Vitamin K-dependent Coagulation Factors Research Articles

Combined factor VIII and IX deficiency: An exceptional case

Familial multiple coagulation factor deficiencies are a group of rare inherited disorders that are characterized by the simultaneous decrease in the levels of two or more coagulation factors. Common synchronous deficiencies are factors VIII and IX and combined deficiency of Vitamin K-dependent coagulation factors (factors II, VII, IX, and X). Here, we report a case of synchronous dual deficiency of factor VIII and IX, which is an extremely rare occurrence and no case report has been mentioned so far to the best of our knowledge. Recognizing such a dual deficiency is very important for proper management of patients.

No Detectable Coagulation Activation After Vitamin K (MK-7) Supplementation in Patients on Dialysis With Functional Vitamin K Deficiency: A One-Year Randomized, Placebo-Controlled Study

Patients on dialysis treatment have poor functional vitamin K status, and this may increase the risk of vascular calcification. Vitamin K supplementation may therefore be relevant in patients on dialysis, but the procoagulant effects have not been studied. We evaluated effects of menaquinone-7 (MK-7) supplementation on biomarkers of coagulation in patients on dialysis. Double-blinded, placebo-controlled study in 123 patients on dialysis randomized to 52weeks of vitamin K (MK-7, 360μg/daily, n=61) or placebo (n=62). Measurements at baseline and after 52weeks of intervention included thrombin generation (endogenous thrombin potential, peak thrombin concentration, time to peak, and lag time); clot activities of vitamin K-dependent coagulation factors (F) II, VII, IX, and X; prothrombin fragment 1+2 (F1+2); and proteins induced by vitamin K absence II (PIVKA-II). Between-group differences (vitamin K vs. placebo) at 52weeks were determined with an analysis of covariance. Within-group changes in vitamin K and placebo groups were analyzed with a paired t-test. Vascular adverse events and serious adverse events were registered based on hospital records, laboratory data, and participant interviews and compared between groups using Fisher's exact test or Pearson's Chi-Squared test. A between-group difference at 52weeks was observed for PIVKA-II (P<.001). PIVKA-II decreased significantly from baseline to 52weeks in the vitamin K group, but not in the placebo group. We observed no between-group differences or within-group changes for biomarkers of coagulation, except for FVII clot activity which was reduced in the placebo group (P=.04), and no between-group differences in adverse events and serious adverse events. One year of vitamin K supplementation in patients on dialysis has no detectable effects on biomarkers of coagulation activation, clot activities of vitamin K-dependent coagulation factors, and vascular events or death, indicating no procoagulant effects of this treatment.

Protein Z ( PROZ) Loss of Function Is Associated with Increased Risk of Ischemic Stroke in 416,767 UK Biobank Participants

Background:The vitamin K-dependent coagulation factor protein Z ( PROZ) is canonically considered to have anticoagulant effects through its negative regulation of factor Xa. However, the true role of protein Z in coagulation is controversial, with conflicting data on the impact of plasma protein Z levels across a range of thrombotic phenotypes. In particular, higher circulating protein Z concentrations have consistently been associated with an elevated risk of acute ischemic stroke, but there is little explanation for this apparent paradox. Because studies of plasma protein levels are especially prone to confounding by environmental factors, a large-scale, adjusted analysis examining the thrombosis risk associated with germline loss-of-function variants in PROZ would represent a novel approach with far greater capacity to address causality. Aims:Perform a large-scale genomic association study to examine the risk associated with functional variants in PROZ across four thrombotic phenotypes. Methods:Using paired clinical and whole exome sequencing data for 416,767 participants in the UK Biobank (UKBB), we identified individuals with rare (MAF&amp;lt;0.1%), in silico predicted function-altering (∆F) variants in PROZ. Using Firth's logistic regression and controlling for age, sex, ancestry, and other known risk factors for thrombosis, we assessed the association between ∆F variants in PROZ and venous thromboembolism (VTE), non-cardioembolic ischemic stroke (NCEIS), peripheral arterial disease (PAD), myocardial infarction (MI), and coagulopathic bleeding. Additionally, we evaluated differences in the levels of 1,472 plasma proteins (Olink Explore 1536 proteomics panel) between carriers and non-carriers of qualifying PROZ variants in a subset of 48,892 UKBB participants. Results: Rare ∆F variants in PROZ were identified in 463 UKBB participants (99.14% heterozygous). The presence of ∆F variants was associated with significantly increased risk of NCEIS (OR=2.34, 95% CI: 1.16-4.15, P=0.02). By contrast, no significant association was seen between PROZ ∆F variants and VTE (OR=0.92, 95% CI: 0.54-1.46, P=0.75), MI (OR=1.03, 95% CI: 0.61-1.61, P=0.93), PAD (OR=0.63, 95% CI: 0.21-1.40, P=0.29), or coagulopathic bleeding (OR=0.96, 95% CI: 0.57-1.52, P=0.88) ( Figure 1). Restricting the analysis to high-confidence loss of function variants (i.e., nonsense, frameshift, and essential splice site mutations) increased the effect size estimate for NCEIS (OR=2.53, 95% CI: 0.96-5.26, P=0.058) without meaningfully changing our findings in the other thrombotic or bleeding phenotypes. Plasma proteomic analysis revealed that carriers of a PROZ ∆F variant had significantly elevated levels of two proteins that have previously been found to mediate brain inflammation after acute ischemic stroke, peroxiredoxin 1 (PRDX1, fold-change = 1.83, P=1.3 x 10 -5) and peroxiredoxin 6 (PRDX6, fold-change = 1.78, P=9.6 x 10- 10) ( Figure 2). Conclusions:In this dataset of over 400,000 participants, we found that rare, germline loss-of-function variants in PROZ are associated with significantly increased risk of NCEIS. Plasma proteomics revealed that PROZ variant carriers also have significantly higher circulating levels of two proteins that are known to play a role in acute ischemic stroke. By contrast, we found no association between PROZ mutations and an increased risk for VTE, MI, PAD, or bleeding. Taken together, these findings suggest that protein Z haploinsufficiency likely confers a uniquely higher risk of ischemic stroke without substantially influencing other forms of thrombosis.

