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Introduction: Congenital factor V (FV) deficiency is a rare clotting disorder affecting ∼1 in 1,000,000, with bleeding severity that ranges broadly for poorly understood reasons. Aim: To help understand the molecular basis of the observed phenotype in FV deficient patients, the genetics and biochemistry causing a patient's FV deficiency were evaluated. Methods and Results: A 71‐year‐old female, who had serious life‐long bleeding upon provocation and profound menorrhagia that lead to hysterectomy, was found to have 3% of normal plasma FV antigen with normal electrophoretic mobility. Platelet FV was similarly low, although the banding pattern was less fragmented than normal. Plasma clotting activity was <1% of normal. Familial inheritance and DNA sequence analysis from peripheral blood leukocytes were consistent with novel compound heterozygosity with missense mutations in exon XVII, Leu1821 to Ser (L1821S) and exon XXV, Gly2192 to Cys (G2192C). The respective single‐mutation variants were expressed and purified. Explaining why the antigen level and activity were inequivalent, thrombin activation of recombinant (r) FV/L1821S was impaired, and rFV/G2192C was unable to bind to a procoagulant phospholipid membrane. Conclusion: These findings are consistent with the observed phenotype, highlighting the importance of understanding FV biochemical function to rationalize clinical bleeding severity when the circulating antigen level is discordant. [ABSTRACT FROM AUTHOR]

Multiple Coagulation Factor Deficiency (MCFD) is an uncommon haematological disorder characterised by simultaneous deficiency of multiple clotting factors, which leads to an increased risk of bleeding and compromised haemostasis. The present series aimed to provide a comprehensive analysis of MCFD, shedding light on its aetiology, clinical manifestations, and potential therapeutic interventions in a case-by-case manner, highlighting the individualised treatment options that are needed for many of these cases. A prospective combined clinical and laboratory study was performed on all patients who presented with bleeding tendencies and were subsequently diagnosed with MCFD. Clinical records and laboratory data of all these patients were reviewed to identify commonalities and variations among affected individuals. Haematological assays were performed to quantify the extent of coagulation factor deficiencies. The present series identified a diverse cohort of eight individuals (2 females and 6 males) with MCFD, showcasing variability in age of onset, severity of symptoms, and potential underlying genetic mutations. Clotting factor assays revealed deficiencies of Factor V and Factor VIII (F5F8D), Vitamin K-dependent clotting factor deficiency, and failure of synthesis of clotting factors of hepatic origin. Clinically, patients presented with a spectrum of bleeding phenotypes, ranging from mild to severe, requiring tailored therapeutic strategies. The present series provides an overview of some of the presentations of MCFD, emphasising the need for a multidisciplinary approach in its diagnosis and management. Clinicians should be vigilant in recognising the varied clinical presentations of MCFD and consider genetic testing for precise diagnosis and management. [ABSTRACT FROM AUTHOR]

Leishmaniases are a collection of neglected tropical diseases caused by kinetoplastid parasites in the genus Leishmania. Current chemotherapies are severely limited, and the need for new antileishmanials is of pressing international importance. Bromodomains are epigenetic reader domains that have shown promising therapeutic potential for cancer therapy and may also present an attractive target to treat parasitic diseases. Here, we investigate Leishmania donovani bromodomain factor 5 (LdBDF5) as a target for antileishmanial drug discovery. LdBDF5 contains a pair of bromodomains (BD5.1 and BD5.2) in an N-terminal tandem repeat. We purified recombinant bromodomains of L. donovani BDF5 and determined the structure of BD5.2 by X-ray crystallography. Using a histone peptide microarray and fluorescence polarization assay, we identified binding interactions of LdBDF5 bromodomains with acetylated peptides derived from histones H2B and H4. In orthogonal biophysical assays including thermal shift assays, fluorescence polarization, and NMR, we showed that BDF5 bromodomains bind to human bromodomain inhibitors SGC-CBP30, bromosporine, and I-BRD9; moreover, SGC-CBP30 exhibited activity against Leishmania promastigotes in cell viability assays. These findings exemplify the potential BDF5 holds as a possible drug target in Leishmania and provide a foundation for the future development of optimized antileishmanial compounds targeting this epigenetic reader protein.

Multiple Coagulation Factor Deficiency (MCFD) is an uncommon haematological disorder characterised by simultaneous deficiency of multiple clotting factors, which leads to an increased risk of bleeding and compromised haemostasis. The present series aimed to provide a comprehensive analysis of MCFD, shedding light on its aetiology, clinical manifestations, and potential therapeutic interventions in a case-by-case manner, highlighting the individualised treatment options that are needed for many of these cases. A prospective combined clinical and laboratory study was performed on all patients who presented with bleeding tendencies and were subsequently diagnosed with MCFD. Clinical records and laboratory data of all these patients were reviewed to identify commonalities and variations among affected individuals. Haematological assays were performed to quantify the extent of coagulation factor deficiencies. The present series identified a diverse cohort of eight individuals (2 females and 6 males) with MCFD, showcasing variability in age of onset, severity of symptoms, and potential underlying genetic mutations. Clotting factor assays revealed deficiencies of Factor V and Factor VIII (F5F8D), Vitamin K-dependent clotting factor deficiency, and failure of synthesis of clotting factors of hepatic origin. Clinically, patients presented with a spectrum of bleeding phenotypes, ranging from mild to severe, requiring tailored therapeutic strategies. The present series provides an overview of some of the presentations of MCFD, emphasising the need for a multidisciplinary approach in its diagnosis and management. Clinicians should be vigilant in recognising the varied clinical presentations of MCFD and consider genetic testing for precise diagnosis and management.

Background: In healthy individuals, plasma levels of clotting proteins naturally vary within a range of 50% to 150% of their mean values. We do not know how these variations modify thrombin generation. Objectives: To assess the impact of protein level variations on simulated thrombin generation in normal and factor (F)VIII-, FIX-, or FXI-deficient blood. Methods: We used a mathematical model of flow-mediated coagulation to simulate thrombin generation with all possible combinations of clotting protein variations within the normal range and for various tissue factor levels. We selected, analyzed, and ranked combinations that enhanced thrombin generation compared with baseline. Results: Protein variations most strongly affected thrombin generation at intermediate tissue factor levels. Low tissue factor levels prevented coagulation initiation, while high tissue factor levels always triggered thrombin generation. At intermediate levels, we identified protein variations that substantially modified thrombin generation. Low-normal FV shortened lag times and increased thrombin generation, whereas high-normal FV lengthened lag times and reduced thrombin generation. With severe FVIII and FIX deficiencies, low-normal tissue factor pathway inhibitor α and antithrombin amplified the effect of low-normal FV. For moderate FVIII and FIX deficiencies, high-normal tissue factor pathway inhibitor α and antithrombin enhanced the impact of high-normal FV in reducing thrombin production. In normal and FXI-deficient blood, high-normal FVIII and FIX significantly boosted thrombin generation. Conclusion: Our mathematical model predicted how variations in clotting protein levels, within the normal range, could contribute to the variability of bleeding phenotypes observed with clotting factor deficiencies. Our study generated experimentally testable hypotheses that could aid in developing new therapies toward normal hemostasis.

