APTT critical value establishment in four different reagent/instrument systems based on single factor deficiencies.

INTRODUCTION: There are significant differences in the activated partial thromboplastin time (APTT) critical values reported in different studies, most of which does not make recommendations for any specific clear detection systems. The International Council for Standardization in Hematology (ICSH) recommends that APTT critical values be established based on the reagent type, coagulation factor sensitivity and heparin response. The objective of this study was to establish APTT critical values by using different reagents and based on single coagulation factor deficiencies. METHODS: The APTT values were determined in commercial endogenous coagulation factor-deficient plasma at concentrations of 1 IU/dL, 2 IU/dL, 5 IU/dL, 10 IU/dL, 20 IU/dL, and 30 IU/dL by using four assay systems. The retrospective collection of data from patients who lacked factor VIII (FVIII), FIX, or FXI alone was performed. Receiver operating characteristic (ROC) curves were constructed to assess the diagnostic accuracy of APTT for identifying patients with an endogenous coagulation factor activity < 5 IU/dL. RESULTS: The APTT values in the plasma samples with the same concentrations of endogenous coagulation factors were significantly different among the four assay systems (P < 0.001). The suggested critical values of APTT were 40.0 s for Sysmex CS5100 (Actin FSL), 58.0 s for Sysmex CS5100 (Actin), 51.8 s for STA-R Evolution (STA-PTTA), and 64.8 s for ACL TOP 700 (HemosIL SynthasIL). On the basis of the ROC curve, the optimal threshold values for APTT (STA-PTTA) were 55.8 s in patients with a simple deficiency of FVIII (sensitivity = 100%, specificity = 85.7%, area under the ROC curve (AUC) = 0.982), 54.3 s in patients with a simple deficiency of FIX (sensitivity = 100%, specificity = 92.9%, AUC = 0.986), and 71.7 s in patients with a simple deficiency of FXI (sensitivity = 100%, specificity = 94.1%, AUC = 0.992), which were closer (difference of 0.6-2.5 s) to the cutoff points for commercial plasma at equal factor levels. CONCLUSIONS: APTT critical values need to be established for different reagents based on the presence of a single coagulation factor deficiency.

Comparative analysis of thrombin generation platforms for patients with coagulation factor deficiencies: A comprehensive assessment.

INTRODUCTION: Thrombin generation assays (TGAs) assess the overall functionality of the hemostatic system and thereby provide a reflection of the hemostatic capacity of patients with disorders in this system. Currently, four (semi-)automated TGA platforms are available: the Calibrated Automated Thrombogram, Nijmegen Hemostasis Assay, ST Genesia and Ceveron s100. In this study, we compared their performance for detecting patients with congenital single coagulation factor deficiencies. MATERIALS AND METHODS: Pooled patient samples, healthy control samples and normal pooled plasma were tested on all four platforms, using the available reagents that vary in tissue factor and phospholipid concentrations. The TGA parameters selected for analysis were peak height and thrombin potential. Results were normalized by using the calculated mean of healthy controls and a correction for between-run variation. Outcomes were presented as relative values, with the mean of healthy controls standardized to 100 %. RESULTS: Across all platforms and reagents used, thrombin potentials and peak heights of samples with coagulation factor deficiencies were lower than those of healthy controls. Reagents designed for bleeding tendencies yielded the lowest values on all platforms (relative median peak height 19-32 %, relative median thrombin potential 19-45 %). Samples representing more severe coagulation factor deficiencies generally exhibited lower relative peak heights and thrombin potentials. CONCLUSIONS: Thrombin generation assays prove effective in differentiating single coagulation factor deficient samples from healthy controls, with modest discrepancies observed between the platforms. Reagents designed for assessing bleeding tendencies, featuring the lowest tissue factor and phospholipid concentrations, emerged as the most suitable option for detecting coagulation factor deficiencies.

A Clot Twist: Extreme Variation in Coagulotoxicity Mechanisms in Mexican Neotropical Rattlesnake Venoms.

