Abnormal proplatelet formation and emperipolesis in cultured human megakaryocytes from gray platelet syndrome patients.

Christian A. Di Buduo,Maria Adele Alberelli,Paola R. Lev,Erica De Candia,Marco Cattaneo,Gianmarco Podda,Paula G. Heller,Raffaele Landolfi,Ana C. Glembotsky,Alessandra Balduini

doi:10.1038/srep23213

Abstract

The Gray Platelet Syndrome (GPS) is a rare inherited bleeding disorder characterized by deficiency of platelet α-granules, macrothrombocytopenia and marrow fibrosis. The autosomal recessive form of GPS is linked to loss of function mutations in NBEAL2, which is predicted to regulate granule trafficking in megakaryocytes, the platelet progenitors. We report the first analysis of cultured megakaryocytes from GPS patients with NBEAL2 mutations. Megakaryocytes cultured from peripheral blood or bone marrow hematopoietic progenitor cells from four patients were used to investigate megakaryopoiesis, megakaryocyte morphology and platelet formation. In vitro differentiation of megakaryocytes was normal, whereas we observed deficiency of megakaryocyte α-granule proteins and emperipolesis. Importantly, we first demonstrated that platelet formation by GPS megakaryocytes was severely affected, a defect which might be the major cause of thrombocytopenia in patients. These results demonstrate that cultured megakaryocytes from GPS patients provide a valuable model to understand the pathogenesis of GPS in humans.

Highlights

Maternal and Paternal platelet α -granules and spleen enlargement
We have recently demonstrated that constitutively released adenosine diphosphate (ADP) by human mature megakaryocytes promotes the activation of Store-Operated Calcium Entry (SOCE) which in turn is responsible for the regulation of platelet formation and interaction with the components of the extracellular matrix[19,25]
Similar results were obtained in different patients, using different sources of hematopoietic progenitor cells to differentiate along the megakaryocytic lineage in culture

Summary

Introduction

Maternal and Paternal platelet α -granules and spleen enlargement. Myelofibrosis was demonstrated in older animals[12]. Nbeal2−/− mice were used to extensively study the role of platelet α -granules constituents in hemostasis, thrombosis, thrombo-inflammatory disease states, megakaryocyte survival and development, platelet production, tissue reconstitution after injury, development of myelofibrosis and cancer metastasis propagation. Some differences among the three Nbeal2−/− mouse strains were reported with regard to megakaryocyte development and differentiation, proplatelet formation and α -granules content. Whether Nbeal[2] loss of function in mice affects megakaryopoiesis and/or proplatelet formation, and how it contributes to thrombocytopenia is unclear. We obtained in vitro differentiated megakaryocytes from peripheral blood or bone marrow hematopoietic progenitor cells of the four GPS patients and five controls and evaluated megakaryocyte maturation and function and proplatelet formation

Methods

Results

Conclusion