Disrupted Signaling through the Fanconi Anemia Pathway Leads to
Dysfunctional Hematopoietic Stem Cell Biology: Underlying Mechanisms and Potential Therapeutic Strategies
Figure 1
Murine models of FA do not have reduced numbers of immunophenotypically defined HSCs but do have lower frequencies of functionally defined HSCs. (a) Representative FACS plots showing the gating scheme which is employed to enumerate long-term HSCs (LT-HSC) in the BM which are defined by the following combination of immunophenotypic markers: Lineage-, c-Kit+, Sca-1+, Flk2−, and CD34−. For illustrative purposes, the compartments enriched for short-term HSC (ST-HSC) and multipotent progenitors (MPP) are also shown. (b) Based on the FACS methodology shown in (a), the frequency of LT-HSCs does not differ significantly in wild-type (WT) mice compared to FA mice. The mean frequency of immunophenotypically defined LT-HSC found in the BM of WT (), (), (), and () mice is shown, ± SEM. NS = by comparison using ANOVA. (c) Schematic representation of the competitive repopulation assay employed to assess the relative frequency of HSCs in WT versus either or mice. BM from FA ( total BM cells) and WT ( total BM cells) mice were coinjected into lethally irradiated (10 Gy total body irradiation) recipient mice at a 3 : 2 ratio, respectively. At four months posttransplant, peripheral blood was harvested from recipient mice and the percentage contribution of FA cells to the peripheral blood was determined by FACS analysis, taking advantage of the differential expression of CD45 subtypes on the surface of FA (CD45.1+ and CD45.2−) and WT (CD45.1+ and CD45.2+) leukocytes. If FA BM contained the same number of functionally defined HSCs as WT BM, then the FA HSCs would be predicted to contribute to 60% of the peripheral blood chimerism at 4 months posttransplant. (d) FA HSCs have a severe engraftment defect compared to WT HSCs. The mean relative frequency that FA or WT cells contributed to peripheral blood leukocyte engraftment at 4 months post-transplantation is shown ± SEM. ** = for comparison of WT versus FA chimerism using ANOVA. for WT versus and for WT versus . , and mice have all been previously described [49].
