Distinct functional effects of NMDA receptors in normal and leukaemic megakaryocytes

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dc.contributor.advisor Kalev-Zylinska, M en
dc.contributor.author Kamal, Tania en
dc.date.accessioned 2017-04-19T21:08:14Z en
dc.date.issued 2016 en
dc.identifier.uri http://hdl.handle.net/2292/32592 en
dc.description.abstract Human megakaryocytes were previously shown to release glutamate and express glutamategated Ca2+-permeable N-methyl-D-aspartate receptors (NMDARs). Their involvement in megakaryocytic maturation has been shown in 2003 but details remain limited. The importance of Ca2+ homeostasis in malignant megakaryocytes has been further highlighted only in 2013 through the discovery of Calreticulin mutations in approximately 30% of patients with megakaryocytic neoplasias. The role of Ca2+ and glutamate pathways in megakaryocytic malignancies remains unknown but their oncogenic potential has already been demonstrated in other cancers. The main objective of this thesis was to characterise the functional role of NMDARs in both normal and leukaemic megakaryocytes. Most of our studies were conducted using three human megakaryocytic cell lines, Meg-01, Set-2 and K-562 that served as models of leukaemic megakaryoblasts. We have characterised NMDAR expression and function as a Ca2+ ion channel in these cells. NMDAR contribution to cell growth and differentiation was examined using well-established NMDAR antagonists (memantine, MK-801, AP5 and riluzole) and agonists (glutamate and NMDA). Cells from patients with megakaryocytic cancers were also used to confirm NMDAR expression. The second part of this thesis used normal mouse megakaryocytes in primary cultures to examine NMDAR contribution to normal megakaryocytic maturation. We found that megakaryocytic leukaemia cells (cell lines and patient-derived) expressed a range of NMDAR subunits, but their combinations differed from normal human megakaryocytes and the brain, suggesting distinct properties of megakaryocytic NMDARs and their deregulation in the context of malignancy. Megakaryocytic NMDARs were functional, contributing Ca2+ entry in Meg-01 cells and amplifying Ca2+ responses to adenosine diphosphate (a known regulator of megakaryocytic maturation). We found that NMDAR agonists increased and NMDAR antagonists reduced growth and proliferation of Meg-01, Set-2 and K-562 cells. Unexpectedly, NMDAR antagonists also facilitated differentiation of leukaemic cells in culture. In contrast to the pro-differentiating effect of the NMDAR inhibition in leukaemia cell lines, NMDAR antagonists inhibited megakaryocytic differentiation of normal mouse megakaryocytes in three primary culture systems: bonemarrow explants, CD41-enriched and lineage-depleted cultures. Finally, NMDAR inhibition was associated with distinct cytoplasmic vacuolation in both leukaemic and normal cells. This was shown not to be due to apoptosis; the nature and significance of this phenomenon will require further elucidation. In conclusion, functional NMDARs support growth of leukaemic megakaryocytes but are not required for leukaemia cells differentiation. In contrast, in normal mouse megakaryocytes, active NMDARs were required to support cell differentiation. These opposing effects of the NMDAR inhibition in normal and leukaemic megakaryocytes suggest that Ca2+ pathways may be subverted in cancerous megakaryocytes away from differentiation towards proliferation. Our results contribute new insights into the importance of glutamate-mediated Ca2+ pathways in megakaryocytic cells, suggesting novel ways to manipulate megakaryocytic differentiation and treat megakaryocytic cancers. en
dc.publisher ResearchSpace@Auckland en
dc.relation.ispartof PhD Thesis - University of Auckland en
dc.relation.isreferencedby UoA99264916410102091 en
dc.rights Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher. en
dc.rights.uri https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm en
dc.title Distinct functional effects of NMDA receptors in normal and leukaemic megakaryocytes en
dc.type Thesis en
thesis.degree.discipline Molecular Medicine and Pathology en
thesis.degree.grantor The University of Auckland en
thesis.degree.level Doctoral en
thesis.degree.name PhD en
dc.rights.holder Copyright: The author en
dc.rights.accessrights http://purl.org/eprint/accessRights/OpenAccess en
pubs.elements-id 622964 en
pubs.record-created-at-source-date 2017-04-20 en

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