The general goal is to define cellular and molecular events involved in thymic microenvironment-thymocyte interactions that lead to normal T cell development or lead to aberrant T cell maturation, such as is seen in congenital and acquired T cell immunodeficiency states, and in T cell malignancies. The studies required to achieve this general goal fall into two main areas--phenotypic and functional characterization of epithelial cells within the human thymic microenvironment, and phenotypic and functional characterization of human T cells and their precursors at defined stages of maturation. To study thymic epithelial maturation keratin subclasses of human endocrine thymic epithelium will be used as markers of thymic micro-environment maturation. Using a newly developed assay of thymic epithelial-T cell binding, we will study and define functionally relevant thymic epithelial surface molecules with regard to thymic epithelial-T cell binding, secretion of thymic hormones and secretion of other immunoregulatory molecules. The phenotype and differentiating capacity of malignant thymic epithelial cells (thymomas) compared to normal and hyperplastic myasthenia gravis will be studied using our recently developed techniques for long term in vitro growth of human thymic epithelium. In addition, EGF-independent transformed thymic epithelial cell lines will be developed by introducing transforming genes (N-ras, c-Myc) into cultured normal thymic epithelial cells. Using a human malignant pluripotent stem cell line (DP.528), we will develop specific monoclonal antibody probes that define surface or cytoplasmic antigens specific for early (pluripotent stem cell, lymphoid stem cell, committed T cell precursor, prothymocyte) stages of T cell maturation. Finally, we will develop in vitro assay systems for cellular, molecular and genetic events that might transpire during early stages of T cell maturation. With these assay systems, the effects of cultured thymic epithelium, other accessory cells, thymic epithelial supernatants, synthetic thymic hormones, purified Interleukin 1 or Interleukin 2 on the differentiating capacity of immature normal and malignant T cells will be studied.
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