Figure 2

Hoxa9 is dispensable for NK differentiation and homeostasis. A) BM cells from wildtype B6 or hoxa9−/− mice were isolated and stained with combinations of antibodies as detailed in the Methods to resolve PreNKP and rNKP progenitor subsets. The Lin+ antibody cocktail includes CDllb/Mac1, CD3ε, NK1.1, CD19, and Ly6D. The gating strategy is depicted from top to bottom with arrows. Data is representative of 6 individual mice of each genotype. B) NKP subset frequencies in BM of wildtype B6 (open bars) and hoxa9−/− (gray bars) mice. Precursor frequencies reflect percentages of CLP, PreNKP, or rNKP within the Lin-CD244+CD27+IL-7R+ gated fractions. C) Absolute numbers of BM NKP subsets in wildtype B6 (open bars) and hoxa9−/− (gray bars) mice. D) Summary of frequencies or E) absolute numbers of BM total CD3ε-CD122+ progenitors, NKP (CD3ε- CD122+NK1.1-Dx5-), immature NK (CD3ε-CD122+NK1.1+ Dx5-), and mature NK (CD3ε-CD122+NK1.1+ Dx5+) cells in wildtype B6 (open bars), hoxa9−/− (light gray bars), and flt3l−/− (dark gray bars). Data reflects the average of each indicated subset pooled from analysis of 7–8 mice per genotype. F) Flow cytometry profile of CD3ε- NK1.1+CD122+ NK cells in spleen. Spleen mononuclear cells were first gated on CD3ε- (top panels). The CD3ε- splenocytes were then analyzed for expression NK1.1 and CD122 (bottom panels). G) Absolute numbers of NK cells (CD3ε-NK1.1+CD122+) in spleen. Precursor frequencies were calculated by multiplying sequential percentages of each gated region as described in Figure1. Absolute numbers were determined by multiplying precursor frequencies x numbers of BM mononuclear cells obtained from 4 hind limb leg bones. The asterisk indicates p < 0.05. Error bars represent SEM.