Prothrombin fragment 1+2 and thrombin–antithrombin complex levels in patients with inherited APC resistance due to factor V Leiden mutation

Inherited activated protein C (APC) resistance is a newly described pathological condition associated with familial thrombophilia. A recent report on a family with APC resistance showed increased levels of prothrombin fragment 1+2 (F1+2) in the affected individuals. No data concerning thrombin–antithrombin complex (TAT) levels in patients with inherited APC resistance are presently available. The aim of this study was to assess the plasma levels of F1+2 and TAT in patients with inherited APC resistance due to factor V (F.V) Leiden mutation and to evaluate F1+2 and TAT levels in symptomatic and asymptomatic patients with the defect (‘carriers’) as compared to their family members having no evidence of F.V Leiden mutation (‘non-carriers’). One hundred and twenty-nine individuals belonging to 30 families with inherited APC resistance due to F.V Leiden mutation were studied. F1+2 and TAT levels were determined using two commercially available ELISA kits and cut-off values were defined as the higher limits of normal ranges obtained in healthy volunteers. Out of the 129 family members investigated, 36 were non-carriers, 85 were heterozygous and eight homozygous for F.V Leiden mutation. Thrombosis had occurred in 2/36 (6%) non-carriers, in 36/85 (42.3%) heterozygous and in 5/8 (63%) homozygous. Median levels of F1+2 and TAT were above cut-off values in carriers, whereas they were below in non-carriers. An overall percentage of 68.8% of carriers exhibited F1+2 levels above the cut-off value as compared to 38.9% of non-carriers. For TAT, an overall percentage of 63.4% of carriers presented with levels above the cut-off compared with 28% of non-carriers. In conclusion, patients with inherited F.V Leiden mutation may exhibit increased levels of F1+2 and TAT. There are no differences in F1+2 and TAT median levels among symptomatic and asymptomatic carriers. The percentage of carriers of F.V Leiden with levels of F1+2 and TAT above cut-off appears to be higher than that found in other clotting inhibitors defects and in this respect the defect might be considered different. However, these findings and the presence of a high percentage of non-carriers presenting with increased F1+2 and TAT levels may suggest the possible coexistence in these families of other unknown defects predisposing to thrombosis.     

Comparative evaluation of five different methods for the measurement of plasma factor II levels in carriers of the 20210A prothrombin variant.

The common genetic G to A variation of the prothrombin gene is associated with elevated levels of prothrombin [factor II (FII)] and is recognized as a risk factor for thrombosis. To determine whether one type of assay for plasma FII measurement was more efficient than other assays in displaying high FII levels in 20210A carriers, we compared five methods of measuring FII levels [i.e. an enzyme-linked immunosorbent assay (ELISA), a standard clotting assay, and three chromogenic methods using three different activators: Ecarin, Oxyuranus, and Textarin] in 30 G20210A patients and 30 G20210G controls. Plasma concentrations of factor X and factor VII + factor X were also determined by a clotting procedure. Functional assays were found to be equally efficient in demonstrating significantly higher FII levels in 20210A carriers than in non-carriers (P < 0.0001). With ELISA, the difference observed was less significant (P < 0.005). The specificity of every assay increased with FII cut-off levels; when a cut-off of 115% was applied, sensitivities of functional assays were between 73 and 93%, while sensitivities of ELISA declined dramatically to 33%. FII/factor X and FII/factor VII + factor X ratios were significantly higher in 20210A carriers (P < 0.0001). In conclusion, functional assays are preferentially required for measurements of FII levels in carriers of the 20210A variant.     

No effect of the prothrombin G20210A mutation on protein C activation in a large kindred with type I protein C deficiency.