Rattlesnakes are a diverse clade of pit vipers (snake family Viperidae, subfamily Crotalinae) that consists of numerous medically significant species. We used validated in vitro assays measuring venom-induced clotting time and strength of any clots formed in human plasma and fibrinogen to assess the coagulotoxic activity of the four medically relevant Mexican rattlesnake species Crotalus culminatus, C. mictlantecuhtli, C. molossus, and C. tzabcan. We report the first evidence of true procoagulant activity by Neotropical rattlesnake venom in Crotalus culminatus. This species presented a strong ontogenetic coagulotoxicity dichotomy: neonates were strongly procoagulant via Factor X activation, whereas adults were pseudo-procoagulant in that they converted fibrinogen into weak, unstable fibrin clots that rapidly broke down, thereby likely contributing to net anticoagulation through fibrinogen depletion. The other species did not activate clotting factors or display an ontogenetic dichotomy, but depleted fibrinogen levels by cleaving fibrinogen either in a destructive (non-clotting) manner or via a pseudo-procoagulant mechanism. We also assessed the neutralization of these venoms by available antivenom and enzyme-inhibitors to provide knowledge for the design of evidence-based treatment strategies for envenomated patients. One of the most frequently used Mexican antivenoms (Bioclon Antivipmyn®) failed to neutralize the potent procoagulant toxic action of neonate C. culminatus venom, highlighting limitations in snakebite treatment for this species. However, the metalloprotease inhibitor Prinomastat substantially thwarted the procoagulant venom activity, while 2,3-dimercapto-1-propanesulfonic acid (DMPS) was much less effective. These results confirm that venom-induced Factor X activation (a procoagulant action) is driven by metalloproteases, while also suggesting Prinomastat as a more promising potential adjunct treatment than DMPS for this species (with the caveat that in vivo studies are necessary to confirm this potential clinical use). Conversely, the serine protease inhibitor 4-(2-aminoethyl)benzenesulfonyl fluoride hydrochloride (AEBSF) inhibited the direct fibrinogen cleaving actions of C. mictlantecuhtli venom, thereby revealing that the pseudo-procoagulant action is driven by kallikrein-type serine proteases. Thus, this differential ontogenetic variation in coagulotoxicity patterns poses intriguing questions. Our results underscore the need for further research into Mexican rattlesnake venom activity, and also highlights potential limitations of current antivenom treatments.

Pharmacological management of rare coagulation factor deficiencies besides hemophilia.

INTRODUCTION: Rare coagulation factor deficiencies are less-known disorders with variable effects on the patient's life. Management of such patients is a challenge due to the paucity of evidence-based data, more so when patients with these rare disorders encounter a more rare, related condition, like inhibitor development or thrombosis. AREA COVERED: A comprehensive literature search related to RCFDs and management was performed in PubMed in order to discuss therapeutic options and challenges, prophylaxis, management of minor and major surgeries, obstetric and gynecological complications, inhibitor development, and thrombosis. EXPERT OPINION: Although significant changes have occurred in the management of RCFDs in recent years, more evidence-based studies besides expert opinion are needed for optimal management.

Thromboelastography Reaction-Time Thresholds for Optimal Prediction of Coagulation Factor Deficiency in Trauma.

BACKGROUND: Coagulopathy is common in multitrauma patients and repletion of procoagulant factor deficiency with fresh frozen plasma (FFP) improves hemostasis. Optimal kaolin-thromboelastography thresholds for FFP transfusion in trauma patients have not been well established. STUDY DESIGN: Adult trauma patients with an Injury Severity Score ≥15 were included in this retrospective observational cohort study. The primary end point was area under the receiver operating characteristic curve (AUROC) for reaction time (R-time) to detect procoagulant factor deficiency, as reflected by an elevated international normalized ratio (INR) or aPTT. Test characteristics for the optimal R-time threshold calculated in our study were compared against thresholds recommended by the American College of Surgeons for FFP transfusion. RESULTS: Six hundred and ninety-four pairs of thromboelastography and conventional coagulation tests were performed in 550 patients, with 144 patients having additional pairs of tests after the first hour. The R-time was able to detect procoagulant factor deficiency (INR ≥1.5 AUROC 0.80; 95% CI, 0.75 to 0.85; aPTT ≥40 seconds AUROC 0.85; 95% 0.80 to 0.89) and severe procoagulant factor deficiency (INR ≥2.0 AUROC 0.82; 95% CI, 0.73 to 0.99; aPTT ≥60 seconds AUROC 0.89; 95% CI, 0.81 to 0.98) with good accuracy. Optimal thresholds to maximize sensitivity and specificity were 3.9 minutes for detection of INR ≥1.5, 4.1 minutes for detection of aPTT ≥40 seconds, 4.3 minutes for detection of INR ≥2.0, and 4.3 for detection of aPTT ≥60 seconds. Currently recommended R-time thresholds for FFP transfusion had 100% specificity for detecting procoagulant factor deficiency, but low sensitivity (3% to 7%). CONCLUSIONS: R-time can detect procoagulant factor deficiency in multitrauma patients with good accuracy, but currently recommended R-time thresholds are highly specific and not sensitive. Use of low-sensitivity thresholds might result in undertreatment of many patients with procoagulant factor deficiency.

