FIG. 8.

Cybr^−/− hematopoietic stems cells have impaired repopulating activity. (A) Three million purified bone marrow stem cells were injected into the lateral tail veins of irradiated rag2^−/− recipients, by using all wild-type stem cells (CD45.2⁺), all Cybr-deficient (CD45.2⁺) stem cells, or a 1:1 mixture of wild-type (CD45.1⁺) and Cybr-deficient (CD45.2⁺) stem cells to produce chimeras. After 8 weeks, thymi, spleens, and lymph nodes were harvested and stained for FACS analysis of T-cell populations. In chimeric mice, CD45.2⁺ cells represent those that have matured from Cybr knockout stem cells. Error bars indicate standard errors of the mean. Asterisks indicate P < 0.05. DP, double positive; SP, single positive; cDC, conventional dendritic cells. (B) Trafficking of wild-type or Cybr-deficient T cells in vivo was analyzed by injecting three million purified CD3⁺ CD45.2⁺ T cells intravenously into congenic CD45.2⁺ recipient mice. Error bars indicate standard errors of the mean. Cell migration was measured by flow cytometry on spleens and lymph nodes 20 h later (n = 5 animals per group). (C) In vivo migration of dendritic cells was also assessed. BMDCs were injected into the footpads of mice, and draining lymph nodes were isolated 24 h later. Cells were stained for the appearance of migrating donor-derived dendritic cells within the draining lymph node. WT, wild type; KO, knockout.