Figure 3
Figure 3. Mast-cell progenitors in SCL−/Δ mice arise from the megakaryocyte-erythroid progenitor cell fraction. (A) Flow cytometric analysis used for isolating common myeloid progenitors (CMPs), megakaryocyte-erythroid progenitors (MEPs), and granulocyte-macrophage progenitors (GMPs) from SCL+/Δ or SCL−/Δ bone marrow cells. The gate used for lineageneg c-kit+ cells is shown in the left dot plot and gates for CMPs, MEPs, and GMPs in the right dot plots. The numbers represent the mean percentage of MEPs of total nucleated cells from 4 mice of each genotype. (B) Mean numbers of colonies from 500 sorted cells from SCL+/Δ or SCL−/Δ mice cultured in methylcellulose or agar containing IL-3, SCF, and Epo. Results are the means from 2 independent experiments. Ery indicates blast forming unit-erythroid; Meg, megakaryocyte colonies; and G/M, granulocyte and granulocyte-macrophage colonies. (C) Acetylcholinesterase stain of a floated and fixed mast cell colony grown from SCL−/Δ MEP cells. (D) Flow cytometric analysis used for isolating myeloid progenitors according to PU.1gfp and FcRIII/II expression on lineageneg Sca-1− IL-7Rα− c-kit+ bone marrow cells from SCL+/Δ or SCL−/Δ mice. The gated regions used for cell sorting and the mean percentage of R3 cells calculated from 3 mice of each genotype are shown. (E) Relative PU.1 mRNA expression in sorted cell fractions measured by real-time PCR. Expression was normalized for HPRT and expression of PU.1 in SCL+/Δ R2 cells was arbitrarily set at 1.0. (F) Mean numbers of colonies from 500 sorted cells from SCL+/Δ or SCL−/Δ mice cultured in methylcellulose or agar containing IL-3, SCF, and Epo. Results are the mean from 3 independent experiments.

Mast-cell progenitors in SCL−/Δ mice arise from the megakaryocyte-erythroid progenitor cell fraction. (A) Flow cytometric analysis used for isolating common myeloid progenitors (CMPs), megakaryocyte-erythroid progenitors (MEPs), and granulocyte-macrophage progenitors (GMPs) from SCL+/Δ or SCL−/Δ bone marrow cells. The gate used for lineageneg c-kit+ cells is shown in the left dot plot and gates for CMPs, MEPs, and GMPs in the right dot plots. The numbers represent the mean percentage of MEPs of total nucleated cells from 4 mice of each genotype. (B) Mean numbers of colonies from 500 sorted cells from SCL+/Δ or SCL−/Δ mice cultured in methylcellulose or agar containing IL-3, SCF, and Epo. Results are the means from 2 independent experiments. Ery indicates blast forming unit-erythroid; Meg, megakaryocyte colonies; and G/M, granulocyte and granulocyte-macrophage colonies. (C) Acetylcholinesterase stain of a floated and fixed mast cell colony grown from SCL−/Δ MEP cells. (D) Flow cytometric analysis used for isolating myeloid progenitors according to PU.1gfp and FcRIII/II expression on lineageneg Sca-1 IL-7Rα c-kit+ bone marrow cells from SCL+/Δ or SCL−/Δ mice. The gated regions used for cell sorting and the mean percentage of R3 cells calculated from 3 mice of each genotype are shown. (E) Relative PU.1 mRNA expression in sorted cell fractions measured by real-time PCR. Expression was normalized for HPRT and expression of PU.1 in SCL+/Δ R2 cells was arbitrarily set at 1.0. (F) Mean numbers of colonies from 500 sorted cells from SCL+/Δ or SCL−/Δ mice cultured in methylcellulose or agar containing IL-3, SCF, and Epo. Results are the mean from 3 independent experiments.

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