Previously, we observed a positive association of prothrombin concentrations with thrombin generation (fragment 1 + 2) and thrombin activity (fibrinopeptide A), but no association with protein C activation peptide levels. We further evaluated a potential beneficial effect of increased prothrombin concentrations on activated protein C generation by assessing the plasma concentration of activated protein C in complex with protein C inhibitor (APC-PCI). Blood samples were used from 195 family members of a large French-Canadian kindred with type I protein C deficiency due to a 3363C insertion in the protein C gene. We utilized a new and highly sensitive assay for measuring the concentration of APC-PCI complex as a measure of the level of activation of protein C. Means of the plasma concentrations of the APC-PCI complex were compared among carriers and non-carriers of the prothrombin G20210A mutation. Protein C activity levels were positively associated with APC-PCI complex plasma concentrations; however, APC-PCI complex levels were not different for carriers of the prothrombin G20210A mutation than for non-carriers. Thus, carriers of the prothrombin G20210A mutation do not have increased protein C activation despite the increased thrombin generation resulting from the higher prothrombin concentrations associated with the G20210A mutation.     

Additional genetic risk factors for venous thromboembolismin carriers of the factor V Leiden mutation

Only a minority of subjects with factor V (FV) Leiden mutation develop venous thromboembolism (VTE), suggesting that additional genetic risk factors may be present in symptomatic carriers. We screened 157 unrelated carriers of the FV Leiden mutation with a first episode of VTE and 291 unrelated asymptomatic FV carriers for the presence of two frequent mutations, i.e. G20210A of the prothrombin gene and C677T of the methylenetetrahydrofolate reductase gene. Carriers with other inherited or acquired thrombophilia-associated abnormalities were excluded from analysis. Heterozygotes for the G20210A mutation were more prevalent among symptomatic carriers than in asymptomatic carriers (10.8% v 2.7%, P  < 0.0001); homozygotes for the C677T mutation were also more prevalent in symptomatic carriers (21.6% v 14.4%, P  = 0.05). Factor V Leiden carriers who had had a VTE episode during oral contraceptive intake were more frequently carriers of the G20210A mutation (14.3%, P  = 0.03). These results further support the idea that VTE in carriers of FV Leiden results from interaction with additional genetic or circumstantial risk factors, and that an accurate search for such factors is required to identify carriers at risk.     

Clinical and laboratory expression of associated thrombophilic conditions (homozygous/heterozygous factor V Leiden mutation and heterozygous prothrombin variant 20210A) in an Italian family.

Prothrombin variant 20210A is maintained to be a mild risk factor for venous thromboembolism (VTE). The association of this defect with other inherited thrombophilic conditions may result in an increased risk of developing VTE. In this article, a family is described in which prothrombin variant was associated with either homozygous or heterozygous factor V Leiden (FV Leiden) mutation. All family members except the proband were asymptomatic despite the presence and the severity of the underlying genetic defect(s). The proband, who carried homozygous FV Leiden mutation and heterozygous prothrombin variant, experienced recurrent VTE during pregnancies, whereas one brother, with the same defect, was asymptomatic. Mean prothrombin antigen and activity levels were higher in carriers of the prothrombin variant as compared with noncarriers. Thrombin generation was assessed in family members, in carriers of prothrombin variant or homozygous FV Leiden mutation and in a control group. Most of the family members presented with increased prothrombin fragment 1+2 levels possibly because of the presence of the FV Leiden mutation. Although it is conceivable that the co-inheritance of prothrombin variant and FV Leiden mutation may increase the risk of VTE, patients with these combined defects may remain asymptomatic. It is likely that acquired triggering conditions play a major role in determining VTE in carriers of a mild genetic predisposition. This has to be taken into account when recommendation for thromboprophylaxis is given.     

The 20210 G-->A mutation in the 3'-untranslated region of the prothrombin gene and the risk for arterial thrombotic disease

