Figure 5.
Older HSC populations from WT GF and IL-1R1KO SPF mice do not generate myeloid-biased output upon transplantation. (A) Schematic experimental approach to test self-renewal and differentiation capacity of 2mo, 1y, and 2y LT-HSCs from WT SPF, IL-1R1KO SPF, and WT GF mice. Phenotypic LT-HSCs (10 or 100 LKS CD34–Flt3–CD48–CD150+ cells) from CD45.2 mice were transplanted together with rescue total BM (300 000 cells from CD45.1 mice) into lethally irradiated mice (CD45.1+ or F1 CD45.1+/CD45.2+). Transplanted mice were bled once per month to follow donor engraftment and lineage repopulation capacities. BM LT-HSC donor chimerism, PB CD45+ donor chimerism, and PB lineage contribution of donor LT-HSCs was assessed 4 months after transplantation (Tx). (B) BM chimerism frequency analysis of donor (CD45.2+) LT-HSCs from 2mo, 1y, and 2y LT-HSCs from WT SPF, IL-1R1KO SPF, and WT GF donor mice 4 months after transplantation. WT SPF 2mo 10/100 cells, n = 8/10; WT SPF 1y 10/100 cells, n = 6/7; WT SPF 2y 10/100 cells, n = 6/7; IL-1R1KO SPF 2mo 10/100 cells, n = 4/5; IL-1R1KO SPF 2y 10/100 cells, n = 3/6; WT GF 2mo 10/100 cells, n = 8/11; WT GF 1y 10/100 cells, n = 5/9. In (B-D), “n” refers to recipient animals. For donors, a minimum of 3 biological replicas (1 biological replica is a pool of 2 mice) were used per condition. Engraftment levels of >0.1% were considered. (C) PB CD45+ chimerism contribution of 10 or 100 LT-HSCs from 2mo, 1y, and 2y LT-HSCs from WT SPF, IL-1R1KO SPF, and WT GF donor mice 4 months after transplantation. WT SPF 2mo 10/100 cells, n = 13/15; WT SPF 1y 10/100 cells, n = 10/17; WT SPF 2y 10/100 cells, n = 14/10; IL-1R1KO SPF 2mo 10/100 cells, n = 11/15; IL-1R1KO SPF 2y 10/100 cells, n = 10/7; WT GF 2mo 10/100 cells, n = 8/13; WT GF 1y 10/100 cells, n = 6/7. Engraftment levels of >0.1% were considered. (D) Lineage contribution of transplanted 10 or 100 LT-HSCs from 2mo, 1y, and 2y WT SPF, IL-1R1KO SPF, and WT GF mice 4 months after transplantation into WT SPF mice, respectively. Transplanted HSC contribution to myeloid cells (CD45.2+ CD11b+CD3e–CD19–) is indicated in red, to T cells (CD45.2+ CD3e+CD19–) in yellow, and to B cells (CD45.2+ CD3e–CD19+) in light green. WT SPF 2mo 10/100 cells, n = 11/12; WT SPF 1y 10/100 cells, n = 5/9; WT SPF 2y 10/100 cells, n = 9/11; IL-1R1KO SPF 2mo 10/100 cells, n = 11/15; IL-1R1KO SPF 2y 10/100 cells, n = 10/7; WT GF 2mo 10/100 cells, n = 8/13; WT GF 1y 10/100 cells, n = 6/7. Dashed line represents 50% cut off for biased lineage repopulation. Error bars represent SD in (B-C) and SEM in (D). P values were calculated using Student t test. *P < .05; **P < .01; ***P < .001; ****P < .0001.

Older HSC populations from WT GF and IL-1R1KO SPF mice do not generate myeloid-biased output upon transplantation. (A) Schematic experimental approach to test self-renewal and differentiation capacity of 2mo, 1y, and 2y LT-HSCs from WT SPF, IL-1R1KO SPF, and WT GF mice. Phenotypic LT-HSCs (10 or 100 LKS CD34–Flt3–CD48–CD150+ cells) from CD45.2 mice were transplanted together with rescue total BM (300 000 cells from CD45.1 mice) into lethally irradiated mice (CD45.1+ or F1 CD45.1+/CD45.2+). Transplanted mice were bled once per month to follow donor engraftment and lineage repopulation capacities. BM LT-HSC donor chimerism, PB CD45+ donor chimerism, and PB lineage contribution of donor LT-HSCs was assessed 4 months after transplantation (Tx). (B) BM chimerism frequency analysis of donor (CD45.2+) LT-HSCs from 2mo, 1y, and 2y LT-HSCs from WT SPF, IL-1R1KO SPF, and WT GF donor mice 4 months after transplantation. WT SPF 2mo 10/100 cells, n = 8/10; WT SPF 1y 10/100 cells, n = 6/7; WT SPF 2y 10/100 cells, n = 6/7; IL-1R1KO SPF 2mo 10/100 cells, n = 4/5; IL-1R1KO SPF 2y 10/100 cells, n = 3/6; WT GF 2mo 10/100 cells, n = 8/11; WT GF 1y 10/100 cells, n = 5/9. In (B-D), “n” refers to recipient animals. For donors, a minimum of 3 biological replicas (1 biological replica is a pool of 2 mice) were used per condition. Engraftment levels of >0.1% were considered. (C) PB CD45+ chimerism contribution of 10 or 100 LT-HSCs from 2mo, 1y, and 2y LT-HSCs from WT SPF, IL-1R1KO SPF, and WT GF donor mice 4 months after transplantation. WT SPF 2mo 10/100 cells, n = 13/15; WT SPF 1y 10/100 cells, n = 10/17; WT SPF 2y 10/100 cells, n = 14/10; IL-1R1KO SPF 2mo 10/100 cells, n = 11/15; IL-1R1KO SPF 2y 10/100 cells, n = 10/7; WT GF 2mo 10/100 cells, n = 8/13; WT GF 1y 10/100 cells, n = 6/7. Engraftment levels of >0.1% were considered. (D) Lineage contribution of transplanted 10 or 100 LT-HSCs from 2mo, 1y, and 2y WT SPF, IL-1R1KO SPF, and WT GF mice 4 months after transplantation into WT SPF mice, respectively. Transplanted HSC contribution to myeloid cells (CD45.2+ CD11b+CD3e–CD19–) is indicated in red, to T cells (CD45.2+ CD3e+CD19–) in yellow, and to B cells (CD45.2+ CD3e–CD19+) in light green. WT SPF 2mo 10/100 cells, n = 11/12; WT SPF 1y 10/100 cells, n = 5/9; WT SPF 2y 10/100 cells, n = 9/11; IL-1R1KO SPF 2mo 10/100 cells, n = 11/15; IL-1R1KO SPF 2y 10/100 cells, n = 10/7; WT GF 2mo 10/100 cells, n = 8/13; WT GF 1y 10/100 cells, n = 6/7. Dashed line represents 50% cut off for biased lineage repopulation. Error bars represent SD in (B-C) and SEM in (D). P values were calculated using Student t test. *P < .05; **P < .01; ***P < .001; ****P < .0001.

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