Patients with type 2B von Willebrand disease (vWD) (caused by gain-of-function mutations in the gene coding for von Willebrand factor) display bleeding to a variable extent and, in some cases, ...thrombocytopenia. There are several underlying causes of thrombocytopenia in type 2B vWD. It was recently suggested that desialylation-mediated platelet clearance leads to thrombocytopenia in this disease. However, this hypothesis has not been tested
The relationship between platelet desialylation and the platelet count was probed in 36 patients with type 2B von Willebrand disease (p.R1306Q, p.R1341Q, and p.V1316M mutations) and in a mouse model carrying the severe p.V1316M mutation (the 2B mouse). We observed abnormally high elevated levels of platelet desialylation in both patients with the p.V1316M mutation and the 2B mice.
, we demonstrated that 2B p.V1316M/von Willebrand factor induced more desialylation of normal platelets than wild-type von Willebrand factor did. Furthermore, we found that N-glycans were desialylated and we identified αIIb and β3 as desialylation targets. Treatment of 2B mice with sialidase inhibitors (which correct platelet desialylation) was not associated with the recovery of a normal platelet count. Lastly, we demonstrated that a critical platelet desialylation threshold (not achieved in either 2B patients or 2B mice) was required to induce thrombocytopenia
In conclusion, in type 2B vWD, platelet desialylation has a minor role and is not sufficient to mediate thrombocytopenia.
Summary
The ristocetin cofactor activity assay (VWF:RCo) is the reference method for assessing von Willebrand factor (VWF) activity but remains difficult to perform, and the coefficient of variation ...of the method is high (about 20–30%). This study evaluated and compared the performance for measuring the VWF activity of two newly commercialised assays VWF:Ac Innovance (VWF:Ac) and VWF:RCo Acustar (VWF:RCo Acu) with the reference VWF:RCo aggregation in 123 pathological plasma samples. The correlation and concordance between both new tests (VWF:RCo-Acu and VWF:Ac) and the reference VWF:RCo were good. The results of the VWF activity to VWF antigen ratio were also comparable whatever the method for the classification of VWF deficiency in all patients. Our results showed that both new tests could replace the “gold standard” VWF:RCo in aggregometry with several benefits: they are fully automated, easier and faster to perform, better adapted to emergency situations if necessary.
Two unrelated families were recruited in the French Reference Center for von Willebrand Disease with moderate bleeding symptoms associated with low von Willebrand factor (VWF) antigen levels, ...decreased collagen binding assay, and no or partial response to desmopressin. Genetic analysis showed the presence of heterozygous mutations in the A3 domain away from the collagen-binding surface: 1 never reported previously (p.L1696R) and another (p.P1824H) described in a Spanish family. The mutations were reproduced by site-directed mutagenesis and mutant VWF was expressed in different expression systems, COS-7 cells, baby hamster kidney cells, and in VWF-deficient mice through hydrodynamic injection. p.L1696R and p.P1824H were associated with very low expression levels both in vitro and in vivo, with intracellular retention for p.P1824H. Both homozygous mutants displayed decreased binding to collagen types I and III but also decreased binding to platelet glycoproteins Ib and IIbIIIa. Co-transfections with wild-type VWF partially corrected these defects, except that collagen binding remained abnormal. The in vivo thrombosis response was severely reduced for both heterozygous mutants. In conclusion, we report 2 VWF A3 domain mutations that induce a combined qualitative and quantitative defect.
•VWF A3 domain mutations inducing defective collagen binding and impaired protein production.
