Abstract
Megakaryocytes are rare hematopoietic cells comprising only about 0.02–0.05% of the bone marrow nucleated cell population. Because of the relative infrequency of megakaryocytes in the bone marrow and their fragility in vitro, studies to characterize expression of platelet-specific genes have mostly been carried out in continuous cell lines originating from leukaemic marrow or blood cells that express a range of megakaryocytic phenotypic properties. HEL cells, which are representative of such cell lines, were instrumental in getting the molecular analysis of megakaryocytes and platelets established, and although these cells are still useful for many studies, it has become clear that they have significant limitations. These limitations include the fact that these lines only approximate megakaryocytes. These lines do not contain α-granules, do not demarcate or release platelets, respond appropriately to thrombopoietin (TPO), or express the high levels of such platelet-specific proteins such as the integrin αIIb and platelet factor 4 (PF4). Other proteins such as the platelet-restricted G protein Gzα has not been detected in any of these cell lines. Further, these cell lines often express a mixture of multiple different lineages and can be easily shifted from one lineage to another.
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References
Kuter, D. J., Gminski, D., and Rosenberg, R. D. (1992) Botrocetin agglutination of rat megakaryocytes: a rapid method for megakaryocyte isolation. Exp. Hematol. 20, 1085–1089.
Ravid, K., Doi, T., Beeler, D. L., Kuter, D. J., and Rosenberg, R. D. (1991) Transcriptional regulation of the rat platelet factor 4 gene: Interaction between an enhancer/silencer domain and the GATA site. Mol. Cell. Biol. 11, 6116–6127.
Block, K. L., Ravid, K., Phung, Q. H., and Poncz, M. (1994) Characterization of regulatory elements in the 5′-flanking region of the rat GPIIb gene by studies in a primary rat marrow culture system. Blood 84, 3385–3393.
Kuter, D. J., Greenberg, S. M., and Rosenberg, R. D. (1989) Analysis of megakaryocyte ploidy in rat marrow cultures. Blood 74, 1952–1962.
Miyazaki, H., Tamura, S., Sudo, T., and Suzuki, T. (1990) Production and characterization of monoclonal antibodies against rat platelet GPIIb/IIIa. Thromb. Res. 59, 941–953.
Hirt, B. (1967) Selective extraction of polyoma DNA from infected mouse cell cultures. J. Mol. Biol. 26, 365–369.
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© 1999 Humana Press Inc.
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Shou, Y., Poncz, M. (1999). Molecular Biology Studies with Primary Megakaryocytes. In: Perry, D.J., Pasi, K.J. (eds) Hemostasis and Thrombosis Protocols. Methods in Molecular Medicine™, vol 31. Humana Press. https://doi.org/10.1385/1-59259-248-1:347
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DOI: https://doi.org/10.1385/1-59259-248-1:347
Publisher Name: Humana Press
Print ISBN: 978-0-89603-419-8
Online ISBN: 978-1-59259-248-7
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