A sequence variation in the 3'-untranslated region of the prothrombin (PT) gene (20210 G-->A) was recently claimed to be associated with elevated plasma prothrombin levels and an increased risk for venous and arterial thrombosis. We examined the prevalence of the 20210 A allele in the prothrombin gene in 400 healthy controls and in 263 patients with proven premature atherosclerotic disease. In addition, we measured prothrombin, prothrombin fragment 1 + 2, thrombin-antithrombin (TAT) complex and D-dimer levels in plasma from carrier and non-carrier patients. The frequency of the variant allele was 1% in the control subjects and 2.7% in the patient group, yielding a relative risk (RR) for the 20210 A allele of 2.7 (95% CI 0.8-9.4). All heterozygotes in the patient group were found to have had a myocardial infarction (MI), yielding a RR for MI of 4.2 (95% CI 1.2-14.6). Plasma prothrombin levels in carriers (126+/-10) were higher than in non-carriers (103+/-1, P=0.02). The levels of TAT complexes (16+/-9 v 6+/-1 microg/ml, P=0.02) as well as of prothrombin fragment 1 + 2 (1.5+/-0.3 v 1.0+/-0.1 nmol/l, P=0.02) were also elevated in carriers of the mutation. Our findings suggest that the 20210 G-->A mutation in the prothrombin gene is a genetic risk factor for MI. In addition, our data provide evidence for an association of the mutation with excessive thrombin generation, which may contribute to the understanding of its role in venous and arterial disease.     

G20210A Prothrombin gene variant in Turkish patients with angiographically documented coronary artery disease

Background G→A substitution at position 20210A in the 3′-untranslated region in prothrombin gene is associated with raised plasma prothrombin. G20210A prothrombin variant has been associated with high prothrombin levels and an increased risk of venous thrombosis. To determine the prevalence of the G20210A prothrombin variant among Turkish and to evaluate the potential relevance of this variant to Turkish patients with angiographically documented coronary artery disease (CAD). Methods We conducted a case-control study on 268 unrelated subjects who referred to the cardiology department of the university hospital for coronary angiography. One hundred-thirty seven patients with angiographically documented CAD and 131 subjects without angiographically documented CAD were studied to examine the association of the G20210A prothrombin variant with CAD. Blood samples from the patients and controls were analyzed for the G20210A prothrombin variant by DNA analysis, using polimeraz chain reaction. Results G20210A prothrombin variant was found in 6 of 137 (4.4%) patients with CAD and 1 of 131 (0.8%) in control subjects (p = 0.064). There were no significant differences in terms of diabetes mellitus, hypertension, dyslipidemia, sex and family history of CAD with and without the G20210A prothrombin variant in the patient group. Conclusions In agreement with the results of recent meta-analyses, our data from northeast Anatolia show a 5-fold higher prevalence of the G20210A prothrombin variant among the patients with angiographically-documented CAD when compared with those without angiographic signs of significant CAD and its variant cannot be considered as a risk factor for CAD in this region.     

The risk of venous thromboembolism in family members with mutations in the genes of factor V or prothrombin or both

Factor V Leiden and the G20210A mutation in the prothrombin gene are the most frequent abnormalities associated with venous thromboembolism. It is unknown whether the risks due to the presence of either mutation are of the same magnitude. We compared the prevalence and incidence rate of venous thromboembolism in relatives with either mutation or both. The finding of different rates might influence the strategies for primary prevention of thrombosis in carriers of these mutations. The study population included 1076 relatives of probands with the prothrombin gene mutation, factor V Leiden or both who underwent screening for inherited thrombophilia and were found to be carriers of single mutations or double mutations or who were non-carriers. The prevalence of venous thromboembolism was 5.7% in relatives with the prothrombin gene mutation, 7.8% in those with factor V Leiden, 17.1% in those with both mutations and 2.5% in non-carriers. Annual incidences of thrombosis were 0.13% [95% confidence interval (CI) 0.06-0.24], 0.19% (0.13-0.25), 0.42% (0.15-0.83) and 0.066% (0.03-0.11), respectively, and the relative risk of thrombosis was two times higher in carriers of the prothrombin gene mutation, three times higher in those with factor V Leiden and six times higher in double carriers than in non-carriers. The incidence of venous thromboembolism in carriers of the prothrombin gene mutation is slightly lower than that observed in carriers of factor V Leiden, whereas in carriers of both mutations it is two or three times higher. These findings suggest that lifelong primary anticoagulant prophylaxis of venous thromboembolism is not needed in asymptomatic carriers of single or double mutations. Anticoagulant prophylaxis seems to be indicated only when transient risk factors for thrombosis coexist with mutations.     

