Figure 3.

Runx binding site mutation leads to reduced numbers of functional HSCs. (A-B) Phenotypic HSCs are unaltered in PU.1-URE-mRunx mice compared with WT (4 months). (A) Flow cytometry of bone marrow samples demonstrating the average percentage ± SD of LSK cells (from lineage-negative population) and the average percentage ± SD of SLAM⁺ LSK cells (from LSK population). (B) Total number of LSK and SLAM⁺ LSK cells in bone marrow (average + SD of 1 femur and 1 tibia; n = 5). (C) Experimental scheme of the limiting dilution competitive repopulation unit (CRU) assay. Indicted numbers of isolated SLAM⁺ LSK cells of WT or PU.1-URE-mRunx mice (CD45.2) were transplanted together with 2 × 10⁵ unselected whole bone marrow cells of competitor mice (CD45.1) into lethally irradiated (1300 rads) recipients (CD45.1). (D) Semilogarithmic plot showing the percentage of negative recipients as a function of the number of transplanted SLAM⁺ LSKs. CRUs are indicated as vertical dashed lines. (E) Table showing the frequency of CRUs and the total number of transplantations per cell dose. Reconstitution was evaluated in blood and bone marrow 6 months after transplantation. Mice with CD45.2 chimerism <0.3% were considered as nonresponders. abs., absolute; Exp., experiment.