Von Willebrand Activity Research Articles

No Increased Incidence of Bleeding in Essential Thrombocythemia Patients with Extreme Thrombocytosis

CONCLUSIONS About 25% of essential thrombocythemia (ET) patients present with extreme thrombocytosis (ExT), defined as having a platelet count ≥1000 x 10^9/L. ExT patients may have an increased bleeding risk, at least in part associated with acquired von Willebrand disease. There are no specific guidelines for managing ExT patients, and many are started on cytoreduction, even if they are younger and otherwise at low thrombotic risk by the revised International Prognostic Score of Thrombosis for ET (R-IPSET). We analyzed the risk of bleeding and thrombosis in ExT vs. non-ExT ET patients to inform treatment decisions. METHODS : We retrospectively analyzed patients who received targeted gene sequencing at Massachusetts General Brigham / Dana-Farber Cancer Institute from 2014-2021 and met WHO 2016 criteria for ET. We performed medical record review and recorded information on treatment, diagnosis, blood counts, and von Willebrand antigen and activity levels. We abstracted the first major bleed, clinically relevant non-major bleed (CRNMB), and thrombotic event in a patient's disease course. Bleeding was categorized using the International Society on Thrombosis & Haemostasis definitions.We determined the cumulative incidence rates of bleeding and thrombosis in ExT and non-ExT patients with death as a competing event and compared these rates using a Gray test. Risk factors for bleeding and thrombosis were evaluated using univariate and multivariate Fine Gray models. We identified 408 patients diagnosed with ET, of which 344 had a platelet count reported at diagnosis. There were 96 ExT patients and 248 non-ExT patients. Median follow-up was 11 and 6 years, and median platelet count at ET diagnosis was 1194 and 651, respectively. ExT patients, compared with non-ExT patients, were more likely to be younger (51 vs. 58 years old, p<.001), have the CALR mutation (49% vs. 23%, p<.001), have lower IPSET-thrombosis risk (65% vs. 49% very low or low risk, p<.001), and have lower von Willebrand activity levels at diagnosis or in disease course (60% vs. 94%, p<.001, Table 1). There were 7 (7%) major bleeds and 7 (7%) CRNMBs in the 96 ExT patients, and 11 (4%) major bleeds and 22 (9%) CRNMBs in the 248 non-ExT patients (p=.29 for major bleeding, p=.83 for CRNMB). 10 year cumulative incidence of major bleed and CRNMB was not different in ExT vs. non-ExT patients (4% vs. 5%, p=.68 for major bleed; 3% vs. 10%, p=.27 for CRNMB, Figure 1). Despite 65% of ExT patients in our sample categorized as very low or low risk for thrombosis, these patients were more likely to be treated with cytoreduction compared to non-ExT patients (63% vs. 50%, p=.21 for very low risk; 81% vs. 51%, p=.022 for low risk). However, there was no difference in the 10 year cumulative incidence of major bleeding and CRNMB in ExT patients who were treated vs. not treated with cytoreduction (6% vs. 0%, p=.13 for major bleed; 2% vs. 8%, p=.13 for CRNMB). There was also no difference in the cumulative incidence of bleeding in ExT and non-ExT patients who were not treated with cytoreduction (untreated or aspirin alone, 0% vs. 4%, p=.34 for major bleed; 8% vs. 9%, p=.75 for CRNMB). In univariate analysis, diabetes mellitus and thrombosis during disease course were significantly associated with increased combined major bleeding and CRNMB. In a multivariable analysis adjusting for these factors in addition to platelet count ≥1000 x 10^9/L and age at ET diagnosis, first von Willebrand factor activity, aspirin and cytoreduction during disease course, and JAK2 mutation status, diabetes mellitus (HR 3.45, 95% CI 1.31-8.72) and thrombosis during disease course (HR 2.92, 95% CI 1.41-6.03) remained as significant risk factors for bleeding. Cytoreduction was not significantly associated with a lower rate of bleeding in multivariable analysis (HR .42, 95% CI .09-3.06). 10 year cumulative incidence of thrombosis was not different in ExT vs. non-ExT patients (22% vs. 26%, p=.94). We found no difference in the cumulative incidence of bleeding and thrombosis in ET patients with ExT and importantly, no clear role for initiation of cytoreductive therapy in these patients who are otherwise low risk by R-IPSET. This confirms the current recommendations to initiate cytoreduction based on thrombotic risk and suggests that ExT alone should not be an indicator for cytoreduction.