Objective To study and compare the effects of different storage temperature and time on coagulation factor after cryoprecipitated antihemophilic factor(CAF) melting, and to provide reference for the establishment of industry standards. Methods From June 2021 to May 2023, a total of 96 bags of CAF were sampled in 4 bags per month, and timely detected in the same month. After the CAF was melted in a 37℃ water bath, the mild to moderate lipemic blood was labeled. Each bag of CAF and two 50 mL transfer bags were divided into two bags and two groups of 20 mL each using a sterile adapter. One group was placed in a 4℃ refrigerator and the other in a 22℃ water bath for 0 h, 4 h, 8 h, 12 h, 24 h and 48 h. Then 2 mL of aseptic sample was taken separately and put into the test tube, and 1mL of sample and 3 mL of buffer were added into the other test tube with the sampling gun and mixed on the machine for testing. The experimental data of 60 bags without mild to moderate lipemic blood cryoprecipitation and coagulation factor were randomly selected and statistically analyzed by SPSS21.0. Results After melting, CAF was stored for 0 h, 4 h, 8 h, 12 h, 24 h and 48 h to detect the average content and growth rate of coagulation factor in the two groups: 1) Storage at 4℃, factor Ⅷ content was 118.62, 111.57(-5.95%), 105.51(-11.05%), 103.30(-12.92%), 94.35(-20.46%) and 83.25(-29.82%) IU/ bag, respectively; Storage at 22℃, the factor Ⅷ content was 118.62, 112.69(-5.00%), 111.41(-6.08%), 109.01(-8.10%), 101.55(-14.39%) and 92.75(-21.81%) IU/ bag, and the storage results of the two groups were compared. At 24 h at 4℃ and 48 h at 22℃, the content of factor Ⅷ had significant statistical significance(P0.05). Conclusion After CAF melting, coagulation factor decreased with the extension of storage time, especially the decrease of factor Ⅷ, followed by factor V, while Fbg basically unchanged. Comparison between the two groups showed that, factor Ⅷ decay rate is slower, factor V decay rate is faster of storage at 22℃. CAF should be transfused as soon as possible after melting. If the delay is unavoidable, for the delay time less than 12 h, storage at 4℃ is recommended, fot the delay time more than 12 h and less than 24 h, storage at 22℃ is recommended.

Hypercoagulable disorders are best described as a group of acquired and hereditary conditions that increase the risk for the development of thrombi within veins or arteries. In the setting of an unprovoked venous thromboembolism, common practice in the inpatient setting has been further investigation via a thrombophilia workup to establish an underlying cause. Current Hematology-Oncology guidelines argue against inpatient workup as the results rarely influence inpatient management. Following American Society of Hematology guidelines (Middledorp), the current study found that only 15% (11/72) of patients met appropriate criteria for thrombophilia testing. There was no relationship between appropriate thrombophilia testing and diagnosis of thrombophilia or initiation of anticoagulation. There was a relationship between appropriate thrombophilia testing and Hematology-Oncology consultation. This demonstrates the need for expert consultation if thrombophilia testing is being considered. The current study provides more evidence that a strong recommendation against inpatient testing should be made as testing does not aid in diagnosis or change management and is an overutilization of healthcare resources. [ABSTRACT FROM AUTHOR]

Introduction: Oral anticoagulant therapy comes at the cost of a significant bleeding risk. However, it is hard to predict which patients are at risk of major bleeding. Previously we found associations of Calibrated Automated Thrombinography (CAT) parameters obtained in the presence of TIX-5 (an inhibitor of the FV activation by FXa), and plasma levels of TFPIα, γ’-fibrinogen and soluble thrombomodulin with major bleeding in the BLEEDS cohort, a cohort especially powered to find new biomarkers of major bleeding during VKA therapy. Methods: To determine and compare the predictive capability for major bleeding in the BLEEDS cohort of the above biomarkers, also in a combined model with clinical risk factors, we performed Univariable Prentice-weighted Cox regression analyses and Bayesian variable selection. Results: The highest predictive value among the laboratory measures were found for thrombin generation lagtime in the presence of TIX-5 (TIX-5 lagtime per 25% increase, hazard ratio (HR) 1.11, 95%CI 1.04–1.18, p=0.001) and full-length tissue factor pathway inhibitor (TFPIα) (per 25% increase HR 1.12, 95%CI 1.03–1.21, p=0.008), which remained significant after correction for multiple testing, and independently associated with major bleeding after Bayesian variable selection. Only the addition of TIX-5 lagtime to the clinical risk factors improved prediction of major bleeding significantly (p

IntroductionProposed mechanisms of acute traumatic coagulopathy (ATC) include decreased clotting potential due to factor consumption and proteolytic inactivation of factor V (FV) and activated factor V (FVa) by activated protein C (aPC). The role of FV/FVa depletion or inactivation in burn-induced coagulopathy is not well characterized. This study evaluates FV dynamics following burn and nonburn trauma.MethodsBurn and trauma patients were prospectively enrolled. Western blotting was performed on admission plasma to quantitate levels of FV antigen and to assess for aPC or other proteolytically derived FV/FVa degradation products. Statistical analysis was performed with Spearman's, Chi-square, Mann-Whitney U test, and logistic regression.ResultsBurn (n = 60) and trauma (n = 136) cohorts showed similar degrees of FV consumption with median FV levels of 76% versus 73% (P = 0.65) of normal, respectively. Percent total body surface area (TBSA) was not correlated with FV, nor were significant differences in median FV levels observed between low and high TBSA groups. The injury severity score (ISS) in trauma patients was inversely correlated with FV (ρ = -0.26; P = 0.01) and ISS ≥ 25 was associated with a lower FV antigen level (64% versus. 93%; P = 0.009). The proportion of samples showing proteolysis-derived FV was greater in trauma than burn patients (42% versus. 16%; P = 0.0006).ConclusionsIncreasing traumatic injury severity is associated with decreased FV antigen levels, and a greater proportion of trauma patient samples exhibit proteolytically degraded FV fragments. These associations are not present in burns, suggesting that mechanisms underlying FV depletion in burn and nonburn trauma are not identical.

Background: Recurrent reproductive failure is a global health issue affecting a significant number of women. Thrombophilias have been implicated as a possible cause. Inherited thrombophilias include a single nucleotide variant on factor V Leiden and prothrombin. Objective: The aim of this study was to evaluate the association between the following single nucleotide variants: factor V Leiden (c.1601G>A), the prothrombin gene (c.*97G>A) and the reproductive failure in the Polish population. Methods: The study was conducted in a group of 545 patients with recurrent pregnancy loss, RPL (≥2 miscarriages), and in a group of 641 patients with infertility. The distribution of genotypes for the selected variants were determined by RFLP-PCR and by the real-time PCR method. Results: A variant of the F5 gene was found in 5.14% of patients with RPL and in 6.08% of infertile women. A variant of the F2 gene was identified in 0.73% of patients with RPL and in 2.03% of women with infertility. The frequency in the study groups did not differ from that in the general population. No association between the studied variants of the F5 gene or the F2 gene and the predisposition to reproductive wastage was found. Conclusions: Recommendations for routine thrombophilia testing in women with recurrent miscarriages should be revisited. The decision regarding testing should be made individually depending on additional factors indicating an increased risk of venous thromboembolism. [ABSTRACT FROM AUTHOR]

