FII G20210A Mutation Research Articles

Role of Genetic Thrombophilia Markers in Thrombosis Events in Elderly Patients with COVID-19

Thrombosis is an extremely dangerous complication in elderly patients with COVID-19. Since the first months of the pandemic, anticoagulants have been mandatory in treatment protocols for patients with COVID-19, unless there are serious contraindications. We set out to discover if genetic thrombophilia factors continue to play a triggering role in the occurrence of thrombosis in patients with COVID-19 with prophylactic or therapeutic anticoagulants. We considered the following genetic markers as risk factors for thrombophilia: G1691A in the FV gene, C677T and A1298C in the MTHFR gene, G20210A and C494T in the FII gene, and (-675) 4G/5G in the PAI-I gene. In a cohort of 176 patients, we did not obtain a reliable result indicating a higher risk of thrombotic complications when taking therapeutic doses of anticoagulants in carriers of genetic markers for thrombophilia except the C494T mutation in the FII gene. However, there was still a pronounced tendency to a higher incidence of thrombosis in patients with markers of hereditary thrombophilia, such as FV G1691A and FII G20210A mutations. The presence of the C494T (Thr165Met) allele in the FII gene in this group of patients showed a statistically significant effect of the mutation on the risk of thrombotic complications despite anticoagulant therapy.

The effect of hereditary thrombotic factors and comorbidities on the severity of COVID-19 disease.

Coronavirus disease 2019 (COVID-19) has rapidly spread worldwide and presents critical challenges for public health. Due to its chronic and systemic course, COVID-19 is currently accepted as a multi-systemic infectious disease. Here we explore the possible association between disease course and hereditary thrombotic factors and comorbidities. The patients admitted to the COVID-19 center in the Istanbul Faculty of Medicine were recruited for the study. The patients were classified according to the clinical course, severe vs. mild. Five polymorphic loci were analyzed by multiplex PCR: Factor V Leiden (FVL), FII G20210A, Beta-fibrinogen G-455A, and methylenetetrahydrofolate reductase (MTHFR) C677T and A1298C. FII G20210A and Beta-fibrinogen G-455A genotypes were significantly higher in the study group compared to the literature. Wildtype genotype (GG) in Factor V Leiden locus was significantly associated with low D-Dimer levels (p =0.013). The GA genotype increased the D-Dimer levels 2.55-times compared to the GG genotype (p =0.003). Moreover, the Beta-fibrinogen G-455G genotype was significantly higher in the LDH>250 group (p =0.046). The presence of solid tumors in patients with COVID-19 was related to the severity of the disease course. No evidence of a correlation between the severity of the disease and all five thrombotic mutations was found, whereas the FII G20210A and Beta-fibrinogen G-455A mutations were significantly high compared to previously reported Turkish population data and global carrier rates. This finding will need to be verified by further studies with larger samples since it may reflect a likelihood of having the COVID-19 disease. The high carrier frequency of FVL mutation was more likely present in the D-dimer high group generating an increase in the D-dimer levels 2.55-times compared to the wildtype.

Association of Inherited Thrombophilia with Recurrent Pregnancy Loss in A Population of Lebanese Women: A Case Control Study.

Recurrent pregnancy loss (RPL) complication is a challenge of reproductive medicine due to its often unknown etiology. A case-control study was carried out between June 2019 and April 2020 to examine the correlation between RPL and inherited thrombophilia (IT), namely mutations in factor V Leiden (FVL G1691A), prothrombin (FII G20210A), and methylenetetrahydrofolate reductase (MTHFR C677T). A total of 120 Lebanese women with RPL was studied and compared, for the frequency of these mutations, to 100 healthy reproductive Lebanese women. The association between the zygosity status of the three tested mutations, the existence of more than one prothrombotic single nucleotide polymorphisms (SNPs), and the increased risk of RPL were examined using Chi-square or two-tailed fisher exact test, and the student t test. The predictive factors of RPL were analyzed using a multiple logistic regression model. P<0.05 was considered to be statistically significant. Our results showed statistically significant higher frequencies of FVL G1691A and FII G20210A mutations among the cases with RPL compared to the control group. Thus, RPL is associated with FVL G1691A and FII G20210A mutations. These mutations seem to increase the risk of RPL in the Lebanese women. Therefore, we suggest thrombophilia screening and adequate genetic counseling for women with RPL and at high-risk to plan for primary prevention, avoiding thromboembolic or obstetric accidents, and reducing the associated morbidity and mortality among Lebanese women.

Unusual pattern of thrombotic events in young adult non-critically ill patients with COVID-19 may result from an undiagnosed inherited and acquired form of thrombophilia.