Hemostasis Biomarkers and Long-Term Risk of Recurrent Cardiovascular Events and Bleeding Complications in Young Patients with an Ischemic Stroke

Background: Patients with ischemic stroke at a young age (18-50 years) have a persistent increased risk of recurrent ischemic cardiovascular events, despite antithrombotic treatment. Additionally, antithrombotic agents result in a higher risk of bleeding complications. Balancing hemostasis to prevent thrombotic events and minimizing the risk of hemorrhagic events is challenging. Previous research observed a hypercoagulable state and a significant degree of inflammation years after an acute ischemic stroke in young adults. It is known that an increased clotting tendency is a risk factor for a first ischemic stroke, especially in young adults. However, the risk profile of a recurrent event in these individuals can be different. To this end, our study aims to investigate the role of plasma pro- and anticoagulation biomarkers, platelet activation markers and thrombin generation parameters in the risk of recurrent ischemic arterial events and bleeding complications after a first ischemic stroke or transient ischemic attack (TIA) at a young age. Method: Patients from a single-center cohort study (the FUTURE cohort) with a first-ever TIA or ischemic stroke event between 1980 and 2010 were included. Blood was collected in 2010 and patients were followed on recurrent ischemic events or bleeding complications between 2010 and 2023. The following coagulation parameters were measured: blood cell count, Von Willebrand factor, active Von Willebrand factor, factor VIII, prothrombin, fragment 1.2, d-dimer, fibrinogen, fibrin degradation products, antithrombin, alpha-2-macroglobulin, serum CXCL-4 and CXCL-7. In addition, thrombin generation (TG) was determined using 1pM and 5pM tissue factor, and thrombomodulin in platelet-poor plasma to assess the functional capacity of plasma to form thrombin. The primary outcome was a composite of recurrent ischemic stroke, TIA, acute coronary syndrome or peripheral artery disease. The secondary outcome was the occurrence of a bleeding complication using the Bleeding Academic Research Consortium (BARC) criteria. We applied Cox proportional hazard models to estimate hazard ratios (HR) with 95% confidence intervals (CI). Results : Among 378 patients (median age 42.1 years, 56% women), 74 experienced a recurrent ischemic event during a mean follow-up of 6.5 years. An increased platelet and leukocyte count, along with elevated levels of fragment 1.2, fibrinogen and D-dimer were associated with a higher risk of ischemic events (table 1). Patients with low plasma antithrombin activity had a higher risk of recurrent ischemic events, especially in women (HR 0.67 [0.47-0.96]). After adjustment for age, sex and traditional risk factors, the association between leukocytes numbers, fragment 1.2 and fibrinogen levels and antithrombin activity and the risk of recurrent ischemic events remained: HR 1.14 [1.05-1.25], 1.07 [1.01-1.14], 0.80 [0.65-0.98], and 1.09 [1.00-1.17], respectively. In patients without vitamin K antagonist treatment, the association between antithrombin activity (HR 0.78 [0.62-0.98]) and leukocytes numbers (HR 1.15 [1.05-1.25]) with recurrent ischemic events were stronger. Thrombin generation parameters were not related to recurrence. Bleeding complications were observed in 16% of patients, mainly among women and predominantly attributable to gynecological bleedings. Hemoglobin, platelets, CXCL-4 and -7, prothrombin, fragment 1.2 and alpha-2-macroglobulin were associated with a bleeding complication (table 2). The association for platelets (HR 0.93 [0.89-0.97]), CXCL-4 (HR 0.90 [0.82-0.99]) and alpha-2-macroglobulin (HR 1.15 [1.03-1.29]) remained after adjustment. Conclusion: Lower antithrombin activity and, higher fragment 1.2, fibrinogen levels, and leukocytes counts were linked to an increased risk of recurrent ischemic events. Bleeding was associated with platelets, CXCL4, and alpha-2-macroglobulin. Overall, especially platelets appear to be significantly involved in the risk of both recurrent arterial thrombosis and bleeding, whereas the observed association with risk of recurrent ischemic events implies a more dominant role of the coagulation system.