Anchoring of coagulation factors to anionic regions of the membrane involves the C2 domain as a key player. The rate of enzymatic reactions of the coagulation factors is increased by several orders of magnitude upon membrane binding. However, the precise mechanisms behind the rate acceleration remain unclear, primarily because of a lack of understanding of the conformational dynamics of the C2-containing factors and corresponding complexes. We elucidate the membrane-bound form of the C2 domain from human coagulation factor V (FV–C2) by characterizing its membrane binding the specific lipid-protein interactions. Employing all-atom molecular dynamics simulations and leveraging the highly mobile membrane-mimetic (HMMM) model, we observed spontaneous binding of FV-C2 to a phosphatidylserine (PS)-containing membrane within 2–25 ns across twelve independent simulations. FV-C2 interacted with the membrane through three loops (spikes 1–3), achieving a converged, stable orientation. Multiple HMMM trajectories of the spontaneous membrane binding provided extensive sampling and ample data to examine the membrane-induced effects on the conformational dynamics of C2 as well as specific lipid-protein interactions. Despite existing crystal structures representing presumed “open” and “closed” states of FV-C2, our results revealed a continuous distribution of structures between these states, with the most populated structures differing from both “open” and “closed” states observed in crystal environments. Lastly, we characterized a putative PS-specific binding site formed by K23, Q48, and S78 located in the groove enclosed by spikes 1–3 (PS-specificity pocket), suggesting a different orientation of a bound headgroup moiety compared to previous proposals based upon analysis of static crystal structures.

Cerebral vein thrombosis (CVT) is a potentially fatal neurovascular disease that most commonly affects young women of child-bearing age. The risk of CVT has been reported to be increased by combined oral contraceptive (COC) use. Inherited thrombophilic conditions including factor V Leiden (FVL) and prothrombin G20210A mutation also increase the risk of CVT. Several studies have shown that a combination of risk factors including COC use plus an inherited thrombophilic disorder increases the risk of CVT in a multiplicative fashion. World Health Organization guidelines advise against the use of COC in women with hereditary thrombophilic defects due to the unacceptably elevated risk of thrombotic events. We report the case of a 16-year-old female with a six-week history of COC use who experienced 48 h of vomiting and ataxia and was found in cardiac arrest. Following cardiopulmonary resuscitation and hospitalization, brain imaging identified subarachnoid and intracranial hemorrhage. After a neurologic examination confirmed brain death, the patient expired. Genetic testing detected heterozygous mutations for both FVL and prothrombin G20210A. A brain only autopsy identified superior sagittal sinus thrombosis, subarachnoid hemorrhage, and foci of ischemic necrosis of the cerebral cortex. This case highlights the increased risk of CVT in patients with hereditary thrombophilia and oral contraceptive use. It is vital for physicians to identify risk factors for venous thrombosis and/or CVT prior to administration of COC.

Coagulation factorⅤ(FⅤ) deficiency is a rare genetic disorder that is primarily characterized by decreased FⅤ activity (FⅤC) and extensive clinical bleeding symptoms. Pregnancy combined with FⅤ deficiency is even more rare, and the risk of poor prognosis is higher. In this article, we report a case of severe homozygote hereditary FⅤ deficiency and postpartum hemorrhage occurring simultaneously. The patient was treated with ongoing high-dose blood product transfusions after being admitted to the hospital with a clear diagnosis of FⅤ deficiency, but the FⅤC did not improve adequately. Later, a live infant was delivered by cesarean section for acute fetal distress, bilateral uterine artery embolization was done for combined postpartum hemorrhage, and postoperative review revealed that the FⅤC had not returned to the normal level, and the patient was discharged with stable condition after symptomatic treatment. We now go over the medical history, the clinical course of this case, and the pertinent literature, with the aim of increasing clinicians′ understanding of this disease and accumulate more clinical experience. [ABSTRACT FROM AUTHOR]

Abstract Background Current quantitative approaches to assess chronic liver disease (CLD) severity have limitations. Further, portal vein thrombosis (PVT) pre-liver transplant (LT) is a major contributor to morbidity in CLD; the means of detecting and/or predicting PVT are limited. We sought to explore whether plasma coagulation factor activity levels can serve as a substitute for prothrombin time/international normalized ratio (PT/INR) in the Model for End-stage Liver Disease (MELD), and/or help assess the risk of PVT. Methods Plasma activity levels of Factor V (FV), Factor VIII (FVIII), Protein C (PC), and Protein S (PS) and the concentrations of D-dimer, sP-selectin, and asTF were assessed in two cohorts of CLD patients (ambulatory, n = 42; LT, n = 43). Results FV and PC activity levels strongly correlated with MELD scores, which enabled the development of a novel scoring system based on multiple linear regressions of the correlations of FV and PC activity with MELD-Na that substitutes PT/INR. Six-month and 1-year follow-up revealed that our novel approach was non-inferior to MELD-Na at predicting mortality. A significant inverse correlation between FVIII activity levels and PVT was found in the LT cohort (p = 0.010); FV and PS activity levels were in-trend (p = 0.069, p = 0.064). We developed a logistic regression-based compensation score to identify patients at risk of PVT. Conclusions We demonstrate that FV and PC activity levels may be used to replace PT/INR in MELD scoring. We also show the potential of using the combination of FV, FVIII, and PS activity levels to assess the risk of PVT in CLD.

Introduction: Fresh Frozen Plasma (FFP) is a blood component separated from whole blood and frozen below -30°C within 8 hours of donation for optimum preservation of coagulation factors. However, logistic and geographical reasons may hamper separation of plasma within 8 hours and the separation may have to be delayed to between 8 and 24 hours and then frozen below -30°C which is called as Frozen Plasma (FP). Plasma separated between 8 and 24 hours is a licensed blood component in the United States of America (USA) for therapeutic use similar to FFP. It is not licensed in India leading to frequent shortage of plasma. Aim: To compare the activity of factors V, VIII and X and the level of fibrinogen between FFP and FP, so as to assess the therapeutic use of FP for formulating recommendation as licensed blood component. Materials and Methods: A prospective observational study was conducted in the Department of Transfusion Medicine at Seth GS Medical College and KEM Hospital, Mumbai, Maharashtra, India. The duration of the study was 10 months, from January 2018 to October 2018. Fifty units each of FFP and FP matched for the camp location, age, gender and blood group were selected. There were 44 males and six females in each of FFP and FP groups. They were compared for the activity of labile coagulation factors (factors V and VIII) and stable factor X. The level of fibrinogen was also measured in both components. It was done within 30 days of preparation of plasma. The mean values of each of the four parameters for FFP and FP were calculated and compared for statistical significance (p) by using unpaired Student's t-test. Microsoft Excel 2016 was used for statistical analysis. The p-value <0.05 was considered statistically significant. Results: The mean age of FFP and FP individuals (blood donors) was 31.2 and 31.3 years respectively, while the median age in years was 31 and 30.5, respectively. The activity/level of all the tested coagulation factors was lower in FP as compared to FFP. The difference was statistically significant for factor VIII (p-value <0.05). It was not significant for factor V, X and fibrinogen. The level/activity of coagulation factors in FP, though lower than that in FFP, fell within normal reference range in 90-95% of units. Conclusion: FP may be used as a therapeutic alternative to FFP excluding patients of haemophilia A in whom factor VIII concentrate and cryoprecipitate are considered better therapeutic modalities. Results of similar multicentre studies will help in formulating recommendations regarding licensing. [ABSTRACT FROM AUTHOR]

Background and Objective: Gene mutations leading to thrombophilic disorders and changes in blood coagulation factors during pregnancy play an important role in the occurrence of recurrent abortions. This study was conducted to determine the relationship between polymorphism of thrombophilic genes and coagulation factors in women with recurrent miscarriage. Methods: In this descriptive-analytical study, 29 women with a history of recurrent abortions (at least three times) were investigated. Demographic data were collected. Total genomic DNA was isolated from peripheral blood. The presence or absence of mutation in factor II (G20210A), factor V Leiden (G1691A), factor V HR2 (H1299R), MTHFR C677T, and MTHFR A1298C polymorphisms were assessed by PCR. Results: Coagulation factors and thrombophilic mutations had no significant association with recurrent miscarriage. Conclusion: The results obtained in this study showed that none of the studied polymorphisms could justify frequent abortions in women. Therefore, it is suggested to evaluate genetic factors in normal individuals or immediate and extended family of the study population with no miscarriage history.