Elevated thrombin generation in patients with congenital disorder of glycosylation and combined coagulation factor deficiencies.

BACKGROUND: Congenital disorders of glycosylation are rare inherited diseases affecting many different proteins. The lack of glycosylation notably affects the hemostatic system and leads to deficiencies of both procoagulant and anticoagulant factors. OBJECTIVE: To assess the hemostatic balance in patients with multiple coagulation disorders by using a thrombin generation assay. METHOD: We performed conventional coagulation assays and a thrombin generation assay on samples from patients with congenital disorder of glycosylation. The thrombin generation assay was performed before and after activation of the protein C system by the addition of soluble thrombomodulin. RESULTS: A total of 35 patients were included: 71% and 57% had low antithrombin and factor XI levels, respectively. Protein C and protein S levels were abnormally low in 29% and 26% of the patients, respectively, whereas only 11% displayed low factor IX levels. Under baseline conditions, the thrombin generation assay revealed a significantly higher endogenous thrombin potential and thrombin peak in patients, relative to controls. After spiking with thrombomodulin, we observed impaired involvement of the protein C system. Hence, 54% of patients displayed a hypercoagulant phenotype in vitro. All the patients with a history of stroke-like episodes or thrombosis displayed this hypercoagulant phenotype. CONCLUSION: A thrombin generation assay revealed a hypercoagulant in vitro phenotype under baseline condition; this was accentuated by impaired involvement of the protein C system. This procoagulant phenotype may thus reflect the risk of severe vascular complications. Further research will have to determine whether the thrombin generation assay is predictive of vascular events.

Plasma contact factors as therapeutic targets.

Direct oral anticoagulants (DOACs) are small molecule inhibitors of the coagulation proteases thrombin and factor Xa that demonstrate comparable efficacy to warfarin for several common indications, while causing less serious bleeding. However, because their targets are required for the normal host-response to bleeding (hemostasis), DOACs are associated with therapy-induced bleeding that limits their use in certain patient populations and clinical situations. The plasma contact factors (factor XII, factor XI, and prekallikrein) initiate blood coagulation in the activated partial thromboplastin time assay. While serving limited roles in hemostasis, pre-clinical and epidemiologic data indicate that these proteins contribute to pathologic coagulation. It is anticipated that drugs targeting the contact factors will reduce risk of thrombosis with minimal impact on hemostasis. Here, we discuss the biochemistry of contact activation, the contributions of contact factors in thrombosis, and novel antithrombotic agents targeting contact factors that are undergoing pre-clinical and early clinical testing.

Venom Protein C Activators as Diagnostic Agents for Defects of Protein C System.

