SAA1(46-112) synergizes with CXCL8 to activate and chemoattract neutrophils and desensitizes the chemotactic response of neutrophils toward cooperating intact SAA1α and CXCL8. (A) The chemotactic potencies of CXCL8 (0.2-10 ng/mL), SAA1(46-112) (100-1000 ng/mL), and a combination of CXCL8 and SAA1(46-112) were evaluated on neutrophils in the Boyden microchamber assay. Data represent the mean CI from 2 to 6 independent experiments. (B) Shape change of neutrophils was determined after 5 minutes of stimulation with CXCL8 (1-25 ng/mL), SAA1(46-112) (300 and 3000 ng/mL), or a combination of CXCL8 and SAA1(46-112). Data (6 independent experiments) are expressed as the net percentage of blebbed (blue bars) and elongated (red bars) neutrophils ± SEM. (C) Neutrophils were preincubated (10 minutes, 37°C) with desensitizing SAA1(46-112) (1000 or 3000 ng/mL) or with control buffer and were subsequently added to the upper wells of the Boyden microchamber. Chemotaxis was induced by adding 3 ng/mL of CXCL8, 300 ng/mL of hu rSAA1α, or a combination of CXCL8 and rSAA1α to the lower wells. Data are shown as mean CI ± SEM and are pooled from 3 to 12 independent experiments. (A-C) Statistically significant synergy (compared with the sum of the values when both chemotactic agents are tested separately) is indicated by daggers (^†P ≤ .05; ^††P ≤ .01; Mann-Whitney U test) and statistically significant inhibition of synergy (compared with synergy on buffer-treated cells) by desensitizing agents is indicated by dollar signs (^$$P ≤ .01; ^$$$P ≤ .001; Mann-Whitney U test).