Proteostasis, i.e., the homeostasis of proteins, responsible for ensuring protein turnover, is regulated by proteases, which also participate in the etiopathogenesis of multiple conditions. The magic of proteases is such that, in blood coagulation, one same molecule, such as coagulation factor V, for example, can perform both a procoagulant and an anticoagulant function as a result of the activity of proteases. However, this magic has an insidious side to it, as it may also prevent the completion of the clinical value chain of factor V deficiency. This value chain encompasses the discovery of knowledge, the transfer of this knowledge, and its translation to clinical practice. In the case of rare and ultra-rare diseases like factor V deficiency, this value chain has not been completed as the knowledge acquisition phase has dragged out over time, holding up the transfer of knowledge to clinical practice. The reason for this is related to the small number of patients afflicted with these conditions. As a result, new indications must be found to make the therapies cost-effective. In the case of factor V, significant research efforts have been directed at developing a recombinant factor V capable of resisting the action of the proteases capable of inactivating this factor. This is where bioethics and health equity considerations come into the equation. [ABSTRACT FROM AUTHOR]

Background: Current quantitative approaches to assess chronic liver disease (CLD) severity have limitations. Further, portal vein thrombosis (PVT) pre-liver transplant (LT) is a major contributor to morbidity in CLD; the means of detecting and/or predicting PVT are limited. We sought to explore whether plasma coagulation factor activity levels can serve as a substitute for prothrombin time/international normalized ratio (PT/INR) in the Model for End-stage Liver Disease (MELD), and/or help assess the risk of PVT. Methods: Plasma activity levels of Factor V (FV), Factor VIII (FVIII), Protein C (PC), and Protein S (PS) and the concentrations of D-dimer, sP-selectin, and asTF were assessed in two cohorts of CLD patients (ambulatory, n = 42; LT, n = 43). Results: FV and PC activity levels strongly correlated with MELD scores, which enabled the development of a novel scoring system based on multiple linear regressions of the correlations of FV and PC activity with MELD-Na that substitutes PT/INR. Six-month and 1-year follow-up revealed that our novel approach was non-inferior to MELD-Na at predicting mortality. A significant inverse correlation between FVIII activity levels and PVT was found in the LT cohort (p = 0.010); FV and PS activity levels were in-trend (p = 0.069, p = 0.064). We developed a logistic regression-based compensation score to identify patients at risk of PVT. Conclusions: We demonstrate that FV and PC activity levels may be used to replace PT/INR in MELD scoring. We also show the potential of using the combination of FV, FVIII, and PS activity levels to assess the risk of PVT in CLD. [ABSTRACT FROM AUTHOR]

Late-onset Fuchs endothelial corneal dystrophy (FECD) is a disease affecting the corneal endothelium (CE), associated with a cytosine-thymine-guanine repeat expansion at the CTG18.1 locus in the transcription factor 4 (TCF4) gene. It is unknown whether CTG18.1 expansions affect global methylation including TCF4 gene in CE or whether global CE methylation changes at advanced age. Using genome-wide DNA methylation array, we investigated methylation in CE from FECD patients with CTG18.1 expansions and studied the methylation in healthy CE at different ages. The most revealing DNA methylation findings were analyzed by gene expression and protein analysis. 3488 CpGs had significantly altered methylation pattern in FECD though no substantial changes were found in TCF4. The most hypermethylated site was in a predicted promoter of aquaporin 1 (AQP1) gene, and the most hypomethylated site was in a predicted promoter of coagulation factor V (F5 for gene, FV for protein). In FECD, AQP1 mRNA expression was variable, while F5 gene expression showed a ~ 23-fold increase. FV protein was present in both healthy and affected CE. Further gene expression analysis of coagulation factors interacting with FV revealed a ~ 34-fold increase of thrombomodulin (THBD). THBD protein was detected only in CE from FECD patients. Additionally, we observed an age-dependent hypomethylation in elderly healthy CE.Thus, tissue-specific genome-wide and gene-specific methylation changes associated with altered gene expression were discovered in FECD. TCF4 pathological methylation in FECD because of CTG18.1 expansion was ruled out. [ABSTRACT FROM AUTHOR]

A 74‐year‐old Pacific Island man with end‐stage renal failure planning to start haemodialysis presented with persistent bleeding after tunnelled dialysis catheter insertion. The laboratory findings revealed a prolonged activated partial thromboplastin time of 118 s, prothrombin ratio of 4.2, factor V activity of <2% and a factor V inhibitor of 40 Bethesda Units. No clear underlying aetiology was identified. The bleeding settled with conservative measures and the factor V inhibitor was successfully treated with oral cyclophosphamide for 6 weeks. [ABSTRACT FROM AUTHOR]

Introduction: Fresh Frozen Plasma (FFP) is a blood component separated from whole blood and frozen below -30°C within 8 hours of donation for optimum preservation of coagulation factors. However, logistic and geographical reasons may hamper separation of plasma within 8 hours and the separation may have to be delayed to between 8 and 24 hours and then frozen below -30°C which is called as Frozen Plasma (FP). Plasma separated between 8 and 24 hours is a licensed blood component in the United States of America (USA) for therapeutic use similar to FFP. It is not licensed in India leading to frequent shortage of plasma. Aim: To compare the activity of factors V, VIII and X and the level of fibrinogen between FFP and FP, so as to assess the therapeutic use of FP for formulating recommendation as licensed blood component. Materials and Methods: A prospective observational study was conducted in the Department of Transfusion Medicine at Seth GS Medical College and KEM Hospital, Mumbai, Maharashtra, India. The duration of the study was 10 months, from January 2018 to October 2018. 50 units each of FFP and FP matched for the camp location, age, gender and blood group were selected. There were 44 males and six females in each of FFP and FP groups. They were compared for the activity of labile coagulation factors (factors V and VIII) and stable factor X. The level of fibrinogen was also measured in both components. It was done within 30 days of preparation of plasma. The mean values of each of the four parameters for FFP and FP were calculated and compared for statistical significance (p) by using unpaired Student’s t-test. Microsoft Excel 2016 was used for statistical analysis. The p-value

Introduction: Thromboembolic events occur due to an imbalance in the hemostasis and some factors associated with this condition can be inherited. In order to evaluate the frequency of genotypes considered to be common hereditary risk factors for thrombophilia associated with venous thrombosis (g.1691G > A and g.20210G > A) and hyperhomocysteinemia (g.677C > T and g.1298A > C), samples from voluntary healthy blood donors at the Hospital de Clínicas de Porto Alegre were tested. Methods: We examined 325 blood samples from blood donors collected from October 2017 to July 2018. Blood was collected on filter paper and the DNA was extracted for single nucleotide polymorphisms (SNPs) analysis using the qualitative real time polymerase chain reaction. Results: The calculated frequencies of each genetic variant in heterozygosity were 4% for the FV gene (g.1691G > A), 4% for the F2 gene (g.20210G > A) and 42% and 39% for methylenetetrahydrofolate reductase (MTHFR), g.677C > T and g.1298A > C, respectively. Only the genetic variants of MTHFR were found in homozygosity, with frequencies of 14% and 6% (g.677C > T and g.1298A > C), respectively. Discussion: Altogether, these results describe the frequencies of genetic variants associated with venous thrombosis and hyperhomocysteinemia in the analyzed group and are important to enhance our current knowledge about the genetic profiles of Brazilian blood donors.