Linkage analysis demonstrates that the prothrombin G20210A mutation jointly influences plasma prothrombin levels and risk of thrombosis

Association studies suggest that the G20210A mutation (G to A substitution at nucleotide position 20210) in the prothrombin gene (PT) is associated with increased plasma prothrombin activity and with increased risk for venous thromboembolism. To test directly for linkage between this PT variant and plasma prothrombin activity we performed a family-based study. The G20210A genotypes and plasma prothrombin activity levels were determined in 435 individuals belonging to 22 extended Spanish families. The sample was composed of 388 homozygous (G/G) normal individuals and 43 heterozygote (G/A) and 4 homozygote (A/A) carriers for the G20210A mutation. The results of variance-component linkage analysis yielded a highly significant lod score of 3.6 (P = 2.4 x 10(-5)) between this mutation and a quantitative trait locus (QTL) that influences prothrombin activity. Importantly, a conditional linkage analysis that simultaneously accounted for association with the G20210A variant completely eliminated the linkage signal, which indicates that this mutation affects the function of the prothrombin gene. Additionally, a bivariate linkage analysis of plasma prothrombin activity and thrombosis significantly improved the linkage signal for prothrombin activity (lod score = 4.7; P = 1.5 x 10(-6)) and provided strong evidence that this QTL has a pleiotropic effect on the risk of thrombosis (lod score = 2.43; P =.0004). These results represent the first direct genetic evidence that a QTL in the PT gene influences prothrombin activity levels and susceptibility to thrombosis and strongly support the conclusion that G20210A is a functional polymorphism. (Blood. 2000;95:2780-2785)     

Familial thrombophilia and the prothrombin 20210A mutation: association with increased thrombin generation and unusual thrombosis.

The 20210A prothrombin mutation has recently been associated with an increased risk of venous thrombosis, but the mechanism of the increased thrombotic risk in affected persons has not been elucidated. We report on a thrombophilic family in which the proband presented with cerebral vein thrombosis and homozygosity for the 20210A prothrombin mutation as her only identifiable risk factor for venous thrombosis. Extended genotyping of family members revealed seven other affected, but asymptomatic, first-degree relatives (one A/A homozygote and six G/A heterozygotes). Plasma levels of prothrombin, prothrombin fragments 1 + 2 and thrombin-antithrombin complexes were highest in A/A homozygotes, intermediate in G/A heterozygotes and lowest in those with the G/G homozygous normal genotype, while D-dimer levels were elevated only in A/A homozygotes. Our results suggest that the 20210A prothrombin mutation is associated with activation of coagulation and increased thrombin generation, not only in patients with a past history of thrombosis but also in otherwise healthy asymptomatic persons. In a similar fashion to the homozygous factor V Leiden mutation, patients with the homozygous 20210A prothrombin mutation could be at highest risk of thrombosis, as suggested by our patient who presented with unusual thrombosis.     

20210G/A mutation of prothrombin gene in a patient with deep venous thrombosis ad pulmonary embolism without other risk factors of thrombosis

A new genetic anomaly predisposing to venous thrombosis was described in 1996, namely the transition of guanine (G) to adenine (A) at position 20210 in the 3-untranslated region of the prothrombin gene. This mutation is associated with high levels of plasma prothrombin and increased risk of thrombotic events in the venous system. We report the case of a man who, lacking known risk factors for thrombosis, suffered a massive pulmonary embolism and deep venous thrombosis in both lower legs. Thrombophilic analysis confirmed that the patient and close relatives were carriers of the heterozygotic 20210G/A variant of the prothrombin gene. Two relatives with the genetic defect had also suffered some type of deep venous thrombosis.     

Factor V Leiden: the venous thrombotic risk in thrombophilic families

Factor V Leiden (FVL) leads to a sevenfold increased risk of venous thrombosis and is present in 50% of individuals from families referred because of unexplained familial thrombophilia. We assessed the association of FVL with venous thromboembolism (VTE) in 12 thrombophilic families of symptomatic probands with FVL in a retrospective follow-up study. We screened 182 first- and second-degree relatives of the 12 unrelated propositi for the FVL mutation and the occurrence of VTE. The incidence rate of VTE in carriers of FVL (0.56%/year) was about six times the incidence for the Dutch population (0.1%/year). The incidence rate in non-carriers also appeared to be higher (0.15% per year). At the age of 50 years, the probability of not being affected by VTE was reduced to 75% for carriers and to 93% for non-carriers (P = 0.009). Identification of carriers of FV Leiden may be worthwhile in young symptomatic individuals and their relatives with a strong positive family history of venous thromboembolism or a history of recurrent venous thrombosis who may be at risk (e.g. pregnancy, use of oral contraceptives). After adjustment for prothrombin G20210A (present in two families), even higher thrombotic incidence rates were found in carriers and non-carriers of FVL. This makes the presence of other unknown prothrombotic risk factors more probable in these families.     

