STAT3 regulates G-CSF–responsive proliferation of immature granulocytes and the kinetics of granulocyte maturation. (A) Immature Gr-1^lo granulocytes were isolated from the bone marrow of wild-type (WT) and STAT3-deficient (KO) mice by fluorescence-activated cell sorting. Cells were labeled with CFSE, cultured in G-CSF (2.5 ng/mL) for 4 days, and analyzed by flow cytometry. Results of a representative experiment (1 of 3) are shown. (B) Gr-1^lo cells were cultured in G-CSF (25 ng/mL) for the indicated times or analyzed directly after isolation (0 hour). Cells were stained with propidium iodide and analyzed by flow cytometry. The percentage of cells in S phase was determined by ModFit LT v3.0 (n = 3 for WT and KO for each condition). (C) Bone marrow Gr-1^lo cells were isolated 4 hours after mice received a single dose of G-CSF (250 μg/kg; +) or BSA carrier alone (−). Gene expression was measured by quantitative PCR, with normalization to 18s RNA (n = 3 for WT and KO for each condition). (D) Gr-1^lo cells were cultured in G-CSF for 2 or 4 days or analyzed directly after isolation (NT). The expression of neutrophil differentiation markers was measured at the indicated times by quantitative PCR (n = 3 for WT and KO for each condition). (B-D) Average values from 3 independent experiments are shown. Error bars represent SEM. *P < .05 compared with untreated or BSA-treated controls. **P < .01, ***P < .001 compared with control.