In 145 previously healthy non-critically ill young adults, coronavirus disease 2019 (COVID-19)-related symptoms, risk factors for thrombosis, coagulation and inflammatory parameters were compared, with 29 patients reporting unusual thrombotic events (UTEs) and 116 not having thrombotic events. The inflammatory indices, coagulation and prothrombotic platelet phenotype (PTPP) were significantly higher in patients with UTEs versus those without. Patients with UTEs were categorised according to detection of thrombophilic genes (TGs), coagulation and inflammatory markers to the non-TG and TG subcohort. A total of 38 UTEs were identified, which included splanchnic vein thrombosis (SVT; 11), stroke (six), cerebral vein thrombosis (five), thrombotic microangiopathy (four), limb ischaemia and inferior vena cava thrombosis (three each), ST-segment elevation myocardial infarction (two), superior vena cava thrombosis (two), upper limb deep venous thrombosis and retinal vein thrombosis, one each. We found a 55% prevalence of TGs mainly heterozygous coagulation factor II, thrombin (FII)-G20210A, Janus kinase 2 (JAK2)-V617F, protein-S, and antithrombin III deficiency with a high (76·9%) prevalence of venous UTEs, multiple vessels thrombosis, and recurrence rate among the TG versus non-TG subcohort. The presence of JAK2-V617F, and FII-G20210A mutations was linked with SVT. Thrombosis in the non-TG subcohort was associated with more haemorrhagic problems, thrombosis progression and a significantly higher level of inflammatory markers, PTPP, mean platelet volume, von Willebrand factor, and factor VIII, which remained high for up to 6 months, as well as elevated D-dimer. Acquired and inherited thrombophilia with endotheliopathy appeared to be a relevant mechanism to explain the occurrence of UTEs that are not correlated to COVID-19 severity.

Hyperbaric oxygen treatment results in a group of Turkish central retinal artery occlusion patients with a combined presence of thrombophilic mutations

Central retinal artery occlusion (CRAO) is a rare ocular-ischemic syndrome causing irreversible blindness. Its pathophysiology has not been clarified, and no targeted therapies are available yet. Hyperbaric oxygen (HBO2) therapy is already an approved therapy for CRAO and has been shown to improve the visual acuity of CRAO patients safely. However, further clinical data are required to classify HBO2 therapy as a type-I general agreement for CRAO. Eleven patients with non-arteritic CRAO were enrolled. Patient demographics, medical history, detailed eye examinations, HBO2 therapy results, pre-/post HBO2 therapy visual acuity measurements and genotypes for common thrombophilic mutations (Factor V G1691A Leiden, Factor II G20210A, MTHFR A1298C, MTHFR C677T, and PAI-1-675 4G/5G) were obtained. Six patients (54%) responded to HBO2 therapy compared to five non-responders (46%). Patients admitted before 12 hours responded well to HBO2 therapy. No systemic diseases nor advanced age were statistically correlated to CRAO. A combination of mutations rather than single mutations for each patient could be seen as responsible for CRAO. No Factor V G1691A Leiden mutations and only one FII G20210A mutation were observed. Eight patients (72%) had MTHFR 677T allele, five patients (45%) had MTHFR 1298C allele, and 10 patients (91%) had the PAI-1-675 4G allele. Not a single mutation but a combination of mutations and other unknown factors probably lead to CRAO, and if intervention is timely, HBO2 therapy offers improvement in visual acuity safely.

Evaluation of Factor V Leiden and prothrombin G20210A mutations in Sudanese women with severe preeclampsia

BackgroundPreeclampsia (PE) is a common pregnancy complication and one of the main causes of maternal and fetal morbidity and mortality, worldwide. While the pathogenesis of PE is unclear, it has been suggested that hypercoagulability due to Factor V Leiden (FVL) and prothrombin gene mutation (FII G20210A) play a role in its progression. PurposeThis study aimed to determine if there is an association between FVL and FII G20210A mutations and severe PE. Patients and methodsThis case-control study enrolled 50 women with severe PE and 50 healthy pregnant women as the control, at Khartoum North Teaching Hospital, in Khartoum State, Sudan, from January 2017 to June 2017. The presence of point mutations in FVL and FII G20210A were determined for each of the participants. Deoxyribonucleic acid (DNA) was extracted, and then an allele-specific polymerase chain reaction (PCR) was used to detect the point mutations in FVL and FII G20210A. ResultsThe results revealed a significant difference between the subjects in the severe PE group and the control group for the means of parity, gestational age/ week and hemoglobin concentration (P < 0.05). No statistically significant body mass index (BMI) differences were found between the groups (P > 0.05). Women with severe PE were found to have a significant difference in FVL (16%; P value = 0.0058; OR: 20.20; 95%CI: 1.132–360.5) and FII G20210A (14%; P value = 0.0125; OR: 17.41; 95%CI: 0.9659–314.0) in comparison to the women in the control group (0%). ConclusionOur findings intensely indicate that there is a statistically proven significant association between FVL, FII G20210A mutations and the development of severe preeclampsia in Sudanese pregnant women.

