Daniela Tormene

Impact of Inherited Thrombophilia on Venous Thromboembolism in Children A Systematic Review and Meta-Analysis of Observational Studies

Background— The aim of the present study was to estimate the impact of inherited thrombophilia (IT) on the risk of venous thromboembolism (VTE) onset and recurrence in children by a meta-analysis of published observational studies. Methods and Results— A systematic search of electronic databases (Medline, EMBASE, OVID, Web of Science, The Cochrane Library) for studies published from 1970 to 2007 was conducted using key words in combination as both MeSH terms and text words. Citations were independently screened by 2 authors, and those meeting the inclusion criteria defined a priori were retained. Data on year of publication, study design, country of origin, number of patients/controls, ethnicity, VTE type, and frequency of recurrence were abstracted. Heterogeneity across studies was evaluated, and summary odds ratios and 95% CIs were calculated with both fixed-effects and random-effects models. Thirty-five of 50 studies met inclusion criteria. No significant heterogeneity was discerned across studies. Although >70% of patients had at least 1 clinical risk factor for VTE, a statistically significant association with VTE onset was demonstrated for each IT trait evaluated (and for combined IT traits), with summary odds ratios ranging from 2.63 (95% CI, 1.61 to 4.29) for the factor II variant to 9.44 (95% CI, 3.34 to 26.66) for antithrombin deficiency. Furthermore, a significant association with recurrent VTE was found for all IT traits except the factor V variant and elevated lipoprotein(a). Conclusions— The present meta-analysis indicates that detection of IT is clinically meaningful in children with, or at risk for, VTE and underscores the importance of pediatric thrombophilia screening programs.

X-Linked Thrombophilia with a Mutant Factor IX (Factor IX Padua)

We report a case of juvenile thrombophilia associated with a substitution of leucine for arginine at position 338 (R338L) in the factor IX gene (factor IX-R338L). The level of the mutant factor IX protein in plasma was normal, but the clotting activity of factor IX from the proband was approximately eight times the normal level. In vitro, recombinant factor IX-R338L had a specific activity that was 5 to 10 times as high as that in the recombinant wild-type factor IX. The R338 substitution causes a gain-of-function mutation, resulting in factor IX that is hyperfunctional.

High vs. low doses of low-molecular-weight heparin for the treatment of superficial vein thrombosis of the legs: a double-blind, randomized trial.

Summary.  In contrast with extensive information on the management of deep vein thrombosis of the lower extremities, little is known on the most appropriate treatment of the superficial vein thrombosis (SVT). In a multicenter, prospective, controlled, double‐blind, double‐dummy clinical trial, 164 consecutive patients with acute SVT of the great saphenous vein were randomized to receive the s.c. administration of either fixed prophylactic doses (2850 a‐Xa IU) or body‐weight adjusted therapeutic doses of nadroparin once daily for 1 month. The main study outcome was to compare the rate of asymptomatic and symptomatic extension of SVT and/or venous thromboembolic (VTE) complications during a 3‐month follow‐up period. Of the 81 patients randomized to the prophylactic doses, seven [8.6%; 95% confidence interval (CI), 3.5–17.0] developed SVT progression or VTE complications as compared with six of the 83 (7.2%; 95% CI, 2.8–15.1) allocated to the treatment group (absolute difference, 1.4; 96% CI, −6.9 to 9.7; P = 0.74). No patient in either group developed major bleeding. Our findings suggest that therapeutic doses of low‐molecular‐weight heparin, administered for 1 month in patients with SVT of the greater saphenous vein do not improve results obtained by prophylactic doses, administered for the same period, over a 3‐month follow‐up period.

Megakaryocytes endocytose and subsequently modify human factor V in vivo to form the entire pool of a unique platelet‐derived cofactor

Summary.  Factor Va (FVa), derived from plasma or released from stimulated platelets, is the essential cofactor in thrombin production catalyzed by the prothrombinase complex. Plasma‐derived factor V (FV) is synthesized in the liver. The source(s) of the platelet‐derived cofactor remains in question. We identified a patient homozygous for the FVLeiden mutation, who received a liver transplant from a homozygous wild‐type FV donor. Eighteen days post‐transplant, phenotypic analysis of the patients platelet‐derived FV indicated that the platelets were acquiring wild‐type FV, consistent with the temporal differentiation of megakaryocytes and subsequent platelet production. Nine months post‐transplant, the platelet‐derived FV pool consisted entirely of wild‐type FV. Consequently, megakaryocyte endocytosis of plasma‐derived FV must account for the entire platelet‐derived pool, because blood‐borne platelets cannot bind or endocytose FV. Subsequent to this endocytic process, the patients platelet‐derived FV was cleaved to a partially active cofactor, and rendered resistant to phosphorylation catalyzed by a platelet‐associated kinase, and hence less susceptible to activated protein C‐catalyzed inactivation. These data provide the first in vivo demonstration of an endocytosed plasma protein undergoing intracellular modifications that alter its function. This process results in the sequestration of active FVa within the platelet compartment, poised for immediate action subsequent to release from platelets at a site of injury.

