NHLBI CLINICAL INVESTIGATOR AWARD PROGRAM

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The events which occur during differentiation along the megakaryocyte-platelet axis will be studied, with emphasis on isolation and characterization of factors which enhance megakaryocyte proliferation and maturation. To facilitate screening of preparations of megakaryocyte colony stimulating factors (meg-CSAs), a rapid quantitative assay for in vitro megakaryocyte growth will be developed using megakaryocyte-platelet membrane specific radiolabeled monoclonal antibodies in a liquid marrow culture technique. Semi-solid medium marrow culture assays will be used to standardize the liquid assay, and to allow evaluation of individual colonies. Rich sources of meg-CSA(s) will be sought among various cell supernatants, cell extracts and patient samples, and individual factors then will be isolated by conventional biochemical separation techniques. Polyclonal and monolonal antibodies will be raised against isolated factors. These antibodies will be used to facilitate further isolations, and to develop radioimmunoassays to be used in patient studies. Isolated meg-CSA(s) will be characterized as to structure, specific cells of origin, and target cells. Coagulation proteins and mature platelets will be tested for a role in feedback regulation of megakaryopoiesis. Rigorous characterization of one discrete regulatory factor in megakaryopoiesis, and the ability to quantitate it in patients, will expand our understanding of clinical disorders in which specific controls of megakaryopoiesis are either inadequate or ineffective and lead to thrombocytopenia or thrombocytosis despite normal numbers of red blood cells and white blood cells.