Regulation of T cell development by Mib1 in the thymic microenvironment. (A) The lethally irradiated CD45.1 mice were injected intravenously with BM cells from the 12–15-wk-old MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice. At 12 wk after transplantation, the thymocytes were analyzed by flow cytometry for the expression of CD4 and CD8 gated on the CD45.2+ populations (top), CD19 and B220 gated on the CD4−CD8− DN thymocytes (middle), and CD24 and c-kit gated on CD4−CD8−CD25−CD44+ cells (bottom). Numbers indicate mean ± SD from four independent experiments. (B) The lethally irradiated MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were transplanted with CD45.1 BM cells. At 5–6 wk after reconstitution with CD45.1 BM cells, the thymocytes were stained for CD4 and CD8 (top) and B220 gated on the CD4−CD8− DN thymocytes (bottom). Numbers indicate mean ± SD from three independent experiments. (C) Genomic DNA was prepared from the CD45− cells sorted from thymi. Quantitative real-time PCR was performed to analyze the Mib1 deletion using deleted allele-specific primers. Expression of nondeleted allele served as a control for relative quantification. Data are mean ± SD from triplicate experiments. *, P < 0.01.