The Hemostatic System in Newborns and the Risk of Neonatal Thrombosis.

Newborns are the most vulnerable patients for thrombosis development among all children, with critically ill and premature infants being in the highest risk group. The upward trend in the rate of neonatal thrombosis could be attributed to progress in the treatment of severe neonatal conditions and the increased survival in premature babies. There are physiological differences in the hemostatic system between neonates and adults. Neonates differ in concentrations and rate of synthesis of most coagulation factors, turnover rates, the ability to regulate thrombin and plasmin, and in greater variability compared to adults. Natural inhibitors of coagulation (protein C, protein S, antithrombin, heparin cofactor II) and vitamin K-dependent coagulation factors (factors II, VII, IX, X) are low, but factor VIII and von Willebrand factor are elevated. Newborns have decreased fibrinolytic activity. In the healthy neonate, the balance is maintained but appears more easily converted into thrombosis. Neonatal hemostasis has less buffer capacity, and almost 95% of thrombosis is provoked. Different triggering risk factors are responsible for thrombosis in neonates, but the most important risk factors for thrombosis are central catheters, fluid fluctuations, liver dysfunction, and septic and inflammatory conditions. Low-molecular-weight heparins are the agents of choice for anticoagulation.

Rare inherited coagulation and fibrinolytic defects that challenge diagnostic laboratories.

Coagulation factors, anticoagulants, and fibrinolytic proteins are important for hemostasis, and mutations affecting these proteins causes some rare inherited bleeding disorders that are particularly challenging to diagnose. This review provides current information on rare inherited bleeding disorders that are difficult to diagnose. A review of the literature was conducted for up to date information on rare and difficult to diagnose bleeding disorders. Some rare bleeding disorders cause an inherited deficiency of multiple coagulation factors (F), such as combined FV and FVIII deficiency and familial vitamin K-dependent clotting factor deficiency. Additionally, congenital disorders of glycosylation can affect a variety of procoagulant and anticoagulant proteins and also platelets. Some bleeding disorders reflect mutations with unique impairments in the procoagulant/anticoagulant balance, including those caused by F5 mutations that secondarily increase the plasma levels of tissue factor pathway inhibitor as well as THBD mutations that increase functional thrombomodulin in plasma or cause a consumptive coagulopathy due to thrombomodulin deficiency. Some bleeding disorders accelerate fibrinolysis due to loss-of-function mutations in SERPINE1 and SERPINF2 or in the case of Quebec platelet disorder, a duplication mutation that rewires PLAU and selectively increases expression in megakaryocytes, resulting in a unique platelet-dependent gain-of-function defect in fibrinolysis. Current information on rare and difficult to diagnose bleeding disorders indicates they have unique clinical and laboratory features, and pathogenic characteristics to consider for diagnostic evaluation. Laboratories and clinicians should consider rare inherited disorders, and difficult to diagnose conditions, in their strategy for diagnosing bleeding disorders.