Background and Objective: Gene mutations leading to thrombophilic disorders and changes in blood coagulation factors during pregnancy play an important role in the occurrence of recurrent abortions. This study was conducted to determine the relationship between polymorphism of thrombophilic genes and coagulation factors in women with recurrent miscarriage. Methods: In this descriptive-analytical study, 29 women with a history of recurrent abortions (at least three times) were investigated. Demographic data were collected. Total genomic DNA was isolated from peripheral blood. The presence or absence of mutation in factor II (G20210A), factor V Leiden (G1691A), factor V HR2 (H1299R), MTHFR C677T, and MTHFR A1298C polymorphisms were assessed by PCR. Results: Coagulation factors and thrombophilic mutations had no significant association with recurrent miscarriage. Conclusion: The results obtained in this study showed that none of the studied polymorphisms could justify frequent abortions in women. Therefore, it is suggested to evaluate genetic factors in normal individuals or immediate and extended family of the study population with no miscarriage history. [ABSTRACT FROM AUTHOR]

The distribution of factor V Leiden G1691A, factor II G20210A, MTHFR C677T, and factor V H1299R polymorphisms known to predispose to thrombophilia in 215 cases and 40 controls admitted with indication of recurrent pregnancy loss (RPL) was investigated. Genotyping was performed by melting curve analysis using simultaneous PCR (RT-PCR). There was no difference between genotype and allele frequencies in the case and control groups in terms of the polymorphisms examined (p>0.05). In the genotype distribution of the Factor V gene G1691A polymorphism, 12.6% GA in the case group and 90.0% GG (wild) genotypes in the control group were found to be higher than the other genotypes. In the genotype distribution of the factor II gene G20210A polymorphism, the GG (wild) genotype was found to be higher in 94.5% and 97.5%, respectively, in the case group and control group than the other genotypes. In the genotype distribution of MTHFR gene C677T polymorphism, 43.7% CC (wild) in the case group and 40.0% CT genotype in the control group were found to be higher than the other genotypes. In the genotype distribution of the factor V gene A4070G polymorphism, 87.4% AA (wild) in the case group and 80% AA (wild) genotype in the control group were found to be higher than the other genotypes. However, the frequency of risk allele A for factor V Leiden G1691A (6.8% and 5%), the frequency of risk allele A for factor II gene G20210A (3% and 1.2%), MTHFR gene C677T were determined in the case and control groups. The frequency of the T allele (35.9% and 42.5%), which is the risk allele for A4070G, and the frequency of the G allele (6.5% and 16.6%), which is the risk allele for the factor V gene A4070G, were determined. When our study results were evaluated, no relationship was found between RPL and factor V Leiden G1691A, factor II G20210A, MTHFR C677T, and factor V A4070G polymorphisms. [ABSTRACT FROM AUTHOR]

OBJECTIVE: The association between recurrent implantation failure and thrombophilia is still controversial depending on the published reports with conflicting results. In this study, we aimed to assess the clinical relevance of screening women with recurrent implantation failure for some thrombophilic variants including factor V H1299R (FV HR2) haplotype. STUDY DESIGN: A total of 279 women were recruited in this case-control study. 229 women with a history of recurrent implantation failure and 50 fertile control with no history of pregnancy losses were screened for eight specific gene mutations, regarding factor V G1691A gene (FV Leiden), FV HR2, factor II prothrombin G20210A, factor XIII V34L, PAI-1 4G/5G, MTHFR C677T, MTHFR A1298C and A3 haplotype of the endothelial cell protein C receptor gene. RESULTS: Recurrent implantation failure group displayed a significantly higher prevalence of FV HR2 heterozygosity than fertile controls while the frequency of FV Leiden mutation was comparable between groups (p=0.011; p=0.619). Additionally, the difference in the prevalence of other specific or total gene mutations among women with recurrent implantation failure was also insignificant. DISCUSSION: The primer outcome of this study was the co-existence of the higher prevalence of FV HR2 haplotype and the insignificant percentage of FV Leiden mutation in women with recurrent implantation failure. Thus, we emphasize that the HR2 haplotype may be associated with recurrent implantation failure particularly in non-carriers for FV Leiden mutation. In the necessity of screening for thrombophilia in recurrent implantation failure, HR2 haplotype should be involved in the searched gene panel particularly in the absence of FV Leiden mutation. Further large-scale prospective studies are needed to investigate whether screening or treatment for HR2 haplotype has any detrimental impact on implantation success in cases of recurrent implantation failure. [ABSTRACT FROM AUTHOR]

The vascular endothelium has several important functions, including hemostasis. The homeostasis of hemostasis is based on a fine balance between procoagulant and anticoagulant proteins and between fibrinolytic and antifibrinolytic ones. Coagulopathies are characterized by a mutation-induced alteration of the function of certain coagulation factors or by a disturbed balance between the mechanisms responsible for regulating coagulation. Homeostatic therapies consist in replacement and nonreplacement treatments or in the administration of antifibrinolytic agents. Rebalancing products reestablish hemostasis by inhibiting natural anticoagulant pathways. These agents include monoclonal antibodies, such as concizumab and marstacimab, which target the tissue factor pathway inhibitor; interfering RNA therapies, such as fitusiran, which targets antithrombin III; and protease inhibitors, such as serpinPC, which targets active protein C. In cases of thrombophilia (deficiency of protein C, protein S, or factor V Leiden), treatment may consist in direct oral anticoagulants, replacement therapy (plasma or recombinant ADAMTS13) in cases of a congenital deficiency of ADAMTS13, or immunomodulators (prednisone) if the thrombophilia is autoimmune. Monoclonal-antibody-based anti-vWF immunotherapy (caplacizumab) is used in the context of severe thrombophilia, regardless of the cause of the disorder. In cases of disseminated intravascular coagulation, the treatment of choice consists in administration of antifibrinolytics, all-trans-retinoic acid, and recombinant soluble human thrombomodulin. [ABSTRACT FROM AUTHOR]