Prothrombin antigen levels in symptomatic and asymptomatic carriers of the 20210A prothrombin variant

A recently discovered variant in the prothrombin gene (20210A) has been found in approximately 5–10% of patients with venous thromboembolism. It has been shown that patients with this variant present with high levels of prothrombin in plasma and this is maintained to be the most likely mechanism by which the risk of thrombosis is increased. We have evaluated prothrombin antigen levels in 50 carriers of the 20210A allele and compared with non‐carriers. 327 subjects were subdivided according to deficiency status and previous thrombosis. 30 symptomatic and 20 asymptomatic carriers had increased mean prothrombin levels as compared to symptomatic (n = 178) or asymptomatic (n = 99) non‐carriers. The percentage of subjects with prothrombin levels above cut‐off values of 1.15 u/ml or 1.30 u/ml was significantly higher in carriers of the prothrombin variant as compared to non‐carriers, regardless of a previous thrombosis. However, among non‐carriers the percentage of those with prothrombin levels above cut‐off values was significantly higher in the group of symptomatic as compared to asymptomatic individuals. In conclusion, increased prothrombin antigen levels, as detected by a specific ELISA, were found among 50 symptomatic and asymptomatic carriers of the 20210A prothrombin variant as well as among a large group of symptomatic non‐carriers. The data are in agreement with those found by using functional tests for the determination of prothrombin levels in these patients.

Hereditary and acquired protein S deficiencies are associated with low TFPI levels in plasma

Summary.  Background: Protein S and tissue factor pathway inhibitor (TFPI) act together in down‐regulating coagulation. Objective: To investigate the TFPI/protein S system in hereditary and acquired protein S deficiency. Methods: Plasma antigen levels of protein S and full‐length TFPI were determined in heterozygous type I protein S‐deficient individuals (n = 35), patients on oral anticoagulant treatment (OAT) (n = 29), oral contraceptive (OC) users (n = 10) and matched controls. Thrombin generation was determined using calibrated automated thrombography. Results: Full‐length TFPI levels were lower in type I protein S‐deficient individuals (76.8 ± 33.8%) than in age‐ and sex‐matched controls (128.0 ± 59.4%, P < 0.001). Among protein S‐deficient individuals with thrombosis, those on OAT had not only lower total protein S levels (25.7 ± 8.2% vs. 54.7 ± 8.2%, P < 0.001), but also lower full‐length TFPI levels (52.6 ± 15.0% vs. 75.4 ± 22.9%, P = 0.009) than those not on OAT. Similarly, OC users had lower protein S (73.8 ± 11.5% vs. 87.9 ± 10.8%, P = 0.005) and full‐length TFPI levels (73.7 ± 27.7% vs. 106.4 ± 29.2%, P = 0.007) than non‐users. When triggered with tissue factor, plasma from protein S‐deficient individuals generated 3–5‐fold more thrombin than control plasma. The difference was only partially corrected by normalization of the protein S level, full correction requiring additional normalization of the TFPI level. Protein S‐immunodepletion experiments indicated that free protein S and full‐length TFPI form a complex in plasma, and the protein S/TFPI interaction was confirmed by surface plasmon resonance analysis. Conclusions: Full‐length TFPI binds to protein S in plasma and is reduced in genetic and acquired protein S deficiency. The concomitant TFPI deficiency substantially contributes to the hypercoagulable state associated with protein S deficiency.

The PAI-I gene 4G/5G Polymorphism and Deep Vein Thrombosis in Patients with Inherited Thrombophilia

Genetic and acquired factors may influence phenotypic expression of inherited thrombophilia. Hypofibrinolysis due to excess PAI-I can be found in patients with deep vein thrombosis (DVT) and 4G/5G polymorphism of the PAI-1 gene may modulate the inhibitors synthesis. In 149 patients with inherited thrombophilia, the possible thrombotic contribution of both 4G/5G polymorphism and PAI-1 plasma levels was evaluated. Sixty-seven patients with idiopathic DVT and 98 normal subjects were also studied. By comparison with controls, a significantly higher prevalence of 4G/4G genotype was seen in idiopathic DVT and in thrombophilia patients, although in this latter group the difference only remained significant in cases symptomatic for thrombosis (p=0.01). The 4G/4G genotype was associated with a greater risk of thrombosis both in symptomatic thrombophilia patients (OR 2.85, 95% CI 1.26-6.46) and in idiopathic DVT patients (OR 3.1, 95% CI 1.26-7.59). The greater frequency of 4G allele in symptomatic thrombophilia patients with respect to controls was statistically significant (p=0.04). Compared to healthy subjects, PAI-i:Ag levels were higher in symptomatic thrombophilia patients and related to the 4G/5G polymorphism, with significantly higher values in the 4G/4G carriers. In conclusion, PAI-1 4G/5G polymorphism may influence PAI-i expression and thrombotic risk in patients with inherited thrombophilia.

