperhomocyst(e)inemia in patients with juvenile venous thrombosis (1994) Arterioscler Thromb, 14, pp. 1080-1083Den Heijer, M., Blom, H.J., Gerrits, W.B.J., Rosendaal, F.R., Haak, H.L., Wijermans, P.W., Bos, G.M.J., Is hyperhomocysteinemia a risk factor for recurrent venous thrombosis? (1995) Lancet, 345, pp. 882-885Den Heijer, M., Koster, T., Blom, H.J., Bos, G.M.J., Briët, E., Reitsma, P.H., Vandenbroucke, J.P., Rosendaal, F.R., Hyperhomocysteinemia as a risk factor for deep-vein thrombosis (1996) N Engl J Med, 334, pp. 759-762Ubbink, J.B., Vermaak, W.J.H., Van Der Merwe, A., Becker, P.J., Vitamin B 12, vitamin B 6, and folate nutritional status in men with hyperhomocysteinemia (1993) Am J Clin Nutr, 57, pp. 47-53Stabler, S.P., Marcell, P.D., Podell, E.R., Allen, R.H., Savage, D.G., Lindenbaum, J., Elevation of total homocysteine in the serum of patients with cobalamin or folate deficiency detected by capillary gas chromatography-mass spectrometry (1988) J Clin Invest, 81, pp. 466-474Kraus, J.P., Molecular basis of phenotype expression in homocystinuria (1994) J Inherit Metab Dis, 17, pp. 383-390Daly, L., Robinson, K., Tan, K.S., Graham, I., Hyperhomocysteinemia: A metabolic risk factor for coronary heart disease determined by both genetic and environmental influences? (1993) Q J Med, 86, pp. 685-689Mudd, S.H., Havlik, Rb., Levy, H.L., McKusick, V.A., Feinleib, M., A study of cardiovascular risk in heterozygotes for homocystinuria (1981) Am J Hum Genet, 33, pp. 883-893Engbersen, A.M.T., Franken, D.G., Boers, G.H.J., Stevens, E.M.B., Trijbels, F.J.M., Blom, H.J., Thermolabile 5,10-methylenetetrahydrofolate reductase as a cause of mild hyperhomocysteinemia (1995) Am J Hum Genet, 56, pp. 142-150Goyette, P., Frosst, P., Rosenblatt, D.S., Rozen, R., Seven novel mutations in the methylenetetrahydrofolate reductase gene and genotype/phenotype correlations in severe methylenetetrahydrofolate reductase deficiency (1995) Am J Hum Genet, 56, pp. 1052-1059Kang, S.S., Wong, P.W.K., Susmano, A., Sora, J., Norusis, M., Ruggie, N., Thermolabile methylenetetrahydrofolate reductase: An inherited risk factor for coronary artery disease (1991) Am J Hum Genet, 48, pp. 536-545Kang, S.S., Wong, P.W.K., Zhou, J., Sora, J., Lessick, M., Ruggie, N., Grcevich, G., Thermolabile methylenetetrahydrofolate reductase in patients with coronay artery disease (1988) Metabolism, 37, pp. 611-613Kang, S.S., Zhou, J., Wong, P.W.K., Kowalisyn, J., Strokosch, G., Intermediate homocysteinemia: A thermolabile variant of methylenetetrahydrofolate reductase (1988) Am J Hum Genet, 43, pp. 414-421Franken, D.G., Boers, G.H.J., Blom, H.J., Trijbels, J.M.F., Kloppernborg, P.W.C., Treatment of mild hyperhomocysteinemia in vascular disease patients (1994) Arterioscler Thromb, 14, pp. 465-470Froost, P., Blom, H.J., Milos, R., Goyette, P., Sheppard, C.A., Matthews, R.G., Boers, G.H.J., Rozen, R., A candidate genetic risk factor for vascular disease: A common mutation in methylenetetrahidrofolate reductase (1995) Nature Genetics, 10, pp. 111-113Kluijtmans, L.A.J., Van Den Heuvel, L.P.W.J., Boers, G.H.J., Frosst, P., Stevens, E.M.B., Van Oost, B.A., Den Heijer, M., Rozen, Rl., Molecular genetic analysis in mild hyperhomocysteinemia: A common mutation in methylenetetrahydrofolate reductase gene is a genetic risk factor for cardiovascular disease (1996) Am J Human Genet, 58, pp. 35-41Motulsky, A.G., Nutritional ecogenetics: Homocysteine-related arteriosclerotic vascular disease, neural tube defects, and folic acid (1996) Am J Hum Genet, 58, pp. 17-20Ubbink, J.B., Vermaak, W.J.H., Delport, R., Van