FIG. 6.

Screening of H. influenzae clinical isolates with 9HTEo− cells for epithelial IL-8 secretion. Monolayers of cultured human tracheal epithelial cells (3 × 10⁶ cells/well in six-well tissue culture plates; serum-free culture media) were coincubated with tissue culture medium alone, IL-1β (20 ng/ml), LOS (0.21 μg/ml), and H. influenzae clinical isolates (5 × 10⁷ CFU/ml). Culture fluid was harvested after 16 h of coincubation with various stimuli, and secreted IL-8 was measured using commercially available ELISAs. The results are shown as nanograms of IL-8 per milliliter of culture fluid. Error bars each represent the standard error of the mean of three different wells each analyzed in triplicate by ELISA. IL-8 concentrations were 0.317 ± 0.006 ng/ml in unstimulated control wells and 14.25 ± 4.87 ng/ml in IL-1β-stimulated control wells. NTHi strains used in this study were derived from wild-type strains AAr176p⁺ (Pili), 11 (Hia), 12 (HMW1/2), N187 (Hap), and Rd. See Table 1 for strain details.