Figure 1
Figure 1. In situ deletion of SHIP does not compromise the capacity of HSCs to mediate long-term multilineage repopulation. (A) Contour plots for CD45.1 versus CD45.2 staining illustrating equal repopulation in PB at 60 days after transplantation before poly I:C treatment in 2 representative MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras. (B) Percentage of donor repopulation in the PB by MxCreSHIPflox/flox (CD45.2) and WT-Ly5.1 (CD45.1) HSCs in MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras at 60 days after transplantation before poly I:C treatment. (C) Representative SHIP Western blot from single-positive CD45.1+CD45.2−- or CD45.2+CD45.1−-sorted cells isolated from splenocytes from MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras 5 months after poly I:C treatment. Single-positive splenocytes were sorted and lysates were probed for SHIP expression. (D) Percentage of global repopulation in the PB by MxCreSHIPflox/flox (CD45.2) and WT-Ly5.1 (CD45.1) HSC in MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras at the indicated times after poly I:C treatment (n ≥ 5). At monthly intervals, the level of donor reconstitution was assessed in PB. (E) RU activity by MxCreSHIPflox/flox (CD45.2) and WT-Ly5.1 (CD45.1) HSCs in the MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras 5 months after poly I:C treatment (n ≥ 5). (F) Contour plots for CD45.1 versus CD45.2 staining that illustrate multilineage repopulation in PB 20 weeks after poly I:C treatment in a representative MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimera. (G) Percentage repopulation of the indicated lymphoid and myeloid cell lineages by MxCreSHIPflox/flox (CD45.2) and WT-Ly5.1 (CD45.1) HSC in MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras 5 months after poly I:C treatment (n ≥ 5). At death, the level of donor reconstitution was assessed in BM. Significance was established using the unpaired Student t test (* P < .05). Errors shown represent the SEM; ■, cells derived from MxCreSHIPflox/flox BM; , cells derived from WT-Ly5.1 BM.

In situ deletion of SHIP does not compromise the capacity of HSCs to mediate long-term multilineage repopulation. (A) Contour plots for CD45.1 versus CD45.2 staining illustrating equal repopulation in PB at 60 days after transplantation before poly I:C treatment in 2 representative MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras. (B) Percentage of donor repopulation in the PB by MxCreSHIPflox/flox (CD45.2) and WT-Ly5.1 (CD45.1) HSCs in MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras at 60 days after transplantation before poly I:C treatment. (C) Representative SHIP Western blot from single-positive CD45.1+CD45.2−- or CD45.2+CD45.1−-sorted cells isolated from splenocytes from MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras 5 months after poly I:C treatment. Single-positive splenocytes were sorted and lysates were probed for SHIP expression. (D) Percentage of global repopulation in the PB by MxCreSHIPflox/flox (CD45.2) and WT-Ly5.1 (CD45.1) HSC in MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras at the indicated times after poly I:C treatment (n ≥ 5). At monthly intervals, the level of donor reconstitution was assessed in PB. (E) RU activity by MxCreSHIPflox/flox (CD45.2) and WT-Ly5.1 (CD45.1) HSCs in the MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras 5 months after poly I:C treatment (n ≥ 5). (F) Contour plots for CD45.1 versus CD45.2 staining that illustrate multilineage repopulation in PB 20 weeks after poly I:C treatment in a representative MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimera. (G) Percentage repopulation of the indicated lymphoid and myeloid cell lineages by MxCreSHIPflox/flox (CD45.2) and WT-Ly5.1 (CD45.1) HSC in MxCreSHIPflox/flox (CD45.2):WT-Ly5.1 (CD45.1) BM chimeras 5 months after poly I:C treatment (n ≥ 5). At death, the level of donor reconstitution was assessed in BM. Significance was established using the unpaired Student t test (* P < .05). Errors shown represent the SEM; ■, cells derived from MxCreSHIPflox/flox BM; , cells derived from WT-Ly5.1 BM.

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