Der Merwe, A., Becker, P.J., Potgieter, H., Effective homocysteine metabolism may protect South African blacks against coronary heart disease (1995) Am J Clin Nutr, 62, pp. 802-808Stein, E.A., Lipids, Lipoproteins, and Apolipoprotrein (1987) Fundamentals of Clinical Chemistry, pp. 449-481. , Tieftz NW, editor. Philadelphia, PA: W. B. SaundersGuidelines on testing for the lupus anticoagulant (1991) J Clin Pathol, 44, pp. 885-889Arruda, V.R., Annichino-Bizzacchi, J.M., Costa, F.F., Reitsma, F.F., Factor V Leiden (FVQ506) is common in Brazilian population (1995) Am J Hematol, 39, pp. 242-243Wolf, M., Boyer-Neumann, C., Martinoli, J.-L., A new functional assay for human protein S activity using activated factor V as substrate (1989) Thromb Haemost, 62, pp. 1144-1145Odegaard, O.R., Try, K., Abilgaard, U., Protein C: A simplified semi-automated activity assay (1986) Thromb Res, 42, pp. 257-262Abildgaard, U., Lie, M., Oldergaard, O.R., Antithrombin (heparin cofactor) assay with new chromogenic substrates (1977) Thromb Res, 11, pp. 549-553Bowie, E.J.W., Owen, J.C.A., Clinical and laboratory diagnosis of hemorrhagic disorders (1991) Disorders of Hemostasis, pp. 48-74. , Ratnoff OD, Forbes CD, editors. Philadelphia: W. B. Saunders CompanyMillar, S.A., Dykes, D.D., Polesky, H.F., A simple salting out procedure for extracting DNA from nucleated cell (1988) Nucleic Acids Res, 16, p. 1215Saiki, R.K., Gelfand, D.H., Stoffel, S., Scharf, S.J., Higuchi, R., Horn, G.T., Mullis, K.B., Erlich, H.A., Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase (1988) Science, 239, pp. 487-491Dean, A.G., Dean, J.A., Coulombier, D., Brendel, K.A., Smith, D.C., Burton, A.H., Dicker, R.C., Arner, T.G., (1994) Epi Info, Version 6: A Word Processing, Database, and Statistics Program for Epidemiology on Microcomputers, , Atlanta: Center for Disease Control and PreventionWilcken, D.E.L., Wang, X.L., Sim, A.S., McCredie, Distribution in healthy and coronary populations of the methylenetetrahydrofolate reductase (MTHFR) C677T mutation (1996) Arterioscler Thromb Vase Biol, 16, pp. 878-88

Prevalence Of The Mutation C677 → T In The Methylene Tetrahydrofolate Reductase Gene Among Distinct Ethnic Groups In Brazil

Vascular disease is a serious public health problem in the industrialized world, and is a frequent cause of death among the adult population of Brazil. Mild hyperhomocysteinemia has been identified as a risk factor for arterial disease, venous thrombosis, and neural tube defects. Individuals homozygous for the thermolabile variant of methylenetetrahydrofolate reductase (MTHFR-T) are found in 5-15% of the general population and have significantly elevated plasma homocysteine levels which represent one of the genetic risk factors for vascular diseases. We have analyzed the prevalence of individuals homozygous for the MTHFR-T in 327 subjects representing the three distinct ethnic groups in Brazil. The prevalence of homozygotes for the mutated allele MTHFR-T was high among persons of Caucasian descent (10%) and considerably lower among Black (1.45%) and Indians persons populations (1.2%). These data suggest that screening for the MTHFR-T allele should help in identifying individuals with a high risk of vascular disease among populations with a heterogeneous background.784332335Arruda, V.R., Von Zuben, P.M., Soares, M.C.P., Menezes, R., Annichino-Bizzacchi, J.M., Costa, F.F., Very low incidence of Arg506 → Gln mutation in the factor V gene among the Amazonian Indians and the Brazilian black population (1996) Thromb Haemost, 75, pp. 860-861Arruda, V.R., Von