Manifestations rares des gammapathies monoclonales : à propos de 2 cas et revue de la littérature

IntroductionMonoclonal gammopathies are common over the age of 50. Patients are usually asymptomatic. However, some patients present with secondary clinical manifestations, which are now grouped under the entity « Monoclonal Gammopathy of Clinical Significance » (MGCS). Case reportHere, we report two rare cases of MGCS: an acquired von Willebrand syndrome (AvWS) and an acquired angioedema (AAE). ConclusionThe discovery of a decrease in von Willebrand activity (vWF:RCo) or angioedema in a patient over 50 years of age, in the absence of a family history, should prompt a search for a hemopathy and in particular, a monoclonal gammopathy.

Von Willebrand factor: from figurant to main character in the scene of inflammation

von Willebrand factor: from figurant to main character in the scene of inflammation

Weight-adjusted dosing of tinzaparin for thromboprophylaxis in obese medical patients

BackgroundThe optimal dose of tinzaparin for prophylaxis in obese medical patients is not well defined. ObjectivesTo evaluate the anti-Xa activity in obese medical patients on tinzaparin prophylaxis adjusted for actual bodyweight. MethodsPatients with a body mass index of ≥30 kg/m2 treated with 50 IU/kg tinzaparin once daily were prospectively included. Anti-Xa and anti-IIa activity; von Willebrand factor antigen and von Willebrand activity; factor VIII activity; D-dimer, prothrombin fragments; and thrombin generation were measured 4 hours after subcutaneous injection between days 1 and 14 after the initiation of tinzaparin prophylaxis. ResultsWe included 121 plasma samples from 66 patients (48.5% women), with a median weight of 125 kg (range, 82-300 kg) and a median body mass index of 41.9 kg/m2 (range, 30.1-88.6 kg/m2). The target anti-Xa activity of 0.2 to 0.4 IU/mL was achieved in 80 plasma samples (66.1%); 39 samples (32.2%) were below and 2 samples (1.7%) above the target range. The median anti-Xa activity was 0.25 IU/mL (IQR, 0.19-0.31 IU/mL), 0.23 IU/mL (IQR, 0.17-0.28 IU/mL), and 0.21 IU/mL (IQR, 0.17-0.25 IU/mL) on days 1 to 3, days 4 to 6, and days 7 to 14, respectively. The anti-Xa activity did not differ among the weight groups (P = .19). Injection into the upper arm compared to the abdomen resulted in a lower endogenous thrombin potential, a lower peak thrombin, and a trend to a higher anti-Xa activity. ConclusionDosing of tinzaparin adjusted for actual bodyweight in obese patients achieved anti-Xa activity in the target range for most patients, without accumulation or overdosing. In addition, there is a significant difference in thrombin generation depending on the injection site.

Functional changes in hemostasis during asexual and sexual parasitemia in a controlled human malaria infection.

Decreased platelet count is an early phenomenon in asexual Plasmodium falciparum parasitemia, but its association with acute or long-term functional changes in platelets and coagulation is unknown. Moreover, the impact of gametocytemia on platelets and coagulation remains unclear. We investigated the changes in platelet number and function during early asexual parasitemia, gametocytemia and convalescence in 16 individuals participating in a controlled human malaria infection study, and studied its relationship with changes in total and active von Willebrand factor levels (VWF) and the coagulation system. Platelet activation and reactivity were determined by flow cytometry, and the coagulation system was assessed using different representative assays including antigen assays, activity assays and global functional assays. Platelet count was decreased during asexual blood stage infection but normalized during gametocytemia. Platelet P-selectin expression was slightly increased during asexual parasitemia, gametocytemia and at day 64. In contrast, platelet reactivity to different agonists remained unchanged, except a marked decrease in reactivity to low dose collagen-related peptide-XL. Thrombin generation and antigen assays did not show a clear activation of the coagulation during asexual parasitemia, whereas total and active VWF levels were markedly increased. During gametocytemia and on day 64, the endogenous thrombin potential, thrombin peak and velocity index were increased and prothrombin conversion and plasma prothrombin levels were decreased. We conclude that the decreased platelet count during asexual parasitemia is associated with increased active VWF levels (i.e. endothelial activation), but not platelet hyperreactivity or hypercoagulability, and that the increased platelet clearance in asexual parasitemia could cause spontaneous VWF-platelet complexes formation.

