Association of factor XII gene C46T polymorphism with cerebral venous thrombosis in the south Indian population

Abstract

To cite this article: Prabhakar P, De T, Nagaraja D, Christopher R. Association of factor XII gene C46T polymorphism with cerebral venous thrombosis in the south Indian population. J Thromb Haemost 2012; 10: 1437–9. To cite this article: Prabhakar P, De T, Nagaraja D, Christopher R. Association of factor XII gene C46T polymorphism with cerebral venous thrombosis in the south Indian population. J Thromb Haemost 2012; 10: 1437–9. Cerebral veno‐sinus thrombosis (CVT), which is considered a rare condition in the developed world, is a common cause of stroke in young women in India [1Panagariya A. Maru A. Cerebral venous thrombosis in pregnancy and puerperium – a prospective study.J Assoc Physicians India. 1997; 45: 857-9PubMed Google Scholar]. Despite the wide range of known causes, the etiology of CVT remains undetermined in approximately 15–35% of cases [2Stam J. Cerebral venous and sinus thrombosis: incidence and causes.Adv Neurol. 2003; 92: 225-32PubMed Google Scholar]. Genetic predisposition resulting from mutations in the coagulation factor genes is linked to an increased risk of venous thrombosis [3Lane D.A. Mollica L.R. Haemostatic gene polymorphisms in venous and arterial thrombosis.Pathophysiol Haemost Thromb. 2002; 32: 213-15Crossref PubMed Scopus (9) Google Scholar], and has been reported to lead to a higher thrombosis risk in the cerebral venous circulation than in the arterial circulation [4Marjot T. Yadav S. Hasan N. Bentley P. Sharma P. Genes associated with adult cerebral venous thrombosis.Stroke. 2011; 42: 913-18Crossref PubMed Scopus (59) Google Scholar]. A comprehensive meta‐analysis of all candidate genes for CVT has shown that factor V Leiden, prothrombin G20201A and MTHFR C677T are associated with an increased risk of CVT [4Marjot T. Yadav S. Hasan N. Bentley P. Sharma P. Genes associated with adult cerebral venous thrombosis.Stroke. 2011; 42: 913-18Crossref PubMed Scopus (59) Google Scholar]. In contrast, studies from India have found no such association [5Dindagur N. Kruthika‐Vinod T.P. Christopher R. Thrombophilic gene polymorphisms in puerperal cerebral veno‐sinus thrombosis.J Neurol Sci. 2006; 249: 25-30Abstract Full Text Full Text PDF PubMed Scopus (19) Google Scholar]. FXII (Hageman factor) activates the intrinsic coagulation pathway and initiates blood clotting through activation of FXI. It also plays a significant role in the fibrinolytic pathway by activating prekallikrein in the kallikrein–kinin system and aiding in the conversion of plasminogen to plasmin [6Fujikawa K. Heimark R.L. Kurachi K. Davie E.W. Activation of bovine factor XII (Hageman factor) by plasma kallikrein.Biochemistry. 1980; 19: 1322-30Crossref PubMed Scopus (87) Google Scholar]. Reduced plasma FXII levels have been reported in patients with venous thrombosis [7Schousboe I. Pharmacological regulation of factor XII activation may be a new target to control pathological coagulation.Biochem Pharmacol. 2008; 75: 1007-13Crossref PubMed Scopus (24) Google Scholar]. A C→T substitution at nucleotide 46 in the 5′‐untranslated region of the F12 gene is associated with low translation efficiency and a decrease in the plasma FXII level [8Kanaji T. Okamura T. Osaki K. Kuroiwa M. Shimoda K. Hamasaki N. Niho Y. A common genetic polymorphism (46 C to T substitution) in the 5′‐untranslated region of the coagulation factor XII gene is associated with low translation efficiency and decrease in plasma factor XII level.Blood. 1998; 91: 2010-14Crossref PubMed Google Scholar]. This gene variant has been reported to account for the lower plasma FXII activity found in Orientals [8Kanaji T. Okamura T. Osaki K. Kuroiwa M. Shimoda K. Hamasaki N. Niho Y. A common genetic polymorphism (46 C to T substitution) in the 5′‐untranslated region of the coagulation factor XII gene is associated with low translation efficiency and decrease in plasma factor XII level.Blood. 