The risk of venous thromboembolism in oral contraceptive users: the role of genetic factors-a prospective cohort study of 240,000 women in the UK Biobank.

BACKGROUND: More than 150 million women worldwide use oral contraceptives. Women with inherited thrombophilia and carriers of certain thrombophilia gene variants, such as factor V Leiden and the prothrombin, are at an increased risk for venous thromboembolism, especially when combined with oral contraceptive use. Venous thromboembolism is a complex disorder involving many genetic risk factors, and recently, polygenic risk scores have been proposed to capture a significant proportion of the genetic risk of venous thromboembolism. OBJECTIVE: The aim of this study was to estimate the risk for developing venous thromboembolism when initiating oral contraceptive use (first 2 years) and during continued use among women with a high genetic liability. STUDY DESIGN: We used a prospective study design in which 244,420 participants from the UK Biobank were followed from birth. The effect of oral contraceptive use during the first 2 years and in the remaining years of oral contraceptive use on the risk of developing venous thromboembolism was estimated using a Cox regression with a time-dependent exposure variable. Women were stratified according to their polygenic risk scores and whether they were carriers of factor V Leiden and/or prothrombin variants. RESULTS: When genetic risk was not considered, an increased risk for venous thromboembolism was observed during the first 2 years of oral contraceptive use (hazard ratio, 3.09; 95% confidence interval, 3.00-3.20) but not during continued use (hazard ratio, 0.92; 95% confidence interval, 0.80-1.05). However, when genetic risk was considered, women in the highest polygenic risk score category had a more pronounced risk of developing a venous thromboembolism during the first 2 years of oral contraceptive use (hazard ratio, 6.35; 95% confidence interval, 4.98-8.09), and a high risk was also observed among factor V Leiden (hazard ratio, 5.73; 95% confidence interval, 5.31-6.17) and prothrombin variant carriers (hazard ratio, 5.23; 95% confidence interval, 4.67 - 5.87). A high polygenic risk score in combination with being a factor V Leiden and prothrombin variant carrier conferred the highest risk for developing a venous thromboembolism during the first 2 years of oral contraceptive use (hazard ratio, 14.8; 95% confidence interval, 9.28-23.6). Women with a high genetic liability also had an increased risk during continued use but it was less pronounced, and the highest risk was conferred to carriers of both factor V Leiden and the prothrombin variant (hazard ratio, 4.93; 95% confidence interval, 3.16-7.7). CONCLUSION: Evaluating polygenic risk can identify additional venous thromboembolism risk that is not captured in the commonly investigated genes for inherited thrombophilia. Our results indicate that oral contraceptive use is associated with an increased risk for developing a venous thromboembolism, particularly among women with a high genetic predisposition, and that oral contraceptive use dramatically increases the risk thereof short after initiation of use, which decreases with continued use. This suggests that the polygenic risk score could be used to identify women who are at high risk for developing a venous thromboembolism and advise them on alternative methods of contraception.

Clinical characteristics and thrombophilia associated gene variants in Egyptians with unprovoked venous thromboembolism: three centers experience.

BACKGROUND: Thrombophilias are characterized by excessive venous and arterial thrombosis at regular or unusual sites. It may result from inherited, acquired, or a combination. Hereditary thrombophilia (HT) is detected in 30-40% of patients with thromboembolism. Venous/arterial thrombosis is considered a multifactorial disorder, some patients may have more than one risk factor which may be transient or permanent. OBJECTIVES: Assess the clinical characteristics of patients with unprovoked thromboembolic events and the role of inherited thrombophilia as a causative or additive risk factor. METHODS: 210 consecutive adult patients with unprovoked thromboembolic events were reviewed in hematology units at three tertiary Egyptian centers between September 2022 and September 2023. The diagnosis of thromboembolic events was confirmed by clinical and radiological findings. Laboratory screening for thrombophilia-associated. RESULTS: Among our patients, 53(25.2%) patients presented with isolated DVT, followed by portal vein thrombosis, 32(15.2%) had a pulmonary embolism, and sagittal sinus thrombosis was developed in 23(10.9%) patients. CONCLUSION: Younger people who experience spontaneous thromboembolism run the chance of having hereditary thrombophilia; the more mutations discovered, the higher the risk of thrombosis; the lower leg and deep vein thrombosis were the most common sites. Lastly, MTHFR C677T was the most common polymorphism in Egyptians, detected in almost half of the cases.

Total shoulder arthroplasty in patients with factor V Leiden.

