FIG 7.

P. stomatis challenge induced specific granule exocytosis and a time- and bacterial-concentration-dependent azurophil granule release. Neutrophils were unchallenged (basal) or challenged with fMLF (5 min), with P. stomatis for 30, 60, or 90 min (MOI, 10), or with different P. stomatis or P. gingivalis MOIs (10, 50, or 100) for 30 min, or pretreated with TNF-α (10 min) followed by fMLF (TNF-α + fMLF). (A to D) Specific and azurophil granule exocytosis activities were quantified by the increase in plasma membrane expression of CD66b or CD63, respectively, by flow cytometry. Data are expressed as the means ± SEM of the mean channel of fluorescence (mcf) from 6 independent experiments (A), 5 independent experiments (B and D), or 10 independent experiments (C). (E and F) Supernatants from all the different experimental conditions were collected, and the release of myeloperoxidase to determine azurophil granule exocytosis was measured by ELISA. Data are expressed as means ± SEM of MPO release in nanograms per 4 × 10⁶ cells from 5 independent experiments. *, P < 0.05; **, P < 0.001; ***, P < 0.0001.