Fig. 6

Functional validation of neutrophil migration enhanced by the combination of IL-17A/F and TNF-α. (A) HBEC-3KT were stimulated with IL-17A/F (50 ng/mL), TNF-α (20 ng/mL) and IFN-γ (30 ng/mL), or the cytomix combinations, as indicated. Tissue culture supernatants were collected after 24 h and used in trans-well cell migration assays, to monitor the migration of neutrophils isolated from human blood. Cell culture medium spiked with human recombinant IL-8 (30 ng/mL) was used as a positive control. (B) Results are shown as boxplots with the median line and IQR, and whiskers show minimum and maximum values. Each data point represents an independent experimental replicate with HBEC supernatant (n = 4), using neutrophil isolated from one donor. Each dot represents the average number of neutrophils that traversed the membrane within two hours in each experiment. Increase in neutrophil migration was calculated after subtraction of neutrophil numbers found with tissue culture supernatant from paired unstimulated cells in trans-well migration assay, for each biological replicate. One-way ANOVA with Bonferroni’s post-hoc test for multiple comparisons was used for statistical significance compared to unstimulated cells as control (***p < 0.001, ****p < 0.0001)