BACKGROUND: Anatomic and reverse total shoulder arthroplasty (TSA) are effective treatment options for end-stage glenohumeral osteoarthritis. However, consideration for pre-existing conditions must be taken into account. Factor V Leiden (FVL), the most common inherited thrombophilia, is one such condition that predisposes to a prothrombotic state and may affect perioperative and longer-term outcomes following TSA. METHODS: Adult patients undergoing primary TSA for osteoarthritis indication were identified in the 2010 through October 2021 PearlDiver M157 database. Patients with or without FVL were matched at a 1:4 ratio based on age, sex, and Elixhauser Comorbidity Index. Ninety-day adverse events and 5-year revision rates were assessed and compared with multivariable logistic regression and rank-log tests, respectively. Finally, the relative use and bleeding/clotting outcomes were assessed based on venous thromboembolic (VTE) prophylactic agents used, with categories defined as (1) warfarin, heparin, or direct oral anticoagulant (DOAC) or (2) aspirin/no prescription found. RESULTS: Of 104,258 TSA patients, FVL was identified for 283 (0.27%). Based on matching, 1081 patients without FVL and 272 patients with FVL were selected. Multivariable analyses demonstrated that those with FVL displayed independently greater odds ratios (ORs) of deep vein thrombosis (DVT, OR = 9.50, P < .0001), pulmonary embolism (PE, OR = 10.10, P < .0001), and pneumonia (OR = 2.43, P = .0019). Further, these events contributed to the increased odds of aggregated minor (OR = 1.95, P = .0001), serious (OR = 6.38, P < .0001), and all (OR = 3.51, P < .0001) adverse events. All other individual 90-day adverse events, as well as 5-year revision rates, were not different between the study groups. When compared to matched patients without FVL on the same anticoagulant agents, FVL patients on warfarin, heparin, or DOAC agents demonstrated lesser odds of 90-day DVT and PE (OR = 4.25, P < .0001, and OR = 2.54, P = .0065) than those on aspirin/no prescription found (OR = 7.64 and OR = 21.95, P < .0001 for both). Interestingly, those on VTE prophylactic agents were not at greater odds of bleeding complications (hematoma or transfusion). DISCUSSION AND CONCLUSIONS: TSA patients with FVL present a difficult challenge to shoulder reconstruction surgeons. The current study highlights the strong risk of VTE that was reduced but still significantly elevated for those with stronger classes of VTE chemoprophylaxis. Acknowledging this risk is important for surgical planning and patient counseling, but also noted was the reassurance of similar 5-year revision rates for those with vs. without FVL.

Total Hip Arthroplasty in Patients Who Have Factor V Leiden: Elevated Risks Isolated to Venous Thromboembolism Events.

BACKGROUND: Total hip arthroplasty (THA) is a common procedure that requires consideration of preexisting comorbidities. Factor V Leiden (FVL), an inherited thrombophilia, is one such condition that predisposes patients to venous thromboembolism (VTE, deep vein thrombosis, and pulmonary embolism). The present study aimed to characterize the risks associated with FVL patients undergoing THA and evaluate the effect of VTE chemoprophylactic agents on these risks. METHODS: A total of 544,022 adult patients who underwent primary THA for osteoarthritis indications between 2010 and October 2021 were identified in an administrative claims database. Of these, FVL was identified in 1,138 (0.21%). Patients who had and did not have FVL were matched at a 1:4 ratio (1,131 with FVL and 4,519 without FVL) based on age, sex, and Elixhauser comorbidity index. Univariable and multivariable analyses were assessed for 90-day complications. Implant survival at 5 years was assessed and compared with log-rank tests. The relative use of different chemoprophylactic agents, including aspirin, warfarin, heparin, or direct oral anticoagulant (DOAC), was assessed. Bleeding events and VTE were compared for those prescribed either aspirin or warfarin, heparin, or DOAC. A Bonferroni correction was applied. RESULTS: On multivariable analysis, FVL patients were found to have increased odds of 90-day deep vein thrombosis (odds ratio (OR) = 9.20), pulmonary embolism (OR = 6.89), and aggregated severe and all adverse events (OR = 4.74 and 1.98, respectively), but not elevated risk of other perioperative adverse events or 5-year reoperations. More potent chemoprophylactic agents (warfarin, heparin, DOAC) reduced, but did not completely eliminate, the increased VTE risks (without increasing bleeding events). CONCLUSIONS: This study quantified the significantly elevated VTE risk associated with FVL patients undergoing THA. The lack of difference in other specific adverse events and 5-year reoperations is reassuring. Clearly, chemoprophylactic agents are important in this population and may need further attention.