Acquired Factor VII Deficiency Associated With Chronic Myeloid Leukemia Blast Crisis.

Factor VII (FVII) is an important, vitamin K-dependent clotting factor. Acquired FVII deficiency is a rare entity that is associated with serious bleeding complications. We report a case of acquired FVII deficiency in a patient with recurrent chronic myeloid leukemia in blast crisis who developed bilateral retinal hemorrhages. The coagulopathy was corrected with the initiation of chemotherapy and subsequent reduction in peripheral blast count.

A case report of Acquired Prothrombin Complex Deficiency in Infant

Introduction: Acquired Prothrombin Complex Deficiency (APCD) is a disorder characterized by spontaneous bleeding caused by decreased activity of vitamin K-dependent coagulation factors (factors II, VII, IX, and X), despite normal fibrinogen and platelet counts. It can occur in newborns as young as 6 months old. This disease is linked to drug use during pregnancy, a lack of vitamin K-containing foods, and other secondary diseases. The manifestations of this disease vary, ranging from skin organs to gastrointestinal organs to the brain, with high morbidity and mortality rates. The aim of this case report is to describe an infant with diagnose of APCD and their management. Case Presentation: A 55-day-old patient had been in a coma and had been unconscious for three days before being admitted to the hospital. The patient vomited, developed a fever, and had a seizure. The laboratory examination showed that haemoglobin levels was 5.6g/dl, prothrombin time (PT) was 21 seconds, activated partial thromboplastin time (APTT) was extended by 54 seconds, and platelet showed abnormal 524 x 10^3µL in the result. The PT and APTT results indicated APCD, but there is an abnormality in platelets, which should be normal in APCD patient. Conclusion: This is a case reported of Acquired Prothrombin Complex Deficiency (APCD) based on laboratory and CT-Scan examination.

High Incidence of Intra-Cardiac Supra-Centimetric Thrombosis and Embolic Complications in Sickle Cell Disease Patients with Dual Lumen Ports

In Sickle Cell Disease (SCD) patients with cerebral vasculopathy and/or other severe complications, a transfusion program is indicated. A sustainable venous access is vital for patients engaged in these programs (5-10% of patients with SCD). Dual lumen ports (Vortextechnology) have emerged has a way to bypass the frequent local complications associated with arteriovenous fistulae. Between July 2015 and December 2020, a dual lumen port was implanted in 27 SCD patients in transfusion program at Necker Hospital. Over a 30 months period, we observed 20 venous thromboembolic events (VTE) in 12 patients (44%): 18 were voluminous intracardiac thrombosis (Fig 1), asymptomatic (n=15) or complicated of pulmonary embolism (n=3), one jugular thrombosis and one superior cellar syndrome, leading to long-term curative anticoagulation, except in cases of contraindication such as Moya-Moya. VTE were diagnosed on average 18 months after implantation (range 1 to 29 months). This observation has considerably limited the use of the device, even though no complications were observed in 56% of patients, suggesting that a subgroup of patients may be at lower risk of thrombosis and could still benefit from a Vortex. We conducted a clinical study aiming to identify potential biological markers related to Vortex thrombosis in SCD patients. Among the biological markers, alterations of thrombin generation and fibrino-formation, as well as potentially procoagulants items altered in SCD, such as microvesicules (MV), red blood cell (RBC) deformability and the intensity of splenic dysfunction were explored. Between January and June 2021, 11 healthy donors and 58 patients with SCD treated by erythrapheresis in the biotherapy department of Necker Hospital were included, 23 patients with a Vortex with (n=12) or without (n=11) intra-cardiac and/or other VTE and 36 patients without a Vortex (including 8 with a history of VTE). Thrombin generation Assay (TGA) in plasma and standard coagulation assays were performed on pre-transfusion sample, including Prothrombin Time (PT), activated partial thromboplastin time (APTT) and fibrinogen, dosage of FII, FV, FVII, FX and FXII, Protein C (PC), Protein C (PS), Antithrombin (AT) dosage. All samples were pretreated by activated carbon for removal of Direct Oral Anticoagulants from plasma. Quantification and characterization of plasma MV, determination of erythrocyte deformability and point of sickling (ektacytometry with oxygenscan) and assessment of splenic function (quantification of pocked RBC and Howell Jolly bodies) were also performed. We found a significant decrease in the mean levels of PC (69.7 vs 109.1%, p=0.0002) and PS (58.4 vs 99.4, p<0.0001) in SCD patients when compared to healthy donors while AT levels were only slightly decreased (91 vs 98,6%, p=0,04). We also observed a significant decrease in vitamin-K dependent coagulation factors: FII (77.5 vs 100.6%, p<0,0001), FVII (61.9 vs 84.5% p=0.005) and FX (71.8 vs 96.8%, p=0.0001) while FV (80.1 vs 93.8% p=0.02) was only slightly decreased and fibrinogen levels were comparable (2.62 vs 2.57g/L p=0.86). No significant differences were noted between SCD patient groups (with or without thrombosis). Thrombin generation Assay (TGA) in plasma showed an increase in endogenous thrombin potential (ETP; 714 vs 451nM.min, p=0.04) and Peak (161 vs 95 nM.min, p=0.03) in SCD patients compared to healthy donors but no differences were found between patient groups. Assessment of splenic function, point of sickling in oxygenscan and deformability of RBC measured by ektacytometry showed no differences between patient groups. Patients with SCD had a significant increase of circulating CD235+ MV (93268 vs 15 207/µL, p<0.0001), Annexin 5 MV (473553 vs 248404 /µL) and CD235+ erythrocyte ghosts (1757 vs 61/µL) compared to healthy donors. Again, no differences were seen between patient study groups. In conclusion, no biological predictive factors for intra cardiac Vortex thrombosis could be demonstrated by our study despite an extensive study of biological factors known to date to favor thrombosis. Given the high incidence of intracardiac thrombosis and the impossibility to date of predicting the risk of occurrence, we recommend that Vortex technology be reserved for patients who are already receiving effective anticoagulant therapy or for whom no other solution can be found, while maintaining a close surveillance of echocardiography.

