Alterations of the platelet proteome in type I Glanzmann thrombasthenia caused by different homozygous delG frameshift mutations in ITGA2B

Stefan Loroch; Katharina Trabold; Stepan Gambaryan; Cora Reiß; Kathrin Schwierczek; Ingrid Fleming; Albert Sickmann; Wolfgang Behnisch; Barbara Zieger; René P. Zahedi; Ulrich Walter; Kerstin Jurk

doi:10.1160/TH16-07-0515

Thrombosis and Haemostasis, Inhaltsverzeichnis

Thromb Haemost 2017; 117(03): 556-569
DOI: 10.1160/TH16-07-0515

Cellular Haemostasis and Platelets

Schattauer GmbH

Alterations of the platelet proteome in type I Glanzmann thrombasthenia caused by different homozygous delG frameshift mutations in ITGA2B

Stefan Loroch

¹Leibniz-Institut für Analytische Wissenschaften – ISAS – e. V., Dortmund, Germany

,

Katharina Trabold

²Center for Thrombosis and Hemostasis (CTH), University Medical Center Mainz, Mainz, Germany

,

Stepan Gambaryan

²Center for Thrombosis and Hemostasis (CTH), University Medical Center Mainz, Mainz, Germany

³Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, S. Petersburg, Russia

,

Cora Reiß

²Center for Thrombosis and Hemostasis (CTH), University Medical Center Mainz, Mainz, Germany

,

Kathrin Schwierczek

²Center for Thrombosis and Hemostasis (CTH), University Medical Center Mainz, Mainz, Germany

,

Ingrid Fleming

⁴Institute for Vascular Signalling, Centre for Molecular Medicine, Goethe-University, Frankfurt, Germany

⁵German Centre for Cardiovascular Research (DZHK), RheinMain, Germany

,

Albert Sickmann

¹Leibniz-Institut für Analytische Wissenschaften – ISAS – e. V., Dortmund, Germany

⁶Medizinisches Proteom Center, Medizin-ische Fakultät, Ruhr Universität Bochum, Bochum, Germany

⁷Department of Chemistry, College of Physical Sciences, University of Aberdeen, Aberdeen, UK

,

Wolfgang Behnisch

⁸Department of Pediatric Oncology, Hematology, Immunology and Pulmonology, University Medical Center, Heidelberg, Germany

,

Barbara Zieger

⁹Department of Pediatrics and Adolescent Medicine, Division of Pediatric Hematology and Oncology, Medical Center – Faculty of Medicine, University of Freiburg, Germany

,

René P. Zahedi

¹Leibniz-Institut für Analytische Wissenschaften – ISAS – e. V., Dortmund, Germany

,

Ulrich Walter

²Center for Thrombosis and Hemostasis (CTH), University Medical Center Mainz, Mainz, Germany

⁵German Centre for Cardiovascular Research (DZHK), RheinMain, Germany

,

Kerstin Jurk

²Center for Thrombosis and Hemostasis (CTH), University Medical Center Mainz, Mainz, Germany

⁵German Centre for Cardiovascular Research (DZHK), RheinMain, Germany

› Institutsangaben

Abstract

Summary

Glanzmann thrombasthenia (GT) is one of the best characterised inherited platelet function disorders but global platelet proteome has not been determined in these patients. We investigated the proteome and function of platelets from two patients with type I GT, caused by different homozygous ITGA2b mutations, from family members and unrelated controls. The global proteome of highly purified washed platelets was quantified by liquid chromatography-mass spectrometry (LC-MS) and targeted MS-methods. Platelet function was analysed by flow cytometry, light transmission aggregometry and flow-based as-says. Platelets from GT patients showed less than 5 % relative levels of the integrin subunit αIIb and 5–9 % fibrinogen compared to controls. These patients demonstrated loss of αIIbβ3-dependent platelet function, but normal platelet granule secretion induced by physiological agonists. Platelets from heterozygous family members of a patient expressed 50–60 % of control αIIb levels which were sufficient for normal αIIbβ3-dependent platelet function. Studying type I GT as model disease we established quantitative LC-MS to detect and clearly distinguish normal platelets, platelets from GT heterozygotes and platelets from GT patients. Diminished levels of factor XIIIB chain, plasminogen and carboxypeptidase 2B were identified in thrombasthenic platelets. Additionally, GT platelets showed up to 2.5-fold increased levels of FcγRIIA and laminin-α4 chain. Elevated levels of platelet FcγRIIA was associated with increased CD63-surface expression after FcγRIIA-crosslinking in one GT-patient which might present a compensatory mechanism of platelet activation in GT. We demonstrate that quantitative LC-MS based proteomics is suitable to validate known but also to identify previously unknown protein level changes of dysfunctional platelets.

Supplementary Material to this article is available online at www.thrombosis-online.com.

Keywords

Glanzmann thrombasthenia - Haemorrhage - αIIbβ3 integrin - platelets - proteomics

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Referenzen

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