At birth, severe thrombocytopenia without context of infection should mainly suggest neonatal alloimmune thrombocytopenia (NAIT), especially in case of a platelet count below 20 GL
. We report two ...cases of severe neonatal thrombocytopenia, first suspected as being NAIT. Both had a platelet count below 20 GL
with platelet clumps. The absence of alloantibodies and failure of platelet transfusion and intravenous immunoglobulins to improve the platelet count led to question the diagnosis and to evoke inherited bleeding disorders. Measurements of Von Willebrand factor (VWF) levels showed a marked reduction of VWF:RCo and a normal VWF:Ag, suggesting a type 2B Von Willebrand disease (VWD2B). Ristocetin-induced platelet aggregation could not be performed because of the very low platelet count. In the first case, after sequencing VWF exon 28, a heterozygous p.Leu1460Pro mutation was found consistent with VWD2B. In the second case, the genetic analysis of VWF exon 28 identified a homozygous mutation: p.Pro1337Leu confirming type VWD2B and also the p.Arg854Gln homozygous mutation in exon 20 confirming type 2N (ratio FVIII/VWF:Ag <0.5). The two cases underline that, even if NAIT remains the most common diagnosis in severe neonatal thrombocytopenia, it should be challenged in the absence of documented incompatibility, chronic evolution, or treatment failure. Diagnosis of VWD2B should be considered in early thrombocytopenia, even without familial history. In the cases presented, genotyping confirmed the subtype of VWD and helped to guide the therapeutic management of bleeding episodes.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
We evaluated the use of the turbidimetric HemosIL von Willebrand Factor (VWF) Activity assay (VWF:Act) on the STA-R automated coagulometer (Stago, Asnières, France) for the diagnosis of von ...Willebrand disease (VWD). For this, we prospectively screened 268 patients. As a second part, we retrospectively assayed 111 patients with well-defined VWD subtype. In the first prospective study, we demonstrate that in most cases of VWD, VWF ristocetin cofactor activity (VWF:RCo) and VWF:Act are highly correlated but that they both cannot be considered a good screening assay when used alone, since they could miss about 25% of VWF abnormalities. However, the association of VWF:Act analysis and the Platelet Function Analyzer-100 (PFA-100) test constitutes an excellent screening strategy. In our second retrospective study concerning VWD subtypes, VWF:RCo and VWF:Act were well correlated but could be very discrepant, especially for some cases of type 2M VWD. We consider that VWF:RCo remains the “reference assay” for VWD subtype classification.
Emicizumab, a bispecific humanised monoclonal antibody restoring to some extent the function of activated FVIII deficient in haemophilia A, represents a major therapeutic advance in the management of ...haemophilia A patients. No dosage adjustment is required, which leads to a major change for patients used to regular biological monitoring which is particularly burdensome in the case of substitution therapy. In some circumstances, such as before an invasive procedure or in case of bleeding, biological monitoring will be necessary and emicizumab's interference with haemostasis tests, particularly those based on an activated partial thromboplastin times (aPTT), must be known to best interpret the tests and to select the most appropriate methods to guide therapy. The normalisation of aPTT in patients treated with emicizumab is not sufficient to consider haemostasis as normalised. In the event of administration of FVIII to a patient receiving emicizumab, the determination of FVIII should use a chromogenic method using non‐human reagents. Coagulation global tests have been proposed to evaluate the biological response when using bypassing agents in patients treated with emicizumab, but the usefulness must be confirmed. The French group BIMHO presents proposals for biological monitoring of a patient treated with emicizumab according to clinical situations.
BACKGROUND:Despite their usefulness in perioperative and acute care settings, factor-Xa inhibitor-specific assays are scarcely available, contrary to heparin anti-Xa assay. We assessed whether the ...heparin anti-Xa assay can (1) be used as a screening test to rule out apixaban, rivaroxaban, fondaparinux, and danaparoid levels that contraindicate invasive procedures according to current guidelines (>30 ng·mL, >30 ng·mL, >0.1 µg·mL, and >0.1 IU·mL, respectively), (2) quantify the anticoagulant level if found significant, that is, if it exceeded the abovementioned threshold.
METHODS:In the derivation cohort then in the validation cohort, via receiver operatingcharacteristics (ROC) curve analysis, we evaluated the ability of heparin anti-Xaassay to detect levels of factor-Xa inhibitors above or below the abovementioned safety thresholds recommended for an invasive procedure (screening test). Among samples with relevant levels of factor-Xa inhibitor, we determined the conversion factor linking the measured level and heparin anti-Xa activity in a derivation cohort. In a validation cohort, the estimated level of each factor-Xa inhibitor was thus inferred from heparin anti-Xa activity. The agreement between measured and estimated levels of factor-Xa inhibitors was assessed.
RESULTS:Among 989 (355 patients) and 756 blood samples (420 patients) in the derivation and validation cohort, there was a strong linear relationship between heparin anti-Xa activities and factor-Xa inhibitors measured level (r = 0.99 95% confidence interval {CI}, 0.99–0.99). In the derivation cohort, heparin anti-Xa activity ≤0.2, ≤0.3, <0.1, <0.1 IU·mL reliably ruled out a relevant level of apixaban, rivaroxaban, fondaparinux, and danaparoid, respectively (area under the ROC curve ≥0.99). In the validation cohort, these cutoffs yielded excellent classification accuracy (≥96%). If this screening test indicated relevant level of factor-Xa inhibitor, estimated and measured levels closely agreed (Lin’s correlation coefficient close to its maximal value95% CI, 0.99–0.99). More than 96% of the estimated levels fell into the predefined range of acceptability (ie, 80%–120% of the measured level).
CONCLUSIONS:A unique simple test already widely used to assay heparin was also useful for quantifying these 4 other anticoagulants. Both clinical and economic impacts of these findings should be assessed in a specific study.
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