Risk Factors, Diagnosis, Pathophysiology and Management of Deep Vein Thrombosis

Deep vein thrombosis occurs most often in the legs, but can form in the veins of the arms, and in the mesenteric and cerebral veins. Common risk factors of deep vein thrombosis includes age 40 years or older, being overweight, a personal or family history of blood clots, birth control pills, hormone replacement therapy, cancer, certain heart cases, stroke, respiratory failure, varicose veins, pregnancy, surgery including hip, knee, or stomach surgery, restricted mobility due to a long illness, injury, or surgery. Vascular injury that may result from major orthopedic surgery (e.g., knee and hip replacement), trauma (especially fractures of the pelvis, hip, or leg), or indwelling venous catheters. The D-dimer blood test measures degraded fibrinogen, which is raised in patients with a clot. The reference range varies and is set by the laboratory. This test is recommended in patients with a low or moderate clinical probability of deep vein thrombosis, as calculated by the Wells score. The two main types of anticoagulants are heparin and warfarin (coumadin) to keep a clot from growing or prevent new clots from forming. Warfarin interfere with hepatic synthesis of the vitamin K-dependent coagulation factors II, VII, IX and X. Warfarin, a vitamin K antagonist, is an effective and cheap oral anticoagulant.

Partial research progress of GGCX pathogenic variation associated phenotypes

γ-glutamyl carboxylase (GGCX), also known as vitamin K-dependent glutamyl carboxylase, catalyzes the posttranslational modification of specific glutamate residues in vitamin K-dependent proteins (VKDPs), and participates multiple biological functions including blood coagulation, bone metabolism, vascular calcification, and cell proliferation. It has been reported originally that GGCX pathogenic variation causes blood coagulation deficiency, which is called as vitamin K-dependent coagulation factor deficiency 1 (VKCFD1). Recently, it has been found that GGCX gene variation results in multiple clinical phenotypes, including dermatological, ophthalmological, skeletal or cardiac abnormalities. Among them, dermatological phenotype is the most common, which is known as pseudoxanthoma elasticum-like syndrome. This paper has reviewed the GGCX pathogenic variation associated phenotypes, in order to increase the recognition of GGCX-related genetic diseases and to help its diagnosis and treatment.

