Receptor expression and proliferation of LKS upon extrinsic stimulation. (A) Quantitative polymerase chain reaction analysis on mRNA expression of the indicated genes in FACS-sorted BM HSCs (LKS Flt3−CD34−CD48−CD150+), MPPs (LKS Flt3+CD34+), GMPs (LK Sca1−FcgR+CD34+), and granulocytes (Mac1+Gr1+), as well as spleen T (CD3+), B (CD19+), and dendritic cells (CD11c+MHCII+). Relative expression of the indicated genes to the expression of Actb is shown in the respective cell populations (mean ± standard error of the mean), data obtained from 3 independent experiments, n = 4. (B) Experimental scheme: steady-state, nonirradiated mice were transplanted with 0.8 to 1.1 × 105 CFSE-labeled LKS cells; 1 week posttransplantation, they were injected over 1 week as indicated in “Study design.” One week after the final injection and 3 weeks after transplantation of CFSE-labeled LKS, mice were analyzed and cells were subsequently sorted for transplantation as indicated. For secondary transplantation, whole BM was used as indicated. (C) Representative dot plot analysis of lineage-depleted BM cells pregated on donor Lin− (upper) or donor Lin−c-Kit+ cells (lower). (D) Percentage of donor LKS in subsequent divisional groups. Graphs show mean ± standard error of the mean (n = 6-24 mice from 3 to 14 independent experiments). Data analyzed with paired, 2-tailed Student t test. ***P < .001, **P < .01, *P < .05. GMPs, granulocyte-macrophage progenitors; ns, not significant; w/o, without.
