Figure 2.
Loss of CD53 does not affect HSC mobilization, but impairs HSC function in response to G-CSF. Mice were treated with 3.75 μg G-CSF every 12 hours for 7 days, with the final dose administered 2 hours before analysis. (A) Shown are absolute number of HSCs in the BM (left) and spleen (right) of vehicle- and G-CSF-treated Cd53−/− and WT mice (n = 13-19 mice per group; age, 6-10 weeks) over 6 independent experiments. (B) Outline of treated chimeric transplants. PBS-treated BM was transplanted at a ratio of 1:1, G-CSF-treated BM was transplanted at a ratio of 3:1, and both PBS- and G-CSF-treated spleens were transplanted at a ratio of 2:1, all competitively transplanted against the untreated whole BM. The donor mice were CD45.2, competitors were CD45.1, and the lethally irradiated recipients were CD45.1/45.2. Chimerism was assessed by peripheral blood flow cytometry analysis every 6 weeks. (C) Engraftment of whole BM from the indicated groups, including total leukocyte chimerism (left) and individual lineages at 24 weeks post-transplant (right). n = 13 to 18 recipients per group over 4 independent transplants, each with 1 donor (4 total donors). (D) Engraftment of whole splenocytes from the indicated groups, including total leukocyte chimerism (left) and individual lineages at 24 weeks post-transplant (right). n = 13 to 18 recipients per group over 4 independent transplants, each with one donor (4 total donors). (E) Engraftment of 100 sorted HSCs from the spleens of G-CSF treated WT or Cd53−/− mice, including total leukocyte chimerism (left) and individual lineages at 24 weeks post-transplant (right). n = 5 to 7 recipients per group over 2 independent transplants, with a total of 3 independent donors. (F) Marrow homing as assessed by transplanting sorted KSL (c-kit+, sca-1+, lineage-) cells from the spleens of G-CSF-treated WT and Cd53−/− mice into lethally irradiated CD45.1 WT mice, and enumerating CD45.2+ cells 20 hours later. n = 7 to 8 recipients in 2 independent transplants. Error bars represent mean ± SEM. ∗P < .05; ∗∗∗P < .001; and ∗∗∗∗P < .0001 by an unpaired Student t test.

Loss of CD53 does not affect HSC mobilization, but impairs HSC function in response to G-CSF. Mice were treated with 3.75 μg G-CSF every 12 hours for 7 days, with the final dose administered 2 hours before analysis. (A) Shown are absolute number of HSCs in the BM (left) and spleen (right) of vehicle- and G-CSF-treated Cd53−/− and WT mice (n = 13-19 mice per group; age, 6-10 weeks) over 6 independent experiments. (B) Outline of treated chimeric transplants. PBS-treated BM was transplanted at a ratio of 1:1, G-CSF-treated BM was transplanted at a ratio of 3:1, and both PBS- and G-CSF-treated spleens were transplanted at a ratio of 2:1, all competitively transplanted against the untreated whole BM. The donor mice were CD45.2, competitors were CD45.1, and the lethally irradiated recipients were CD45.1/45.2. Chimerism was assessed by peripheral blood flow cytometry analysis every 6 weeks. (C) Engraftment of whole BM from the indicated groups, including total leukocyte chimerism (left) and individual lineages at 24 weeks post-transplant (right). n = 13 to 18 recipients per group over 4 independent transplants, each with 1 donor (4 total donors). (D) Engraftment of whole splenocytes from the indicated groups, including total leukocyte chimerism (left) and individual lineages at 24 weeks post-transplant (right). n = 13 to 18 recipients per group over 4 independent transplants, each with one donor (4 total donors). (E) Engraftment of 100 sorted HSCs from the spleens of G-CSF treated WT or Cd53−/− mice, including total leukocyte chimerism (left) and individual lineages at 24 weeks post-transplant (right). n = 5 to 7 recipients per group over 2 independent transplants, with a total of 3 independent donors. (F) Marrow homing as assessed by transplanting sorted KSL (c-kit+, sca-1+, lineage-) cells from the spleens of G-CSF-treated WT and Cd53−/− mice into lethally irradiated CD45.1 WT mice, and enumerating CD45.2+ cells 20 hours later. n = 7 to 8 recipients in 2 independent transplants. Error bars represent mean ± SEM. ∗P < .05; ∗∗∗P < .001; and ∗∗∗∗P < .0001 by an unpaired Student t test.

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