BACKGROUND: Protein C is a vitamin K dependent plasma zymogen. It inhibits clotting by inhibiting clotting by inactivating factor V and factor VIII. Protein C activation pathway involves three steps: (i) Activation of protein C; (ii) Inhibition of coagulation through inactivating factor V and VIII by activated protein C and (iii) Inhibition of activated protein C by plasma protease inhibitors specific for this enzyme. Proteinases converting the zymogen protein C (PC) of vertebrates into activated PC have been detected in several snake venoms. Most PC activators have been purified from venom of snake species belonging to the genera of the Agkistrodon complex. Unlike the physiological thrombin-catalyzed PC activation reaction which requires thrombomodulin as a cofactor, most snake venom activators directly convert the zymogen PC into the catalytically active form which can easily be determined by means of coagulation or chromogenic substrate techniques. CONCLUSION: The fast-acting PC activator Protac® from Agkistrodon contortrix contortrix (Southern copperhead snake) venom has found a broad application in diagnostic practice for the determination of disorders in the PC pathway. Recently, screening assays for the PC pathway have been introduced, based on the observation that the PC pathway is probably the most important physiological barrier against thrombosis.

Congenital Factor Deficiencies in Children: A Report of a Single-Center Experience.

Congenital factor deficiencies (CFDs) refer to inherited deficiency of coagulation factors in the blood. A total of 481 patients with CFDs, who were diagnosed and followed at our Pediatric Hematology and Oncology Clinic between 1990 and 2015, were retrospectively evaluated. Of the 481 cases, 134 (27.8%) were hemophilia A, 38 (7.9%) were hemophilia B, 57 (11.8%) were von Willebrand disease (vWD), and 252 (52.3%) were rare bleeding disorders (RBDs). The median age of the patients at the time of diagnosis and at the time of the study was 4.1 years (range: 2 months to 20.4 years) and 13.4 years (range: 7 months to 31.3 years), respectively. The median duration of the follow-up time was 6.8 years (range: 2.5 months to 24.8 years). One hundred nineteen (47.2%) of 252 patients with RBDs were asymptomatic, 49 (41.1%) of whom diagnosed by family histories, 65 (54.6%) through preoperative laboratory studies, and 5 (4.2%) after prolonged bleeding during surgeries. Consanguinity rate for the RBDs was 47.2%. Prophylactic treatment was initiated in 80 patients, 58 of whom were hemophilia A, 7 were hemophilia B, 13 were RBDs, and 2 were vWD. Significant advances have been achieved during the past 2 decades in the treatment of patients with CFDs, particularly in patients with hemophilias. The rarity and clinical heterogeneity of RBDs lead to significant diagnostic challenges and improper management. In this regard, multinational collaborative efforts are needed with the hope that can improve the management of patients with RBDs.

Performance of Activated Partial Thromboplastin Time (APTT): Determining Reagent Sensitivity to Factor Deficiencies, Heparin, and Lupus Anticoagulants.

The activated partial thromboplastin time (APTT) is a useful global assay for the assessment of the contact factor pathway of hemostasis and its inhibitors. The test is usually performed on fully automated analyzers using commercially prepared reagents. The three main clinical areas of interest are detection of factor deficiencies, detection of lupus anticoagulants and in the monitoring of therapy with unfractionated heparin. Methods are described here for assessing APTT reagents for their sensitivity to clotting time prolongation in each of these areas of interest.

Performance and Interpretation of Mixing Tests in Coagulation.

Mixing tests in coagulation studies allow the laboratory to correctly differentiate factor deficiency from inhibitors as the cause of a prolonged clotting time. This is important in terms of subsequent laboratory follow-up and clinical patient management. In this chapter, a suggested method of performing mixing tests is given, with procedures applying two different formulas for interpretation, the percent correction and the index of circulating anticoagulant.

Rare bleeding disorders-old diseases in the era of novel options for therapy.

Rare diseases are defined as life-threatening or chronically debilitating diseases with a prevalence of less than one per 2000 according to the European Union or one per 1250 according to the USA. Congenital rare bleeding disorders RBD are reported in most populations, with incidence varying from 1 in 5000 (Hemophilia A), 1:30,000 (Hemophilia B) to much rarer (1:500,000 for FVII deficiency, 1-3 million for Prothrombin or FXIII deficiency). Acquired Hemophilia A is also a rare bleeding disorder with estimated frequency of 1 in million. Most RBDs are inherited as autosomal recessive (AR); however, heterozygous carriers with varying degrees of corresponding factor deficiency may render an unpredictable propensity for bleeding. In patients with bleeding symptoms, laboratory assessment and especially molecular techniques currently enable accurate diagnosis and may provide tools for prenatal and family counseling. Currently hemostasis control is mainly based upon replacement of the missing coagulation factors (unless presence of inhibitors renders it impossible), however future gene therapy and disruptive, non-replacement alternatives may be promising for patients with RBD.

