Acquired hemophilia A revealed by spontaneous bleeding in an 80-year-old man: a marginal diagnosis?

This case report illustrates the difficulty associated with diagnosing acquired hemophilia A by reviewing the case of an 80-year-old man admitted to the hospital for anemia. A prolonged activated partial thromboplastin time (aPTT) was not noticed until the patient developed a severe hemorrhagic syndrome.

Inherited deficiency of stress granule ZNFX1 in patients with monocytosis and mycobacterial disease.

Human inborn errors of IFN-γ underlie mycobacterial disease, due to insufficient IFN-γ production by lymphoid cells, impaired myeloid cell responses to this cytokine, or both. We report four patients from two unrelated kindreds with intermittent monocytosis and mycobacterial disease, including bacillus Calmette-Guérin-osis and disseminated tuberculosis, and without any known inborn error of IFN-γ. The patients are homozygous for ZNFX1 variants (p.S959* and p.E1606Rfs*10) predicted to be loss of function (pLOF). There are no subjects homozygous for pLOF variants in public databases. ZNFX1 is a conserved and broadly expressed helicase, but its biology remains largely unknown. It is thought to act as a viral double-stranded RNA sensor in mice, but these patients do not suffer from severe viral illnesses. We analyze its subcellular localization upon overexpression in A549 and HeLa cell lines and upon stimulation of THP1 and fibroblastic cell lines. We find that this cytoplasmic protein can be recruited to or even induce stress granules. The endogenous ZNFX1 protein in cell lines of the patient homozygous for the p.E1606Rfs*10 variant is truncated, whereas ZNFX1 expression is abolished in cell lines from the patients with the p.S959* variant. Lymphocyte subsets are present at normal frequencies in these patients and produce IFN-γ normally. The hematopoietic and nonhematopoietic cells of the patients tested respond normally to IFN-γ. Our results indicate that human ZNFX1 is associated with stress granules and essential for both monocyte homeostasis and protective immunity to mycobacteria.

Is There Any Improvement of the Coagulation Imbalance in Sickle Cell Disease after Hematopoietic Stem Cell Transplantation?

Several components of the clotting system are modified towards hypercoagulability in sickle cell disease (SCD). To date, hematopoietic stem cell transplantation (HSCT) is the only validated curative treatment of SCD. Here, we investigated the changes in the hemostatic potential of SCD children who've received a successful HSCT. Seventeen children with severe SCD were enrolled in the study. Thrombin generation (TG) was performed on citrated platelet-poor plasma, obtained before and 3, 6, 9, 12 and 15 months after HSCT. TG was triggered using 1 pM tissue factor and 4 µM phospholipids with or without thrombomodulin (TM). Before the HSCT, SCD children showed a higher endogenous thrombin potential (ETP), higher peak, higher velocity and shorter time-to-peak of TG than the normal controls (NC). ETP did not significantly change following the HSCT. However, the peak, velocity and time-to-peak of TG reversed to normal ranges from 3 months post-HSCT and remained so up to 15 months post-HSCT. The reduction of ETP after the addition of thrombomodulin (RETP) was dramatically reduced in SCD children before HSCT as compared with the NC. A partial reversal of RETP was observed from 3 months through 15 months post-HSCT. No statistical difference was observed for patient age or donor hemoglobinopathy status. In summary, successful HSCT improves the kinetics of TG but not the total thrombin capacity in SCD children.

Different profile of thrombin generation in children with acute lymphoblastic leukaemia treated with native or pegylated asparaginase: A cohort study.

BACKGROUND: Asparaginase (Asp) and corticosteroid (CS) treatment in patients with acute lymphoblastic leukaemia (ALL) is associated with an increased risk of thrombotic events. OBJECTIVE: Characterization of global haemostatic phenotypes of patients with ALL during Asp therapy. PROCEDURE: Thrombin generation (TG) was monitored in platelet-poor plasma of 56 children treated for a B lineage ALL (36 with native, 20 with PEG Asp) using 1 pM tissue factor and 4 µM phospholipids, with and without thrombomodulin. Protein C activity (PC), free protein S (PS), antithrombin (AT) and fibrinogen levels were also measured. RESULTS: Elevated endogenous thrombin potential (ETP) and peak of TG were noted at diagnosis, throughout the Induction phase and Late Intensification but was significantly less for PEG than for native Asp (P < 0.001), while age, sex, type of corticosteroid during Induction and molecular response had no significant effect. The reduction of ETP after addition of thrombomodulin was significantly lower in ALL children compared with that in controls, suggesting impairment in PS/PC pathway. Three patients experienced thrombosis: two treated with native and one with PEG Asp. The two patients with native Asp had, at the time of thrombosis, a prothrombotic profile. CONCLUSIONS: Treatment with Asp, in combination with CS, enhances TG in children with ALL, more significantly with native than PEG Asp, which is present early at diagnosis, persists during Induction and reappears during Late Intensification. This is consistent with the high incidence of thrombotic events described during these phases of therapy. The less pronounced effect of PEG Asp remains to be elucidated.

