Pathological events in platelets of Wiskott‐Aldrich syndrome patients

Abstract

Summary. The Wiskott-Aldrich syndrome (WAS) is a severe X-linked platelet/immunological disorder arising from mutations of the gene WASP. At the clinical level, the major platelet abnormalities are small size and low number, both partially correctable by splenectomy. To identify underlying pathological events, we examined WAS platelets at various stages of their lifetime. In spleen sections from WAS patients, uorescence microscopy showed dramatic co-localization of markers of platelets (CD41) and macrophages (CD68) compared to non-thrombocytopenic controls, suggesting that WAS splenic macrophages are involved in platelet removal. Study of isolated WAS blood platelets by¯ow cytometry showed substantial enhancement of surface exposure of phosphatidylserine (PS), a signal for engulfment by macrophages. Isolated resting WAS platelets were also aberrantly susceptible to microparticle release, and plasma samples of WAS patients contained > 5 times normal numbers of platelet-derived microparticles which may explain the small size of circulating platelets. Measurements with the Ca 2+ sensitive dye¯uo-3 revealed signi®cantly increased Ca 2+ levels, 310 6 13 nmol/l for WAS platelets versus 106 6 12 nmol/l for normal platelets, and also prolongation of agonist-induced Ca 2+¯u x. Cumulatively, these studies identify abnormal events occurring in WAS platelets: increased Ca 2+ levels and enhancement of two Ca 2+ dependent processes, PS exposure and microparticle release; these abnormal events may contribute to the in vivo decrease of platelet number and reduction of platelet size in this disease.

Rapid platelet turnover in WASP(−) mice correlates with increased ex vivo phagocytosis of opsonized WASP(−) platelets

“…There is evidence in favor of reduced or abnormal platelet production as well [15][16][17], but not all the relevant studies have supported this [18]. Reported abnormalities in the function of WASP(−) platelets include a slight increase in collagen-induced aggregation and a significant increase in the duration of calcium influx after thrombin stimulation [19,20]. Structural and biochemical abnormalities that have been demonstrated in WASP(−) platelets include reduced numbers of both α and dense granules, increased microparticle release, and increased surface phosphatidyl serine [12][13][14]19].…”

“…Reported abnormalities in the function of WASP(−) platelets include a slight increase in collagen-induced aggregation and a significant increase in the duration of calcium influx after thrombin stimulation [19,20]. Structural and biochemical abnormalities that have been demonstrated in WASP(−) platelets include reduced numbers of both α and dense granules, increased microparticle release, and increased surface phosphatidyl serine [12][13][14]19]. However, no consistent abnormalities in overall actin polymerization, platelet shape change, or Arp2/3 complex activation after platelet stimulation have been observed [20][21][22][23].…”

Wiskott–Aldrich syndrome protein is required for NK cell cytotoxicity and colocalizes with actin to NK cell-activating immunologic synapses

“…Diagnosis of WAS was based on the presence of established clinical findings (4) and mutation in the WASP gene. WASP sequence analysis was performed as described (19)(20)(21). Pre-bone marrow transplant immunophenotypes of all WAS patients diagnosed and treated at Children's Hospital between 1992 and 2002 were reviewed retrospectively.…”

Tributyltin (TBT) induces ultra-rapid caspase activation independent of apoptosome formation in human platelets

“…Similar results have been demonstrated for calcium ionophore in human platelets. Initial observations reported that calcium ionophore induces the degradation of caspase-3, thus claiming a calcium ionophore-mediated activation of caspases (Vanags et al, 1997;Shcherbina and Remold O'Donnell, 1999). A detailed study by Wolf et al (1999) could demonstrate that human platelets contain all elements of the mitochondrial death pathway, that is, Apaf-1, cytochrome c, caspase-9 and -3 and that this pathway could be activated in vitro via cytochrome c plus dATP.…”

“…However, since addition of cathepsin inhibitors only slightly reduced TBT-induced Figure 1 Kinetic analysis of TBT-mediated processing of caspases and catalytic activity in human platelets. Freshly isolated platelets from normal healthy donors were prepared as described by Shcherbina and Remold O'Donnell (1999). (a) A total of 5 Â 10 7 platelets were pretreated with or without 100 mm zVAD-fmk for 45 min and subsequently stimulated with 2 mm TBT for the indicated duration of time.…”