Diagnóstico laboratorial de coagulopatías hemorrágicas en pacientes del Hospital Nacional de Itauguá / Laboratory diagnosis of hemorrhagic coagulopathies in patients of the National Hospital of Itauguá

Introducción: la hemostasia es el conjunto de sistemas que actúan coordinadamente para mantener la integridad de los vasos sanguíneos y la fluidez de la sangre; la alteración puede desencadenar trastornos trombóticos o hemorrágicos, dependiendo de la naturaleza de la falla. Objetivos: describir las coagulopatías hemorrágicas que se registraron en el departamento de laboratorio del Hospital Nacional de Itauguá, desde julio 2014 hasta diciembre 2015, obtener frecuencia y datos demográficos, edad, sexo, procedencia y clasificar las coagulopatías según deficiencias de factores de la coagulación, vía extrínseca, vía intrínseca y vía común final; sospecha de inhibidores adquiridos y enfermedad de von Willebrand. Material y Métodos: diseño observacional, descriptivo, retrospectivo de corte trasverso; incluyéndose pacientes de ambos sexos, todas las edades, derivados de médicos hematólogos. Resultados: se registraron 77 pacientes con coagulopatías hemorrágicas en el Laboratorio, 43 fueron del sexo masculino, de 1 a 75 años, mediana 18 años; 31% (24/77) con deficiencias del factor VII, todos leves, edades 7 a 75 años. En la vía intrínseca, la deficiencia del factor VIII o Hemofilia A, fue la más observada 29%(22/77), mayoría severas (13/22), mientras que déficit de factor IX, Hemofilia B, en 4 pacientes 5%(4/77). 85% Hemofilia A y 15% Hemofilia B, edades 1 y 64 años, mediana 13,5 años, todos del sexo masculino; no se registraron deficiencias de FXI y FXII en el periodo de estudio. De la vía común final, se encontraron 3 pacientes con hipofibrinogenemia, 1 con probable disfibrinogenemia, 1 con déficit de Factor II, 2 de Factor V y 2 de Factor X, 2 adultas, y el resto pediátricos; estas deficiencias son muy poco frecuentes, Factor I, V y X de 1/1.000.000 y FII 1/2.000.000 personas. Quince pacientes con sospecha de inhibidores, dos de ellas con inhibidor específico anti-FVIII, y probables inhibidores de interferencia. Se confirmó el primer déficit de factor Von Willebrand, en una mujer de 47 años. Conclusiones: entre las coagulopatías hemorrágicas de mayor frecuencia, se encuentran las Hemofilias A y B, seguida de deficiencias del factor VII y probables inhibidores de interferencia, los dos casos de inhibidores específicos anti Factor VIII fueron en pacientes con Hemofilia A severa. Fue relevante también el hallazgo de deficiencias de la vía común de la coagulación, a pesar de ser poco frecuentes. Algunos pacientes fueron diagnosticados en edad adulta, reflejando lo tardío que se llega al diagnóstico en el país.