PREVALENCE OF FACTOR V LEIDEN AND PROTHROMBIN G20210A IN WOMEN WITH VON WILLEBRAND DISEASE TYPE 1

Background.Von Willebrand disease is a hereditary malfunction of the blood coagulation system caused by waveform quantitative and/or qualitative deficiency of von Willebrand factor (vWF).Aim.To evaluate the frequency of occurrence of FVLeiden and FII G20210A mutations in female patients with von Willebrand type 1 disease.Materials and methods.136 women aged from 18 to 45 years (mean 31.7 ± 0.5 years) were enrolled in a study conducted during the January 2011 — December 2017 period. Questionnaire was used to reveal hemorrhagic diathesis. Inclusion criteria were as follows: no less than 3 positive responses to questions 1–7, or 2 positive responses to questions 1–7 plus no less than 100 points of the evaluated menstrual blood loss. An independent inclusion criterion was 180 points or more in the question concerning menstrual blood loss. A mandatory inclusion criterion was the confirmation of absence of thromboembolic events in a proband and first line relatives. The study included assessment of such parameters as ristocetin-cofactor activity of von Willebrand factor (vWF:RCo), von Willebrand factor antigen (vWF:Ag), factor VIII (FVIII:C), platelet aggregation induced with ADP, ristomycin, collagen, as well as molecular-genetic assay of factor V (FVLeiden) and gene (FII G20210A) polymorphism using allele-specific polymerase chain reaction.Results.No mutations of FVLeiden and FII G20210A were revealed in 102 women with von Willebrand disease type 1. Heterozygous mutation of FVLeiden was found in 12 (8.8 %) subjects with von Willebrand disease type 1 (vWF:RCo from 27 to 47 % (mean 37.3 ± 0.8 %), vWF:Ag from 25 to 46 % (mean 37.5 ± 0.8 %), FVIII:C from 29 to 49 % (mean 44.1 ± 0.5 %). Homozygous mutation of FVLeiden was identified in 3 (2.2%) women with von Willebrand disease type 1, with vWF:RCo being 40, 43 and 45 %, vWF:Ag — 39, 44 and 42 %, FVIII:C — 47, 45 and 48 %, respectively. Heterozygous mutation FII G20210A was detected in 19 (13.9 %) subjects with von Willebrand disease type 1 (vWF:RCo from 36 to 49 % (mean 43.0 ± 0.4 %), vWF:Ag from 32 to 46 % (mean 42.2 ± 0.6 %), FVIII:C from 30 to 49 % (mean 45.1 ± 0.4 %).Conclusion.By means of diminishing the coagulation potential of the blood coagulation system, a decrease in the activity of VIII and von Willebrand factors may compensate possible negative effects associated with FVLeiden and FII G20210A gene mutations in female patients with von Willebrand type 1 disease.

Prevalence of Factor V Leiden and Prothrombin Mutation G20210A in Patients with a History of Thrombotic Phenomena in Temuco, Araucanía Region

Thrombotic phenomena are multifactorial phenomena with both acquired and inherited factors. Factor V Leiden (FVL) and G20210A mutation of prothrombin (FIIG20210A) are the most studied inherited factors, with a prevalence that varies by population and race, high in Europe and almost nil in Asian countries. Information concerning the prevalence of these mutations in Chile is scarce. The aim of this study was to determine the prevalence of FVL and FII G20210A mutation in patients with a history of thrombotic events in the city of Temuco, Araucanía region. Genetic analysis for FVL Thrombophilia and FIIG20210A mutation was performed from January 2012 to December 2013. Three hundred thirty-three screenings by PCR in real time and the genotype of each mutation were determined and analyzed by Melting curve. We found an average prevalence of 2.28% forthe FVL, whereas FIIG20210A mutation was 6.49%. Nomutations were found in homozygous state. Both mutations are present in our population, it is therefore important to evaluate the risk associated with their presence in patients with a history of thrombotic events. In our specific population FII G20210A mutation is highly prevalent, thus genetic studies should be realized or requested.

Оценка протромботического фенотипа у асимптомных носителей мутации G1691A в гене фактора V и/или мутации G20210A в гене протромбина с помощью теста генерации тромбина