Ex Vivo Assessment of Different Oral Anticoagulant Regimens on Pump Thrombosis in a HeartWare Ventricular Assist Device

In light of decreased intracranial hemorrhage with direct oral anticoagulants and concerns about their safety in continuous flow left ventricular assist devices, we conducted an ex vivo study of thrombus formation using multiple anticoagulation agents. A continuous flow left ventricular assist device (HeartWare ventricular assist device) hemocompatibility loop was run using human blood under 7 conditions: control (no anticoagulation or antiplatelet); in vitro addition of aspirin; in vitro addition of apixaban at low dose (equivalent 2.5 mg twice daily); addition of apixaban at high dose (equivalent 5 mg twice daily); patients on warfarin; patients on apixaban (5 mg twice daily); and patients on dabigatran (150 mg twice daily). The primary outcome was time to formation of intrapump thrombosis. Secondary outcomes were reduction in clotting times over 1 hour, hemolysis, reduced platelet aggregation, and von Willebrand activity. Twenty-one runs were completed. Times to thrombosis in median (interquartile range) were control, 131 (127-134.5); in vitro aspirin, 124 (114.5-137); and patients on dabigatran, 131 (130.5-135.5) minutes, respectively. Times in patients on warfarin were, 137 (136.5-143.5); in vitro low-dose apixaban, 141 (138.5-142); and patients on apixaban, 140 (138-142.5) minutes, respectively. No thrombus formed in the in vitro high-dose apixaban group. There were no significant differences between the individual groups. When all apixaban groups were compared with nonapixaban groups, the time to thrombosis formation was significantly longer, 143 (137-150) versus 133.5 (128.5-140) minutes, P=0.02. There were similar changes in lactate dehydrogenase levels and other secondary end points. In an in vitro study of anticoagulation using human blood in a mock loop with a HeartWare HVAD, we demonstrated similar thrombosis times for apixaban and warfarin. Time to clotting was longer in the combined apixaban groups compared with combined other groups, but thrombosis times between individual groups were not significantly different.

Transient Ischemic Attack in a Hemophilia Patient with Severe Preeclampsia after Preoperative Administration of Tranexamic Acid and Factor VIII Replacement for Cesarean Section

Hemophilia A in females accounts for few cases due to hemophilia A and B having X-linked recessive inheritance patterns. Hemostatic changes in pregnancy include an increase in coagulation factors and von Willebrand activity, placing hemophilia patients at an increased risk for Postpartum Hemorrhage (PPH). General recommendations include considering pharmacologic prophylaxis, including tranexamic acid and factor replacement when necessary. The ultimate goal is to prevent uncontrolled bleeding during vaginal or operative delivery. Benefits of prophylactic therapies must be weighed with relevant risk profiles of each intervention. We present a case where a parturient with hemophilia prophylactically treated with TXA and FVIII experienced a transient ischemic attack. We discuss the background information known regarding tranexamic acid and factor replacement, and the subsequent recommendations for their use in this patient population. We consider recommendations to expand the multidisciplinary team incorporated in the assessment and planning for the peripartum care of such a patient.

Ischemic Stroke, Inflammation, and Endotheliopathy in COVID-19 Patients.

[Figure: see text].