Introduction: hemostasis is the set of systems that work in concert to maintain the integrity of blood vessels and blood flow; alteration can trigger thrombotic disorders or bleeding, depending on the nature of the fault Objective: describe hemorrhagic coagulopathy registered in the Medical Laboratory Department diagnosis at the National Hospital of Itauguá, from July 2014 to December 2015, obtain frequency and demographics, age, sex, origin and classify coagulopathy in to deficiencies of coagulation factors of the extrinsic pathway, intrinsic pathway, and common pathway; suspicion acquired inhibitors and von Willebrand's disease. Material and Methods: the design was an observational, descriptive, retrospective cross sectional study; being including patients of both sexes, all ages, referred by hematologists. Results: 77 patients with hemorrhagic coagulopathies, were female 43 male and 34 female, from 1 to 75 years, median age of 18 years; 31% (24/77) with factor VII deficiency, all mild, ages 7 to 75, 1 / 500,000 appears. In the intrinsic pathway, the factor VIII deficiency or hemophilia A, was the most observed 29% (22/77) severe majority (13/22), while Factor IX deficit, Hemophilia B, in 5% of patients (4/77). 85% Hemophilia A, and 15% Hemophilia B, ages 1 to 64 years, median age of 13.5 years, all male; no FXI and FXII deficiencies were recorded in the study period.Of the final common pathway, 3 patients with hipofibrinogenemia were found, 1 probable dysfibrinogenaemia, 1 deficiency of FII, 2 FV and FX 2, 2 were adult and the rest were pediatric; these deficiencies are rare, FI, V and X of 1 / 1,000,000 and FII 1 / 2,000,000 people. Fifteen patients with suspected inhibitors adquired, two of them with anti-FVIII specific inhibitor, and probable interference inhibitors. The first von Willebrand, factor deficiency was confirmed in a woman of 47 years. Conclusions: among the most frequent hemorrhagic coagulation disorders, we found hemophilia A and B, followed by deficiencies of factor VII and probable interference inhibitors both cases of specific inhibitors of factor VII were found in patients with severe Hemophilia A. Some patients were diagnosed in adulthood

Treatment of rare factor deficiencies in 2016.

Rare bleeding disorders (RBDs) are a heterogeneous group of coagulation disorders characterized by fibrinogen, prothrombin, factors V, VII, X, XI, or XIII (FV, FVII, FX, FXI, or FXIII, respectively), and the combined factor V + VIII and vitamin K-dependent proteins deficiencies, representing roughly 5% of all bleeding disorders. They are usually transmitted as autosomal, recessive disorders, and the prevalence of the severe forms could range from 1 case in 500 000 for FVII up to 1 in 2-3 million for FXIII in the general population. Patients affected with RBDs may present a wide range of clinical symptoms, varying from mucocutaneous bleeding, common to all types of RBDs to the most life-threatening symptoms such as central nervous system and gastrointestinal bleeding. Treatment of these disorders is mainly based on the replacement of the deficient factor, using specific plasma-derived or recombinant products. In countries where these facilities are not available, bleedings could be managed using cryoprecipitate, fresh frozen plasma (FFP), or virus-inactivated plasma. Minor bleedings could be managed using antifibrinolytic agents. Recently, 2 novel drugs, recombinant FXIIIA and a plasma-derived FX, have been added to the list of available specific hemostatic factors; only prothrombin and FV deficiencies still remain without a specific product. Novel no-replacement therapies, such as monoclonal antibody anti-tissue factor pathway inhibitor, RNA interference, and a bispecific antibody that is an FVIIIa mimetic, enhancing thrombin generation through different mechanisms, were developed for patients with hemophilia and may in the future be a good therapeutic option also in RBDs.

Point-of-care PT and aPTT in patients with suspected deficiencies of coagulation factors.

INTRODUCTION: There are several clinical settings and patient conditions especially in intensive care units, emergency departments, and operating theaters, where the coagulation status of a patient must be known immediately and point-of-care (POC) systems are beneficial due to low time to result. METHODS: This noninterventional, single-blinded, multicenter study with prospectively collected whole blood samples was performed to evaluate the diagnostic accuracy of the CoaguChek PT Test (POC PT) and CoaguChek aPTT Test (POC aPTT) compared to standard laboratory testing in patients with suspected deficiencies of coagulation factors. RESULTS: In total, 390 subjects were included. Both POC PT and POC aPTT showed concordance with the laboratory PT and aPTT. Lot-to-lot variation was below 2% both for POC PT and for POC aPTT. The mean relative difference of capillary blood compared to venous blood was 0.2 % with POC PT and 8.4% with POC aPTT. The coefficients of variation for repeatability of POC PT using whole blood were found to be between 2% and 3.6%. CONCLUSION: Our findings suggest reliable quantitative results with this POC system to support on-site decision-making for patients with suspected deficiencies of coagulation factors in acute and intensive care.

In vitro sensitivity of different activated partial thromboplastin time reagents to mild clotting factor deficiencies.