Application of fresh frozen plasma transfusion in the management of excessive warfarin-associated anticoagulation

Warfarin is a commonly used oral anticoagulant. Patients with artificial valve replacement, atrial fibrillation, pulmonary embolism, deep vein thrombosis, and other diseases require long-term anticoagulant oral treatment with warfarin. As warfarin exhibits prompt action with long maintenance time, it has become a key drug for the treatment of patients at risk of developing thrombosis or thromboembolism. Warfarin is a bican coumarin anticoagulant, that exhibits competitive action against vitamin K as its mechanism of action, thereby inhibiting the synthesis of coagulation factors—predominantly the vitamin K-dependent coagulation factors II, VII, IX, and X—in hepatocytes. Long-term warfarin is known to significantly increase the risk of organ bleeding in some patients, while some patients may need to reverse the anticoagulation effect. For instance, patients scheduled for emergency or invasive surgery may require rapid anticoagulation reversal. During such medical circumstances, fresh frozen plasma (FFP) is clinically used for the reversal of excess warfarin-associated anticoagulation, as it contains all the coagulation factors that can alleviate the abnormal blood anticoagulation status in such patients. Accordingly, this article aims to perform an in-depth review of relevant literature on the reversal of warfarin with FFP, and insightful deliberation of the application and efficacy of this clinical intervention.

Reversing direct factor Xa or thrombin inhibitors: Factor V addition to prothrombin complex concentrate is beneficial in vitro

BackgroundProthrombin complex concentrate (PCC) is a human plasma‐derived mixture of partially purified vitamin K‐dependent coagulation factors (VKCF). Current therapeutic indication is treatment and perioperative prophylaxis of bleeding in acquired VKCF deficiency. Off‐label uses include treatment of direct factor Xa‐ or thrombin inhibitor‐associated bleeds, treatment of trauma‐induced coagulopathy, and hemorrhagic complications in patients with liver disease. ObjectiveConsidering PCC as a general prohemostatic drug, we argued that its clinical efficacy can benefit from supplementation with coagulation factors that are absent in the current PCC formulation. In this study, we focused on factor V. MethodsWe mimicked a coagulopathy in vitro by spiking whole blood or derived plasma with the direct oral anticoagulants (DOAC) rivaroxaban or dabigatran. We studied DOAC reversal by PCC and factor V concentrate (FVC) using a thrombin generation assay, thromboelastography, fibrin generation clot lysis test, and microfluidic thrombus formation under flow. ResultsIn DOAC‐treated plasma, PCC increased the amount of thrombin generated. The addition of FVC alone or in combination with PCC caused a partial correction of the thrombin generation lag time and clotting time. In DOAC‐treated whole blood, the combination of PCC and FVC synergistically improved clotting time under static conditions, whereas complete correction of fibrin formation was observed under flow. Clot strength and clot resistance toward tissue plasminogen activator‐induced lysis were both increased with PCC and further enhanced by additional FVC. ConclusionOur in vitro study demonstrates a beneficial effect of the combined use of PCC and FVC in DOAC reversal.

A case of late-onset vitamin K deficiency bleeding in an exclusively breastfed infant emphasizing the need for revision of vitamin K prophylaxis policy

A 4-month-old male child presented with a history of seizure and vomiting. There was no history of fever, fall, or any other trauma. On examination, there was anisocoria, and anterior fontanelle was bulging and nonpulsating. Computerized tomography revealed an acute on chronic bilateral subdural hematoma (left > right) with a midline shift. He had no relevant past or family history suggestive of any bleeding disorder. Blood investigations revealed anemia with prolonged prothrombin time (PT)/international normalized ratio/and activated partial thromboplastin time (APTT). Mixing study was done and it showed a correction with pooled normal plasma. The possibility of coagulation factor deficiency was confirmed. Further, mixing studies and factor assays revealed normal levels of fibrinogen and Factor VIII, with a reduced level of Factor IX. Since both PT and APTT were prolonged and Factor IX level was low, a deficiency of Vitamin K-dependent coagulation factors was suspected. The child was immediately given intravenous Vitamin K injection. The diagnosis of Vitamin K deficiency bleeding (VKDB) was confirmed by protein induced by Vitamin K absence testing. Unfortunately, the child succumbed to the complications of the intracranial bleed.

Profile of the single-use, multiple-pass protein A adsorber column in immunoadsorption.

Immunoadsorptions (IA) are used to remove autoantibodies from the plasma in autoimmune disorders. In this study, we evaluated the effects of a single-use, recombinant staphylococcal protein A-based immunoadsorber on blood composition of the patient. In a cohort of patients with myasthenia gravis or stiff-person syndrome, essential parameters of blood cell count, coagulation, clinical chemistry or plasma proteins and immunoglobulins (Ig) were measured before and after IA (n= 11). In average, IA reduced the levels of total IgG, IgG1, IgG2 and IgG4 by approximately 60%, the acetylcholine receptor autoantibody levels by more than 70%. IgG3, IgA or IgM were diminished to a lower extent. In contrast to fibrinogen or other coagulation factors, the column markedly removed vitamin K-dependent coagulation factors II, VII, IX and X by approximately 40%-70%. Accordingly, international normalized ratio and activated partial thromboplastin time were increased after IA by 59.1% and 32.7%, respectively. Coagulation tests almost returned to baseline values within 24 h. Blood cell count, electrolytes, total protein or albumin were not essentially affected. No clinical events occurred. The single-use, multiple-pass protein A adsorber column is highly efficient to remove IgG1, IgG2 and IgG4 or specific acetylcholine receptor autoantibodies from the plasma. Coagulation parameters should be monitored, since the column has the capacity to largely reduce vitamin K-dependent factors.

