The inhibition effect of anti-GPIIIa49–66 antibody on megakaryocyte differentiation

Ruimin Pan; Jianhui Wang; Michael A. Nardi; Zongdong Li

doi:10.1160/TH11-03-0153

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2011; 106(03): 484-490
DOI: 10.1160/TH11-03-0153

Platelets and Blood Cells

Schattauer GmbH

The inhibition effect of anti-GPIIIa49–66 antibody on megakaryocyte differentiation

Ruimin Pan^*

¹Department of Medicine, New York University School of Medicine, New York, New York, USA

,

Jianhui Wang^*

¹Department of Medicine, New York University School of Medicine, New York, New York, USA

,

Michael A. Nardi

²Departments of Pediatrics and Pathology, New York University School of Medicine, New York, New York, USA

,

Zongdong Li

¹Department of Medicine, New York University School of Medicine, New York, New York, USA

› Author Affiliations

Abstract

Summary

We previously reported that patients with early-onset HIV-1 ITP developed a unique anti-platelet integrin GPIIIa antibody against the GPIIIa49–66 epitope. Anti-GPIIIa49–66 antibody-induced platelet fragmentation requires sequential activation of the platelet 12-lipoxygenase (12-LO) and NADPH oxidase to release reactive oxygen species (ROS). 12-LO is upstream of the NADPH oxidase pathway and 12(S)- HETE, the product of 12-LO, induces the same oxidative platelet fragmentation as anti-GPIIIa49–66. Since the megakaryocyte (MK) is the progenitor cell for platelets, we have investigated the effect of anti- GPIIIa49–66 on MK differentiation and, in particular, the potential role of anti-GPIIIa49–66 induced ROS in this process. We first show that polyclonal anti-GPIIIa49–66 antibody isolated from HIV-1 ITP patients inhibits MK proliferation 2.5-fold in in vitro culture of human cord blood CD34⁺ cells driven by thrombopoietin (TPO). We also observe a three-fold decrease in the number of MK colony-forming units in the presence of a human monoclonal anti-GPIIIa49–66 antibody. However, we could not detect ROS release in DCFH-loaded mouse megakaryoblastic cells L8057 treated with anti-GPIIIa49–66 antibody. In addition, 12(S)-HETE does not inhibit the in vitro differentiation of L8057 cells induced by TPO. In fact, we found a dose dependent increase in the percentage of CD41 positive cells (from 17.1% to 48.7%) in in vitro culture of L8057 cells treated with various concentrations of H₂O₂ (from 5 to 20 μM). We therefore conclude that the anti-GPIIIa49–66 antibody inhibits MK differentiation through β3 integrin signalling independent of ROS release.

Keywords

Thrombocytopenia - HIV - autoantibody

Full Text

References

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