Введение. К числу безусловных наследственных детерминант тромбофилии относятся мутации в генах факторов II (G20210A) и V (G1691A). Однако их наличие, особенно в гетерозиготной форме, не всегда приводит к развитию тромботических осложнений, что свидетельствует о целесообразности поиска новых объективных методов выявления протромботического фенотипа у носителей рассматриваемых мутаций. Тест генерации тромбина (ТГТ) отражает состояние гемостаза индивидуума в совокупности взаимодействия прокоагулянтных и антикоагулянтных механизмов и позволяет эффективно выявлять гиперкоагуляцию. Цель исследования: установить целесообразность использования ТГТ для оценки гемостатического потенциала у асимптомных носителей мутаций в генах фактора V (G1691A) и/или протромбина (G20210A). Материалы и методы. Обследовано 42 асимптомных носителя мутации FV G1691A и/или FII G20210A (14 мужчин и 28 женщин, средний возраст — 37,0 ± 15,0 лет). Постановка ТГТ выполнялась в бедной тромбоцитами плазме. В тромбинограмме оценивали следующие показатели: PT (Peak thrombin — максимальное количество тромбина, образующегося в образце), ETP (endogenous thrombin potential — эндогенный тромбиновый потенциал), Lag-time — время инициации свёртывания и V (скорость образования тромбина). Результаты. Согласно полученным данным, гетерозиготное носительство мутаций FV G1691A и/или FII G20210A может ассоциироваться как с протромботическим, так и с нормальным фенотипом. У асимптомных носителей Лейденской мутации повышенные значения ЕТР и РТ обнаружены в 44% случаев, в то время как резистентность к активированному протеину С — в 67%. У большей части носителей мутации в гене протромбина отмечены превышающие норму значения ЕТР и РТ (71% и 57% случаев, соответственно). Скорость генерации тромбина превышала нормальные величины у 57% носителей мутации FV G1691A и у 64% носителей мутации FII G20210A. Значительную часть обследованных можно охарактеризовать как имеющих промежуточный протромботический фенотип, учитывая наличие у них одновременно как нормальных, так и патологических показателей ТГТ. Заключение. ТГТ является перспективным методом в плане создания индивидуального профиля риска для асимптомных носителей мутаций, ассоциированных с наследственной тромбофилией. Introduction. Mutations in genes of factors II (G20210A) and V (G1691A) are well known determinants of hereditary thrombophilia, but their presence, especially in the heterozygous form, does not always lead to the development of thrombotic complications.This indicates the importance of objective methods for the detection of prothrombotic phenotype in carriers of these mutations. Thrombin generation test (TGT) demonstrates the state of individual’s hemostasis in the interaction of procoagulant and anticoagulant mechanisms and allows to detect the hypercoagulability. Aim: to assess the hemostatic potential in asymptomatic carriers of FV G1691A or/and FII G20210A mutations by TGT. Materials and methods. We examined 42 asymptomatic carriers of FV G1691A mutation and/or G20210A prothrombin gene mutation (14 men and 28 women, average age — 37,0 ± 15,0 years). TGT was performed in platelet-poor plasma. In thrombinogram we assessed following parameters: PT (Peak thrombin is the maximum amount of thrombin formed in the sample), ETP (endogenous thrombin potential), Lag-time (coagulation initiation time) and V (thrombin generation rate). Results. According to data obtained, heterozygous carriage of FV G1691A and/or FII G20210A mutations can be associated with both prothrombotic and normal phenotypes. In asymptomatic carriers of Leiden mutation we revealed elevated values of ETP and RT in 44% of cases, while resistance to activated protein C — in 67%. The majority of FII G20210A mutation carriers demonstrated increased values of ETP and PT (71% and 57%, respectively). Thrombin generation rate exceeded normal values in 57% and 64% carriers of FV G1691A and FII G20210A mutations, respectively. The majority of examined patients can be characterized as having intermediate prothrombotic phenotype, taking into account that they had both normal and pathological values of TGT parameters. Conclusion. TGT is a perspective method for creating an individual risk profi le in asymptomatic carriers of mutations associatedwith hereditary thrombophilia.

VENOUS THROMBOSIS IN CHILDREN, ADOLESCENTS AND YOUNG ADULTS WITH LYMPHOMAS: INCIDENCE, CHARACTERISTICS, RISK FACTORS AND PROGNOSTIC VALUE

Objective . To define the epidemiology of venous thrombosis (VT) in children, adolescents and young adults with lymphomas, to identify the thrombotic risk factors and VT’s prognostic value. Materials and methods. We reviewed the medical records of 513 lymphoma patients (284 with Hodgkin's lymphoma (HL) and 229 with non-Hodgkin lymphoma (NHL)) aged from 0,6 to 26 years (median – 14 years), diagnosed with lymphomas in 2005–2017. Risk thrombotic factors were assessed using binary Cox regression in univariate and multivariate models with calculation of the odds ratio (OR) with 95 % CIs. Results . Twenty-eight of 513 patients (5.5 %, 95 % CI, 4.5–6.5 %) were diagnosed as having VT with a total of 32 thrombotic events. The incidence of VT in NHL patients (23/229, 10.5 %, 95 % CI, 8.5–12.5 %) was higher than in HL patients (5/284, 1.8 %, 95 % CI, 1.0–2.6 %, p &lt;0.0001). VT was associated with venous catheterization (VC) in 53.6 % and with local compressive effect in 32.1 % cases. VT occurred during the first 6 weeks after lymphoma diagnosis in 57.1 % patients. Genetic predisposition to thrombosis (FV Leiden и FII G20210A mutations) was revealed in 3 (10.7 %) patients. The recurrence rate of VT was 14.3 %. Age older than 13 years (ОR 2.5, 95 % CI, 1.1–6.0, p &lt;0.05) and lymphoblastic lymphoma (LL) subtype (ОR 5.5, 95 % CI; 2.3–13.6, p &lt;0.01) independently associated with the occurrence of VT in NHL patients. In HL group all cases of VT were detected in patients older than 15 years. The only risk factor predisposing patients with HL to VT was significant mediastinal lymphadenopathy (bulky disease) (ОR 7.0, 95 %; 1.2–42.3, p&lt;0.05). The presence of VT did not influence OS of NHL patients (73.7 %, SE 9.2 % (n = 23) versus 83.4 %, SE 9.2 % (n = 206), p &gt;0.05), but had a negative impact on the OS of HL patients (60.0 %, SE 21.9 % (n = 5) versus 94.8 %, SE 1.5 % (n = 279), p &lt;0.001). HL patients older than 15 years with bulky disease (38/284, 13.4 %), had a high thrombotic risk with lower OS of 78.8 ± 8.3 %, p &lt;0.001. Conclusion. Cumulative incidence of VT in NHL patients was higher than in HL patients. Age older than 13 years and LL were independent thrombotic risk factors for NHL patients. Age older than 15 years and bulky disease increased risk of VT in HL patients. VT occurrence decreased OS of young HL patients. It may be necessary to delay the central VC of vena cava superior in children with massive mediastinal tumor. Prophylactic anticoagulation for high thrombotic risk patients might be warranted.

