American Society of Hematology

Figure 3

$Figure 3. Selective deficiencies in GMPs and preGMPs in FL-deficient mice. (A) Representative FACS profiles and gating strategies for myeloid progenitors in WT and Fl −/− mice. preGMPs (Lin−Kit+Sca-1−CD41−CD150−FcγR−CD105low), GMPs (Lin−Kit+Sca-1−CD41−CD150−FcγR+), MkP (Lin−Kit+Sca-1−CD41+), and CFU-E (Lin−Kit+Sca-1−CD41−CD150−FcγR−CD105high). Numbers represent frequencies of gated populations relative to total BM cells, and are mean values from more than 3 experiments. (B) Absolute number of total BM cells, preGMPs, GMPs, MkPs, and CFU-Es per tibia in WT and Fl −/− mice. Each dot represents an individual mouse. More than 3 litters were analyzed for each genotype and progenitor subset. ns indicates not significant; **P < .01, ***P < .001. (C) Representative FACS profiles of peripheral blood analysis of WT and Fl−/− mice. Cells were first gated as CD45+NK1.1−. For myeloid lineage analysis cells were as indicated first gated as CD19−CD3− cells and then analyzed for Gr-1 and Mac-1 expression. (D) Peripheral blood cell analysis in WT and Fl −/− mice based on same gating strategy as shown in panel C. Data shown are total number of cells per milliliter of blood, B cells (CD19+), T cells (CD3+), granulocytes (Mac-1+Gr-1hi), and monocytes (Mac-1+Gr-1neg-lo). Each dot represents an individual mouse. Data are from 3 experiments. ns indicates not significant; *P < .05, **P < .01. (E) CFU-GM cultures: 10 000 unfractionated BM cells from WT or Fl−/− mice were cultured in methylcellulose supplemented with FL, IL-3, GM-CSF, and G-CSF. Clones were scored after 7 to 8 days. Each dot represents data from 1 mouse (means of 2 replicates). Horizontal bars indicate mean values of all mice in each genotype. Data are from at least 3 experiments. ***P < .001. (F) Effect of FL on CFU-GM cultures; 10 000 unfractionated BM cells from WT or Fl−/− mice were added to methylcellulose supplemented with IL-3, GM-CSF, and G-CSF with or without FL. Clones were scored after 7 to 8 days. Data represent mean (SD) values from 6 mice/genotype, 2 experiments. Statistical significance was tested with the paired Mann-Whitney, between WT, with and without FL, and Fl−/− with and without FL, respectively. *P < .05.$

Selective deficiencies in GMPs and preGMPs in FL-deficient mice. (A) Representative FACS profiles and gating strategies for myeloid progenitors in WT and Fl^−/− mice. preGMPs (Lin⁻Kit⁺Sca-1⁻CD41⁻CD150⁻FcγR⁻CD105^low), GMPs (Lin⁻Kit⁺Sca-1⁻CD41⁻CD150⁻FcγR⁺), MkP (Lin⁻Kit⁺Sca-1⁻CD41⁺), and CFU-E (Lin⁻Kit⁺Sca-1⁻CD41⁻CD150⁻FcγR⁻CD105^high). Numbers represent frequencies of gated populations relative to total BM cells, and are mean values from more than 3 experiments. (B) Absolute number of total BM cells, preGMPs, GMPs, MkPs, and CFU-Es per tibia in WT and Fl^−/− mice. Each dot represents an individual mouse. More than 3 litters were analyzed for each genotype and progenitor subset. ns indicates not significant; **P < .01, ***P < .001. (C) Representative FACS profiles of peripheral blood analysis of WT and Fl^−/− mice. Cells were first gated as CD45⁺NK1.1⁻. For myeloid lineage analysis cells were as indicated first gated as CD19⁻CD3⁻ cells and then analyzed for Gr-1 and Mac-1 expression. (D) Peripheral blood cell analysis in WT and Fl^−/− mice based on same gating strategy as shown in panel C. Data shown are total number of cells per milliliter of blood, B cells (CD19⁺), T cells (CD3⁺), granulocytes (Mac-1⁺Gr-1^hi), and monocytes (Mac-1⁺Gr-1^neg-lo). Each dot represents an individual mouse. Data are from 3 experiments. ns indicates not significant; *P < .05, **P < .01. (E) CFU-GM cultures: 10 000 unfractionated BM cells from WT or Fl^−/− mice were cultured in methylcellulose supplemented with FL, IL-3, GM-CSF, and G-CSF. Clones were scored after 7 to 8 days. Each dot represents data from 1 mouse (means of 2 replicates). Horizontal bars indicate mean values of all mice in each genotype. Data are from at least 3 experiments. ***P < .001. (F) Effect of FL on CFU-GM cultures; 10 000 unfractionated BM cells from WT or Fl^−/− mice were added to methylcellulose supplemented with IL-3, GM-CSF, and G-CSF with or without FL. Clones were scored after 7 to 8 days. Data represent mean (SD) values from 6 mice/genotype, 2 experiments. Statistical significance was tested with the paired Mann-Whitney, between WT, with and without FL, and Fl^−/− with and without FL, respectively. *P < .05.

This Feature Is Available To Subscribers Only