Heparin Reversal After Cardiopulmonary Bypass: Are Point-of-Care Coagulation Tests Interchangeable?

OBJECTIVE: Protamine is used to neutralize heparin after patient separation from cardiopulmonary bypass (CPB). Different bedside tests are used to monitor the adequacy of heparin neutralization. For this study, the interchangeability of the activated coagulation time (ACT) and thromboelastometry (ROTEM; Tem Innovations GmbH, Basel, Switzerland) clotting time (CT) ratios in children undergoing cardiac surgery was assessed. DESIGN: Single-center, retrospective, cohort study between September 2010 and January 2012. SETTING: University children's hospital. PARTICIPANTS: The study comprised children 0 to 16 years old undergoing elective cardiac surgery with CPB. Exclusion criteria were preoperative coagulopathy, Jehovah's witnesses, and children in a moribund condition (American Society of Anesthesiologists score 5). INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: After heparin neutralization with protamine, the ratio between ACT, with and without heparinase, and the CT measured with INTEM/HEPTEM (intrinsic test activated with ellagic acid was performed without heparinase [INTEM] and with heparinase [HEPTEM]) using tests of ROTEM were calculated. Agreement was evaluated using Cohen's kappa statistics, Passing-Bablok regression, and Bland-Altman analysis. Among the 173 patients included for analysis, agreement between both tests showed a Cohen's kappa statistic of 0.06 (95% CI: -0.02 to 0.14; p = 0.22). Bland-Altman analysis showed a bias of 0.01, with a standard deviation of 0.13, and limits of agreement between -0.24 and 0.26. Passing-Bablok regression showed a systematic difference of 0.40 (95% CI: 0.16-0.59) and a proportional difference of 0.61 (95% CI: 0.42-0.86). The residual standard deviation was 0.11 (95% CI: -0.22 to 0.22), and the test for linearity showed p = 0.10. CONCLUSION: ACT, with or without heparinase, and the INTEM/HEPTEM CT ratios are not interchangeable to evaluate heparin reversal after pediatric patient separation from CPB. Therefore, the results of these tests should be corroborated with the absence/presence of bleeding and integrated into center-specific treatment algorithms.

Performances of the H-Score for Diagnosis of Hemophagocytic Lymphohistiocytosis in Adult and Pediatric Patients.

OBJECTIVES: In this study, we compared the performances of adapted hemophagocytic lymphohistiocytosis (HLH)-2004 guidelines with those of the new diagnostic H-score to identify patients with HLH in a multicenter cohort consisting of adult and pediatric cases of suspected HLH. METHODS: The study sample consisted of 147 cases, including 20 adults and 16 children with HLH. Two sets of biological data were evaluated: at presentation and the maximal values reached during the episode. RESULTS: At presentation, for both children and adults, the H-score was more efficient than adapted HLH-2004 guidelines to identify HLH. The diagnostic sensitivity and specificity were respectively 100% and 80% for children and 90% and 79% for adults. However, for adults, performances became comparable between adapted HLH-2004 guidelines and H-score as patient clinical status worsened. The specificity decreased to 73% for the same sensitivity. CONCLUSIONS: The adapted HLH-2004 guidelines seem less powerful and H-score seems to be more appropriate for children, which may be due to less significantly marked biological features. For adults, H-score performances are better when determined at presentation. The cutoff value of the H-score should be adapted depending on the target population to obtain optimal specificity.

Effective tranexamic acid concentration for 95% inhibition of tissue-type plasminogen activator induced hyperfibrinolysis in children with congenital heart disease: A prospective, controlled, in-vitro study.