Background: All current X-ray structures of factor (F)Xa are devoid of the γ-carboxyglutamate (Gla) domain and fail to reveal the overall conformation of the free protein. The recent cryogenic electron microscopy (cryo-EM) structure of FXa in the prothrombinase complex is the only structure of full-length FXa and shows that the Gla domain is positioned at an angle relative to the epidermal growth factor 1 domain., Objectives: Establish if the curved conformation of FXa revealed by cryo-EM is also present in solution., Methods: The conformation of FXa in solution was studied by single-molecule Förster resonance energy transfer., Results: The conformation of full-length FXa in solution is resolved for the first time. The conformation is curved and extremely sensitive to Ca 2+ . It does not differ significantly from its zymogen form or from that present in the prothrombinase complex free or bound to the physiologic substrates prothrombin and meizothrombin., Conclusion: Measurements by single-molecule Förster resonance energy transfer reveal that FXa has a curved conformation in solution, free or bound to physiologic ligands, and validate the recent cryo-EM structures of prothrombinase. The drastic conformational changes observed in the absence of Ca 2+ suggest that the structural architecture of FXa changes upon administration of vitamin K antagonists that perturb the interaction of the Gla domain with divalent cations., Competing Interests: Declaration of competing interests There are no competing interests to disclose., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Factor (F)V is pivotal in both procoagulant and anticoagulant mechanisms. The present report describes a novel F5 mutation in a FV-deficient patient (FV activity, 6 IU/dL; FV antigen, 32 IU/dL) complicated by recurrent deep vein thrombosis. The patient demonstrated activated protein C resistance (APCR) with compound heterozygous mutations consisting of FV-Y1961C (FV Kanazawa ) and FV-1982&#95;1983del., Objectives: To clarify thrombotic mechanisms associated with this FV abnormality., Methods and Results: Levels of FV-1982&#95;1983del were below the detection sensitivity in our expression experiments using human embryonic kidney 293T cells, and analyses were targeted, therefore, on the FV-Y1961C mutation. Activated partial thromboplastin time-based clotting assays demonstrated that FV-Y1961C exhibited APCR and that the reduced activated protein C (APC) susceptibility in FVa-Y1961C resulted in a marked depression of APC-catalyzed inactivation with delayed cleavage at Arg506 and little cleavage at Arg306 with or without protein S. The APC cofactor activity of FV-Y1961C in APC-catalyzed FVIIIa inactivation promoted by Arg336 cleavage in FVIII was impaired. The binding affinity of FVa-Y1961C to phospholipid membranes was reduced in reactions involving APC/protein S-catalyzed inactivation and in prothrombinase activity. Furthermore, the addition of FVa-Y1961C to plasma failed to inhibit tissue factor-induced procoagulant function. These characteristics were similar to those of FV-W1920R (FV Nara ) and FV-A2086D (FV Besançon )., Conclusion: We identified a compound heterozygous FV-Y1961C mutation in the C1 domain representing a novel FV mutation (FV Kanazawa ) resulting in not only APCR due to impaired FVa susceptibility and FV cofactor activity for APC function but also impaired inhibition of tissue factor-induced procoagulant function. These defects in anticoagulant function associated with FV in FV-Y1961C contributed to a prothrombotic state., Competing Interests: Declaration of competing interests There are no competing interests to disclose., (Copyright © 2024 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)

The STA R Max3 (Stago, France) and Cobas t511 (Roche Diagnostics, Germany) are two automated hemostasis analysers that can be used to perform a wide range of tests. The STA R Max3 uses a mechanical clot detection system to measure coagulation times, while the Cobas t511 uses optical detection. The aim of this study was to compare the analytical performance of these two analysers using fresh plasma samples with or without a risk of interference due to hemolysis or lipemia. For plasma samples without interference, acceptable agreement was observed for prothrombin time (PT) (n = 55), activated partial thromboplastin time (APTT) (n = 56), fibrinogen (n = 56) and factor II (n = 43) with R² of 0.907, 0.963, 0.979 and 0.968 respectively. For factor V (n = 43) and D-dimers (n = 45), agreement was less acceptable, with respective R² of 0.756 and 0.887, but with no clinical impact. For hemolysed samples, acceptable results were observed for PT, fibrinogen and D-dimers, but three results were clinically discordant for APTT, and STA R Max3 offered greater robustness for processing highly lipemic samples.

Background: In healthy individuals, plasma levels of clotting proteins naturally vary within a range of 50% to 150% of their mean values. We do not know how these variations modify thrombin generation., Objectives: To assess the impact of protein level variations on simulated thrombin generation in normal and factor (F)VIII-, FIX-, or FXI-deficient blood., Methods: We used a mathematical model of flow-mediated coagulation to simulate thrombin generation with all possible combinations of clotting protein variations within the normal range and for various tissue factor levels. We selected, analyzed, and ranked combinations that enhanced thrombin generation compared with baseline., Results: Protein variations most strongly affected thrombin generation at intermediate tissue factor levels. Low tissue factor levels prevented coagulation initiation, while high tissue factor levels always triggered thrombin generation. At intermediate levels, we identified protein variations that substantially modified thrombin generation. Low-normal FV shortened lag times and increased thrombin generation, whereas high-normal FV lengthened lag times and reduced thrombin generation. With severe FVIII and FIX deficiencies, low-normal tissue factor pathway inhibitor α and antithrombin amplified the effect of low-normal FV. For moderate FVIII and FIX deficiencies, high-normal tissue factor pathway inhibitor α and antithrombin enhanced the impact of high-normal FV in reducing thrombin production. In normal and FXI-deficient blood, high-normal FVIII and FIX significantly boosted thrombin generation., Conclusion: Our mathematical model predicted how variations in clotting protein levels, within the normal range, could contribute to the variability of bleeding phenotypes observed with clotting factor deficiencies. Our study generated experimentally testable hypotheses that could aid in developing new therapies toward normal hemostasis., (© 2024 The Author(s).)

Background: Protein S (PS) is an anticoagulant that functions as a cofactor for activated protein C and the tissue factor pathway inhibitor. PS deficiency is a risk factor for venous thromboembolism. PS activity is commonly measured using clot-based assays involving fibrin and thrombin production, but improvements are needed., Objectives: To develop a new assay for measuring plasma PS activity by quantifying the amount of activated coagulation factor (F)V cleaved by activated protein C., Methods: We designed a recombinant, modified FV (FVm) that mimicked FVa. We analyzed 160 purposively selected plasma samples from the Biobank of the National Cerebral and Cardiovascular Center., Results: The assay using mixed normal and PS-deficient plasma detected FVm cleavage in a PS concentration-dependent manner. The correlation between PS activity, measured using the FVm cleavage assay, and free PS antigen levels was relatively weak. We then sequenced all exons of PROS1 from 47 subjects with <60% activity in either the FVm cleavage assay or the clot-based assay. Nonsynonymous variants were identified in 12 of 24 subjects with <60% activity in both assays and in 2 of 7 subjects with <60% activity in the FVm cleavage assay alone. No variants were identified in 16 subjects with <60% activity in the clot-based assay alone. Unlike the clot-based assay, the FVm cleavage assay was not affected by the presence of rivaroxaban in the plasma., Conclusion: An assay using the FVm substrate may be less susceptible to interference and provide a more accurate evaluation of plasma PS activity than clot-based assays., Competing Interests: Declaration of competing interests There are no competing interests to disclose., (Copyright © 2024 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)

Abstract A State of the Art lecture titled “Cryo‐EM structures of coagulation factors” was presented at the ISTH Congress in 2022. Cryogenic electron microscopy (cryo‐EM) is a revolutionary technique capable of solving the structure of high molecular weight proteins and their complexes, unlike nuclear magnetic resonance (NMR), and under conditions not biased by crystal contacts, unlike X‐ray crystallography. These features are particularly relevant to the analysis of coagulation factors that are too big for NMR and often recalcitrant to X‐ray investigation. Using cryo‐EM, we have solved the structures of coagulation factors V and Va, prothrombinase on nanodiscs, and the prothrombin‐prothrombinase complex. These structures have advanced basic knowledge in the field of thrombosis and hemostasis, especially on the function of factor V and the molecular mechanism for prothrombin activation, and set the stage for exciting new lines of investigation. Finally, we summarize relevant new data on this topic presented during the 2022 ISTH Congress.