The Etiology of the Thrombotic Phenomena Involved in the Process of Coronary Artery Disease-What Is the Role of Thrombophilic Genes in the Development of This Pathology?

Cardiovascular diseases, among which includes coronary artery disease, represent one of the most important causes of mortality and morbidity worldwide. Research aimed at determining the risk factors involved recognizes a group of "traditional" risk factors, but also more recent studies identified over 100 "novel" ones which may have a role in the disease. Among the latter is the thrombophilia profile of a patient, a pathology well-established for its involvement in venous thromboembolism, but with less studied implications in arterial thrombosis. This paper reviews the literature, explaining the pathophysiology of the thrombophilia causes associated most with coronary thrombosis events. Results of several studies on the subject, including a meta-analysis with over 60,000 subjects, determined the significant involvement of factor V Leiden, prothrombin G20210A mutation, plasminogen activator inhibitor-1 and antiphospholipid syndrome in the development of coronary artery disease. The mechanisms involved are currently at different stages of research, with some already established and used as therapeutic targets.

Contribution of genotypes in Prothrombin and Factor V Leiden to COVID-19 and disease severity in patients at high risk for hereditary thrombophilia.

Thrombotic and microangiopathic effects have been reported in COVID-19 patients. This study examined the contribution of the hereditary thrombophilia factors Prothrombin (FII) and Factor V Leiden (FVL) genotypes to the severity of COVID-19 disease and the development of thrombosis. This study investigated FII and FVL alleles in a cohort of 9508 patients (2606 male and 6902 female) with thrombophilia. It was observed that 930 of these patients had been infected by SARS-CoV-2 causing COVID-19. The demographic characteristics of the patients and their COVID-19 medical history were recorded. Detailed clinical manifestations were analyzed in a group of cases (n = 4092). This subgroup was age and gender-matched. FII and FVL frequency data of healthy populations without thrombophilia risk were obtained from Bursa Uludag University Medical Genetic Department's Exome Databank. The ratio of males (31.08%; 27.01%) and the mean age (36.85 ± 15.20; 33.89 ± 14.14) were higher among COVID-19 patients compared to non-COVID-19 patients. The prevalence of FVL and computerized tomography (CT) positivity in COVID-19 patients was statistically significant in the thrombotic subgroup (p < 0.05). FVL prevalence, CT positivity rate, history of thrombosis, and pulmonary thromboembolism complication were found to be higher in deceased COVID-19 patients (p < 0.05). Disease severity was mainly affected by FVL and not related to genotypes at the Prothrombin mutations. Overall, disease severity and development of thrombosis in COVID-19 are mainly affected by the variation within the FVL gene. Possible FVL mutation should be investigated in COVID-19 patients and appropriate treatment should be started earlier in FVL-positive patients.

Tranexamic acid is safe and effective in patients with heterozygous factor V Leiden mutation during total joint arthroplasty.

INTRODUCTION: Patients with an inherent hypercoagulable state are at a higher risk of venous thromboembolism (VTE) following total joint arthroplasty (TJA). Further administration of tranexamic acid (TXA) during TJA may increase the risk of VTE in these high-risk patients. There is no study that specifically analyzes the safety and efficacy of TXA during TJA in patients with factor V Leiden (FVL) mutation; therefore, the purpose of this study was to evaluate the safety and efficacy of TXA use on the risk of VTE and bleeding in patients carrying FVL mutation. MATERIALS AND METHODS: A total of 42 patients with FVL mutation (22 hips, 20 knees) and 40 control patients (20 hips, 20 knees) who underwent TJA were retrospectively reviewed. All patients received 1 g TXA intravenously 15 min before the skin incision and 2 g of TXA was administered locally at the surgical site as a periarticular injection. Pharmacological thromboprophylaxis (low-molecular-weight heparin) was administered to all patients. Estimated blood loss and in-hospital thromboembolic complications were compared between the groups. RESULTS: In both total knee arthroplasty (TKA) and total hip arthroplasty (THA) patients, there was no significant difference in the amount of estimated blood loss among the groups (p = 0.980, and p = 0963, respectively). None of the patients in the THA group received a blood transfusion. The transfusion rate was similar in the TKA group (p = 0.756, one patient in each group). No VTE, myocardial infarction, or any other complications related to TXA use were observed in any of the patients. CONCLUSIONS: The combined local and systemic administration of TXA could be safely used in patients with heterozygous FVL mutation receiving pharmacological thromboprophylaxis during TJA without increasing the risk of VTE.

