CELLULAR MODULATION OF HEMOPOIESIS

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We will evaluate factors involved in the humoral and cellular modulation of hemopoiesis in normal individuals and those with hemopoietic diseases, including acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS). Major emphasis will be placed on evaluating (1) the in vitro interactions between purified recombinant human polypeptide hemopoietic growth: factors (HGFs) and human hemopoietic cells, and (2) the roles of enriched populations of accessory marrow and blood cells in generating HGFs under the influence of interleukin 2 (IL2) and other immune modulators. We are using a library of recombinant HGFs and monoclonal antibodies and have developed a variety of cell separation techniques, ligand binding procedures and serum-free cell proliferation, differentiation and clonogenic assays to perform these studies. Specific objectives regarding (1) hormonal interactions, are to determine, at the level of specific cell surface receptors, the role of the mesodermal stimulatory hormones insulin and insulin-like growth factors (IGFI & II) for modulating hemopoietic cell proliferation and differentiation, either directly or via alteration of receptor binding action for colony stimulating factor (GM-CSF) or erythropoietin. We will evaluate the hypothesis that multiple growth factors, including insulin and IGFs, interact in situ to provide cellular competence and progression factors for hemopoiesis. As hemopoietic progenitor cell (HPC) survival is partly mediated through HGF effects on glucose transport and ATP generation, we will examine the interactions between the metabolically active mitogenic hormones insulin, IGFs and CSFs to probe mechanisms of hemopoietic regulation. Studies using supplemented serum-free medium will be performed with human myeloid and erythroid cell lines and enriched HPCs as target cells to more precisely determine the functional effects and relationships between HGFs. Further, we will assess production of IGFs by these cells to evaluate the possible autocrine or paracrine roles of these hormones for leukemic vs normal cell growth. (2) We will examine the role of IL2 in enhancing provision of HGFs by enriched T and non T- cell subsets, with particular emphasis on the effector function of natural killer (NK) cells. We will assess production of proliferative (GM-CSF, BPA) and differentiative factors (G-CSF, DF) by resting T and NK cells (which lack TAC+ IL2 receptors) vs such activated Tac+) cells to evaluate the role of IL2 receptors in generating HGFs. After standardizing techniques to define HGF production, binding and responsiveness by normal myeloid cells and leukemic cell lines, we will (3) evaluate these parameters for cells from our patients with hemopoietic diseases, particularly AML and MDS, and will classify these disorders according to such basic cellular defects in growth regulation.