The proposed project is directed towards elucidating those factors that regulate early events in human megakaryocytic differentiation. The following studies will be performed: (1) Examination of the effect of platelet microparticles present in human serum and plasma on in vitro megakaryocytopoiesis. (2) Examination of the effect of aplastic anemia serum on the release of megakaryocyte colony stimulating activity by peripheral blood cells. (3) Identification of megakaryocyte colony stimulating activity in media conditioned by various human tissues and human continuous cell lines. (4) Isolation and characterization of megakaryocyte colony stimulating activity. (5) Production of polyclonal and monoclonal antibodies to megakaryocyte colony stimulating activity. (6) Development of a radioimmunoassay for megakaryocyte colony stimulatilng activity. (7) Determination of the specificity of action of megakaryocyte colony stimulating activity. (8) Use of polyclonal and monoclonal antibodies to localize the site of production of megakaryocyte colony stimulating activity immunocytochemical techniques. The completion of this project will result in important information concerning both hematopoietic stem cell differentiation in general and in specific megakaryocytic differentiation. These studies will determine if the stimulating activity observed in aplastic anemia serum is the result of the presence of a regulatory molecule or merely the result of the absence of an inhibitor. Purification of megakaryocyte colony stimulating activity will allow more careful studies of its biological effects and hopefully will allow one to evaluate the use of this hematopoietic regulatory macromolecule as a therapeutic tool in thrombocytopenic cancer patients. The development of a radioimmunoassay for megakaryocyte colony stimulating activity will allow us to study perturbations of this regulatory substance in disease states. Localization of the site of production will hopefully result in the isolation of its cell of origin and investigation of the regulation of the production of this factor at the cellular level. This information will be of use to both biologists and clinicans who are interested in cancer biology and treatment.
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