Interfering with Protein Z Dependent Inhibition of Factor Xa Improves Coagulation in Hemophilia

Blood coagulation involves a highly regulated balance between pro-coagulant and anticoagulant activities, allowing for the abatement of bleeding while limiting clot formation, which, if unchecked, could result in thrombosis. Hemostasis is triggered upon vascular injury when extravascular tissue factor (TF) binds plasma coagulation factor VIIa (FVIIa) forming a FVIIa/TF complex that generates limited amounts of activated factors IX (FIXa) and X (FXa). In normal hemostasis, FIXa with FVIIIa dramatically amplifies the production of FXa; FXa with FVa cleaves prothrombin to thrombin, and thrombin cleaves fibrinogen to form a fibrin clot. FXa generated by FVIIa/TF alone is insufficient for normal coagulation as FVIIa/TF activity is limited due to tissue factor pathway inhibitor (TFPI) and TFPI and other plasma protease inhibitors suppress the action of the initially generated FXa (and thrombin). Lacking FVIII or FIX, hemophiliacs fail to generate the additional FXa that is critical for normal hemostasis. Several methods designed to reduce the activity of endogenous inhibitors of coagulation are currently being investigated as a means to ameliorate the bleeding in hemophilia. These include antibody-mediated reduction in the activities of TFPI and protein S, and siRNA-mediated reduction in antithrombin (AT).Protein Z (PZ) is a vitamin K-dependent coagulation factor that serves as the cofactor for FXa inhibition by the serpin, PZ-dependent protease inhibitor (ZPI), at phospholipid surfaces. If inhibition of the action of PZ/ZPI significantly enhanced coagulation in hemophilia, it might be associated with a reduced risk of thrombosis than other approaches. In the latter regard it is of interest that TFPI, PS and AT deficiency are associated with clinical thrombotic disease and mice lacking these proteins die in utero or perinatally of thromboembolic disease. In contrast, PZ and ZPI KO mice appear healthy.Re-bleeding is a hallmark of clinical hemophilia. A tail vein re-bleeding (TVRB) assay, adapted from the saphenous vein re-bleeding assay of Whinna, was developed to assess the effects of PZ and ZPI deficiency on re-bleeding in the hemophilia A mouse model. In this assay, the lateral vein of the tail (diameter 2.25 mm) of an anesthetized mouse is lacerated to a depth of 1 mm with scalpel and the tail placed in warm saline. When bleeding stops, the wound is wiped with a balled Kimwipe to re-induce bleeding and the tail replaced in warm saline. This process is repeated and the number of wound disruptions occurring within 15 minutes is recorded. In normal mice, the period of bleeding following each wound disruption remains relatively constant, whereas in hemophilia mice the bleeding periods tend to progressively prolong with each subsequent disruption leading to a reduced number of total disruptions over 15 minutes.Disruptions in the mouse TVRB assay were: for C57Bl/6 25.8+1.7 (mean+SEM); for FVIII KO 4.2+1.3; for PZ/FVIII double KOs 15.2+3.2; for ZPI/FVIII doubles KOs 14.6+3.5; for WT/KO littermates 4.0+2.6; and for FVIII KO with 100% human FVIII replacement 25.4+3.7. Based on FVIII dose-response experiments, the results in PZ/FVIII and ZPI/FVIII double KOs were equivalent to ~15% FVIII replacement. This outcome of the TVRB assays was confirmed in tests of the survival of sibling WT/FVIII KO (0%, n=5) and ZPI/FVIII double KO (75%, n=12) mice following tail transection 10 mm from the tip.To assess the translation of the PZ/ZPI results in mice to humans, the effect of an anti-PZ monoclonal antibody (Mab) was compared to FVIII replacement on TF (1 pM) induced thrombin generation in human hemophilia A plasma. The Mab enhanced thrombin generation in hemophilia plasma to a similar degree as ~15% FVIII.These studies provide proof of concept, both in vivo in mice and in vitro in human plasma, that blockade of the PZ/ZPI system may be sufficient to alter the phenotype of severe hemophilia to that of mild hemophilia, perhaps with limited thrombotic risk. DisclosuresNo relevant conflicts of interest to declare.