Pro-Coagulant and Pro-Inflammatory Effect of Interferon Alpha in Myeloproliferative Neoplasms

Myeloproliferative neoplasms (MPN) are associated with an increased risk of arterial and venous thrombosis. Besides clinical factors (previous history of thrombosis, age > 60 years, cardiovascular risk factors), procoagulant phenotype, proadhesive properties of the endothelium and secretion of inflammatory cytokines, contribute to the pathogenesis of thrombosis in MPN. Pegylated interferon alpha (IFN) has obviously a great therapeutic efficacy on hematopoietic cell proliferation but its other biological effects particularly on hemostasis and inflammation have not been determined.The aim of the study was to determine whether IFN impacts the biological profile including endothelial and platelet markers compared to MPN patients treated by hydroxyurea (HU) and non-treated (NT) patients.Patients with polycythemia vera (PV) or essential thrombocythemia (ET) treated with IFN or HU or without any cytostatic drug were included. Platelet membrane glycoproteins and platelet activation were measured by flow cytometry. We evaluated shear-induced platelet aggregation (SIPA) at 4000 sec-1 that mimics stenotic arteries. We measured coagulation proteins by their activity and endothelial parameters (von Willebrand factor (VWF) antigen and activity, soluble thrombomodulin). Thrombin generation was measured in platelet poor plasma and platelet rich plasma. Platelet accumulation on immobilized collagen was measured using whole blood perfused at 1500 sec-1 in flow chambers. Statistical analysis was performed by ANOVA to compare the 3 groups of patients (p<0.05) and post-test Newman-Keuls to compare groups side-by-side (p<0.01).Eighty-eight patients were included: 28 treated by IFN (20 PV and 8 ET), 38 treated by HU (27 PV and 11 ET) and 22 NT (6 PV and 16 ET). All patients were treated with aspirin 75 or 100 mg/day. Prior history of thrombosis occurred in 25.7% of ET and in 31.5% of PV. During the follow up (> 6months), only 1 arterial and 2 venous thrombosis were observed.Hematological parameters and JAK2 V617F status was in the expected range in ET and PV patients.Surprisingly, we observed some significant effect of IFN on 1) platelets with a decreased GPVI expression compared to HU- and non-treated patients and increased levels of SIPA at 4000 sec-1. 2) endothelial cells with significantly increased levels of Willebrand antigen and activity. 3) coagulation parameters with decreased protein S activity and increased levels of factor VIII:C and fibrinogen (table 1). All the other parameters tested were similar in the 3 groups of patients.We had the opportunity to test 10 patients at least 6 months after IFN discontinuation. IFN was stopped for side effects in 6 patients, for complete molecular response in 2 and for normalization of hematological parameters in 2 others. GPVI expression, SIPA, VWF activity and antigen, protein S activity, FVIIIC and fibrinogen returned to levels similar to those of non-treated patients and were significantly different from the levels in IFN-treated patients (table 2). These results indicate that IFN was responsible for these biological modifications.In conclusion, these unexpected data showing an increase of procoagulant and inflammatory markers in IFN-treated MPN patients could be considered as a part of IFN biological activity. Though minimal, these alterations could require more intensive preventive anticoagulation than aspirin in patients with heterozygous FV Leiden or FII G20210A mutation. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

The frequencies of FV Leiden and FII G20210A mutations in patients with different clinical manifestations of venous thromboembolism: Experience from large Serbian cohort

