Figure 1.

FOXO1 deletion impairs bacteria-induced neutrophil recruitment. (A,B) Porphyromonas gingivalis was inoculated into the connective tissue of the scalp and mice were examined at the indicated time points. (A) The mice was euthanized at 12 h and number of cells in the inoculated tissue and (B) neutrophils in peripheral blood were measured by immunofluorescent flow cytometry using specific antibodies for neutrophils (Ly6G), T cells (CD3), B cells (B220), and macrophages (F4/80). FOXO1-deleted LyzM.Cre⁺FOXO1^L/L mice and littermate control LyzM.Cre⁻FOXO1^L/L mice were inoculated with bacteria at the indicated time points. The number of neutrophils in the inoculated tissue (C), peripheral blood (D), and bone marrow (E) was measured by immunofluorescent flow cytometry using specific antibodies. (F) Neutrophil mobilization was calculated as described in methods to estimate the percentage of total neutrophils in the blood. RNA was isolated from mouse neutrophils from FOXO1-deleted LyzM.Cre⁺FOXO1^L/L mice and littermate control LyzM.Cre⁻FOXO1^L/L mice. FOXO1 mRNA levels were measured by RT-PCR and normalized to ribosomal protein L32 (G). (H) Neutrophils from experimental LyzM.Cre⁺FOXO1^L/L mice and littermate control LyzM.Cre⁻FOXO1^L/L mice were analyzed by Western blots for FOXO1 expression with actin as loading control. The data are representative of two or three independent experiments. Data are presented as the mean ± SEM from triplicate samples. *Significant difference between neutrophils from FOXO1-deleted LyzM.Cre⁺FOXO1^L/L mice and littermate control LyzM.Cre⁻FOXO1^L/L mice (P < 0.05).