Screening for thrombophilia and antithrombotic prophylaxis in pregnancy: Guidelines of the Italian Society for Haemostasis and Thrombosis (SISET)

The term thrombophilia describes an increased tendency to develop thrombosis and many laboratory markers with different strengths of association with thrombosis have been identified. The main causes of maternal mortality and morbidity in developed countries is venous thromboembolism (VTE) and obstetric complications. During pregnancy and puerperium the risk for VTE increases due to hemostatic imbalance towards a prothrombotic state, and it is further increased in women carriers of thrombophilia; recent studies have also demonstrated an association between thrombophilia and obstetric complications. These complications are, therefore, considered potentially preventable with the prophylactic administration of anticoagulant drugs, although their efficacy is not proven by data from randomized controlled trials. After a systematic comprehensive literature review and using a rigorous methodology, the expert panel formulated recommendations regarding the usefulness of screening for thrombophilia in pregnancy to identify high-risk women and for the management of antithrombotic prophyalxis. When evidence is lacking, consensus-based recommendations are provided.

The risk of first venous thromboembolism during pregnancy and puerperium in double heterozygotes for factor V Leiden and prothrombin G20210A

Background: The risk of venous thromboembolism (VTE) during pregnancy in double heterozygous carriers of factor (F) V Leiden and prothrombin G20210A is not established. Hence, whether or not these women deserve antithrombotic prophylaxis when pregnant is unknown.Patients and methods: In the frame of a multicenter family study, 52 double heterozygous carriers of FV Leiden and prothrombin G20210A who had remained pregnant at least once before knowledge of thrombophilia, were retrospectively investigated with respect to the occurrence of first VTE during pregnancy and puerperium. They were compared with 104 heterozygous carriers of FV Leiden, 104 of prothrombin G20210A and 104 women without thrombophilia. Results: Double heterozygotes were similar to single heterozygous carriers and non‐carriers for the age at first pregnancy, age at testing and rate of full‐term pregnancies. No VTE during pregnancy was observed in the four groups of women, whereas in the puerperium it occurred in two double carriers (1.8% of pregnancies, 95% CI: 0.5–6.3), three single FV Leiden carriers (1.5%, 0.5–4.3), two single prothrombin G20210A carriers (1%, 0.2–3.6) and one non‐carrier (0.4%, 0–2.5).Conclusions: The risk of first VTE during pregnancy and puerperium in double heterozygous carriers of FV Leiden and prothrombin G20210A is low and similar to that of single carriers. As for single heterozygotes, antithrombotic prophylaxis in asymptomatic double heterozygous carriers appears to be justified only in puerperium.

Differential effects of high prothrombin levels on thrombin generation depending on the cause of the hyperprothrombinemia.

Summary.  Background: Hyperprothrombinemia, resulting from the prothrombin G20210A mutation or other causes, is associated with activated protein C (APC) resistance and increased thrombosis risk. When high prothrombin levels are a result of increased hepatic biosynthesis, these effects may be counteracted by concomitantly increased levels of the anticoagulant factors (particularly protein S). Differently, in prothrombin G20210A carriers only prothrombin levels are elevated. Objective: To investigate whether prothrombin G20210A carriers have a more severe hypercoagulable state than non‐carriers with comparable prothrombin levels. Patients/methods: Coagulation factor levels, thrombin generation (Calibrated Automated Thrombogram in the presence and absence of APC) and APC resistance were measured in normal (n = 132), heterozygous (n = 167) and homozygous (n = 3) individuals. Results: Prothrombin levels, thrombin generation and APC resistance were higher in carriers of the prothrombin G20210A mutation (especially those who had experienced venous thrombosis) than in non‐carriers, whereas protein S and antithrombin levels were similar among genotype groups. Because individuals with high prothrombin levels in the absence of the prothrombin G20210A mutation tend to have all liver‐synthesized factors elevated, carriers of the mutation had lower protein S and antithrombin levels than non‐carriers with equally high prothrombin levels. Accordingly, they also generated more thrombin and showed a tendency toward higher APC resistance. Analogous effects, but less pronounced, were observed in homozygotes for the prothrombin A19911G polymorphism, which also upregulates prothrombin levels.Conclusions: Individuals with hyperprothrombinemia as a result of prothrombin gene mutations generate more thrombin and tend to be more APC‐resistant than individuals with comparable prothrombin levels because of other causes.