Glycosylation of autoantibodies: Insights into the mechanisms of immune thrombocytopenia

Tamam Bakchoul; Kathrin Walek; Annika Krautwurst; Mathias Rummel; Gregor Bein; Sentot Santoso; Ulrich J. Sachs

doi:10.1160/TH13-04-0294

Thrombosis and Haemostasis, Inhaltsverzeichnis

Thromb Haemost 2013; 110(06): 1259-1266
DOI: 10.1160/TH13-04-0294

Platelets and Blood Cells

Schattauer GmbH

Glycosylation of autoantibodies: Insights into the mechanisms of immune thrombocytopenia

Tamam Bakchoul

¹Institute for Clinical Immunology and Transfusion Medicine, Justus Liebig University Giessen, Germany

²Institute for Immunology und Transfusion Medicine, Ernst-Moritz-Arndt University, Greifswald, Germany

,

Kathrin Walek

¹Institute for Clinical Immunology and Transfusion Medicine, Justus Liebig University Giessen, Germany

,

Annika Krautwurst

¹Institute for Clinical Immunology and Transfusion Medicine, Justus Liebig University Giessen, Germany

,

Mathias Rummel

³Department of Hematology-Oncology, University Hospital Giessen and Marburg, Giessen Campus, Germany

,

Gregor Bein

¹Institute for Clinical Immunology and Transfusion Medicine, Justus Liebig University Giessen, Germany

⁴Center for Transfusion Medicine and Hemotherapy, University Hospital Giessen and Marburg, Marburg Campus, Germany

,

Sentot Santoso

¹Institute for Clinical Immunology and Transfusion Medicine, Justus Liebig University Giessen, Germany

,

Ulrich J. Sachs

¹Institute for Clinical Immunology and Transfusion Medicine, Justus Liebig University Giessen, Germany

⁴Center for Transfusion Medicine and Hemotherapy, University Hospital Giessen and Marburg, Marburg Campus, Germany

› Institutsangaben

Abstract

Summary

Immune thrombocytopenia (ITP) is a bleeding disorder caused by IgG autoantibodies (AAbs) directed against platelets (PLTs). IgG effector functions depend on their Fc-constant region which undergoes post-translational glycosylation. We investigated the role of Asn279-linked N-glycan of AAbs in vitro and in vivo. AAbs were purified from ITP patients (n=15) and N-glycans were enzymatically cleaved by endoglycosidase F. The effects of native AAbs and deglycosylated AAbs were compared in vitro on enhancement of phagocytosis of platelets by monocytes and complement fixation and activation applying flow cytometry, laser scanning microscopy, and a complement consumption assay. AAb-induced platelet phagocytosis was inhibited by N-glycan cleavage (median phagocytic activity: 8% vs 0.8%, p=0.004). Seven out of 15 native AAbs bound C1q and activated complement. N-glycan cleavage significantly reduced both effects. In vivo survival of human PLTs was assessed after co-transfusion with native or N-glycan cleaved AAbs in a NOD/SCID mouse model. Injection of AAbs resulted in rapid clearance of human platelets compared to control (platelet clearance after 5h (CL_5h) 75% vs 30%, p<0.001). AAbs that were able to activate complement induced more pronounced platelet clearance in the presence of complement compared to the clearance in the absence of complement (CL_5h 82% vs 62%, p=0.003). AAbs lost their ability to destroy platelets in vivo after deglycosylation (CL_5h 42%, p<0.001). N-glycosylation of human ITP AAbs appears to be required for platelet phagocytosis and complement activation, reducing platelet survival in vivo. Posttranslational modification of AAbs may constitute an important determinant for the clinical manifestation of ITP.

Keywords

Thrombocytopenia - autoantibody - ITP - animal model - complement

Volltext

Referenzen

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