Cocaine Abuse as an Immunological Trigger in a Case Diagnosed with Eales Disease.

Background: Eales disease is a clinical syndrome affecting the mid-peripheral retina with an idiopathic occlusive vasculitis and possible subsequent retinal neovascularization. The disease can develop into visually threatening complications. Case Presentation: We report the case of a 40-year-old Caucasian male with a history of cocaine abuse who presented with blurred vision in the left eye (LE). Fundus examination showed vitreous hemorrhages, peripheral sheathing of venous blood vessels, areas of retinal neovascularization in the LE, and peripheral occlusive phlebitis in the right eye. The full serologic panel was negative except for the heterozygous mutation of factor V Leiden. Clinical and biochemical parameters suggested a diagnosis of Eales disease. Therapy with dexamethasone, 1 mg per kg per day, tapered down slowly over 4 months, and peripheral laser photocoagulation allowed a regression of clinical signs and symptoms. Conclusion: This case shows an uncommon presentation of Eales disease associated with cocaine abuse. Both cocaine abuse and a thrombophilic pattern, as cofactors, might have sensitized the retinal microcirculation on the pathogenetic route to this retinal pathology. Furthermore, in view of this hypothesis, a thorough ocular and general medical history investigating drug abuse and coagulation disorders is recommended for ophthalmologists in such cases.

Heterozygous factor V Leiden mutation manifesting with combined central retinal vein occlusion, cilioretinal artery occlusion, branch retinal artery occlusion, and anterior ischaemic optic neuropathy: a case report.

BACKGROUND: Our purpose was to describe a patient who developed combined central retinal vein occlusion (CRVO), cilioretinal artery occlusion, branch retinal artery occlusion (BRAO), and anterior ischaemic optic neuropathy (AION) followed by CRVO in the second eye because of the heterozygous factor V Leiden (FVL) mutation. CASE PRESENTATION: A 39-year-old female with a history of recurrent pregnancy losses presented with acute blurred vision in the right eye (RE), with visual acuity limited to counting fingers. She was diagnosed with combined impending CRVO, cilioretinal artery occlusion, BRAO, and AION. The results of thrombophilia testing, not including the FVL mutation, were negative. Retinal atrophy with vascular attenuation and optic disc pallor developed after resolution of acute retinal findings. Nine months after initial presentation, the patient developed an impending CRVO in the left eye (LE), with a secondary progression to a complete CRVO causing a decrease in best corrected visual acuity (BCVA) to 20/40. The patient was determined to be heterozygous for the FVL mutation. She subsequently was treated with acenocoumarol. At the last follow-up visit, the BCVA was 20/400 in the RE and 20/20 in the LE, and there was a complete resolution of the acute CRVO findings in the LE. CONCLUSION: Our case shows that the heterozygous FVL mutation may manifest with combined retinal vascular occlusion involving multiple sites in both eyes. Early recognition of such an inherited thrombophilic disorder is important because it implies the need for long-term anticoagulative therapy to reduce the patient's risk of recurrent, sight-threatening and life-threatening thrombotic events.

The prognostic role of extended preoperative hypercoagulability work-up in high-risk microsurgical free flaps: a single-center retrospective case series of patients with heterozygotic factor V Leiden thrombophilia.

INTRODUCTION: Hypercoagulability is associated with an increased risk of microvascular complications and free flap failures. The authors present their experience and approach to diagnosing and treating patients with heterozygotic factor V Leiden (hFVL) thrombophilia undergoing free flap reconstruction. METHODS: Between November 2009 and June 2018, 23 free flap surgeries were performed in 15 hypercoagulable patients with hFVL. According to the timing of perioperative hypercoagulability work-up, they were grouped into flaps with established diagnoses prior to surgery (Group A) versus flaps with unknown diagnoses prior to surgery (Group B). Baseline characteristics and perioperative complications were compared between both groups, including revision surgeries due to microvascular thromboses, acute bleedings, hematomas, flap necroses, and reconstructive failures. RESULTS: HFVL mutations had been confirmed preoperatively in 14 free flap surgeries (61%, Group A), whereas in 9 free flap surgeries (39%, Group B), mutations were only diagnosed postoperatively after the occurrence of microvascular thromboses had warranted extended hypercoagulability work-up. The overall rate of intraoperative flap thromboses was 9% (n = 2), whereas the overall rate of postoperative flap thromboses was 43% (n = 10). The corresponding salvage rates were 100% (n = 2/2) for intraoperative and 40% (n = 4/10) for postoperative pedicle thromboses. A total of five free flaps were lost (22%). Upon comparison, flaps with an unconfirmed diagnosis prior to surgery were at ten times higher risk for developing total necroses (flaps lost in Group B = 4/9 versus Group A = 1/14; OR: 10.4; 95% CI 1.0, 134.7; p = 0.03). CONCLUSION: Meticulous preoperative work-up of patients with any history of hypercoagulability can help reduce free flap loss rates, thus improving surgical outcomes and increasing patient safety.

