NOD1L administration restores numbers of HSPCs in GF mice to the levels observed in SPF mice. (A) Levels of HSC, MPP, and CLP in SPF vs GF mice. Each symbol indicates the absolute number of LSK or CLP in BM from individual SPF (n = 6) and GF mice (n = 11). The horizontal bars represent the mean values for each group. (B) Serum levels of NOD1 or NOD2 ligand in naïve SPF and GF mice. Bars represent the mean ± standard error of the mean (SEM) of the values obtained in the reporter gene assay used (n = 8 per group). (C) Levels of Nod1 and Nod2 mRNA expression in indicated in BM HSPC populations isolated from WT, NOD1^−/−, or NOD2^−/− mice (n = 10). BMMϕ from WT animals were included as positive controls. Bars represent the mean ± standard deviation (SD) of triplicate values of Nod1 or Nod2 expression levels relative to the Rplp2 housekeeping gene. (D) NOD1 ligand levels in GF mice (n = 5-9) given either synthetic NOD1 ligand (C12-iE-DAP, 100 μg) or phosphate-buffered saline by gavage every 2 or 3 days for 2 weeks. (E-F) HSPC populations in GF mice after gavage with NOD1 ligand (C12-iE-DAP, 100 μg) as noted before. (E) The representative dot plots shown were gated on lineage^neg and IL-7Rα^neg (upper panels) or IL-7Rα⁺ (lower panels) cells, respectively. (F) Absolute numbers of LT-HSC, ST-HSC, MPP, CLP, and CMP in SPF mice, GF mice, and GF mice treated with NOD1 ligand. The symbols represent the cell numbers for the individual animals in each group (n = 7-12). The bars represent the means ± SEM of these values. *P < .05; **P < .01; ***P < .001 denote the statistical significance of the differences in group means. The data shown in (A), (B), (D), and (F) are pooled from 2 to 3 independently performed experiments, whereas the values shown in (C) are from an experiment representative of 2 performed.