Zuben, P.M., Annichino-Bizzacchi, J.M., Costa, F.F., The mutation Ala677 → Val in the methylenetetrahydrofolate reductase gene: A risk factor for premature arterial disease and venous thrombosis (1997) Thromb Haemost, 77, pp. 818-821Black, F.L., Salzano, F.M., Layrisse, Z., Franco, M.H.L.P., Harris, N.S., Weimer, T.A., Restriction and persistence of polymorphisms of HLA and other blood genetic traits in the Parakanā Indians of Brazil (1980) Am J Phys Anthr, 52, pp. 119-132Boushey, C.J., Beresford, S.A.A., Omenn, G.S., 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a risk factor for deep-vein thrombosis (1996) N Engl J Med, 334, pp. 759-762Duncan, B.B., Schmidt, M.I., Polanczyk, C.A., Mengue, S.S., High mortality rates in Brazilian adult populations-an international comparison (1992) Rev Ass Med Brasil, 38, pp. 138-144Engbersen, A.M.T., Franken, D.G., Boers, G.H.J., Stevens, E.M.B., Trijbels, F.J.M., Blom, H.J., Thermolabile 5,10-methylenetetrahydrofolate reducttase as a cause of mild hypermocysteinemia (1995) Am J Hum Genet, 56, pp. 142-150Falcon, C.R., Cattaneo, M., Panzeri, D., Martinelli, I., Mannucci, P.M., High prevalence of hyperhomocyst(e)inemia in patients with juvenile venous thrombosis (1994) Arterioscler Thromb, 14, pp. 1080-1083Franken, D.G., Boers, G.H.J., Blom, H.J., Trijbels, J.M.F., Kloppernborg, P.W.C., Treatment of mild hyperhomocysteinemia in vascular disease patients (1994) Arterioscler Thromb, 14, pp. 465-470Froost, P., Blom, H.J., Milos, R., Goyette, P., Sheppard, C.A., Matthews, R.G., Boers, G.H.J., Rozen, R., A candidate genetic risk factor for vascular disease: A common mutation in methylenetetrahydrofolate reductase (1995) Nature Genet, 10, pp. 111-113Gallagher, P.M., Meleady, R., Shields, D.C., Tan, K.S., McMaster, D., Rozen, R., Evans, A., Whitehead, A.S.D., Homocysteine and risk of premature coronary heart disease (1996) Circulation, 94, pp. 2154-2158Kang, S.S., Wong, P.W.K., Susmano, A., Sora, J., Norusis, M., Ruggie, N., Thermolabile methylenetetrahydrofolate reductase: An inherited risk factor for coronary artery disease (1991) Am J Hum Genet, 48, pp. 536-545Kluijtmans, L.A.J., Van Den Heuvel, L.P.W.J., Boers, G.H.J., Frosst, P., Stevens, E.M.B., Van Oost, B.A., Den Heijer, M., Rozen, R.I., Molecular genetic analysis in mild hyperhomocysteinemiaa common mutation in methylenetetrahydrofolate reductase gene is a genetic risk factor for cardiovascular disease (1996) Am J Human Genet, 58, pp. 35-41McCully, K.S., Vascular pathology of homocysteinemia (1969) Am J Pathol, 56, pp. 111-128McCully, K.S., Homocysteine and vascular disease (1996) Nature Medicine, 2, pp. 386-389Millar, S.A., Dykes, D.D., Polesky, H.F., A simple salting out procedure for extracting DNA from nucleated cell (1988) Nucleic Acids Res, 16, p. 1215Motulsky, A.G., Nutritional ecogenetics: Homocysteine-related arteriosclerotic vascular disease, neural tube defects, and folic acid (1996) Am J Hum Genet, 58, pp. 17-20Ou, C.Y., Stevenson, R.E., Brown, V.K., Schwartz, C.E., Allen, W.P., Khoury, M.J., Rozen, R., Adams Jr., M.J., 5,10 Methylenetetrahydofolate reductase genetic polymorphism as a risk factor for neural tube defects (1996) Am J Med Genet, 63, pp. 610-614Rees, D.C., Cox, M., Clegg, J.B., World distribution of factor V Leiden (1995) Lancet, 346, pp. 1133-1134Saiki, R.K., Gelfand, D.H., Stoffel, S., Scharf, S.J., Higuchi, R., Horn, G.T., Mullis, K.B., Erlich, H.A., Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase (1988) Science, 239, pp. 487-491Stevenson, R.E., Schwartz, C.E., 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