Activated protein C resistance impact on Syrian candidates for in vitro fertilisation and the benefit of anticoagulation therapy: a retrospective cohort study.

Activated protein C resistance (APCR) is a common thrombophilia, caused mainly by a mutation. The impact of APCR on the efficacy of In Vitro Fertilization (IVF) are still unclear, and no solid recommendations for its management were published. To investigate the effect of APCR on IVF outcomes and assess the efficacy of our management protocol, we retrospectively scanned the medical records of women who were tested with APCR assay in 2019 at our fertility centre. The 66 women (12%) positive for APCR had lower odds of reaching clinical pregnancies after IVF 0.18 [95% CI: 0.07-0.47] and fewer live births. The administration of low-molecular-weight heparin and aspirin associated with more implantation in treated compared to untreated APCR-positive women with an odds ratio of 43.2 [7.51-248.6]. In conclusion, APCR negatively affects the number of clinical pregnancies after IVF, but anticoagulation therapy can mitigate this effect and significantly increase clinical pregnancies.Impact StatementWhat is already known on this subject? The evidence about the impact of APCR on IVF outcomes is still inconclusive. According to the Canadian guideline, routine screening for thrombophilia in patients with recurrent pregnancy loss is not recommended. No clear recommendations regarding the management of APCR in the planning for IVF are yet available.What do the results of this study add? APCR significantly increases implantation failure among infertile women who conduct IVF. Management of APCR using LMWH and aspirin was effective in mitigating this effect and increasing successful implantation.What are the implications of these findings for clinical practice and/or further research? Our findings can support the recommendation to include APCR assay in the routine tests for infertile women conducting IVF, and suggest the combination between LMWH and aspirin as an effective therapy to increase successful implantation in APCR positive candidates. However, more controlled clinical trials are still needed to confirm our results.

RISK OF PLACENTA-ASSOCIATED COMPLICATIONS AT PREECLAMPSIA IN PREGNANT WOMEN WITH THROMBOPHILIA.

OBJECTIVE: The aim: To study the distribution and influence of coagulation factor gene polymorphisms, endothelial dysfunction, blood pressure regulator on the development of obstetric and perinatal complications in women with preeclampsia (PE). PATIENTS AND METHODS: Materials and methods: The prospective cohort study included 46 women with PE and maternal or fetal complications and 87 pregnant women with PE, without complications. Genetic polymorphisms of coagulation factors and fibrinolysis (1691 G→A FVL, 20210 G→A prothrombin, 675 5G/4G PAI-1, 455 G→A fibrinogen ß), endothelial dysfunction (192 Q→R PON-1, 677 C→T MTHFR) and blood pressure regulator (235 M→T angiotensinogen II) were studied with the help of allele-specific polymerase chain reaction. RESULTS: Results: Markers of predisposition to the development of obstetric and perinatal complications in pregnant women with PE are the following genotypes: 1691 GA by V Leiden factor gene - increases the risk in 2.9 times (95% CI 1.94-4.33), 20210 GA by prothrombin gene - in 2.36 times (95% CI 1.54-3.6), 20210 AA by prothrombin gene - in 3.12 times (95% CI 2.4-4.0). Pathological polymorphisms in the genes of angiotensinogen II 235 M→T, PAI-1 5G/4G, fibrinogen ß 455 G→A, paraoxonase-1 192 Q→R do not significantly affect the development of complications during preeclampsia. CONCLUSION: Conclusions: The development of PE against the background of the existence of acquired and hereditary types of thrombophilia is associated with a more severe course, early-onset and the development of life-threatening complications for a mother and fetus.

Hereditary thrombophilia and recurrent pregnancy loss: a systematic review and meta-analysis.