Ex Vivo Perfusion and In Vivo Xenotransplantation of Pig Lungs with Humanized Von Willebrand Factor Demonstrate Reduced Platelet Sequestration

<h3>Purpose</h3> In normal hemostasis, activated von Willebrand factor (vWF) stimulates platelet adhesion and aggregation, but porcine vWF (pvWF) is "always active" when exposed to human blood. We hypothesized that humanized vWF (h*pvWF) would reduce this nonphysiologic platelet adhesion in lung xenotransplantation models. <h3>Methods</h3> GTKO.hCD46.h*pvWF pig lungs were perfused with human blood <i>ex vivo</i> and compared to reference GTKO.hCD46 lungs. Anti-GPIb Fab, 1-BIA (thromboxane synthase inhibitor), and antihistamines were used in all perfusions. One lung from each pair (n=5 h*pvWF, n=5 ref) also received integrin (H52/IB4) and selectin (GMI-1271, PSGL-1) blockade. Lungs surviving 8 hrs of perfusion were terminated electively. Platelets were measured by flow cytometry. h*pvWF lungs were also transplanted into baboon recipients (n=3 h*pvWF, n=10 ref), with platelets measured by CBC. <h3>Results</h3> All <i>ex vivo</i> groups had similar survival, with most electively terminated. Pulmonary vascular resistance was very low and constant in all groups. Platelet sequestration was significantly reduced and delayed in h*pvWF lungs compared to pvWF (p<0.01), not further modulated by selectin and integrin blockade (Fig.1). No significant differences were seen in BTG (platelet activation) levels, F1+2 (thrombosis) levels, or neutrophil counts. h*pvWF lung baboon recipients trended toward longer survival (p=0.07) with attenuated thrombocytopenia compared to reference lungs (p<0.01)(Fig.2). <h3>Conclusion</h3> When perfused with human blood, h*pvWF lungs had reduced sequestration of platelets compared to pvWF lungs, with similar reductions in lung xenotransplant recipients. "Humanization" of porcine vWF attenuates the nonphysiologic platelet adhesion and aggregation associated with xenograft rejection. Further study will focus on Fc-receptors, adenosine and coagulation pathway dysregulation, and activated neutrophils through additional drugs and genetic modifications.

Vascular activation is a strong predictor of mortality in coronavirus disease 2019 patients on the ICU

Respiratory failure in coronavirus disease 2019 (COVID-19) patients is one of the most frequent causes for referral to the ICU. A significant percentage of these patients does not survive the infection due to thromboembolic complications. Furthermore, the vascular system seems also to be involved in the pathogenesis. To investigate the role of hemostasis and endothelium on the outcome of COVID-19 patients admitted to the ICU. Blood was drawn from 16 ICU COVID-19 patients for hemostatic analysis. Patients were followed-up till discharge (n = 11) or death (n = 5). Parameters related to both coagulation and fibrinolysis, though disturbed, were not associated with mortality. Contrarily, activated Von Willebrand factor was increased and ADAMTS13 levels were decreased by two-fold in nonsurvivors compared with survivors. Our data established the involvement of the Von Willebrand factor-ADAMTS13 axis in the COVID-19 pathogenesis, thereby demonstrating that these plasma proteins seem to be strong predictors for ICU mortality.

Targeting shear gradient activated von Willebrand factor by the novel single-chain antibody A1 reduces occlusive thrombus formation in vitro.

Intraluminal thrombus formation precipitates conditions such as acute myocardial infarction and disturbs local blood flow resulting in areas of rapidly changing blood flow velocities and steep gradients of blood shear rate. Shear rate gradients are known to be pro-thrombotic with an important role for the shear-sensitive plasma protein von Willebrand factor (VWF). Here, we developed a single-chain antibody (scFv) that targets a shear gradient specific conformation of VWF to specifically inhibit platelet adhesion at sites of shear rate gradients (SRG) but not in areas of constant shear. Microfluidic flow channels with stenotic segments were used to create SRG during blood perfusion. VWF-GPIbα interactions were increased at sites of SRG compared to constant shear rate of matched magnitude. The scFv-A1 specifically reduced VWF-GPIbα binding and thrombus formation at sites of SRG but did not block platelet deposition and aggregation under constant shear rate in upstream sections of the channels. Significantly, the scFv A1 attenuated platelet aggregation only in the later stages of thrombus formation. In the absence of shear, direct binding of scFv-A1 to VWF could not be detected and scFVA1 did not inhibit ristocetin induced platelet agglutination. We have exploited the pro-aggregatory effects of SRG on VWF dependent platelet aggregation and developed the shear gradient-sensitive scFv-A1 antibody that inhibits platelet aggregation exclusively at sites of SRG. The lack of VWF inhibition in non-stenosed vessel segments places scFV-A1 in an entirely new class of anti-platelet therapy for selective blockade of pathological thrombus formation while maintaining normal hemostasis.