Thrombin generation (TG) is known as a physiological approach to assess the hemostatic function. Although it correlates well with thrombosis and bleeding, in the current setup it is not sensitive to the effects of fluctuations in single coagulation factors. We optimized the calibrated automated thrombinography (CAT) method to quantify FII, FV and FX activity within the coagulation system. The CAT assay was fine-tuned for the assessment of FII, FV and FX by diluting the samples in FII-, FV-, or FX-deficient plasma, respectively, and measuring TG. Plasma FII levels correlated linearly with the ETP up to a plasma concentration of 100% FII. FV and FX levels correlated linearly with the peak height up to a plasma level of 2.5% FV and 10% FX, respectively. Sensitized CAT protocols were designed by adding a fixed volume of a pre-diluted patient sample to FII, FV, and FX deficient plasma in TG experiments. This approach makes the TG measurement dependent on the activity of the respective coagulation factor. The ETP or peak height were quantified as readouts for the coagulation factor activity. The intra- and inter-assay variation coefficients varied from 5.0 to 8.6%, and from 3.5 to 5.9%, respectively. Reference values were determined in 120 healthy subjects and the assays were clinically validated in 60 patients undergoing coronary artery bypass grafting (CABG). The sensitized CAT assays revealed that the contribution of FII, FV, and FX to the TG process was reduced after CABG surgery, leading to reduced prothrombin conversion and subsequently, lower TG.

Objectives: Absolute or relative protein (P)C pathway abnormalities (PC deficiency, PS deficiency, antiphospholipid syndrome (APS), factor (F)V‐abnormality, and high FVIII level) cause thrombophilia. Although screening assays for these thrombophilias are available, one utilizing clot waveform analysis (CWA) remains unknown. We aimed to establish a CWA‐based screening assay to distinguish PC pathway abnormality‐related thrombophilia. Methods: Samples were reacted with tissue factor (TF)/phospholipids and recombinant thrombomodulin (rTM; optimal 20 nM), followed by CWA measurement. The peak ratio (with/without rTM) of the first derivative curve of clot waveform was calculated. Results: The peak ratio in healthy plasmas (n = 35) was 0.36 ± 0.13; hence, the cutoff value was set to 0.49. The peak ratios in plasmas with PC deficiency, PS deficiency, high‐FVIII (spiked 300 IU/dl), and APS were higher than the cutoff values (0.79/0.97/0.50/0.93, respectively). PC‐deficient plasma or PS‐deficient plasma mixed with normal plasma (25%/50%/75%/100% PC or PS level) showed dose‐dependent decreases in the peak ratios (PC deficient: 0.85/0.64/0.44/0.28; PS deficient: 0.69/0.53/0.40/0.25), suggesting that the peak ratio at ≤50% of PC or PS level exceeded the cutoff value. The peak ratio in FV deficiency with FV ≤25% was higher than the cutoff value. FV‐deficient plasma spiked with 40 IU/dl rFV‐R506Q (FVLeiden) or rFV‐W1920R (FVNara) showed >90% peak ratios. Conclusions: rTM‐mediated TF‐triggered CWA might be useful for screening PC pathway abnormality‐related thrombophilia. [ABSTRACT FROM AUTHOR]

Proposed mechanisms of acute traumatic coagulopathy (ATC) include decreased clotting potential due to factor consumption and proteolytic inactivation of factor V (FV) and activated factor V (FVa) by activated protein C (aPC). The role of FV/FVa depletion or inactivation in burn-induced coagulopathy is not well characterized. This study evaluates FV dynamics following burn and nonburn trauma. Burn and trauma patients were prospectively enrolled. Western blotting was performed on admission plasma to quantitate levels of FV antigen and to assess for aPC or other proteolytically derived FV/FVa degradation products. Statistical analysis was performed with Spearman's, Chi-square, Mann–Whitney U test, and logistic regression. Burn (n = 60) and trauma (n = 136) cohorts showed similar degrees of FV consumption with median FV levels of 76% versus 73% (P = 0.65) of normal, respectively. Percent total body surface area (TBSA) was not correlated with FV, nor were significant differences in median FV levels observed between low and high TBSA groups. The injury severity score (ISS) in trauma patients was inversely correlated with FV (ρ = −0.26; P = 0.01) and ISS ≥ 25 was associated with a lower FV antigen level (64% versus. 93%; P = 0.009). The proportion of samples showing proteolysis-derived FV was greater in trauma than burn patients (42% versus. 16%; P = 0.0006). Increasing traumatic injury severity is associated with decreased FV antigen levels, and a greater proportion of trauma patient samples exhibit proteolytically degraded FV fragments. These associations are not present in burns, suggesting that mechanisms underlying FV depletion in burn and nonburn trauma are not identical. [ABSTRACT FROM AUTHOR]

Recurrent pregnancy loss (RPL) is defined as the loss of two or more pregnancies, affecting approximately 1 to 3% of women worldwide. Scientific data highlight a possible correlation between thrombophilic genetic variants and RPL. H1299R variant in the factor V gene would lead to an increased thrombotic risk associated with frequent miscarriages. However, the data are often conflicting, making this an interesting question for further investigations by evaluating genotype-phenotype correlations to improve the clinical management and genetic counseling of couples. A systematic review and meta-analysis will follow the preferred reporting elements for systematic review and meta-analysis protocols (PRISMA-P). The Pubmed (MEDLINE) and Embase (OVID) databases will be explored to identify suitable articles based on inclusion and exclusion criteria. Inclusion criteria are: (a) H1299R genotyping with clear data reported, referred to as Heterozygous (Het) and/or Homozygous (Hom); (b) articles written in English; (c) analyses of only RPL female patients having at least two or more previous pregnancy losses and compared with a control group. This analysis will present selected scientific evidence, addressing the questions concerning the association between the H1299R variant and RPL, hoping to clarify this still unresolved issue. PROSPERO registration number: CRD42022330077. [ABSTRACT FROM AUTHOR]

Sarah O John-Olabode,1 Kehinde S Okunade,2 Ayorinde James,3 Gbenga Olorunfemi,4 Obiefuna I Ajie,5 Akinniyi A Osuntoki,6 Alani S Akanmu1 1Department of Hematology and Blood Transfusion, College of Medicine, University of Lagos, Lagos, Nigeria; 2Department of Obstetrics and Gynecology, College of Medicine, University of Lagos, Lagos, Nigeria; 3Department of Biochemistry and Nutrition, Nigerian Institute of Medical Research, Lagos, Nigeria; 4Division of Epidemiology and Biostatistics, School of Public Health, University of Witwatersrand, Johannesburg, South Africa; 5Department of Clinical Pathology, College of Medicine, University of Lagos, Lagos, Nigeria; 6Department of Biochemistry, College of Medicine, University of Lagos, Lagos, NigeriaCorrespondence: Sarah O John-OlabodeDepartment of Hematology and Blood Transfusion, College of Medicine, University of Lagos, Lagos, NigeriaTel +234 8096608152Email sarahajibola@yahoo.comIntroduction: Inherited thrombophilia and venous thromboembolism (VTE) have been closely linked to adverse pregnancy outcomes such as preeclampsia/eclampsia contributing to increased maternal and perinatal morbidity and mortality. There is, however, little genetic data from Africa including Nigeria that explores the prevalence of common VTE genetic risk markers such as factor V Leiden mutation (FVL G1691A) and prothrombin gene mutation (F2 G20210A) among pregnant women in Nigeria.Purpose: To determine the prevalence and distribution of FVL G1691A and F2 G20210A in pregnant women in Lagos, Nigeria.Patients and Methods: This hospital-based cross-sectional pilot study was conducted among pregnant women between 1 July 2019 and 31 August 2020. The genotype of interest was determined through amplification by polymerase chain reaction using G1691A of FV and prothrombin A20210G specific primers. Descriptive data were presented using Stata version 15 (Stata Corp) statistical software.Results: Of the 400 recruited participants, 397 and 389 samples were successfully processed for FVL G1691A and F2 G20210A mutations, respectively. Three participants had FVL heterozygous mutation; thus, the prevalence of heterozygous mutation of FVL among the study participants was 0.76%, 95% CI: 0.002– 0.023%, n=3/397. There was no F2 G20210A mutation detected among the study participants.Conclusion: This study indicates that screening for factor V Leiden mutation and prothrombin gene mutation in pregnancy might not be of any clinical significance among Nigerian women. However, carrying out a genome-wide associated study is recommended to determine the true impact of these two common inherited thrombophilias in this population.Keywords: factor V Leiden G1691A, prothrombin G20210A gene mutation, factor 2, factor V, preeclampsia, venous thromboembolism