STUDY QUESTION: Is there an association between hereditary thrombophilia in pregnant women and risk of recurrent pregnancy loss (RPL)? SUMMARY ANSWER: Pregnant women with hereditary thrombophilia have an increased risk of RPL, especially for pregnant women with the G1691A mutation of the factor V Leiden (FVL) gene, the G20210A mutation of the prothrombin gene (PGM), and deficiency of protein S (PS). WHAT IS KNOWN ALREADY: Prior studies have suggested that pregnant women with hereditary thrombophilia have a higher risk of RPL, however, the results are inconsistent; furthermore, a complete overview is missing. This lack of information is an obstacle to the risk assessment of RPL in pregnant women with hereditary thrombophilia. A comprehensive meta-analysis on the relation between hereditary thrombophilia and the risk of RPL is needed. STUDY DESIGN, SIZE, DURATION: A systematic review and meta-analysis was performed using observational studies published in English before 1 April 2020 to evaluate the relation between hereditary thrombophilia and risk of RPL. PARTICIPANTS/MATERIALS, SETTING, METHODS: Relevant studies were identified from PubMed, Web of Science, and EMBASE searches and complemented with perusal of bibliographies of retrieved articles. The exposure of interest was hereditary thrombophilia, including FVL mutation, PGM, deficiency of antithrombin (AT), deficiency of protein C (PC), and deficiency of PS. The overall risk estimates were pooled using random effects models. Subgroup and sensitivity analyses were carried out to explore possible sources of heterogeneity and assess the robustness of the results. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 89 studies involving 30 254 individuals were included. Results showed that women with FVL mutation (odds ratio (OR): 2.44, 95% CI: 1.96-3.03), PGM (OR: 2.08, 95% CI: 1.61-2.68), or deficiency of PS (OR: 3.45, 95% CI: 1.15-10.35) had higher risks of developing RPL. Compared with the reference group, there was no observed relation between a deficiency in AT or PC and RPL (all P > 0.05). Heterogeneity in the risk estimates of RPL was partially explained by geographic region, definitions of RPL, types of RPL, and controlled confounders. Sensitivity analyses validated the robustness of the findings. LIMITATIONS, REASONS FOR CAUTION: Only 39 of the included studies controlled for one or more confounders, and the heterogeneity across all included studies was high. Based on the data available, we cannot determine whether this association is confounded by other potential risk factors of RPL. WIDER IMPLICATIONS OF THE FINDINGS: This systematic review and meta-analysis show a possible association between hereditary thrombophilia and an increased risk of RPL, suggesting that testing for hereditary thrombophilia should be considered in individuals with RPL. STUDY FUNDING/COMPETING INTEREST(S): The study was funded by the Hunan Provincial Key Research and Development Program (Grant number: 2018SK2062) and National Natural Science Foundation Program (Grant number: 81973137). There are no conflicts of interest. REGISTRATION NUMBER: N/A.

A case-control study on factor V Leiden: an independent, gender-dependent risk factor for venous thromboembolism.

BACKGROUND: Activated protein C resistance (APCR) due to factor V Leiden (FVL) mutation (R506Q) is a major risk factor in patients with venous thromboembolism (VTE). The present study investigated the clinical manifestations and the risk of venous thromboembolism regarding multiple clinical, laboratory, and demographic properties in FVL patients. MATERIAL AND METHODS: A retrospective cross-sectional analysis was conducted on a total of 288 FVL patients with VTE according to APCR. In addition, 288 VET control samples, without FVL mutation, were also randomly selected. Demographic information, clinical manifestations, family and treatment history were recorded, and specific tests including t-test, chi-square and uni- and multi-variable regression tests applied. RESULTS: APCR was found to be 2.3 times significantly more likely in men (OR: 2.1, p < 0.05) than women. The risk of deep vein thrombosis (DVT) and pulmonary embolism (PE) in APCR patients was 4.5 and 3.2 times more than the control group, respectively (p < 0.05). However, APCR could not be an independent risk factor for arterial thrombosis (AT) and pregnancy complications. Moreover, patients were evaluated for thrombophilia panel tests and showed significantly lower protein C and S than the control group and patients without DVT (p < 0.0001). CONCLUSION: FVL mutation and APCR abnormality are noticeable risk factors for VTE. Screening strategies for FVL mutation in patients undergoing surgery, oral contraceptive medication, and pregnancy cannot be recommended, but a phenotypic test for activated protein C resistance should be endorsed in patients with VTE.

Prevalence of rare F5 variants in general population from Bosnia and Herzegovina.

