BackgroundSickle cell disease is characterized by a hypercoagulable state as a result of multiple factors, including chronic hemolysis and circulating cell-derived microparticles. There is still no consensus on the cellular origin of such microparticles and the exact mechanism by which they may enhance coagulation activation in sickle cell disease. Design and MethodsIn the present study, we analyzed the origin of circulating microparticles and their procoagulant phenotype during painful crises and steady state in 25 consecutive patients with sickle cell disease. ResultsThe majority of microparticles originated from platelets (GPIIIa,CD61) and erythrocytes (glycophorin A,CD235), and their numbers did not differ significantly between crisis and steady state. Erythrocyte-derived microparticles strongly correlated with plasma levels of markers of hemolysis, i.e. hemoglobin (r=-0.58, p<0.001) and lactate dehydrogenase (r=0.59, p<0.001), von Willebrand factor as a marker of platelet/endothelial activation (r=0.44, p<0.001), and D-dimer and prothrombin fragment F1+2 (r=0.52, p<0.001 and r=0.59, p<0.001, respectively) as markers of fibrinolysis and coagulation activation. Thrombin generation depended on the total number of microparticles (r=0.63, p<0.001). Anti-human factor XI inhibited thrombin generation by about 50% (p<0.001), whereas anti-human factor VII was ineffective (p>0.05). The extent of factor XI inhibition was associated with erythrocyte-derived microparticles (r=0.50, p=0.023). ConclusionsWe conclude that the procoagulant state in sickle cell disease is partially explained by the factor XI-dependent procoagulant properties of circulating erythrocyte-derived microparticles.Key words: microparticles, sickle cell disease, coagulation activation, hemolysis. Citation

Substantial heterogeneity within mutant TP53 AML and MDS-EB precludes the exact assessment of prognostic impact for individual patients. Here we performed in-depth clinical and molecular analysis of mutant TP53 AML and MDS-EB to dissect the molecular characteristics in detail and determine its impact on survival. We performed next-generation sequencing (NGS) on 2,200 AML/MDS-EB specimens and assessed the TP53 mutant allelic status (mono or bi-allelic), the number of TP53 mutations, mutant TP53 clone size, concurrent mutations, cytogenetics and mutant TP53 molecular minimal residual disease, and studied the associations of these characteristics with overall survival (OS). TP53 mutations were detected in 230 (10.5%) AML/MDS-EB patients with a median variant allele frequency (VAF) of 47%. Bi-allelic mutant TP53 status was observed in 174 (76%) patients. Multiple TP53 mutations were found in 49 (21%) patients. Concurrent mutations were detected in 113 (49%) patients. No significant difference in any of the aforementioned molecular characteristics of mutant TP53 were detected between AML and MDS-EB. Mutant TP53 patients have a very poor outcome (2-year OS, 12.8%), however no survival difference among AML and MDS-EB was observed. Importantly, none of the molecular characteristics were significantly associated with survival. In the majority of patients, TP53 mutations remained detectable in complete remission (CR) by deep sequencing (73%). Detection of residual mutant TP53 was not associated with survival. In conclusion, mutant TP53 AML and MDS-EB do not differ with respect to molecular characteristics and survival. Therefore, mutant TP53 AML/MDS-EB should be considered a distinct molecular disease entity.

1. Disseminated intravascular coagulation frequently accompanies Gram-negative sepsis and may contribute to widespread deposition of microthrombi. Besides the endotoxin-induced activation of coagulation, an important role for the fibrinolytic system has been postulated. The precise mechanisms underlying these fibrinolytic changes during endotoxaemia are not known but have been suggested to be mediated directly by cytokines or secondary to thrombin generation. 2. In the present study we have delineated in detail the fibrinolytic response to a bolus injection of endotoxin in non-human primates and analysed the contribution of cytokines and thrombin generation to the endotoxin-induced release of tissue-type plasminogen activator and plasminogen activator inhibitor 1. Chimpanzees received a bolus injection of endotoxin alone or in combination with blocking monoclonal antibodies directed against tumour necrosis factor or interleukin 6 or in combination with pentoxifylline. Furthermore, to assess the effect of coagulation activation on the activation of fibrinolysis, another group of chimpanzees received endotoxin in combination with either anti-tissue factor antibodies or recombinant hirudin. 3. Infusion of endotoxin induced a rapid increase in plasminogen activator activity and tissue-type plasminogen activator antigen levels and subsequent plasmin generation, reaching peak levels 2h after endotoxin administration. Plasminogen activator inhibitor 1 levels remained constant for the first 2 h, after which time a steep increase was observed. Plasminogen activator activity and plasmin generation decreased simultaneously with the rise in plasminogen activator inhibitor 1 levels. Fibrinolytic activity remained suppressed during the remainder of the study owing to sustained increased levels of plasminogen activator inhibitor 1. The administration of pentoxifylline strongly attenuated the release of tissue-type plasminogen activator and plasminogen activator inhibitor 1, whereas the antitumour necrosis factor antibodies blocked the fibrinolytic response entirely. In contrast, interleukin 6-neutralizing antibodies did not affect the fibrinolytic response. Although endotoxin-induced generation of thrombin was completely prevented by the administration of tissue factor-neutralizing antibodies or by hirudin, no effect on the fibrinolytic response was observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Background. We investigated the effect of inhibition of plasminogen activator inhibitor-1

The Netherlands Trial Register #NTR166 (www.trialregister.nl) and the International Standard Randomised Controlled Trial, #ISRCTN95796863 (http://isrctn.org).