1998; 91: 2010-14Crossref PubMed Google Scholar]. Some studies have found an association of this mutation with deep vein thrombosis [9Tirado I. Soria J.M. Mateo J. Oliver A. Souto J.C. Santamaria A. Felices R. Borrell M. Fontcuberta J. Association after linkage analysis indicates that homozygosity for the 46C‐‐>T polymorphism in the F12 gene is a genetic risk factor for venous thrombosis.Thromb Haemost. 2004; 91: 899-904Crossref PubMed Google Scholar], although this has been contradicted by others [10Bertina R.M. Poort S.R. Vos H.L. Rosendaal F.R. The 46C‐‐>T polymorphism in the factor XII gene (F12) and the risk of venous thrombosis.J Thromb Haemost. 2005; 3: 597-9Crossref PubMed Scopus (25) Google Scholar]. After a study in 78 German CVT patients, Reuner et al. suggested that the F12 46TT genotype may be a new risk factor for CVT, and could be added to the panel of established risk factors after confirmation in an independent study [11Reuner K.H. Jenetzky E. Aleu A. Litfin F. Mellado P. Kloss M. Juttler E. Grau A.J. Rickmann H. Patscheke H. Lichy C. Factor XII C46T gene polymorphism and the risk of cerebral venous thrombosis.Neurology. 2008; 70: 129-32Crossref PubMed Scopus (26) Google Scholar]. Reuner’s study was replicated in a Spanish study that included five CVT patients [9Tirado I. Soria J.M. Mateo J. Oliver A. Souto J.C. Santamaria A. Felices R. Borrell M. Fontcuberta J. Association after linkage analysis indicates that homozygosity for the 46C‐‐>T polymorphism in the F12 gene is a genetic risk factor for venous thrombosis.Thromb Haemost. 2004; 91: 899-904Crossref PubMed Google Scholar]. To the best of our knowledge, there are no published reports for Asian CVT patients. As genetic risk factors may vary among ethnic groups, we hypothesized that F12 genetic variants could be associated with aseptic CVT in the Indian population, and evaluated the association in south Indians. The study protocol was approved by the National Institute of Mental Health and Neuro Sciences Ethics Committee, and all participants/spouses gave written informed consent. Two hundred and six consecutive patients (82 men, 124 women, mean age 32.5 ± 8.7 years) admitted to the Stroke Unit and diagnosed with aseptic CVT were investigated. The diagnosis was confirmed by magnetic resonance (MR) imaging/MR venography. Patients were excluded when: (i) neuroimaging studies were inconclusive; (ii) CVT was secondary to head trauma, invasive procedures, sepsis, neuroinfection, or malignancy; or (iii) clinical and laboratory evidence of renal or hepatic dysfunction was present. The control group consisted of 196 clinically normal, age‐ and gender‐matched subjects (86 men, 110 women, mean age 33.9 ± 9.6 years), from the same south Indian region as the cases. Previous thrombosis was excluded by clinical history. As puerperal CVT was diagnosed in 44 cases, we included as controls 38 healthy women who were in the same puerperal period. The baseline demographic data, history of conventional vascular risk factors and family history of vascular events were recorded. Women were considered to be on oral contraceptives if they had taken them until a week or less before the thrombotic event for patients, or at the time of blood collection for controls. Puerperal CVT was diagnosed when CVT occurred during the first 4 weeks after childbirth. All subjects underwent detailed neurologic examination, and routine hematologic and biochemical profiling. DNA was extracted from peripheral blood leukocytes, and genotyping for the F12 C46T polymorphism carried out by amplification of a 142‐bp fragment followed by BsaHI digestion [12Zito F. Drummond F. Bujac S.R. Esnouf M.P. Morrissey J.H. Humphries S.E. Miller G.J. Epidemiological and genetic associations of activated factor XII concentration with factor VII activity, fibrinopeptide A concentration, and risk of coronary heart disease in men.Circulation. 