Human gene F5, encoding coagulation factor V, was previously reported to be highly polymorphic. Apart from FV Leiden, several other rare variants have been detected in clinical practice and associated with thrombotic events, especially in cases when patient's phenotype and FV Leiden genotype were not in agreement. In this study, the prevalence of 17 rare F5 variants has been studied on a sample of 130 healthy adult individuals from the general Bosnian-Herzegovinian population. DNA was isolated from buccal swab samples, while genotyping was performed using MALDI-TOF MS method. The results have shown that Asp2194Gly and Met2120Thr are polymorphic in the study population with minor allele frequencies of 0.077 and 0.073, respectively. Additionally, these two variants were mutually exclusive with FV Leiden and none of them was positively associated with participants' family history of cardiovascular or cerebrovascular diseases. While the obtained results are in agreement with previously reported data for the general Caucasian populations, it is worth noting that only two rare F5 variants were detected in the study population, albeit at considerable frequencies. Still, scientific information on rare F5 variants is rather scarce and further studies aiming to assess functional importance of these variants, as well as their role as prothrombotic factors are necessary.

MTHFR (C677T, A1298C), FV Leiden polymorphisms, and the prothrombin G20210A mutation in arterial ischemic stroke among young tunisian adults.

Arterial ischemic stroke (AIS) in young adults is less common in older adults, but the underlying pathogenesis and risk factors are more multi-faceted. The role of inherited thrombophilia such as 5, 10-methylenetetrahydrofolate reductase (MTHFR) gene polymorphism, (C677T and A1298C), factor V of Leiden (FVL) polymorphism, and the prothrombin G20210A mutations remains unclear. This study aims to evaluate the role of prothrombin genetic factor in AIS among young adults in Tunisia and to assess the synergistic effect between thrombogenic mutations in the pathogenesis of AIS. In this case-control study, blood samples were collected from patients and healthy controls, all matched for age and gender. The difference between them is evaluated by using the chi-square test. The odds ratio (OR) was carried out to evaluate the associations between each polymorphism and AIS risk using a binary logistic regression model. Values were considered statistically significant when p < 0.05. Patients carrying simultaneously the MTHFR polymorphisms (677T and 1298C) have a higher risk to develop AIS compared to controls. The heterozygous variants FVL increased the risk of AIS only when it is associated with MTHFR C677T or MTHFR A1298C polymorphisms. In conclusion, our study confirmed the involvement of MTHFR polymorphisms as AIS's important risk factors. The existence of FVL polymorphism or prothrombin G20210A mutation alone doesn't correlate with the occurrence of stroke. We assume that the presence of both MTHFR and FVL polymorphisms has a synergistic effect and increased the risk of the AIS.

Risk Factors of Venous Thromboembolism in Sudanese Pregnant Women.

Venous thromboembolism (VTE) is one of the major causes of pregnancy-related mortality and morbidity. This study aimed to determine the frequency of factor V Leiden (FVL) and prothrombin G20210A polymorphisms and measure the plasma levels of protein C (PC), protein S (PS) and antithrombin (AT) in pregnant women with VTE and healthy pregnant women. This prospective case-control study determined the frequencies of FVL G1691A and prothrombin G20210A polymorphisms and measured the plasma levels of PC, PS and AT in 198 pregnant women with VTE and 198 healthy pregnant women. Allele-specific polymerase chain reaction (ASPCR) was used to detect the FVL G1691A polymorphisms and prothrombin G20210A gene mutations. The FVL G1691A polymorphism and prothrombin G20210A gene mutations were detected only in pregnant women with VTE, with frequencies of 4.0 and 0.5%, respectively. The highest frequency of FVL G1691A polymorphism was observed in patients with deep vein thrombosis (DVT) and positively associated with contraceptive use and termination. Pregnant women with VTE had significantly lower levels of PC, PS and AT than those of controls. In conclusion, among the VTE cases, FVL G1691A polymorphism and PC, PS and AT deficiencies were the most common findings in patients presenting with DVT. Antithrombin deficiency was more common than PC and PS deficiencies. Contraceptive use, high body mass index (BMI) and termination correlated strongly with FVL G1691A polymorphism and PC and PS deficiencies in patients with VTE.

ARMS TaqMan real-time PCR for genotyping factor V Leiden mutation in Han Chinese.