Risk of venous thrombosis in carriers of the prothrombin G20210A variant and factor V Leiden and their interaction with oral contraceptives

BACKGROUND AND OBJECTIVES: The prothrombin G20210A mutation and factor V Leiden have been found to be associated with an increased risk of venous thrombosis, but the reported prevalences of the prothrombin gene variant both in the normal population and in patients with deep venous thrombosis (DVT) vary greatly in the literature. Moreover, the influence of oral contraceptives (OC) on thrombotic events in patients with the prothrombin G20210A variant has not been well established. In this study we evaluate both circumstances. DESIGN AND METHODS: A case-control study was run on 229 patients with DVT and 246 healthy controls. The patients' history of thrombosis and acquired thrombotic risk factors, especially OC, were recorded. Prothrombin G20210A mutation, factor V Leiden, antithrombin, heparin II cofactor, plasminogen and proteins C and S were evaluated. RESULTS: Seven and a half percent of the patients and 2.9% of the controls were carriers of the prothrombin mutation, while 12.2% of the patients and 1.6% of the controls had factor V Leiden. Among the 229 DVT patients there were 130 patients with clinically suspected thrombophilia (first thrombotic event occurring before the age of 45 years or positive family history of thrombosis or recurrent venous thrombosis). Ten percent of these 130 patients were carriers of the prothrombin G20210A mutation and 18.5% had the factor V Leiden mutation. The odds ratios (OR) for DVT risk were: 2.4 (95% CI, 1.0-6.3) for the total DVT patients and 5.2 (95% CI, 1.4-19.5) for the patients with clinically suspected thrombophilia with the prothrombin mutation. The risk of thrombosis was 6.9 (95% CI, 2.3-20.6) for the DVT patients and 14.3 (95% CI, 3.3-64.6) for the patients with clinically suspected thrombophilia with factor V Leiden. Fifty-five percent of the patients with combined congenital defects (prothrombin mutation G20210A plus another congenital defect) had recurrent thrombosis. In women receiving OC the risk of DVT was 3.5 (95% CI, 1.5-8.2) that of the patients not receiving OC. When women with combined defects were also taking OC, the risk of thrombosis increased significantly. INTERPRETATION AND CONCLUSIONS: The prevalence of the prothrombin G20210A mutation in the healthy population in our study is similar to that observed in other southern European countries. The prothrombin G20210A mutation does not by itself seem to be a high thrombotic risk factor. However, when it is present together with other thrombotic risk factors, the predicted risk of thrombotic events increases. The use of OC by women with the prothrombin G20210A variant or FV Leiden, either alone or combined with other thrombotic risk factors, was associated with a significant increase in the risk of venous thrombosis.     

Contribution of the glycoprotein Ia 807TT, methylene tetrahydrofolate reductase 677TT and prothrombin 20210GA genotypes to prothrombotic risk among factor V 1691GA (Leiden) carriers

The cooperative effects of the GPIa 807TT, MTHFR 677TT and prothrombin 20210GA genotypes with the FV Leiden 1691GA (FVL) genotype were evaluated by comparing these genotype frequencies in 77 asymptomatic and 156 symptomatic heterozygous FVL carriers. The GPIa 807TT and MTHFR 677TT genotypes did not segregate within the symptomatic FVL carrier group and did not contribute to venous thrombotic risk in this patient cohort. There was no difference in the prothrombin 20210GA genotype frequency between asymptomatic FVL carriers and a random Caucasian control group; however, the prothrombin 20210GA genotype was nearly 5 times as prevalent (19/156 v 2/77; P < 0.02) in the symptomatic FVL carriers (odds ratio 5.21; 95% confidence interval 1.20-47.62), demonstrating that this important prothrombotic risk factor acts synergistically with FVL.     