2000; 102: 2058-62Crossref PubMed Scopus (75) Google Scholar]. Statistical analysis was performed with IBM spss statistics 17.0 (IBM Corporation, NY, USA) and Graph Pad Prism version 5.0.1 (Graph Pad Software, La Jolla, CA, USA). Differences in baseline characteristics between patients and healthy controls were assessed by the chi‐squared test for categorical variables and the t‐test for continuous parameters. The frequencies of the alleles and genotypes were compared between patient and control groups with the chi‐squared test. Hardy–Weinberg equilibrium (HWE) for each genotype polymorphism was determined for control subjects with the chi‐squared test. Univariate odds ratios (ORs) and 95% confidence intervals (CIs) were estimated, and a multiple logistic regression analysis was used to adjust ORs for age and sex. The combined effect of F12 46TT genotype and oral contraceptive use was determined by the generation of a contingency table for logistic regression analysis. Because use of oral contraceptives is incompatible with being pregnant or in the postpartum state, the proportion of women taking oral contraceptives was calculated after exclusion of pregnant or postpartum women. P‐values of < 0.05 were considered to be statistically significant. CVT patients did not differ significantly from controls with respect to smoking (13.3% vs. 14.6%, P = 0.77) and alcohol consumption (11.2% vs. 12.2%, P = 0.76). However, oral contraceptive use (7.28% vs. 1.3%, P = 0.001) was more prevalent in patients than in controls. The common symptoms observed in CVT patients were headache (91.8%), seizures (63.7%), altered sensorium (58.2%), and limb weakness (33.8%). Routine investigations and vasculitis workup were negative in all cases. FV Leiden was present in six cases and four controls only. The prothrombin gene G20210A polymorphism was not detected in any subject. Genotype frequencies of F12 C46T were in HWE. Genotype and allele frequency distributions are presented in Table 1. The differential distribution of F12 C46T genotypes between patients and controls was statistically significant. However, the T allele frequency was not significantly different. When the two groups were compared by use of a recessive model (CC + CT/TT), the TT genotype was associated with an increased risk of CVT (adjusted OR 2.89, 95% CI 1.61–5.19). A higher prevalence of the TT genotype was observed in patients aged ≤ 45 years than in those aged > 45 years (96% vs. 4%, P < 0.0001). The OR for CVT increased eight‐fold (OR 8.34, 95% CI 0.96–72.35) in women with the TT genotype who used oral contraceptives (Table 2).Table 1F12 C46T genotype and allele distribution with odds ratio (OR) F12 C46TControls (n = 196)Cases (n = 206) P‐valueAlleleT (frequency)0.570.690.329GenotypesCC, n (%)47 (23.9)43 (20.1)0.844CT, n (%)75 (38.1)42 (20.3)0.002TT, n (%)74 (38.0)121 (58.7)0.003OR at 95% CICC + CT vs. TT genotype2.89 (1.61–5.19)*<0.001CI, confidence interval. *Adjusted OR at 95% CI for age and gender. Open table in a new tab Table 2Combined risk associated with F12 TT genotype and oral contraceptive use in womenCombinationCVT n = 80Controls n = 56Odds ratio at 95% CI P‐valueNo. (%)No. (%)No 46TT, no OC26 (32.5)31 (55.3)1 (reference)–No 46TT, OC8 (10.0)2 (3.5)4.76 (0.92–24.47)0.0846TT, no OC39 (48.7)22 (39.3)2.11 (1.01–4.42)0.0646TT, OC7 (8.7)1 (1.8)8.34 (0.96–72.35)0.02*CI, confidence interval; CVT, cerebral veno‐sinus thrombosis; OC, oral contraceptive use. *Significant. Open table in a new tab CI, confidence interval. *Adjusted OR at 95% CI for age and gender. CI, confidence interval; CVT, cerebral veno‐sinus thrombosis; OC, oral contraceptive use. *Significant. Kanaji et al. [8Kanaji T. Okamura T. Osaki K. Kuroiwa M. Shimoda K. Hamasaki N. Niho Y. A common genetic polymorphism (46 C to T substitution) in the 5′‐untranslated region of the coagulation factor XII gene is associated with low translation efficiency and decrease in plasma factor XII level.Blood. 