INTRODUCTION: Activated partial thromboplastin time (aPTT) is a routine clotting assay that is widely used to globally screen for coagulation abnormalities. It is commonly admitted that a prolonged test result, may trigger the need for specific assays to be performed, particularly factor measurement. However, the sensitivity of aPTT reagents to deficiencies of clotting factors varies. METHODS: We evaluated, according to the recommendation of the CLSI H47-A2 guideline, the responsiveness to single factor levels of five aPTT reagents by using factor-deficient plasmas spiked with a calibration plasma to produce individual factor activities ranging from <1 to ~100 Unit (U)/dL. Test results were expressed as the sample-to-control ratio, the latter was defined as the clotting time obtained in the calibration plasma containing ~100 U/dL factor activity. The factor activity producing a prolongation of aPTT above the upper limit of its specific normal range (in ratio) was assigned as the factor responsiveness in U/dL to that reagent. RESULTS: Responsiveness ranged from 34 to 47 U/dL to FVIII: C, from 18 to 57 U/dL to FIX, from 38 to 52 U/dL to FXI, from 29 to 50 U/dL to FXII, from 40 and 59 U/dL to FV, from 7.5 to 49 U/dL to FX, and from 9.1 to 10.5 U/dL to FII. CONCLUSIONS: These results suggest that the sensitivity of the tested aPTT reagents to single factor deficiency is highly variable. Moreover, for one given aPTT reagent, its sensitivity was very different depending on the deficient factor. This must be considered when analyzing clinical materials.

[Pseudoxanthoma elasticum-like disease with deficiency of vitamin K-dependent clotting factors and cutis laxa features]. / Pseudoxanthome élastique variant avec déficit en facteurs de coagulation vitamine K-dépendants et perte d'élasticité cutanée.

BACKGROUND: Pseudoxanthoma elasticum (PXE)-like syndrome is characterized by the association of PXE and cutis laxa (CL) features with a deficiency of vitamin K-dependent clotting factors. It was first described in 1971 and was identified as a distinct genetic entity in 2007 with analysis of the GGCX (γ-glutamyl carboxylase) gene, which is involved in congenital deficiency in vitamin K-dependent clotting factors. Here we report a new case of this extremely rare syndrome. PATIENTS AND METHODS: A 23-year-old female patient was seen for the emergence of loose and redundant skin following extensive weight loss. She also presented a deficiency of vitamin K-dependent clotting factors. Physical examination revealed excessive, leathery skin folds in the axillary and neck regions. A skin biopsy revealed polymorphous and fragmented elastic fibers in the reticular dermis. These were mineralized, as was demonstrated by Von Kossa staining. The clinical features of CL associated with the histopathological features of PXE and vitamin K-dependent clotting factor deficiency led us to a diagnosis of PXE-like syndrome. A molecular study of the GGCX gene showed compound heterozygosity. DISCUSSION: The GGCX gene is usually responsible for PXE-like syndrome. GGCX encodes a γ-glutamyl carboxylase necessary for activation of gla-proteins. Gla-proteins are involved both in coagulation factors in the liver and in the prevention of ectopic mineralization of soft tissues. Uncarboxylated forms of gla-proteins in fibroblast would thus enable mineralization and fragmentation of elastic fibers.

Applying a direct aPTT ratio (PlatelinLS/ActinFS) permits to identify rapidly and reliably a bleeding-related factor deficiency or a lupus anticoagulant sequential to an isolated prolongation of aPTT in paediatric pre-operative screening.

OBJECTIVES: An isolated prolongation of activated partial thromboplastin time (aPTT) found in paediatric pre-operative screening could be due to bleeding or non-bleeding aetiologies. The aim of our study was to evaluate clinical benefits of an additional ActinFS and/or a mixing aPTT study to identify a bleeding-related factor deficiency (BRFD). METHODS: Over a 4-yr period, isolated prolongation of aPTT with PlatelinLS was detected in 308 paediatric patients and confirmed in 161 cases by a 2nd sample. ActinFS, a mixing study, FVIII, FIX, FXI, FXII and lupus anticoagulant (LA) were performed. Three different aPTT ratios between PlatelinLS and ActinFS were analysed. RESULTS: We found 17 BRFD, 31 FXII deficiencies and 64 positive LA. A prolonged ActinFS had a significant association with BRFD (P < 0.0001) while a corrected mixing study did not. The direct aPTT ratio had a significant relationship with positive LA (P < 0.05), and with BRFD (P < 0.0001). Using this ratio, the sensitivity of BRFD's and LA's detection could be increased, respectively, to 82% from 59% using ActinFS alone, and to 86% from 55% using mixing study. CONCLUSIONS: Applying this direct aPTT ratio allows to quickly and reliably identify both BRFD and LA sequential to an isolated prolongation of aPTT.