Impact of the factor II: G20210A variant on the risk of venous thromboembolism in relatives from families with the factor V: R506Q mutation

OBJECTIVES: Factor V: R506Q mutation and the prothrombin G20210A variant (factor II: G20210A variant) are associated with an increased risk of venous thromboembolism (VTE). In cohorts of unrelated patients a cosegregation of both mutations has been shown to be associated with an increased risk of developing VTE. The aim of this study was to investigate the impact of the coinheritance of both mutations on the risk of VTE in relatives of symptomatic carriers of the factor V: R506Q mutation and the factor II: G20210A variant. PATIENTS AND METHODS: Four families with 48 family members were investigated for the presence of the factor V: R506Q mutation and the factor II: G20210A mutation, and their clinical history was evaluated. RESULTS: VTE was more frequent in family members with a combined defect (3/10; 30%) compared to those with a single mutation (1/16; 6%) or without a defect (1/12; 8%). The probability for VTE for 40-yr-old individuals with both mutations, a single mutation and no mutation was 56%, 12% and 20%, respectively. CONCLUSIONS: These data suggest that the G to A transition at position 20210 of the prothrombin gene leads to an increase in the risk of VTE in carriers of the factor V: R506Q mutation. The determination of the factor II: G20210A variant in index patients carrying a factor V: R506Q mutation and, if present, in family members may help to identify individuals who are at high risk for VTE.     

Arterial wall thickness and the risk of recurrent ischemic events in carriers of the prothrombin G20210A mutation with clinical manifestations of atherosclerosis

The G20210A mutation in the prothrombin gene is an established risk factor for venous thrombosis. There is controversy about the role of this mutation in arterial thrombotic disease and atherosclerosis. We determined the presence of the prothrombin mutation and examined its influence on carotid and femoral artery intima-media thickness (IMT) and the occurrence of new ischemic events during follow-up in 277 patients with clinically manifest atherosclerotic disease: ischemic stroke, myocardial infarction or peripheral arterial disease. The mean age at entry was 63 years. Mean IMT was significantly higher in carriers of the prothrombin mutation (1.17 (SD 0.29) mm versus 0.97 (SD 0.25) mm: (delta)IMT=0.20, P=0.02). The increase in IMT was not attributable to differences in age, type of arterial disease or cardiovascular risk factors between carriers and non-carriers. During a mean follow-up of 3.5 years, a strong trend for more ischemic events was observed: 4 of the 11 carriers suffered from a recurrent ischemic event, compared with 30 of the 164 male non-carriers (36 versus 18%; P=0.06). These results suggest that the G20210A mutation contributes to the process of arterial wall thickening and is associated with the occurrence of ischemic events in a cohort of elderly persons with established atherosclerosis.     

Prothrombin A19911G and G20210A polymorphisms' role in thrombosis

The prothrombin G20210A polymorphism, which correlates with the plasmatic prothombin levels, is the second genetic risk factor for deep venous thrombosis (DVT), although its prothrombotic role is mild. Recently, the prothrombin A19911G polymorphism, also associated with slight variations of the prothrombin level, has been suggested to modulate the thrombotic risk of the G20210A polymorphism in a preliminary study including few patients and controls. Our study evaluated the effect of the A19911G polymorphism in the arterial and venous thrombotic risk of the prothrombin 20210G/A genotype, analysing 204 consecutive DVT patients and 204 matched controls. Moreover, we analysed 213 carriers of the 20210G/A genotype (152 with DVT, 26 with arterial thrombosis and 35 healthy subjects) and 10 homozygous 20210 A/A carriers. We developed a simple method to simultaneously determine the genotype of both polymorphisms. In accordance with our case/control study, the A19911G polymorphism did not play a significant role in the development of DVT. Analysis of 120 20210 A alleles demonstrated a complete linkage disequilibrium with the 19911 A allele. These polymorphisms (alone or combined) did not modify the risk of arterial thrombosis. However, the 19911A/G genotype slightly increased the risk of developing DVT in carriers of the 20210G/A genotype (OR 3.34 vs 5.86), supporting that the prothrombin 19911 polymorphism could modulate the risk of the G20210A polymorphism in developing DVT.     

