The proposed research program makes use of T-cell colonies grown in semi-solid culture media as an in vitro model system to study T-lymphocyte differentiation and the development of the specificity repertoire. Conditioned medium from phytohemagglutinin stimulated leukocytes will induce the growth of murine T-colonies from early T-cell precursors. The colony system permits the detection of an immature T-cell precursor which is capable of giving rise to functional cytotoxic-T-lymphocyte precursors (CLP). As CLP of several different specificities can be generated from a single pre-CLP, there appears to be diversification of the repertoire occurring in vitro in the colonies. The differentiation of pre-CLP requires the interaction with another cell which we have termed the """"""""inducer"""""""" cell. I propose to further characterize the colony-forming cells and to use the system as an in vitro model for testing some of the current ideas concerning the influence of the environment on the generation of the T-cell repertoire. More specifically, a) I will examine the ontogeny of the pre-CLP and determine whether or not the pre-CLP is a thymus-processed cell. b) I will carry out a comparative analysis of the specificity repertoire of the CLP generated in the colonies and the in vivo repertoire of CLP. c) I will examine the role of the inducer cell and test its ability to modulate the specificity repertoire of the CLP in three different experimental approaches: 1) elimination of anti-self reactivity; 2) maintenance of neonatally-induced tolerance; 3) imposition of H-2 restriction. The long-term objective of my research is aimed at understanding the development of the specificity repertoire. My strategy is to use in vitro model systems of T-lymphocyte differentiation to facilitate the dissection of cellular and microenvironmental elements which control and influence the generation of the repertoire.
