No abstract

The source and significance of blood-borne tissue factor (TF) are controversial. TF mRNA, protein, and TF-dependent procoagulant activity (PCA) have been detected in human platelets, but direct evidence of TF synthesis is missing. Nonstimulated monocyte-free plate-lets from most patients expressed TF mRNA, which was enhanced or induced in all of them after platelet activation. Immunoprecipitation assays revealed TF protein (mainly of a molecular weight [Mr] of approximately 47 kDa, with other bands of approximately 35 and approximately 60 kDa) in nonstimulated platelet membranes, which also increased after activation. This enhancement was con-comitant with TF translocation to the plasma membrane, as demonstrated by immunofluorescence-confocal micros-copy and biotinylation of membrane proteins. Platelet PCA, assessed by factor Xa (FXa) generation, was induced after activation and was inhibited by 48% and 76% with anti-TF and anti-FVIIa, respectively , but not by intrinsic pathway inhibi-tors. Platelets incorporated [ 35 S]-methi-onine into TF proteins with Mr of approximately 47 kDa, approximately 35 kDa, and approximately 60 kDa, more intensely after activation. Puromycin but not actinomycin D or DRB (5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole) inhibited TF neosynthesis. Thus, human platelets not only assemble the clotting reactions on their membrane, but also supply their own TF for thrombin generation in a timely and spatially circum-scribed process. These observations simplify, unify, and provide a more coherent formulation of the current cell-based model of hemostasis. (Blood. 2007;109:5242-5250)

Summary.  Objectives and patients: We compared the template bleeding time (BT) and closure time (CT) in the PFA‐100® as screening tests in 148 consecutive patients with unequivocal mucocutaneous bleeding and positive family history. Exclusion criteria: drug intake, concomitant diseases including minor infections, low platelet count, diseases of secondary hemostasis.Results: Type 1 von Willebrand disease (VWD‐1) was diagnosed in 26 patients, primary platelet secretion defect (PSD) in 33, VWD‐1 + PSD in nine, whereas 80 patients did not comply with the criteria for known hemostatic disorders (UD, unknown diagnosis). BT and CT were prolonged in 35.8% and 29.7% of all the patients, respectively (P = 0.23). Sensitivity increased to 48% if an abnormality of BT and/or CT was considered. Same comparisons for BT and CT in each diagnostic category were, respectively: 42 vs. 61.5% in VWD‐1 (P = 0.18), 42 vs. 24% in platelet secretion defects (P = 0.11), 67 vs. 89% in VWD‐1 + PSD (P = 0.50), and 27.5 vs. 15% in UD (P = 0.06). Conclusion: Both tests were relatively insensitive and not significantly different in detecting incoming patients with mucocutaneous hemorrhages. In patients with VWD‐1, the PFA‐100® performed slightly better, whereas the opposite occurred in those patients with platelet secretion defects. In the UD group, both tests lost sensitivity, but the BT detected 1.8 times more patients than the PFA‐100®. Given the large proportion of undiagnosed bleeders and the overall low sensitivity of these tests, clinical decisions still rely on the medical history and etiological diagnosis of the bleeding disorder.

SummarySeveral parameters of primary hemostasis and markers of activation of coagulation and fibrinolysis were measured in 48 patients with severe (creatinine clearance <20 ml/min) chronic renal failure (CRF) without dialysis and diseases or drugs affecting hemostasis. Bleeding time (BT) was prolonged in 25/48 patients, and was correlated with age of patients, severity of renal failure, hematocrit, impairment in platelet aggregation-secretion and decrease in platelet ATP content. Defects in von Willebrand factor played no role in the prolongation of the BT. Multivariate analysis showed that only platelet dysfunction and severity of renal disease were independent predictors of the BT in uremia. The platelet functional disorder was significantly correlated with a reduction in platelet ATP and ADP.High levels of plasma thrombin-antithrombin complexes (TAT), prothrombin fragment F1+2, fibrinogen and factor VIIc were observed in patients with CRF, as described in prethrombotic states. Plasmin-antiplasmin complexes (PAP), fibrinogen and fibrin degradation products (FgDP, FnDP) were significantly increased, and the activity of plasminogen activator inhibitor (PAI-1) was slightly reduced, denoting an activation of fibrinolysis.A negative correlation was found between platelet levels of ATP and ADP with plasma TAT, F1+2 and PAP. Furthermore, plasma PAI-1 activity was negatively correlated with the BT and was lower in patients with prolonged BT as compared with controls and patients with normal BT. These links between primary hemostasis and activation of coagulation and fibrinolysis suggest that increased intravascular generation of thrombin and/or plasmin is an important mediator of the defects in primary hemostasis, prolongation of the BT and, probably, bleeding in CRF.

The major advances from research on platelet molecular and cell biology, physiology, and pathophysiology over the past decades have not been adequately translated to clinical laboratory diagnosis. Hereditary platelet function disorders (PFDs) are at least as prevalent in the general population as von Willebrand disease (VWD) although PFDs tend not be as well recognized or evaluated. Clinical mucous and skin bleeding in patients with PFDs is prototypic of primary hemostasis disorders, and the bleeding pattern is not distinguishable from that of other primary hemostasis disorders such as VWD. However, different treatment needs, between these discrete disorders, make a precise diagnosis mandatory. Currently, clinicians receive reliable laboratory reports when testing patients with severe PFDs, such as Glanzmann thrombasthenia and Bernard-Soulier syndrome, due to the distinctive laboratory defects that these disorders present, together with the availability of differential diagnostic tests. This is not the case for the majority of PFDs generically classified as "platelet secretion disorders," which are a heterogeneous group of "mild bleeding disorders," for which there are not universally accepted diagnostic criteria. An important reason for robust diagnostic tests is the high proportion (more than 50% in some reports) of patients with unequivocal bleeding who have no precise diagnosis established after a complete laboratory workup. It is paradoxical that the current "gold standard" test for PFD diagnosis, light transmission aggregometry (LTA), has not been standardized after more than four decades of worldwide clinical use. This review describes current diagnostic assays for PFD in a clinical hemostasis laboratory, relating these with current knowledge on platelet function and pathophysiology. Special emphasis will be given to LTA and platelet secretion tests, as well as to the reasons why sensitive tests are needed to explore the lesser known participation of platelets in blood clotting and fibrinolytic processes.