OBJECTIVE: To determine whether electronic health record (EHR) tools improve documentation of pre- and postanalytic care processes for genetic tests ordered by nongeneticists. METHODS: We conducted a nonrandomized, controlled, pre-/postintervention study of EHR point-of-care tools (informational messages and template report) for three genetic tests. Chart review assessed documentation of genetic testing processes of care, with points assigned for each documented item. Multiple linear and logistic regressions assessed factors associated with documentation. RESULTS: Preimplementation, there were no significant site differences (P > 0.05). Postimplementation, mean documentation scores increased (5.9 (2.1) vs. 5.0 (2.2); P = 0.0001) and records with clinically meaningful documentation increased (score >5: 59 vs. 47%; P = 0.02) at the intervention versus the control site. Pre- and postimplementation, a score >5 was positively associated with abnormal test results (OR = 4.0; 95% CI: 1.8-9.2) and trainee provider (OR = 2.3; 95% CI: 1.2-4.6). Postimplementation, a score >5 was also positively associated with intervention site (OR = 2.3; 95% CI: 1.1-5.1) and specialty clinic (OR = 2.0; 95% CI: 1.1-3.6). There were also significantly fewer tests ordered after implementation (264/100,000 vs. 204/100,000; P = 0.03), with no significant change at the control site (280/100,000 vs. 257/100,000; P = 0.50). CONCLUSIONS: EHR point-of-care tools improved documentation of genetic testing processes and decreased utilization of genetic tests commonly ordered by nongeneticists.Genet Med 19 1, 112-120.

Factor V, encoded by the F5 gene, is a procoagulant blood clotting factor that increases the production of thrombin, the central enzyme that converts fibrinogen to fibrin, which leads to the formation of a blood clot. The F5 gene is localized to 1q24.2 chromosome and consists of 25 exons. There are various mutations in the F5 gene that lead to resistance of activated protein C (APC) (elimination of the APС cleavage site in factor V and factor Va), which can lead to arterial and venous thrombosis. The aim of the present study was to analyze variants of the F5 gene in patients diagnosed with coronary atherosclerosis without acute coronary syndrome with stable functional class II–IV angina pectoris, confirmed by coronary angiography data, using the method of whole exome sequencing. Material and methods. The study was conducted in the framework of the Program of joint research work IIPM — branch of the ICG SB RAS and the FSBI «Research Institute of Circulation Pathology named after E.N. Meshalkin» Ministry of Health of Russian Federation. The study included 30 men aged 40–70 years with coronary angiography-­verified coronary atherosclerosis, without ACS, with stable angina pectoris of the II–IV FC. Patients were admitted for coronary bypass surgery, and endarteriaectomy from the coronary artery (s) was performed during the operation according to intraoperative indications. Whole exome sequencing (SureSelectXT Human All Exon v.6+UTR) was carried out on an Illumina NextSeq 500 instrument (USA). Results. In 30 patients, 29 single-­nucleotide variants were found in the F5 gene. In patients with coronary atherosclerosis, rs9332701 of the F5 gene is 3.33 times more common, and rs6027 is 1.67 times more common than in the population. And rs184663825 was found in 3.33% of cases, while its occurrence in the population is 0.05%. For variants rs6034 and rs144979314, a possible damaging effect on the protein product is shown. Conclusion. The single-­nucleotide variants rs9332701, rs6027, rs184663825, rs6034, rs144979314 of the F5 gene are of interest for inclusion in the genetic panels for the analysis of risk factors for the development of acute coronary syndrome.

Abstract Acquired factor V inhibitor represents a rare condition, described only in case reports. The use of activated bypassing agents in bleeding control could be of aid and improve the survival in symptomatic patients with acquired factor V inhibitor.

Proteostasis, i.e., the homeostasis of proteins, responsible for ensuring protein turnover, is regulated by proteases, which also participate in the etiopathogenesis of multiple conditions. The magic of proteases is such that, in blood coagulation, one same molecule, such as coagulation factor V, for example, can perform both a procoagulant and an anticoagulant function as a result of the activity of proteases. However, this magic has an insidious side to it, as it may also prevent the completion of the clinical value chain of factor V deficiency. This value chain encompasses the discovery of knowledge, the transfer of this knowledge, and its translation to clinical practice. In the case of rare and ultra-rare diseases like factor V deficiency, this value chain has not been completed as the knowledge acquisition phase has dragged out over time, holding up the transfer of knowledge to clinical practice. The reason for this is related to the small number of patients afflicted with these conditions. As a result, new indications must be found to make the therapies cost-effective. In the case of factor V, significant research efforts have been directed at developing a recombinant factor V capable of resisting the action of the proteases capable of inactivating this factor. This is where bioethics and health equity considerations come into the equation.

Coagulation factor V (or FV for the purpose of medical safety) is an essential cofactor of coagulation factor X in the common pathway of coagulation; severe FV deficiency leads to a bleeding tendency. Although both congenital and acquired FV deficiencies are widely recognized, FV deficiency also presents as an autoimmune disorder. A nationwide survey on autoimmune coagulation factor deficiencies (AiCFDs) conducted in Japan by our Japanese Collaborative Research Group identified 24 new patients with autoimmune FV deficiency (AiFVD) in the past 5 years. Furthermore, our extensive literature search confirmed that 177 AiFVD cases have been reported in previous articles published from Japan. Patients with AiFVD in Japan were predominantly men, with age similar to those with other AiCFDs. AiFVD was confirmed as a relatively mild type of bleeding diathesis, associated with lower mortality rate than that for AiFVD and other AiCFDs reported in previous studies. Patients with AiFVD had variable FV inhibitor titers and both neutralizing anti-FV autoantibodies and nonneutralizing counterparts. Although spontaneous resolution occurs in some patients, timely initiation of hemostatic and immunosuppressive therapies helps arrest the bleeding and eliminate anti-FV antibodies, resulting in a high cumulative recovery rate. Immunological anti-FV antibody detection is recommended to avoid missing AiFVD cases for the presence of nonneutralizing anti-FV autoantibodies. Further investigation is necessary to clarify the long-term prognosis and optimal management of AiFVD. [ABSTRACT FROM AUTHOR]