Studies employing in vitro assays for a murine megakaryocytic progenitor cell (CFU-M) have increased the understanding of the regulation of platelet production. These studies have suggested that the megakaryocytic lineage is structured on at least two levels, with early and late megakaryocytopoiesis being independently regulated. We propose to study further the regulation of late murine megakaryocytopoiesis in vitro. A liquid culture system which rapidly quantitates murine megakaryocytopoisis on the basis of synthesis of acetylcholinesterase will be used as the primary means of identifying materials which influence the latter aspects of megakaryocytopoiesis. This assay will be used to screen for possible sources of megakaryocytic maturation activities including thrombocytopenic plasmas, and the supernatants of various cell lines. As an additional assay for the action of late regulatory factor(s), ploidy measurements will be performed on megakarycytes derived from these liquid cultures. Preliminary purification studies will be performed on materials demonstrated to have late regulatory activity. These studies will be performed in order to obtain a useful biological reagent for the further study of megakaryocytic maturation, and to determine the feasibility of complete purification. Finally, cell affinity chromatography using an acetylcholinesterase-binding ligand will be performed in an attempt to purify populations of megakaryocytes.
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