The positive selection of T cell development ensures that only those thymocytes expressing TCR capable of binding self MHC are retained, which results in MHC restriction. Studies in both mice and humans have demonstrated the existence of alloreactive T cells, capable of recognizing peptide in the context of foreign MHC molecules. So we address the question of how the specific MHCI molecule mediating thymic positive selection impacts on mature epitope specific CD8+T cell population and their ability to respond to cognate peptide-MHCI complex.
Using the tetramers based magnetic enrichment technique, we identified influenza-specific H2b-restiricted naïve CTL precursors (CTLps) in H2 mismatched mice BALB/c(H2d) and C3H(H2k). In order to investigate the specific role of positive selection in shaping these naïve epitope-specific CTLps, we have generated mixed bone marrow chimeras. (B6x C3H) F1 bone marrow cells were injected into lethally irradiated B6 (H2b) or C3H (H2k) hosts to produce F1>B6 or F1>C3H chimeras. Because negative selection is mediated mainly by radio-sensitive bone marrow derived cells and positive selection by the radio-resistance thymic epithelium, the naïve epitope-specific CTLp populations in these mice differ only in the MHCI molecule on which they have been positively selected. The functional consequences of this differential positive selection are being assessed by challenged with influenza virus. Tetramer staining and intracellular cytokine staining for IFN-g, TNF-a and IL-2 will be analyzed. The frequencies and TCRab repertoire in these chimeras mice will be assessed by tetramer-based magnetic enrichment and single cell multiplexed RT-PCR. These data will provide information on how positive selection shapes naïve epitope-specific CD8+ T cell populations.