The differentiation of myelomonocytic cells from pluripotent stem cells to mature, functioning monocytes/macrophages and granulocytes is an orderly, multi-step process beginning from rare pluripotent stem cells present in the bone marrow. This process is accompanied by histochemical changes as well as the appearance and/or disappearance of proteins on the cell surface. The long term goal of this project is to understand the mechanisms that control the differentiation of myeloid cells, and to elucidate the functions of myeloid differentiation antigens. This project will initially focus on CD14, a membrane protein whose expression is induced upon the differentiation of stem cells to mature monocytes and macrophages. The role of soluble forms of CD14 on lymphoid and myeloid cell differentiation and/or function will be examined in hematopoietic progenitor assays and in in vitro functional assays of monocytes and B cells. Furthermore, the murine gene will be isolated and used to study the function of CD14 in mice. In addition, the DNA sequences that are responsible for regulating the tissue-specific expression of CD14 will be identified using transient transfection systems and transgenic mice. Finally, the function of CD14 will be analyzed using several molecular approaches. The effects of CD14 expression on the growth, morphology and phenotype of immature myeloid cells, which normally do not express CD14, will be determined by transfecting them with expressible CD14 constructs. In addition, the effects of abnormal expression of CD14 in transgenic mice will be determined. Transgenic mice which overexpress CD14 will be produced, and changes in cellularity and migration of cells in a variety of tissues including hematopoietic tissue will be analyzed. In addition, the immunoglobulin micro enhancer will be ligated to the CD14 gene and transgenic mice which have lymphoid cells expressing CD14 will be produced. The effects of this abnormal expression on hematopoiesis and the cellularity of various tissues will be analyzed.
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