High risk of thrombosis in patients with severe SARS-CoV-2 infection: a multicenter prospective cohort study.

PurposeLittle evidence of increased thrombotic risk is available in COVID-19 patients. Our purpose was to assess thrombotic risk in severe forms of SARS-CoV-2 infection.MethodsAll patients referred to 4 intensive care units (ICUs) from two centers of a French tertiary hospital for acute respiratory distress syndrome (ARDS) due to COVID-19 between March 3rd and 31st 2020 were included. Medical history, symptoms, biological data and imaging were prospectively collected. Propensity score matching was performed to analyze the occurrence of thromboembolic events between non-COVID-19 ARDS and COVID-19 ARDS patients.Results150 COVID-19 patients were included (122 men, median age 63 [53; 71] years, SAPSII 49 [37; 64] points). Sixty-four clinically relevant thrombotic complications were diagnosed in 150 patients, mainly pulmonary embolisms (16.7%). 28/29 patients (96.6%) receiving continuous renal replacement therapy experienced circuit clotting. Three thrombotic occlusions (in 2 patients) of centrifugal pump occurred in 12 patients (8%) supported by ECMO. Most patients (> 95%) had elevated D-dimer and fibrinogen. No patient developed disseminated intravascular coagulation. Von Willebrand (vWF) activity, vWF antigen and FVIII were considerably increased, and 50/57 tested patients (87.7%) had positive lupus anticoagulant. Comparison with non-COVID-19 ARDS patients (n = 145) confirmed that COVID-19 ARDS patients (n = 77) developed significantly more thrombotic complications, mainly pulmonary embolisms (11.7 vs. 2.1%, p < 0.008). Coagulation parameters significantly differed between the two groups.ConclusionDespite anticoagulation, a high number of patients with ARDS secondary to COVID-19 developed life-threatening thrombotic complications. Higher anticoagulation targets than in usual critically ill patients should therefore probably be suggested.Electronic supplementary materialThe online version of this article (10.1007/s00134-020-06062-x) contains supplementary material, which is available to authorized users.

Impact of Intraoperative Cell Salvage on Blood Coagulation Factor Concentrations in Patients Undergoing Cardiac Surgery.

Cardiothoracic surgery is associated with major blood loss and allogeneic transfusion of red blood cell concentrates. To minimize allogeneic red blood cell (RBC) transfusion, intraoperative cell salvage has been effectively used for years. The objective of this study was to evaluate the impact of cell salvage on blood coagulation factors. We enrolled 30 patients scheduled for cardiac surgery in a prospective single-center observational cohort study at an academic hospital. Blood samples from the cell salvage system were obtained from both the reservoir and the processed red blood cell concentrate. Coagulation factors, fibrinogen, antithrombin and von Willebrand activity, and antigen were assessed before and after cell salvage. Statistical analysis was performed using Wilcoxon matched-pairs signed rank test. Our results revealed a significant decrease of fibrinogen (P < .001), coagulation factors II (P = .004), factors VII, X, and XIII (P < .001), and all other measured coagulation factor concentrations/activities in the processed red blood cell concentrate, when compared to the concentrations/activities of the reservoir. The results of the present study revealed a significant reduction of coagulation factor concentrations/activities by the washing process. Therefore, physicians need to consider adequate management of coagulation in patients with major blood loss and the need of large volumes of RBC transfusion.

