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Cytomatrix 
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Engineering a Thymus for Efficent Production of Human T Cells |
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Deborah J. Marshall, Thomas Meyer, Julie Reitter, James Bagley, James Kinchsular, Mark J. Pykett, David T. Scadden* and Michael Rosenzweig
Cytomatrix, Woburn MA, 01801 *Experimental Hematology, Harvard-MGH, Boston MA 02129 |
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The immune system is a potentially powerful tool for the treatment of many diseases including cancer, AIDS, autoimmunity and complications arising from organ transplants. However, the production of human T cells in vitro for use in clinical applications has been problematic. To this end, we have developed a culture system whereby seeding of a biocompatible inorganic matrix (CellfoamTM) with either murine thymic stroma or human skin supports the in vitro differentiation of human hematopoietic progenitors into mature T cells. Briefly, human skin tissue or murine thymic stroma was cultured in Cellfoam for 2-3 weeks. AC133+CD2-CD3- cells isolated from human bone marrow were then co-cultured on the stroma in the absence of cytokines for an additional two weeks. FACS analysis demonstrated the development of CD4+/CD8+ double positive as well as CD4+ and CD8+ single positive cells, the majority of which were CD45RA+ (naïve) and TCRab+. These T cells arose from de novo synthesis as demonstrated by the detection of T cell receptor excisional circles (TRECs) by PCR. In addition, spectratype analysis demonstrated the presence of a diverse T cell repertoire that can respond to a variety of antigens presented on autologous dendritic cells including CMV and measles antigens. Thus, we have demonstrated the ability to recapitulate the thymic microenvironment in vitro both phenotypically and functionally. This system is currently being used to investigate 1) the production of self-tolerant T cells 2) the generation of antigen-specific Tcells and 3) positive and negative selection within a thymic organoid. |
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