Platelets from patients with the Quebec platelet disorder contain and secrete abnormal amounts of urokinase-type plasminogen activator

Abstract

The Quebec platelet disorder (QPD) is an autosomal dominant platelet disorder associated with delayed bleeding and ␣-granule protein degradation. The degradation of ␣-granule, but not plasma, fibrinogen in patients with the QPD led to the investigation of their platelets for a protease defect. Unlike normal platelets, QPD platelets contained large amounts of fibrinolytic serine proteases that had properties of plasminogen activators. Western blot analysis, zymography, and immunodepletion experiments indicated this was because QPD platelets contained large amounts of urokinase-type plasminogen activator (u-PA) within a secretory compartment. u-PA antigen was not increased in all QPD plasmas, whereas it was increased more than 100-fold in QPD platelets (P < .00009), which contained increased u-PA messenger RNA. Although QPD platelets contained 2-fold more plasminogen activator inhibitor 1 (PAI-1) (P < .0008) and 100-fold greater u-PA-PAI-1 complexes (P < .0002) than normal platelets, they contained excess u-PA activity, predominantly in the form of two chain (tcu-PA), which required additional PAI-1 for full inhibition. There was associated proteolysis of plasminogen in QPD platelets, to forms that comigrated with plasmin. When similar amounts of tcu-PA were incubated with normal platelet secretory proteins, many ␣-granule proteins were proteolyzed to forms that resembled degraded QPD platelet proteins. These data implicate u-PA in the pathogenesis of ␣-granule protein degradation in the QPD. Although patients with the QPD have normal to increased u-PA levels in their plasma, without evidence of systemic fibrinogenolysis, their increased platelet u-PA could contribute to bleeding by accelerating fibrinolysis within the hemostatic plug. QPD is the only inherited bleeding disorder in humans known to be associated with increased u-PA. IntroductionCongenital platelet disorders are usually associated with defective primary hemostasis. [1][2][3] The Quebec platelet disorder (QPD) is an autosomal dominant platelet disorder that has unusual clinical features: it is associated with moderate to severe delayed bleeding, that typically begins 12 to 24 hours after surgery or trauma, and its hemorrhagic manifestations can be controlled with fibrinolytic inhibitors but not with platelet transfusions. 1,[4][5][6] This disorder was initially designated as factor V Quebec because of the abnormalities found in platelet factor V of these patients. 7 Two families from Quebec have been identified with this condition, which is now known to be associated with other platelet abnormalities that include reduced to low-normal platelet counts, proteolytic degradation of soluble and membrane proteins stored in platelet ␣-granules, an apparent quantitative deficiency of the ␣-granule protein multimerin, and defective aggregation with epinephrine. 1,[4][5][6]8 Although patients with the QPD have elevated levels of fibrinogen degradation products (FDPs) in their serum (because of platelet fibrinogen degradation), their plasma contains normal amo...

Leukocyte- and endothelial-derived microparticles: a circulating source for fibrinolysis

“…Accordingly, plasminogen activators have not been described on red blood cells and their expression by human platelets has not been clearly demonstrated, except for the ectopic production of uPA in the platelet Quebec syndrome. 35 It is, however, possible that platelet microparticles may, in some cases, develop plasminogen activator activity. Indeed, since platelets bind plasminogen, [36][37][38] platelet microparticles may be a source of substrate for enhanced fibrinolysis by single-chain uPA 39 via a fibrinolytic crosstalk mechanism we demonstrated recently.…”

Bleeding risks associated with inheritance of the Quebec platelet disorder

“…Affected or unaffected status of study participants was based on platelet glycoprotein analyses, and for other individuals in the pedigree their presumed affected or unaffected status was assigned on the basis of historical data provided by other family members, evidence that they had or had not transmitted QPD to offspring, and QPD blood test results (if available) from previous studies. [12][13][14][15][16] …”

“…[11][12][13][14][15][16] The cases initially reported had moderate to severe delayed bleeding problems that responded to treatment with fibrinolytic inhibitors. [11][12][13] Although these bleeding problems were initially attributed to a deficiency of functional platelet factor V, and hence the designation factor V Quebec, the disorder is now known to be associated with a complex spectrum of unique platelet and fibrinolytic abnormalities that include increased megakaryocyte expression and storage of urokinase-type plasminogen activator (u-PA), ␣-granule protein degradation associated with intraplatelet generation of plasmin, normal to increased plasma u-PA without increased plasma D-dimers, normal to reduced platelet counts, absent platelet aggregation responses to 6 M epinephrine, and normal to abnormal aggregation responses to collagen and adenosine diphosphate (ADP).…”

“…[11][12][13] Although these bleeding problems were initially attributed to a deficiency of functional platelet factor V, and hence the designation factor V Quebec, the disorder is now known to be associated with a complex spectrum of unique platelet and fibrinolytic abnormalities that include increased megakaryocyte expression and storage of urokinase-type plasminogen activator (u-PA), ␣-granule protein degradation associated with intraplatelet generation of plasmin, normal to increased plasma u-PA without increased plasma D-dimers, normal to reduced platelet counts, absent platelet aggregation responses to 6 M epinephrine, and normal to abnormal aggregation responses to collagen and adenosine diphosphate (ADP). [11][12][13][14][15][16][17] Access to affected and unaffected individuals within families with QPD, the large pedigree sizes, and definitive blood tests for QPD [12][13][14] provided a unique opportunity to investigate bleeding risks for an autosomal dominant trait and to gather data on responses of QPD bleeding to fibrinolytic inhibitor therapy. With use of a standardized medical history questionnaire (completed by affected and unaffected blood relatives) and blood tests to independently assign affected or unaffected status, odds ratios and 95% confidence intervals were calculated for a spectrum of bleeding symptoms.…”

Thrombopoietin levels in Quebec platelet disorder—Implications for the mechanism of thrombocytopenia

“…We postulated that a detailed analysis of TPO levels in Quebec platelet disorder (QPD) might provide insights into why platelet counts are reduced in this inherited bleeding disorder3 which is caused by a duplication mutation of PLAU , the urokinase plasminogen activator (uPA) gene 4, 5. QPD markedly and selectively increases the production of normal PLAU transcripts by the disease chromosome in megakaryocytes but not in leukocytes through unknown mechanisms,6 and this increases megakaryocyte and platelet uPA levels by more than 100‐fold 7, 8. As QPD megakaryocytes and platelets sequester uPA in α‐granules,8 the disorder results in a unique, platelet activation‐dependent, gain‐of‐function defect in fibrinolysis, without systemic fibrinolysis 9.…”