ADAMTS-13 and Von Willebrand Factor Levels and Activities in Children With Cirrhosis and/or Portal Hypertension

ADAMTS-13 and Von Willebrand Factor Levels and Activities in Children With Cirrhosis and/or Portal Hypertension

Implant stability in patients treated with platelet-rich fibrin and bovine bone substitute for alveolar ridge preservation is associated with peripheral blood cells and coagulation factors.

AimsThe aim of the present study was to assess the association between dental implant stability and peripheral blood cell composition and levels of coagulation factors in patients treated with alveolar ridge preservation with platelet‐rich fibrin (PRF) and bovine bone substitute.Materials and methodsFifty patients were included between 2015 and 2017. PRF was prepared from autologous blood, in which blood cells and coagulation factor levels were measured. PRF and bovine bone were placed in the socket, followed by closure with PRF membrane. Implants were placed 14 (±2.5) weeks postextraction. The implant stability quotient was measured at t = 0, t = 10 days, t = 7 weeks, and t = 17 weeks by resonance frequency analysis.ResultsErythrocyte count was inversely associated with PRF membrane length, but not with implant stability. Conversely, platelet count did not correlate with membrane size but inversely correlated with implant stability at 7 and 17 weeks. In addition, implant stability was directly correlated with levels FXIII (t = 0, p < .01), active von Willebrand factor (VWF; t = 0 and 7 weeks, p < .05), and total VWF (t = 7 weeks, p = .012).ConclusionImplant stability following alveolar ridge preservation with PRF and bovine bone substitute is associated with circulating blood cells and coagulation factors. In particular, fibrin structure, VWF, and FXIII may be important modulators of implant stability.

The Platelet Millionaire: A Rare Cause of Thrombocytosis in an Adolescent

Objectives: Primary myelofibrosis is rare in pediatrics, often manifesting as persistent idiopathic thrombocytosis.Transitions from pediatric to adult medical care can be complicated by workup requiring invasive procedures. J.M., an 18-year-old healthy male, presented for excessive gingival bleeding after wisdom tooth extraction. Workup revealed persistent thrombocytosis to 1,165K, prompting a referral to hematology-oncology. A peripheral smear was notable for many platelets but normal RBC morphology. He had splenomegaly on abdominal ultrasound and a decreased von-Willebrand's activity to antigen ratio, suggesting acquired vWD. A bone marrow biopsy was advised; however, J.M. became lost to follow up for over 9 months owing to self-reported anxiety about the procedure. He remained asymptomatic in this interim until he re-presented to clinic for easy bruising, with no other evidence of bleeding at the time. The biopsy was pursued, revealing hypercellular marrow for age with left shifted granulocytic and erythroid maturation, abnormal megakaryocytes, and 3% blasts. This was consistent with primary early myelofibrosis (PMF), positive for MF-1, CALR, and TP53 mutations and negative for JAK2 and BCR-ABL. He was transitioned to adult hematology, maintained on baby aspirin, and referred for potential allogeneic hematopoietic stem cell transplant (HSCT). PMF is characterized by marrow fibrosis due to secretion of fibroblast growth factor by clonally proliferative megakaryocytes. It is a disease of adulthood, with 67 years being the median age at diagnosis. Only 100 cases have been reported in children, most of which are secondary to AML, ALL or other malignancies.1 Most patients present with complications of extramedullary hematopoiesis or bleeding.2 Diagnosis is suggested by a leukoerythroblastic picture on peripheral smear and confirmed with a bone marrow biopsy "dry tap" revealing marrow fibrosis.3 Prognosis in pediatric PMF is difficult to predict but outcomes tend to be worse;4 TP53 mutation is rare and based on limited adult studies may portend a poorer prognosis.5 Our young patient with this rare mutation was therefore referred for HSCT evaluation. Further complicating this case was J.M.'s anxiety, which delayed definitive diagnosis by biopsy. He only agreed to it when, at the med-peds clinic, the concept of local pain management was discussed. Anticipation of upcoming procedures by primary care physicians and close follow-up is especially important for patients transitioning from pediatric to adult providers. Disclosures No relevant conflicts of interest to declare.

Circulating active von Willebrand factor levels are increased in chronic kidney disease and end-stage renal disease.