Venous thromboembolism is a multifactorial disorder with two manifestations: deep-vein thrombosis and pulmonary embolism. Pulmonary embolism is usually considered as the complication of deep-vein thrombosis, but there are reported cases of isolated pulmonary embolism. FV Leiden and FII G20210A mutations are most common genetic risk factors for the venous thromboembolism. Several studies reported "FV Leiden paradox": lower prevalence of FV Leiden mutation among patients with isolated pulmonary embolism than among those with deep-vein thrombosis. The aim of this study was to determine FV Leiden and FII G20210A mutations frequency in thrombophilic patients in Serbian population. We tested prevalence of these mutations carriers in 1427 individuals divided in three groups of patients (with deep-vein thrombosis, deep-vein thrombosis/ pulmonary embolism and isolated pulmonary embolism) and control group. All subjects were tested for these mutations using PCR-RFLP analysis. Detected frequency of FV Leiden heterozygous carriers in patients with isolated pulmonary embolism was 6.9% (for FII G20210A 11.6%), while in other two groups of patients with deep-vein thrombosis and deep vein thrombosis/pulmonary embolism, frequency was 18.6% (for FII G20210A mutation were 11.6% and 8.3%, respectively). Our results showed that FV Leiden mutation is less frequent in patients with isolated pulmonary embolism than in patients with deep-vein thrombosis or deep-vein thrombosis accompanied with pulmonary embolism, confirming "FV Leiden paradox". On the other hand, detected frequency of FII G20210A mutation carriers was similar in all three groups of patients.

Thrombophilic Mutations as Risk Factor for Retinal Vein Occlusion

Retinal vein occlusion (RVO) is the second most common retinal vein disease and an important cause of blindness and visual morbidity. Many conditions are associated with RVO but the real role of the thrombophilic mutations is still unclear. To evaluate the potential role of thrombophilic mutations in RVO. We have evaluated 113 patients with RVO and compared with 104 volunteer controls. The controls were all healthy blood donors without previous venous thromboembolism episode or arterial thromboembolism episode. All patients were tested for 5 gene variants (here all named as mutations): factor V (FV) Leiden (G1691A), factor II (FII; G20210A), 5,1-methylenetetra-hydrofolate reductase (MTHFR; C677T), plasminogen activator inhibitor 1 (PAI-1; 4G/5G), and angiotensin-converting enzyme (ACE; Del/Ins). Statistical analysis were performed by the 2-tailed chi-square test. Statistical test showed that TT homozygous patients of the MTHFR C677T mutation (P = .017) and heterozygous GA patients of the FII G20210A mutation (P = .018) were significantly higher than that in controls. For FV Leiden, even if the values were higher in patients than in controls, P value was not statistically significant. Conversely, for the ACE (Ins/Del) and PAI-1 (4G/5G) mutations, no difference was observed among genotypes of patients with RVO and control participants. In our study, the FII G20210A and the MTHFR C677T mutations resulted significantly higher in patients than in controls; in contrast, thrombophilic mutation of FV, ACE, and PAI-1 genes was not statistically correlated with RVO. In spite of having found an association between some thrombophilic mutations and RVO, more studies with a major number of patients are necessary to determine the final role of these gene variants.

Relationship between Methylenetetrahydrofolate Reductase (C677T), Factor V Leiden (G1691A), Prothrombin Mutation (G20210A) and Severe Preeclampsia in a Brazilian Population

Objectives: To better understand the etiologic factors that can influence preeclampsia, we inves- tigated hereditary factors for thrombosis, FV Leiden, F II 20210A mutation and the polymorphism C677T of the MTHFR, as singly and as in association, in a group of women from Ceará state-Northeast Brazil with severe preeclampsia. Material and Methods: We conducted a case-control study. 101 cases of severe preeclampsia were recruited from School Maternity Assis Chateaubriand, a reference Maternity of State of Ceará, Brazil, from December 2009 to December 2010. For clinical correlations, women were interrogated about fetal low weight (less than2500 grams) and fetal loss in previous pregnancies. Low weight, fetal loss and necessity of neonatal intensive care unit (neonatal ICU) related to current pregnancy were registered. 245 healthy voluntary women were recruited from the blood bank donors to verify the frequency of FV Leiden, FII and MTHFR as a control group. The study was approved by the Committee on Ethics in Human Research of Maternidade Escola Assis Chateubriand and all individuals gave their informed consent to participate in the study. Standard veinipuncture, with EDTA as anticoagulant, was used to collect blood samples. Genomic DNA was extracted, soon after, using a salting out method. Agarose gel electrophoresis with ethidium bromide staining was performed to ensure the quality of DNA extraction. Results: The frequencies of FV Leiden and FII mutation carrier were 0.99% (1/101) for both factors in the preeclampsia patients and 1.86% (4/214) and 0.93% (2/214), respectively, in control group. All mutates were heterozygous and concomitance of both mutations was not found. The genotype distribution of the MTHFR C677T in the patients and controls frequencies was in Hardy-Weinberg equilibrium (p ≤ 0.05). No statistical difference was observed between cases and controls in MTHFR genotypes or alleles. Conclusions: The FV Leiden, FII G20210A mutation and MTHFR C677T were not risks for preeclampsia development. FV Leiden and FII G20210A mutations had low frequency in the population studied, which may justify the absence of association.

