Paroxysmal nocturnal haemoglobinuria (PNH) is characterized pathophysiologically by intravascular lysis of blood cells and clinically by thromboembolic events, often atypical in localization. In this study, we examined the plasmatic coagulation system of PNH patients to investigate a potential relation between coagulation alterations and disease intensity (PNH clone size). We found evidence for both an increase in procoagulant and in fibrinolytic activity, resulting in increased fibrin generation and turnover. Whereas a positive association of the procoagulant potential with PNH clone size was notable, fibrinolytic activity showed an inverse association with clone size. As a possible cause, a growing impairment of fibrinolytic activation and/or an increasing displacement of fibrinolytic activity is assumed. These mechanisms are most likely caused by the detachment of the glycosyl-phosphatidyl-inositol-anchored urokinase plasminogen activator receptor from cell surfaces, causing a progressive resistance to fibrinolytic stimuli, together with a probable shift of the fibrinolytic potential from cell surfaces to soluble, circulating complexes, resulting in a cellular fibrinolysis-steal phenomenon. Together, these processes are accused of mediating an increased thrombophilic risk in PNH. As hereditary prothrombogenic defects were found more frequently in patients suffering ischaemic complications, genetic thrombophilia seems to confer an additional thromboembolic risk in PNH, and should therefore be screened for.

Paroxysmal nocturnal haemoglobinuria (PNH) is a rare, acquired stem cell disorder, characterised by an abnormal susceptibility of red blood cells to complement induced lysis, resulting in repeated episodes of intravascular haemolysis and haemoglobinuria, thromboembolic events at atypical locations and, to a much lesser extent, bleeding complications. Platelet function is assumed to be abnormal, however, a defect has not yet been characterised and underlying mechanisms remain elusive. To explore these issues, we investigated platelet function in PNH patients using assays for clot formation under low and high shear force (thrombelastography and PFA100 device), adhesion to glass beads in native whole blood (Hellem method), aggregometry using various agonists (Born method), and flow cytometric assays for baseline and agonist-induced surface expression density of alpha-granule (CD62P) and lysosomal granule proteins (CD63), ligand binding to surface receptors (thrombospondin), and expression density of activation-induced neoepitopes of the fibrinogen receptor complex (PAC-1). Platelet PNH clone size determined by CD55 and CD59 labelling was compared to the clone sizes of granulocytes, monocytes, erythrocytes, and reticulocytes. A profound reduction of platelet reactivity was observed in PNH patients for all "global function" assays (clot formation, adhesion, aggregation). Platelet hyporeactivity was confirmed using flow cytometric assays. Whereas baseline levels of flow cytometrically determined platelet activation markers did not differ significantly between controls and PNH patients, agonist-induced values of all markers were distinctly reduced in the PNH group. Moreover, significantly reduced white blood cell counts (3.1/nl vs. 5.9/nl), haemoglobin values (9.5 vs. 14.3/g per dl), and platelet counts (136 vs. 219/nl) delineate profound tricytopenia in PNH patients. The fraction of particular cell types lacking the surface expression of GPI-anchored glycoproteins is referred to as the respective PNH clone; median PNH clone sizes of cells with short life spans (reticulocytes, platelets, granulocytes) was 50-80% of total cell populations compared to 20% of red blood cells. The results of our laboratory investigations show, that in PNH, reduced platelet counts coincide with reduced platelet reactivity. The foremost clinical complication in PNH, however, is venous thromboembolism, very probably induced by an activated and dysregulated plasmatic coagulation system. From these seemingly contradictory findings we infer, that part of the platelet hyporeactivity is probably due to reactive downregulation of platelet function in response to chronic hyperstimulation. The overall result is thought to be an unsteady balance, associated with thromboembolism in a larger proportion of patients, and with bleeding in a smaller proportion.

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