Vitamin K2 (Menaquinone-7) supplementation does not affect vitamin K-dependent coagulation factors activity in healthy individuals.

Background:Vitamin K has long been regarded as a procoagulant drug by physicians, and concerns have been raised with regard to its effects on hemostasis. Although many studies have shown that vitamin K supplementation is safe for thrombotic events, the effect of vitamin K supplementation on the activities of vitamin K dependent procoagulation factors in healthy individuals is not available.Objectives:This study aimed to investigate whether vitamin K2 supplementation at recommended doses affects the activity of vitamin K dependent procoagulation factors in healthy individuals without any anticoagulation treatment.Design:Forty healthy volunteers between 25 and 40 years of age were recruited. Menaquinone-7 (MK-7) was administrated at 90 μg for 30 days. Prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), and blood coagulation factors II, VII, IX, and X activities and Protein induced by vitamin K absence or antagonist-II (PIVKA-II) were measured on days 0 and 30 after MK-7 administration.Results:PT, APTT, and TT showed no significant differences on day 30 when compared with baseline. The activities of coagulation factors II, VII, IX, and X on day 30 showed no significant differences with those at baseline. PIVKA-II levels were unchanged after 30 days of MK-7 supplementation.Conclusions:MK-7 supplementation at recommended dosage does not affect vitamin K-dependent coagulation factors’ coagulation activity, and does not enhance the carboxylation of prothrombin in healthy individuals. This indicated that MK-7 administration does not alter hemostatic balance in healthy populations without anticoagulation treatment.

An in vitro study of canine cryopoor plasma to correct vitamin K-dependent coagulopathy in dogs.

To compare the efficacy of fresh frozen plasma (FFP) with cryopoor plasma (CPP) to treat vitamin K-dependent factor deficiency in a canine in vitro setting. In vitro laboratory study. University veterinary medical teaching hospital. Seven units of FFP and 6 units of CPP from unique canine donors from the university veterinary blood bank. Canine FFP was adsorbed by oral barium sulfate suspension to mimic vitamin K-dependent coagulopathy. A sequential mixing study was completed by adding FPP or CPP to the adsorbed plasma. Measurements of prothrombin time (PT), activated partial thromboplastin time (aPTT), fibrinogen, and factor activities of factors II, VII, and IX (FII, FVII, and FIX) were compared between the 2 treatment groups. When comparing the sequential addition of CPP or FPP to adsorbed plasma, the following had no statistical significance: PT (P=0.94), aPTT (P=0.66), FII (P=0.05), and FIX (P=0.90). There was a dose-dependent decrease with PT and aPTT and a dose-dependent increase with FII and FIX. In contrast, after the addition of either CPP or FFP, there was a significant difference between the treatment groups for the concentration of fibrinogen (P=0.005) and activity of FVII (P=0.044), with FFP resulting in a greater concentration of fibrinogen and CPP resulting in a greater concentration of FVII. Measurements of factor X (FX) were initially included in the study but were later excluded because FX appeared to be continually adsorbed even after the addition of CPP or FFP. CPP partially corrected the coagulation times and concentration of vitamin K-dependent coagulation factors to the same degree as FFP. CPP, generally less expensive than FFP, may provide an alternative treatment option for vitamin K-dependent coagulopathies, although in vivo testing is needed.