1998; 91: 2010-14Crossref PubMed Google Scholar] have reported that the T allele is relatively frequent in Orientals (0.80/0.20) as compared with Caucasians (0.27/0.73). Other studies have found a wide variation in the frequency, ranging from 0.18 in the Spanish population [13Santamaria A. Martinez‐Rubio A. Mateo J. Tirado I. Soria J.M. Fontcuberta J. Homozygosity of the T allele of the 46 C‐‐>T polymorphism in the F12 gene is a risk factor for acute coronary artery disease in the Spanish population.Haematologica. 2004; 89: 878-9PubMed Google Scholar] to 0.67 among Japanese [14Oguchi S. Ito D. Murata M. Yoshida T. Tanahashi N. Fukuuchi Y. Ikeda Y. Watanabe K. Genotype distribution of the 46C/T polymorphism of coagulation factor XII in the Japanese population: absence of its association with ischemic cerebrovascular disease.Thromb Haemost. 2000; 83: 178-9Crossref PubMed Scopus (30) Google Scholar]. In our population, the T allele frequency was 0.57, which is similar to that in Japanese. Although there are limited studies in CVT, the role of F12 C46T has been extensively studied in peripheral venous thrombosis. In a French cohort of 32 463 pregnant women (the Nimes Obstetricians and Hematologist cohort of first intended pregnancies study), the 46TT genotype was strongly predictive for peripheral venous thrombosis, with a relative risk of 5.99 [15Cochery‐Nouvellon E. Mercier E. Lissalde‐Lavigne G. Daures J.P. Quere I. Dauzat M. Mares P. Gris J.C. Homozygosity for the C46T polymorphism of the F12 gene is a risk factor for venous thrombosis during the first pregnancy.J Thromb Haemost. 2007; 5: 700-7Crossref PubMed Scopus (28) Google Scholar]. In contrast, in the Leiden Thrombophilia Study, Bertina et al. [10Bertina R.M. Poort S.R. Vos H.L. Rosendaal F.R. The 46C‐‐>T polymorphism in the factor XII gene (F12) and the risk of venous thrombosis.J Thromb Haemost. 2005; 3: 597-9Crossref PubMed Scopus (25) Google Scholar] found no association of the 46TT genotype or T allele with venous thrombosis. A meta‐analysis of evidence from observational studies has shown a weak association between F12 C46T and venous thromboembolism [16Johnson C.Y. Tuite A. Morange P.E. Tregouet D.A. Gagnon F. The factor XII ‐4C>T variant and risk of common thrombotic disorders: A HuGE review and meta‐analysis of evidence from observational studies.Am J Epidemiol. 2011; 173: 136-44Crossref PubMed Scopus (20) Google Scholar]. Differences in study design, patient selection criteria and the number and ethnicity of the subjects could account for these discrepancies. Moreover, variables predisposing to thrombosis in one venous territory may differ from those in another. Our study showed that the 46TT genotype was a strong, independent risk factor for CVT in the south Indian population, conferring a 2.9‐fold increase in risk. Previous studies have shown not only a very high relative risk of CVT in users of oral contraceptives, but an even stronger effect of contraceptives among women who carry hereditary prothrombotic conditions [17Calderwood C.J. Greer I.A. The role of factor V Leiden in maternal health and the outcome of pregnancy.Curr Drug Targets. 2005; 6: 567-76Crossref PubMed Scopus (18) Google Scholar]. In this study, the combined effect of F12 46TT and the use of oral contraceptives increased the risk of CVT eight‐fold. As the prevalence of F12 C46T is high in Indians, this finding raises the question of whether screening should be carried out in women before the prescription of oral contraceptives. The feasibility of this measure needs to be examined in a larger study. We conclude that the F12 46TT genotype is strongly associated with CVT in the south Indian population. The magnitude of risk is substantially increased in women who use oral contraceptives. Further studies in other populations are warranted before inclusion of the F12 C46T gene polymorphism in the routine diagnostic workup of CVT can be considered. The authors state that they have no conflict of interest.