Routine tests of haemostasis and low-oestrogen oral contraceptives: what to expect?

Frequent requests from gynaecologists for testing of haemostatic parameters in women using hormonal contraception in order to predict development of thromboembolic disease, along with advancement of testing quality and accuracy, created the need to assess and compare the overall effect of oral contraceptives on today's routine haemostatic tests. This 6-month prospective study included 195 women, first time users of oral contraceptives or with at least a 3-month pause before their entry in the study. Haemostatic changes were assessed by the following tests performed at three study time-points: at baseline, after three and after six months of taking oral contraceptives: prothrombin time, activated partial thromboplastin time, fibrinogen, resistance to activated protein C ratio, protein C activity, protein S activity, FVIII activity, antithrombin activity, plasminogen activity, alpha2-antiplasmin activity, inhibitor of plasminogen activator type 1 (PAI-1), and D-dimers. In addition, study subjects underwent genetic testing for FV Leiden mutation, FII G20210A mutation, and PAI-1 4G/5G polymorphism. Significant changes were found in all haemostatic variables except for FVIII activity. With in-depth analysis, we showed that PAI-1 concentration decreased only in subjects with PAI-1 4G/5G genotype. None of the subjects developed venous thromboembolic disease during the study. As most changes were within reference ranges, routine laboratory testing still does not appear to be recommendable in women without a known risk of thromboembolic disease.

Risk factors and recurrent thrombotic episodes in patients with cerebral venous thrombosis.

The prevalence of thrombophilic abnormalities in patients with cerebral vein thrombosis has been reported to be similar to that in patients with deep vein thrombosis of the lower limb. The role of gender-specific risk factors (pregnancy, oral contraceptives) is well established, whereas that of other acquired risk conditions is debated. We screened 56 patients with cerebral vein thrombosis and 184 age- and sex-matched apparently healthy controls for prothrombin (factor II, FII) G20210A and factor V Leiden polymorphisms; protein S, protein C, and antithrombin deficiency; anticardiolipin antibodies; hyperhomocysteinaemia and other putative risk factors. The G20210A polymorphism was found in 29.1% of patients and in 5.7% of controls (odds ratio [OR] 7.1; P<0.0001; adjusted OR 12.67, P<0.0001). Frequencies of factor V Leiden and hyperhomocysteinaemia were not significantly different in patients and controls, nor were the other thrombophilic tests and some established cardiovascular risk factors, such as smoking, obesity or overweight and arterial hypertension. Conversely, 53.7% of the women who developed cerebral vein thrombosis did so while assuming oral contraceptives (OR 6.12; P<0.0001), with a further increase of risk in FII G20210A carriers (OR 48.533). Some associated diseases (onco-haematological disorders and infections) also had a significant role. Over a median 7-year follow-up, irrespective of the duration of antithrombotic treatment, 9/56 (16%) patients had further episodes of venous/arterial thrombosis. No significant risk factor for recurrent thrombosis was identified. In spite of the limitations of the sample size, our data confirm the role of FII G20210A mutation in this setting and its interactions with acquired risk factors such as oral contraceptives, also highlighting the risk of recurrent thrombosis in cerebral vein thrombosis patients.

G1691A factor V and G20210A FII mutations, acute ischemic stroke of unknown cause, and patent foramen ovale

BackgroundGenetic polymorphisms of haemostatic factors such as G1691A factor V (FV Leiden) and G20210A prothrombin (FII) may be involved in the onset of patent foramen ovale (PFO)-related cerebral ischaemia. We assessed the possible association between such inherited thrombophilic alterations and right-to-left shunt in patients with stroke. MethodsWe investigated the presence of G20210A FII and FV Leiden mutations in 340 Caucasian patients consecutively evaluated by our Angiology Unit for stroke of unknown cause. PFO was assessed in all patients with Transcranial Doppler with intravenous injection of agitated saline. Stroke patients were divided into two groups: patients with PFO (n=136), and patients without PFO (n=204). As control group, we studied 272 subjects with early venous insufficiency. ResultsThe prevalence of FII G20210A mutation was significantly higher in patients with PFO vs. controls (OR: 2.90; 95% CI: 1.19–7.07) and in patients without PFO vs. controls (OR: 2.88; 95% CI: 1.25–6.60) but was similar in patients with and without PFO (OR: 1.11; 95% CI: 0.51–2.44). The frequency of FV Leiden mutation was similar in the three groups. Across the population the presence of the FII G20210A mutation (OR: 2.97;95% CI: 1.32–6.69), a history of DVT (OR: 1.04; 95% CI: 1.02–1.06), and oestrogen-containing contraceptive therapy (OR: 1.14; 95% CI: 1.09–1.18) were all associated with stroke of unknown cause after adjustment for other risk factors, This was not the case with PFO. ConclusionsOur data do not support the assumption that these inherited thrombophilic alterations are associated with PFO in patients with cryptogenic stroke. FII G20210A mutation may be associated with cryptogenic stroke irrespective of the presence of PFO.

