Figure 3
Figure 3. MEm1/m1 mice lack long-term hematopoietic repopulating activities. (A) HPP-CFC assay performed on WT and MEm1/m1 marrow cells. (B) Noncompetitive transplantation of whole BM cells from either WT or from MEm1/m1 mice (CD45.2 isotype) injected into syngeneic CD45.1 recipients. The 2-dimensional scatter plots analyze CD45.1 (recipient) compared with CD45.2 (donor), where the contribution of the donor BM is in the bottom right quadrant. Whereas WT marrow maintains a high percentage of contribution, the MEm1/m1 marrow is not able to persist. (C) Competitive transplantation of WT CD45.1 donor marrow cells together with CD45.2 marrow from either WT (left) or MEm1/m1 (right) donors, transplanted into lethally irradiated CD45.1 syngeneic recipient mice. Flow analysis of peripheral blood at 1 month and 3 months was performed, quantitating CD45.1+ and CD45.2+ cells. The cells of interest are in the bottom right quadrant: WT donors persisted over time, whereas MEm1/m1 cells were not maintained. (D) Summary of competitive transplantation data (n = 9 for each group, error bars indicate SD) on the percentage of donor reconstitution over time. (E) Flow cytometric analysis of BM from recipients at 20 weeks, analyzing for Sca-1 (FL2, y-axis) and c-Kit (FL3, x-axis). Boxed area in top right quadrant represents LSK progenitors, which were present in mice reconstituted with WT BM, but not in recipients reconstituted with homozygous BM. (F) CD45.1 donor engraftment into nonmyeloablated recipients 3 months after injection of 2 × 106 BM cells (n = 10 for each group, error bars indicate SD).

MEm1/m1 mice lack long-term hematopoietic repopulating activities. (A) HPP-CFC assay performed on WT and MEm1/m1 marrow cells. (B) Noncompetitive transplantation of whole BM cells from either WT or from MEm1/m1 mice (CD45.2 isotype) injected into syngeneic CD45.1 recipients. The 2-dimensional scatter plots analyze CD45.1 (recipient) compared with CD45.2 (donor), where the contribution of the donor BM is in the bottom right quadrant. Whereas WT marrow maintains a high percentage of contribution, the MEm1/m1 marrow is not able to persist. (C) Competitive transplantation of WT CD45.1 donor marrow cells together with CD45.2 marrow from either WT (left) or MEm1/m1 (right) donors, transplanted into lethally irradiated CD45.1 syngeneic recipient mice. Flow analysis of peripheral blood at 1 month and 3 months was performed, quantitating CD45.1+ and CD45.2+ cells. The cells of interest are in the bottom right quadrant: WT donors persisted over time, whereas MEm1/m1 cells were not maintained. (D) Summary of competitive transplantation data (n = 9 for each group, error bars indicate SD) on the percentage of donor reconstitution over time. (E) Flow cytometric analysis of BM from recipients at 20 weeks, analyzing for Sca-1 (FL2, y-axis) and c-Kit (FL3, x-axis). Boxed area in top right quadrant represents LSK progenitors, which were present in mice reconstituted with WT BM, but not in recipients reconstituted with homozygous BM. (F) CD45.1 donor engraftment into nonmyeloablated recipients 3 months after injection of 2 × 106 BM cells (n = 10 for each group, error bars indicate SD).

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