Do PT and APTT sensitivities to factors' deficiencies calculated by the H47-A2 2008 CLSI guideline reflect the deficiencies found in plasmas from patients?

INTRODUCTION: Prothrombin time (PT) and activated partial thromboplastin time (APTT) sensitivity for detecting isolated factor deficiencies varies with different reagents and coagulometers. The Clinical and Laboratory Standards Institute (CLSI) H47A2 guideline proposed a method to calculate these sensitivities, but some inconsistency has been reported. This study aimed to calculate factor sensitivities using CLSI guideline and to compare them with those obtained from single factor-deficient patients' data. METHODS: Different mixtures of normal pooled and deficient plasmas were prepared (<1IU/dL to 100 IU/dL) according to the CLSI H47A2 guideline. PT with rabbit brain (RB) and human recombinant (HR) thromboplastins, APTT and factors' activities were measured in an ACL TOP coagulometer. Sensitivities (maximum factor concentration that produces PT or APTT values out of the reference range) were calculated from mixtures and from patients with single-factor deficiencies: 17 factor FV, 36 FVII, 19 FX, 39 FVIII, 15 FIX 15 FXI and 24 FXII. RESULTS: PT sensitivity with RB was as follows: FV 38 and 59, FVII 35 and 58, FX 56 and 64 IU/dL; PT sensitivity with HR was as follows: FV 39 and 45, FVII 51 and 50, FX 33 and 61 IU/dL; and APTT sensitivity was as follows: FV 39 and 45, FX 32 and 38, FVIII 47 and 60, FIX 35 and 44, FXI 33 and 43, FXII 37 and 46 IU/dL, respectively. CONCLUSIONS: Reagent-coagulometer combination has adequate sensitivities to factor deficiencies according to guideline recommendations (>30 IU/dL). These should not be considered as actual sensitivities because those obtained by analysing patients' plasmas with single-factor deficiencies were higher for most factors and could induce misinterpretation of the basic coagulation test results.

First-Step Results of Children Presenting with Bleeding Symptoms or Abnormal Coagulation Tests in an Outpatient Clinic.

OBJECTIVE: Mild bleeding symptoms are commonly seen in the general population. The aim of this study was to determine the final clinical and laboratory features of children referred for a first evaluation with a suspected bleeding disorder in the pediatric outpatient clinic of Istanbul University. MATERIALS AND METHODS: The medical records of 26,737 outpatients who were admitted to the Division of Ambulatory Pediatrics between 31 October 2011 and 31 October 2012 were evaluated retrospectively. Ninety-nine patients were initially diagnosed as having probable bleeding disorders and were followed up. The symptoms of bleeding in addition to coagulation tests were analyzed. RESULTS: Of the 99 patients, 52 (52.5%) were male and 47 were female, and the mean age of the entire study group was 9.1±4.1 years (minimum-maximum: 2-18 years). Major bleeding symptoms were epistaxis in 36 patients (36.4%), easy bruising in 32 (32.3%), and menorrhagia in 6 (6.1%). After initial tests ordered by the pediatrician, 36 of 99 patients (36.4%) were diagnosed as having bleeding disorders that included von Willebrand disease in 12 (12.1%), hemophilia A or B in 9 (9.1%), and other rare factor deficiencies in 9 (9.1%). Six patients (6.1%) were found to have combined deficiencies. Seven of 36 patients had a family history of bleeding. CONCLUSION: Among the patients referred for bleeding disorders, 36.4% were diagnosed with a bleeding disorder with the help of primary screening tests ordered in the outpatient clinic.