BACKGROUND: Although recent studies have assessed tranexamic acid (TXA) pharmacokinetics in different subgroups, the effective concentration of TXA required to completely inhibit fibrinolysis remains to be determined. OBJECTIVE: An in-vitro determination of the effective TXA concentration needed for 95% inhibition (EC95) of tissue-type plasminogen activator (t-PA) activated fibrinolysis, using an experimental model designed for thromboelastometry (ROTEM). DESIGN: A prospective interventional study. SETTING: Department of Anaesthesiology, Queen Fabiola Children's University Hospital and Laboratory of Haematology and Haemostasis, Brugmann University Hospital. Patients were enrolled between June 2013 and October 2014. PATIENTS AND VOLUNTEERS: Twenty children, aged between 1 and 10 years, undergoing elective cardiac catheterisation were included (10 with cyanotic and 10 with noncyanotic diseases). Exclusion criteria were child requiring a procedure in a moribund state. Ten adult volunteers were also included as controls. INTERVENTION: Citrated whole blood samples were obtained from children and volunteers. MAIN OUTCOMES MEASURES: The extrinsic coagulation pathway was activated by tissue factor using the EXTEM test on ROTEM. The degree of lysis measured 30 min (LI30) after the clotting time (CT), and clot amplitudes measured at different times were recorded at baseline, after addition of 1535 units t-PA ml(-1), and following the addition of increasing TXA concentrations in t-PA activated samples. RESULTS: The concentration-effect analysis performed with lysis index after 30 min (LI30) allowed the determination of TXA efficacy concentration 50% (EC50), and calculation of the EC95, which was significantly lower in cardiac surgery children than in adults [8.6 µg ml(-1); 95% confidence interval (95% CI) 6.9 to 14.9 versus 11.3 µg ml(-1); 95% CI 10.6 to 12.9, P < 0.001]. CONCLUSION: In this in-vitro study, we observed that the EC95 TXA concentration that completely inhibited t-PA induced hyperfibrinolysis in children with congenital heart was significantly lower than the concentration required in healthy adult volunteers. Further studies are needed to confirm that this plasma concentration can effectively inhibit fibrinolysis activation in children undergoing cardiac surgery.

Experimental model of hyperfibrinolysis designed for rotational thromboelastometry in children with congenital heart disease.

We assessed an in-vitro model of hyperfibrinolysis using rotational thromboelastometry (ROTEM) by the addition of increasing concentrations of tissue-type plasminogen activator (t-PA) on whole blood obtained from children undergoing cardiac surgery. We assessed the relevance of this model by repeating the tests in the same population after tranexamic acid (TXA) infusion. In addition, we determined the sensitivity and specificity of ROTEM parameters to detect the different degrees of fibrinolysis. Blood samples obtained from 20 children were analyzed at two predefined timepoints: after induction of anesthesia, before TXA (baseline), and at the end of surgery during TXA infusion (end surgery). At baseline, an extrinsic activation with tissue factor (EXTEM) test was performed without and with increasing concentration of t-PA (102, 255, 512, 1024, 1535, and 2539 units t-PA/ml). At the end of surgery, a second EXTEM test was performed without and with two different t-PA concentrations (1535 and 2539 units t-PA/ml). At baseline, increasing t-PA concentrations in the EXTEM test induced a gradual increase of hyperfibrinolysis characterized by a reduction in clot firmness and stability parameters. In the presence of TXA, t-PA-induced hyperfibrinolysis was completely abolished. Lysis-onset time (LOT) and degree of fibrinolysis measured at 30 min (LI30) best assessed the degree of fibrinolysis. This in-vitro model of t-PA-induced hyperfibrinolysis using the EXTEM test of ROTEM may represent a promising tool to assess hyperfibrinolysis in the pediatric population. In addition, we observed that LOT and LI30 should be considered as the best parameters to detect different degrees of fibrinolysis.

Influence of dabigatran and rivaroxaban on routine coagulation assays. A nationwide Belgian survey.

The Belgian national External Quality Assessment Scheme performed a nationwide survey using lyophilised plasma samples spiked with dabigatran or rivaroxaban to demonstrate to the Belgian clinical laboratories how these drugs affect their routine coagulation assays prothrombin time (PT), activated partial thromboplastin time (aPTT), fibrinogen and antithrombin. Virtually all Belgian laboratories performing routine coagulation testing (189/192) participated in the survey. Both, dabigatran and rivaroxaban significantly prolonged the PT and aPTT in a concentration- and reagent-dependent manner. PT reagents were more influenced by rivaroxaban than by dabigatran and aPTT reagents more influenced by dabigatran than by rivaroxaban. Among PT reagents, Neoplastin R® was the most sensitive to rivaroxaban and Innovin® and Thromborel S® the least sensitive. Converting PT results to INR only increased the variability between reagents. Among aPTT reagents, Actin FSL® was the least sensitive to dabigatran while the other aPTT reagents showed slightly higher sensitivities. The presence of dabigatran led to falsely reduced fibrinogen concentrations when measured with a low thrombin concentration reagent. The presence of dabigatran caused an overestimation of the antithrombin level when measured with a thrombin-based activity assay and the presence of rivaroxaban an overestimation of the antithrombin level when measured with a FXa-based assay. Instrument-related differences were found for all tested parameters. In conclusion, this paper provides detailed information on the effect of dabigatran and rivaroxaban on routine coagulation assays as performed with a large number of reagent/instrument combinations.