CLINICAL APPLICATION OF PLASMA PROTEIN C DETERMINATION

Objective: Protein C, a vitamin K-dependent coagulation factor, is involved in blood coagulation. Activated protein C inactivates Va and VIIIa and stimulates fibrinolysis. In this process, protein S serve as an important factor for activated protein C. Furthermore, excess protein S drives cancer cell proliferation and cell survival through oncogenic receptor Axl(Anexelekto). We determined ranges of protein C both in healthy individuals and distinct hospitalized patients.&#x0D; Methods: A total of 100 patients with different diagnostic diseases and 50 healthy individuals were included in their plasma protein C determination. A rabbit antibody against human protein C was used for the quantitative estimation of plasma protein C antigen by using rocket immunoassay.&#x0D; Results: In healthy individuals protein C antigen (PC:Ag) ranges o.6439- 1.4752 µ/ml. The mean coefficient of variation (CV) of length of rocket was calculated to be 12.45%. PC:Ag within laboratory variation was 11.47%. Plasma protein C antigen was destroyed at 56℃ for 30 minutes, whereas no significant decrease of protein C was found at 4℃ refrigerator for one week.&#x0D; Conclusion: The results showed that plasma protein C antigen was considerably high in 22 diabetes mellitus. On the other hand, the PC:Ag was significantly decreased in 19 liver cirrhosis(p&lt; 0.001) and was positively correlation with serum albumin levels(p&lt; 0.05). In 20 acute leukemias, on the average,there was slightly lower values in PC:Ag, and accompanied with significant decrease of PC:Ag in 5 M5 subtype and in 9 hyper- leukocytes acute leukemias. However, the 3 acute promyelocytic leukemia (APL) with overt laboratory picture of DIC(disseminated intravascular coagulation) had protein C concentration no lower than the remaining 2 patients with infectious DIC, which suggested the coagulopathy in APL might be due to mechanisms different from other forms of DIC.&#x0D; &#x0D; Peer Review History: &#x0D; Received 17 November 2020; Revised 12 Decembe; Accepted 1 January, Available online 15 January 2021&#x0D; UJPR follows the most transparent and toughest ‘Advanced OPEN peer review’ system. The identity of the authors and, reviewers will be known to each other. This transparent process will help to eradicate any possible malicious/purposeful interference by any person (publishing staff, reviewer, editor, author, etc) during peer review. As a result of this unique system, all reviewers will get their due recognition and respect, once their names are published in the papers. We expect that, by publishing peer review reports with published papers, will be helpful to many authors for drafting their article according to the specifications. Auhors will remove any error of their article and they will improve their article(s) according to the previous reports displayed with published article(s). The main purpose of it is ‘to improve the quality of a candidate manuscript’. Our reviewers check the ‘strength and weakness of a manuscript honestly’. There will increase in the perfection, and transparency.&#x0D; Received file: Comments of reviewer(s): &#x0D; Average Peer review marks at initial stage: 5.5/10&#x0D; Average Peer review marks at publication stage: 7.5/10&#x0D; Reviewer(s) detail:&#x0D; Dr. Idoko Alexander, Caritas University, Enugu, Nigeria, idokoalexander1@gmail.com&#x0D; Dr. Rawaa Souhil Al-Kayali, Aleppo University, Syria, rawah67@hotmail.com&#x0D; Similar Articles:&#x0D; THE ASSOCIATION BETWEEN LEVELS OF HEPCIDIN, IRON STATUS AND MICRO-INFLAMMATION MARKERS AMONG HAEMODIALYSIS &#x0D; COUMARIN ANALOGUES AS A POTENTIAL INHIBITOR OF LEISHMANIASIS: A MULTI-TARGETING PROTEIN INHIBITION APPROACH BY MOLECULAR DOCKING &#x0D; VITAMIN A, RETINOIC ACID AND TAMIBAROTENE, A FRONT TOWARD ITS ADVANCES: A REVIEW

Treatment of rare bleeding disorders

Rare bleeding disorders include inherited deficiencies of fibrinogen, factors (F)II, FV, FVII, FX, FXI, FXII, and FV + FVIII, as well as a multiple deficiency of vitamin K-dependent coagulation factors. Some of these deficiencies are more studied, due to the large number of patients, some are extremely rare, and so at this stage it is quite difficult for them to develop a universal approach to therapy and prophylactic treatment. The purpose of this review was to evaluate treatment options for these deficiencies.

Effect of Citrofortunella microcarpa (Calamansi) Peelings on Whole Blood Coagulation Using Blood Samples from Albino Mice

Citrofortunella microcarpa, locally known as calamansi in the Philippines, is an intergeneric hybrid between Citrus reticulata and Fortunella japonica. This fruit is widely cultivated in the Philippines for its fruit juice as an abundant source of vitamin C and as a condiment in many local foods in the country. Sadly, only the pulp is needed for squeezing while the peels are thrown after extracting the juice. Previous studies revealed that the peels of Citrus, as member of the Rutaceae family, can synthesize both coumarins and furanocoumarins wherein their derivates are used as oral anticoagulants which can inhibit vitamin K from functioning as a cofactor in the hepatic synthesis of the vitamin K-dependent coagulation factors II, VII, IX, and X. In this study, the extract of calamansi peelings were proven to have an anticoagulant property on blood samples from albino mice. This study will pave the way for scientists to allot time in studying calamansi peelings for it may be another source of medicine to help patients who are prone to have stroke, myocardial infarction, and other blood clotting diseases.