Reconstitution of T cell immunity following bone marrow transplantation is critical for the protection of the host from both infectious pathogens and tumor recurrence but this process is slow and is limited by parameters which are poorly defined. Further characterization of the mechanisms limiting immune reconstitution is restricted by available experimental systems. To address this issue, we have developed a novel model of in vitro T lymphopoiesis that recapitulates the differentiation of functional T cell from lineage negative bone marrow progenitor cells. In this project we propose to use this model system to examine T lymphopoiesis after allogeneic stem cell transplantation and to develop therapeutic strategies based on ex vivo T cell generation. We propose to adapt the in vitro system to provide a semi-quantitative measure of total and subset specific lymphopoietic output and repertoire complexity from input stem cells. We will test samples derived from patients undergoing transplantation and correlate in vitro findings with clinical parameters of immune reconstitution and T cell generation. These studies will determine whether stem cell lymphopoietic capacity dictates immune reconstitution and may provide a method by which the immunologic potential of a stem cell graft may be defined prior to transplantation. In addition, we will assess the capability of the in vitro system to provide T cell neogenesis and determine if reactivity to defined antigens can be achieved. Determination of whether negative selection permits expansion of a fully tolerized T cell population may enable adoptive transfer for host defense.
Specific Aims : 1. To optimize an in vitro model system that supports the differentiation of human hematopoietic stem cells into functional polyclonal T cells. 2. To establish methods for positive and negative selection of donor T cells in vitro. 3. To develop in vitro correlates of in vivo T cell lymphopoiesis. 4, To develop methods for in vitro expansion of mature polyclonal na ve T cells for adoptive T cell infusion after allogeneic BMT.
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