Transient leukemia in a newborn without Down syndrome: case report and review of the literature.

UNLABELLED: Transient neonatal leukemia occurs almost exclusively in Down syndrome babies. We report here the unusual case of a newborn without Down syndrome who presented neonatal transient leukemia and who achieved spontaneously complete remission. Trisomy 21 and GATA1 mutation were both present in leukemic cells. While close follow-up is advised since true leukemia may develop later, the patient is still in remission for 2.5 years. We performed a literature review of 15 other similar cases. CONCLUSION: Our case of transient leukemia without Down syndrome and the literature review highlight the important role of trisomy 21 and GATA1 mutation in the development of transient neonatal leukemia.

Evaluation of the procoagulant activity in the plasma of cancer patients using a thrombin generation assay.

INTRODUCTION: Circulating microparticles are reported to play a role in cancer hypercoagulability. The procoagulant properties of microparticles derive from the amount of tissue factor and/or phosphatidylserine that they can expose. The aim of our study is to assess the procoagulant activity, including microparticles' activity, in the plasma of newly diagnosed cancer patients with a simple assay, easy to implement in the laboratory. MATERIAL AND METHODS: Newly diagnosed cancer patients (n=31) before the start of anticoagulant or chemotherapy were compared to matched controls. We used a thrombin generation assay in four conditions: 1: addition of 1 pM tissue factor and 4 µM procoagulant phospholipids, 2: without any trigger, 3 and 4: addition of tissue factor or procoagulant phospholipids alone respectively. RESULTS: When we added only phospholipids, so that thrombin generation is dependent upon endogenous tissue factor, the lag time was significantly shorter in cancer patients. When we added only tissue factor, i.e. made the results dependent upon phospholipids, the endogenous thrombin potential, the peak, and the velocity index were significantly higher and the time-to-peak was significantly shorter. This suggests that the plasma of cancer patients contained a higher activity of endogenous phospholipids and/or tissue factor which may be borne by microparticles. CONCLUSION: This new simple methodology can demonstrate a procoagulant activity in the plasma of newly diagnosed cancer patients which can be explained by higher procoagulant phospholipids and tissue factor activity and thus, brings potentially useful information that current coagulation tests cannot provide.

Plasma osteoprotegerin is an independent risk factor for mortality and an early biomarker of coronary vascular calcification in chronic kidney disease.

BACKGROUND: Cardiovascular disease is the major cause of morbidity and mortality in chronic kidney disease (CKD) and early biomarkers are required which can predict disease and death in such patients. The aim of our study was to investigate if osteoprotegerin (OPG) could be a predictor of coronary artery calcification (CAC) and mortality in CKD. METHODS: A total of 77 outpatients (32 with pre-dialysis CKD and 45 undergoing hemodialysis) were followed-up during 2 years. Measurements of CAC were performed using Siemens Multidetector CT software and calcium scores were measured according to the Agatston method. RESULTS: OPG was an independent predictor of the Agatston score for CAC and correlated with the degree of CAC in pre-dialysis patients. A two-sample t-test characterized survivors as having a better glomerular filtration rate, lower Agatston scores, and lower serum levels of OPG. Kaplan-Meier survival curves separated survivors from non-survivors at plasma OPG cut-off levels of <3.1 ng/mL. A multivariable logistic regression analysis showed that OPG was an independent predictor of mortality from all causes in CKD patients. CONCLUSIONS: OPG predicted mortality in CKD patients and could be a valuable biomarker in early detection of CAC in these patients.

Management of refractory essential thrombocythemia with anagrelide in a patient undergoing hemodialysis.

