Figure 6.

Serum amyloid A (SAA)3 induces neutrophil expression of IL-22. Neutrophils from bone marrow of wild-type C57BL/6 mice (both male and female, 8–10 weeks of age) were treated with recombinant SAA3 (rSAA3) (1 µM) or PBS (Con) for 24 h before analysis. (A) Representative flow cytometry dot plots showing the effects of rSAA3 on the CD11b⁺Ly6G^high neutrophil population (left two panels) and IL-22-producing neutrophils (right two panels). Quantification data, based on five mice in each group, are shown in bar charts on the right. *p < 0.05. (B) Bone marrow neutrophils were treated with mouse rSAA3 (mSAA3, left) or human rSAA1 (hSAA, right) for 24 h. IL-22 release was quantified by ELISA. Bone marrow neutrophils (C) or splenocytes (D) were incubated with an anti-IL-23 antibody (1 µg/ml) or IgG isotype control (Isotype) for 1 h before adding rSAA3 (1 µM) for 24 h. The culture medium was collected and IL-22 concentration was determined by ELISA. Splenocytes (right) from the same mice were pre-incubated with the IL-23 Ab or IgG isotype control for 1 h before stimulation with 20 ng/ml of IL-23 for 24 h. IL-22 release was measured by ELISA. (E) IL-23 production by neutrophils stimulated with indicated concentrations of rSAA3 for 24 h. All quantitative data shown are mean ± standard margin of error based on triplicate measurement, using neutrophils from three to five mice. *p < 0.05, **p < 0.01 between different samples compared.