Circulating active von Willebrand factor levels are increased in chronic kidney disease and end-stage renal disease.

Non-inferiority of a new advanced dry plasma thawing system compared to leading standard method: Preservation of coagulation and anticoagulation proteins necessary for use in the clinic.

e14675 Background: Cancer can affect the bone marrow, cause anemia and internal or excess bleeding, requiring blood or plasma transfusion. Fresh frozen plasma (FFP) should be thawed before transfusing to patients but it must be monitored closely to avoid contamination, uncontrolled or prolonged thawing which may subsequently lead to plasma protein denaturation. In this study, we compared and evaluated the effect of two different thawing methods on clotting factor activities as well as factors such as throughput and turnaround time. Methods: Over a period of approximately one month, November 6 to December 3, 2018, our testing laboratory at the UCSD School of Medicine received three sets of samples for a total of 197 freshly collected (pre-freeze) or freshly thawed plasma samples (standard commercial thawing device and the new dry ZipThaw device) that had been collected and processed in accordance with a San Diego Blood Bank protocol described elsewhere. The Lab conducted over 1,782 tests on these plasma for the following factors: Prothrombin time (PT), International Normalized Ratios (INR), Activated Partial Thromboplastin time (aPTT) Factor VIII activity, Factor V activity, Protein C activity, Protein S antigen, Von Willebrand activity and Thrombin Antithrombin Complex (TAT) concentration. The assays were carried out on an ACL TOP 700 Hemostasis Analyzer (Instrumentation Laboratory, Bedford, MA, USA) using manufacturer’s reagents and automated protocols. The ACL TOP 700 has validated precision CV of 5- 10%. All assays were completed within 4 hours of collection or, if frozen, within 4 hours of thawing. The statistical analysis method employed is two-tailed, student t-test. Results: We observed 100% concordance with no statistically significant differences between the pre-freeze group and the two different post thaw groups, except for a slight increase in the mean TAT complex concentration. Overall, the means of all three groups were well within the normal range defined by the assay kits (1-10 ng/ml). Further, the increased mean TAT is more notable with standard method (33%, p = 0.004) than with the ZipThaw (18%, p = 0.054). Conclusions: We report for the first time non-inferiority between the new ZipThaw device and the leading commercial predicate in plasma thawing while preserving coagulation and anticoagulation proteins and minimizing activation of clotting cascade. The results provide support for the use of ZipThaw device in clinical settings.

Analytical characterization and reference interval of an enzyme-linked immunosorbent assay for active von Willebrand factor.

BackgroundInteraction of von Willebrand factor (VWF) with platelets requires a conformational change that exposes an epitope within the VWF A1 domain, enabling platelet glycoprotein Ibα binding. Quantification of this ‘‘active” conformation of VWF has been shown to provide pathophysiological insight into conditions characterized by excessive VWF-platelet interaction.MethodsWe developed an immunosorbent assay based on a variable heavy chain antibody fragment against the VWF A1 domain as a capture antibody. Assay performance in terms of specificity (binding to active R1306W- and sheared VWF), precision, accuracy, linearity, limits of detection and stability were determined. Active VWF, VWF antigen, VWF ristocetin cofactor activity, VWF:GP1bM and VWF propeptide were measured in citrated plasma and platelet-VWF binding in whole blood from 120 healthy individuals to establish a reference interval for active VWF and to assess associations with other VWF parameters.ResultsIntra- and inter-assay CVs were between 2.4–7.2% and 4.1–9.4%, depending on the level. Mean recovery of spiked recombinant R1306W VWF was 103±3%. The assay was linear in the range of 90.1–424.5% and had a limit of quantification of 101%. The reference interval for active VWF was 91.6–154.8% of NPP. Significant, positive correlations between active VWF and all other VWF parameters were found, with the strongest correlation with VWF:GP1bM binding.ConclusionsWe developed and validated an immunosorbent assay for the accurate detection of active VWF levels in plasma. The assay fulfilled all analytical criteria in this study and a reference interval was established, allowing its use to quantify active VWF in pathological conditions for future research.