BACKGROUND: Management of essential thrombocythemia (ET) in high-risk patients is difficult because high platelet numbers can lead to vascular occlusive events and bleeding. Therapeutic interventions in ET are limited to hydroxyurea and anagrelide; however, in Europe, anagrelide is contraindicated in patients with chronic renal disease. OBJECTIVE: The aim of this case report was to describe the use of anagrelide in a patient with ET and renal impairment. CASE SUMMARY: A 73-year-old white female patient with severe renal impairment who was diagnosed with ET was receiving treatment with hydroxyurea 1 g/d since 2001. At this time she was also receiving aspirin 80 mg/d; calcium carbonate 1 g/d; pravastatin 40 mg/d; folic acid 5 mg/d; furosemide 40 mg/d; cetirizine 10 mg/d; erythropoietin 10,000 U once monthly; a vitamin B complex, 1 tablet a day; and iron tablets 105 mg/d. In February 2007, because her white blood cell count fell to 2.1 x 10(9)/L, myelodepression was suspected and hydroxyurea was stopped. This led to enhanced platelet levels and the introduction of anagrelide at an initial dose of 0.5 mg/d that was steadily increased to 2.5 mg/d. All other treatments were continued with some dosage adjustments. Sodium bicarbonate 1 g/d and vitamin D were added to her regimen. After 18 months of anagrelide treatment, a sudden but moderate fall of platelets to 142 x 10(3)/microL with severe anemia (hemoglobin, 6.5 g/dL) was observed. The patient had anemia since 2004, but the condition worsened due to bleeding related to an ulcer at the cecal valve. The patient refused blood and platelet transfusions and surgical intervention for religious reasons. Because of hemodynamic instability, she was admitted to the intensive care unit in December 2008 and died 24 hours after admission. CONCLUSION: We report a case of ET and chronic renal failure treated with anagrelide and low-dose aspirin in a patient who did not receive transfusion and surgical intervention due to religious reasons, and had a fatal outcome.

OPG, RANK-L, bone metabolism, and BMD in patients on peritoneal dialysis and hemodialysis.

OBJECTIVES: Bone turnover is regulated locally by osteoprotegerin (OPG) and receptor activator of NFkappaB ligand (RANK-L); it is not known how the circulating concentrations of these cytokines reflect renal osteodystrophy. METHODS: We measured serum OPG, RANK-L, parathyroid hormone (iPTH), collagen C-terminal cross-linked telopeptide (betaCrossLaps), and bone densitometry (BMD) in 79 patients with end-stage renal disease (ESRF) undergoing dialysis. A hand X-ray of these patients was also analyzed. Controls were 65 healthy subjects. RESULTS: ESRF patients had high OPG and RANK-L levels; RANK-L was higher in hemodialysis than in peritoneal dialysis. OPG and RANK-L did not depend on iPTH. The bone markers were significantly increased and correlated with serum iPTH, but not with OPG or RANK-L; neither OPG nor RANK-L correlated significantly with BMD. OPG was significantly higher in patients with acro-osteolysis. CONCLUSIONS: OPG and RANK-L serum concentrations do not strongly reflect bone status in ESRF. However, OPG was significantly higher in patients with acro-osteolysis.

Bone cytokines and renal osteodystrophy in peritoneal dialysis patients.

Bone turnover is regulated by local concentrations of cytokines such as osteoprotegerin (OPG) and receptor activator of nuclear factor kappaB ligand (RANKL). It is not known whether these cytokines can predict renal osteodystrophy in peritoneal dialysis (PD) patients. We measured serum levels of OPG, RANKL, intact parathyroid hormone (iPTH), calcium, phosphorus, and biologic parameters of bone turnover [carboxy-terminal propeptide of type I procollagen (PICP) and beta-crosslaps (betaCL)] in 21 PD patients and 42 healthy subjects matched for age and sex, who served as controls. Bone mineral density (BMD) was also evaluated (Z-scores) in the PD patients. Circulating levels of OPG were significantly higher in PD patients than in healthy subjects (p < 0.001). Mean levels of RANKL did not differ from normal. However, RANKL levels were increased in the group of patients with iPTH levels above 322 pg/mL. Biologic parameters of bone turnover (PICP and betaCL) were significantly increased in PD patients (p < 0.001). We found a positive correlation between serum levels of betaCL and iPTH. At several skeletal sites, betaCL also correlated with the BMD Z-score. No correlations were observed between OPG, RANKL, PICP, betaCL, CaxP, or time on dialysis. Circulating levels of OPG and RANKL do not reflect bone status in PD patients. The value of betaCL is a good marker of bone resorption that correlates with iPTH and BMD.

Late onset of clinical symptoms and recurrent ecchymotic skin lesions in a 12-year-old girl with a severe double heterozygous protein C deficiency.

The authors describe a 12-year-old girl with late-onset clinical symptoms due to severe protein C deficiency. Protein C gene analysis showed double heterozygosity for two distinct mutations, associated with type I protein C deficiency. Her parents and only brother were also evaluated. Coumarin-induced skin necrosis was a recurrent feature during oral anticoagulation therapy, forcing her physicians to treat her with nadroparin (Fraxiparin) for only a few months.