Factor V Leiden G1691A and factor II G20210A point mutations and pregnancy in North-West of Iran

The roles of several hereditary predispositions for venous thromboembolism have been evaluated in women with habitual abortion. We studied the prevalence of FV Leiden G1691A and FII G20210A mutations in women with habitual abortion and healthy controls. 60 unrelated fertile females, as controls, and 70 unrelated women with at least three consecutive pregnancy losses entered at the present study. MAS-PCR was carried out for detection of FV Leiden G1691A and FII G20210A mutations. FV Leiden G1691A mutation was not found in the studied cases and controls, that is, all of the cases and the controls had normal FV Leiden 1691GG genotype. FII 20210AA genotype was not found in any of patients or controls. 2.5% of alleles (3 out of 120 chromosomes) in controls and 15.714% of alleles (22 out of 140 chromosomes) in cases had FII 20210A mutation. The FII G20210A allele frequency was 0.157 in cases and 0.025 in controls. Regarding FII G20210A mutation, the distribution of GG, GA and AA genotypes were 48 (68.57%), 22 (31.43%) and 0 (0%) in the cases and 95 (95%), 5 (5%) and 0 (0%) in the controls, respectively. Significant differences in both FII G20210A alleles and FII G20210A genotypes frequencies were observed in the cases versus the controls. FII G20210A mutation is significantly associated with habitual abortion.

Clinical Characteristics and Type of Thrombophilia in Women with Pregnancy-Related Venous Thromboembolic Disease

Background: Normal pregnancy is characterized by numerous changes in the hemostatic system, creating the hypercoagulable state which increases the risk of venous thromboembolic event (VTE) occurrence. The risk is further increased by the presence of inherited or acquired thrombophilia. Objective: In this study, we aimed to determine the prevalence of different types of thrombophilia in women with pregnancy-related VTE, and to investigate the possible connection between the type of thrombophilia and localization of VTE as well as the gestational age of VTE occurrence. Participants and Methods: Two hundred and two women with the first episode of pregnancy-related VTE and 130 controls were investigated. The antithrombin, protein C and protein S activity, APC resistance, FVG1691A, and FIIG20210A were determined. None of the investigated women was pregnant at the time of thrombophilia testing, and none was using oral contraceptives. Results: Thrombophilia was diagnosed in 95 patients (47%) and 7 controls (5.4%). The prevalence of FV Leiden, FIIG20210A mutations, antithrombin, PC and PS deficiencies taken together and combined thrombophilia was 22.3, 10.4, 6.9 and 6.9%, respectively. Significantly more frequent antepartum occurrence of VTE (11 vs. 3, p < 0.05) was found in women with natural coagulation inhibitor deficiency. Pulmonary embolism occurred more frequently in nonthrombophilic women (25 vs. 3, p < 0.001). Conclusion: Inherited thrombophilia was found to be considerably more frequently present in women with pregnancy- and puerperium-related VTE compared to healthy controls. Women with thrombophilia are at higher risk of developing thromboses localized in the iliacofemoral region, and women without thrombophilia are at higher risk of developing pulmonary embolism. Deficiency in natural coagulation inhibitors is associated with antepartum VTE occurrence.

FV Leiden, FII G20210A and MTHFR C677T mutations in patients with lower or upper limb deep vein thrombosis

Deep vein thrombosis (DVT) is a multifactorial disease that occurs with frequency of 1/1000 per year. The FV Leiden, FII G20210A and MTHFR C677T mutations represent genetic factors for the occurrence of vein thrombosis. The goal of this study was to determine the frequency of these mutations in patients with DVT of upper and lower limbs. The study encompassed 119 patients divided in two groups. The group of patients with the lower limbs thrombosis included 77 patients, while the upper limbs thrombosis group included 42 patients. The presence of FV Leiden, FII G20210A and MTHFR C677T mutations was detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. In patients with DVT of lower limbs, the frequency of FV Leiden mutation was 26,0% in heterozygous form and 1,3% in homozygous form. In the group of patients with DVT of upper limbs, the frequency of heterozygous carriers was 7.2%. In patients with DVT of lower limbs, FII G20210A mutation occurred in heterozygous form in 15.6% subjects, and in the group with DVT of upper limbs the frequency was 7.2% in heterozygous and 2.3% in homozygous form. The frequency of MTHFR C677T mutation in patients with lower limbs DVT was 42.8% in heterozygous form and 13% in homozygous form, while in the group of patients with upper limbs DVT, the frequency was 52.4% in heterozygous form and 9.5% in homozygous form. The FV Leiden and FII G20210A mutations represent significant risk factors for the occurrence of DVT of lower limbs. These mutations are less frequent in DVT of upper limbs and more extensive further studies are needed to determine their potential role. The MTHFR C677T mutation represents less significant risk factor for lower limb DVT and should be taken into account only in cases when it occurs in combination with other risk factors.