Factor V Leiden (FVLeiden ) is a missense mutation of 1691 position (G1691A) in exon 10 of FV gene, and being a genetic risk for venous thrombosis. Currently, there are several PCR-based methods for detecting FVLeiden mutation; however, these methods have disadvantages such as time-consuming, cumbersome steps and potentially hazardous gels. The aims of present study were to develop a simple, time-saving, accurate, and gel-free method, called amplification refractory mutation system (ARMS) TaqMan real-time PCR, for detecting FVLeiden mutation. We severally designed two specific reverse primers for mutant and wild-type through intentional introduction of mismatched nucleotide at the penultimate 3' position. Although target amplicon amplification efficiency is reduced, but another corresponding amplicon is almost completely inhibited. Then, specific TaqMan-probe was designed to detect target amplicon. Established method was used to detect 500 unselected samples in Han Chinese, the results showed 499 cases of wild-type and one heterozygote. Afterward, 50 randomly picked wild-type cases and one heterozygote were reexamined by bidirectional DNA sequencing, which is considered as "Gold standard method." Exhilaratingly, the results detected by the two methods were completely consistent. At last, allelic frequency of FVLeiden was calculated the in Han Chinese. Given the above results, A FVLeiden heterozygote has been found in 500 random samples in Han Chinese, and the allelic frequency was 0.1%. In conclusion, the ARMS TaqMan real-time PCR is an ideal detecting system for genotyping FVLeiden mutation in clinical application, and FVLeiden mutation exists in Han Chinese despite extremely low prevalence.

DOAC-Remove abolishes the effect of direct oral anticoagulants on activated protein C resistance testing in real-life venous thromboembolism patients.

Background Direct oral anticoagulants (DOACs) may cause false results of activated protein C resistance (APC-R) ratio. DOAC-Remove, a new reagent based on activated carbon, has been designed to eliminate the interference of DOACs on coagulation assays. The aim of the study was to investigate whether the use of DOAC-Remove enables to determine APC-R in patients treated with DOACs. Methods We assessed 74 venous thromboembolism (VTE) patients, including 25 on rivaroxaban, 25 on apixaban and 24 taking dabigatran. APC-R was determined using the Russell Viper Venom Time (RVVT)-based clotting test. APC-R and DOAC concentrations were tested at baseline and following DOAC-Remove. Thrombophilia, including factor V Leiden (FVL) mutation was tested. Results FVL mutation was found in 20 (27%) patients. The APC-R ratio at baseline was measurable in 43 patients (58.1%), including 20 (80%) on rivaroxaban, 19 (76%) on apixaban and four (16.7%) on dabigatran. In patients with measurable APC-R at baseline, the ratio >2.9 was found in 23 patients (53.5%). In 16 (37.2%) subjects APC-R ratio <1.8 suggested FVL mutation which was genetically confirmed. Four (9.3%) FVL carriers on dabigatran showed negative/equivocal APC-R results. In 11 (14.9%) patients taking rivaroxaban or apixaban, in whom blood was collected 2-5 h since the last dose, we observed unmeasurable APC-R. DOAC-Remove almost completely eliminated all plasma DOACs. After addition of DOAC-Remove all APC-R ratios were measurable. In four FVL carriers on dabigatran with false negative APC-R, DOAC-Remove resulted in APC-R ratios <1.8. Conclusions DOAC-Remove effectively reduces DOACs concentration in plasma, which enables FVL testing using APC-R.

Validation and standardization of the ETP-based activated protein C resistance test for the clinical investigation of steroid contraceptives in women: an unmet clinical and regulatory need.

Background Regulatory bodies recommend the use of an assay based on the assessment of the endogenous thrombin potential (ETP) for the investigation of the activated protein C resistance (APCr) in the development of steroid contraceptives in women. However, the assays described in the literature are home-made and not standardized regarding the method, the reagents, the reference plasma and the quality controls. In the absence of any commercially available method, we aimed at validating the ETP-based APCr assay. Methods The validation was performed according to regulatory standards. The method targets a 90% inhibition of the ETP in healthy donors in the presence of APC compared to the same condition in the absence of APC. As a large-scale production of a pool of plasma from well-selected healthy donors is impossible, algorithms were applied to a commercial reference plasma to correlate with the selected pool. Results Repeatability and intermediate precision passed the acceptance criteria. The assay demonstrated a curvilinear dose response to protein S and APC concentrations (R2 > 0.99). Analysis of plasma samples from 47 healthy individuals (22 women not taking combined hormonal contraceptives [CHC], and 25 men not Factor V Leiden carriers) confirmed the validity of the test, with a mean inhibition percentage of 90%. Investigations in 15 women taking different contraceptives and in two subjects with Factor V Leiden confirmed the good sensitivity and performance of the assay. Conclusions This validation provides the pharmaceutical industry, the regulatory bodies and physicians with a reproducible, sensitive and validated gold-standard ETP-based APCr assay.