Figure 3.
RXRα;RXRβ-deficient HSCs are functionally defective. (A-C) Competitive transplantation assay (1:1 mix) of unfractionated BM cells from WT or VavCre+Rxrabfl/fl mice (CD45.2+) and C57BL/6 mice (CD45.1+) into lethally irradiated C57BL/6 mice (CD45.1+) (data are representative of 3 independent experiments). (A) Experimental design. (B) Fluorescence-activated cell sorting (FACS) analysis of the percentage of total donor chimerism in PB 4, 8, 12, and 16 weeks after transplantation (n = 9-10). (C) FACS analysis of cell-specific chimerism in PB, BM, and spleen 16 weeks after competitive repopulation assay. (D-F) Homing assay of total BM cells from WT or VavCre+Rxrabfl/fl mice (CD45.2+), transplanted into lethally irradiated C57BL/6 mice (CD45.1+). (D) Experimental design. (E) Representative FACS plots showing the gating strategies for analysis of CD45.2+ HSCs. (F) Total numbers of CD45.2+ HSCs found in the BM of recipient mice. (G-H) Transplantation of purified LT-HSCs from WT or VavCre+Rxrabfl/fl mice (CD45.2+) along with LKs from C57BL/6 mice (CD45.1+) into lethally irradiated C57BL/6 mice (CD45.1+) (n = 19-20; data pooled from 2 independent experiments). (G) Experimental design. (H) Kaplan-Meier survival plot. (I-J) Serial CFU plating assay after 48-hours in vitro treatment of purified LT-HSCs from 5-month-old WT or VavCre+Rxrabfl/fl mice with 9c-RA or vehicle. (I) Experimental design. (J) Number of CFUs normalized to control vehicle treatment of each corresponding plating (data pooled from 2 independent experiments). All data are shown as means ± SEM, and dots represent individual animals. Significance was determined by 2-way analysis of variance (ANOVA) followed by the Sidak multiple comparisons test (B), unpaired Student t test (C and F), log-rank (Mantel-Cox) test (H), or ordinary 1-way ANOVA (J), and is represented as follows: ∗P ≤ .05, ∗∗P ≤ .01, ∗∗∗P ≤ .001, and ∗∗∗∗P ≤ .0001.

RXRα;RXRβ-deficient HSCs are functionally defective. (A-C) Competitive transplantation assay (1:1 mix) of unfractionated BM cells from WT or VavCre+Rxrabfl/fl mice (CD45.2+) and C57BL/6 mice (CD45.1+) into lethally irradiated C57BL/6 mice (CD45.1+) (data are representative of 3 independent experiments). (A) Experimental design. (B) Fluorescence-activated cell sorting (FACS) analysis of the percentage of total donor chimerism in PB 4, 8, 12, and 16 weeks after transplantation (n = 9-10). (C) FACS analysis of cell-specific chimerism in PB, BM, and spleen 16 weeks after competitive repopulation assay. (D-F) Homing assay of total BM cells from WT or VavCre+Rxrabfl/fl mice (CD45.2+), transplanted into lethally irradiated C57BL/6 mice (CD45.1+). (D) Experimental design. (E) Representative FACS plots showing the gating strategies for analysis of CD45.2+ HSCs. (F) Total numbers of CD45.2+ HSCs found in the BM of recipient mice. (G-H) Transplantation of purified LT-HSCs from WT or VavCre+Rxrabfl/fl mice (CD45.2+) along with LKs from C57BL/6 mice (CD45.1+) into lethally irradiated C57BL/6 mice (CD45.1+) (n = 19-20; data pooled from 2 independent experiments). (G) Experimental design. (H) Kaplan-Meier survival plot. (I-J) Serial CFU plating assay after 48-hours in vitro treatment of purified LT-HSCs from 5-month-old WT or VavCre+Rxrabfl/fl mice with 9c-RA or vehicle. (I) Experimental design. (J) Number of CFUs normalized to control vehicle treatment of each corresponding plating (data pooled from 2 independent experiments). All data are shown as means ± SEM, and dots represent individual animals. Significance was determined by 2-way analysis of variance (ANOVA) followed by the Sidak multiple comparisons test (B), unpaired Student t test (C and F), log-rank (Mantel-Cox) test (H), or ordinary 1-way ANOVA (J), and is represented as follows: ∗P ≤ .05, ∗∗P ≤ .01, ∗∗∗P ≤ .001, and ∗∗∗∗P ≤ .0001.

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