Hyperprothrombinemia associated with prothrombin G20210A mutation inhibits plasma fibrinolysis through a TAFI-mediated mechanism

The prothrombin gene mutation G20210A is a common risk factor for thrombosis and is associated with increased prothrombin levels. However, the mechanism whereby hyperprothrombinemia predisposes to thrombosis remains unclear. Because thrombin is the physiologic activator of TAFI (thrombin activatable fibrinolysis inhibitor), the precursor of an antifibrinolytic carboxypeptidase (TAFIa), we evaluated the influence of hyperprothrombinemia on fibrinolysis. Thirty-two heterozygous carriers of the G20210A mutation and 30 noncarriers were studied. Plasma fibrinolytic factors and TAFI levels were similar in the 2 groups. Mean lysis time of tissue factor-induced plasma clots exposed to 25 ng/mL exogenous tissue-type plasminogen activator (t-PA) was significantly longer in 20210A carriers than in control donors. This difference disappeared on addition of a specific inhibitor of TAFIa. Determination of thrombin and TAFIa activity, generated during clot lysis, revealed that G20210A mutation was associated with a significant enhancement of late thrombin formation and an increase in TAFI activation. Plasma prothrombin level was highly significantly correlated with both clot lysis time and TAFI activation. The addition of purified prothrombin, but not of factors X or VIIa, to normal plasma caused a concentration-dependent, TAFI-mediated inhibition of fibrinolysis. These findings provide a new mechanism that might contribute to the thrombotic risk in prothrombin 20210A carriers.     

The prothrombin gene G20210A variant: prevalence in a U.K. anticoagulant clinic population

We have investigated the prevalence of a recently reported genetic variation in the prothrombin gene (G20210A) in patients with an objectively confirmed history of venous thrombosis. 12/219 patients (5.5%) were found to be heterozygous carriers of the 20210 A allele. The incidence of the 20210 A allele in a group of 164 healthy controls was 1.2% (allele frequency 0.61%, 95% CI 0.08–2.19). When patients with a known alternative hereditary risk factor for venous thrombosis (factor V Leiden mutation or deficiency of antithrombin, protein C or protein S) were excluded, the G20210A variant was found to increase the risk for venous thrombosis by approximately 5-fold (odds ratio 5.4, 95% CI 1.16–25.0). This prothrombin gene sequence variation adds further to the list of recognized genetic risk factors for thrombophilia.     

Prevalence of mild hyperhomocysteinaemia and association with thrombophilic genotypes (factor V Leiden and prothrombin G20210A) in Italian patients with venous thromboembolic disease

Mild hyperhomocysteinaemia is an established risk factor for deep vein thrombosis (DVT); few data concerning its potential interaction with thrombophilic genotypes are available at the present time. We investigated 121 thrombosis-free individuals and 111 patients with at least one objectively confirmed episode of DVT. A thrombophilic condition (deficiency in antithrombin, protein C and S, factor V Leiden, prothrombin G20210A) was detected in 25.2% of the patients; mutant factor V or prothrombin genotypes were present in 6.6% of the controls. Hyperhomocysteinaemia was found in 14.4% of patients and 3. 3% of the controls, with a 3.7-fold increase in risk for DVT (95% CI 1.1-12.3). Adoption of different cut-off levels for definition of hyperhomocysteinaemia did not substantially change the magnitude of the risk. Carriership of both hyperhomocysteinaemia and factor V Leiden or prothrombin G20210A was detected in 2.7% of patients for each combination and in none of the controls. An approximate estimate of 30-fold increased risk in carriers of both hyperhomocysteinaemia and factor V Leiden and 50-fold increased risk in carriers of both hyperhomocysteinaemia and prothrombin G20210A was calculated, suggesting a synergistic interaction between hyperhomocysteinaemia and such thrombophilic genotypes. Yet statistical analysis is highly unstable due to the small number of individuals with combined defects. Further investigations on large series of patients are needed.