Figure 5. PVL enhances PMN bactericidal activity.

(A–C) Surface expression of CD11b. PMNs (1×10⁶) were incubated with or without 1 nM PVL or iPVL (B), and surface expression of CD11b was measured by flow cytometry. Bars indicate the mean ± se of three or four separate experiments. *P < 0.05 for the indicated comparisons using a one-way ANOVA and Tukey's post-test. (C) PMNs pretreated for 30 min with 10 μM SB203580 were incubated ± 1 or 2 nM PVL or 100 ng/ml LPS for an additional 30 min at 37°C. Surface expression of CD11b was measured by flow cytometry. Bars indicate the mean ± se of five to six separate experiments. *P < 0.05 using a paired t test. −incub., PMNs not preincubated for 30 min, as needed for treatment with SB203580. (D) Redistribution of NADPH oxidase components following exposure of PMNs to 1 nM PVL. Immunoblots containing proteins from the plasma membrane-enriched fractions of PMNs, stimulated ± 1 nM PVL, were probed with antibodies specific for gp91phox (αgp91phox) or p47phox (αp47phox). Results shown are representative of three separate experiments. Cyt., Cytosol-enriched fraction. (E) Binding/ingestion of USA300 by human PMNs following priming by 1 nM PVL or 100 ng/mL LPS for 30 min. Following priming, bacteria were combined with PMNs at a 1:1 ratio and rotated gently for 2 h, at which time, an aliquot of the assay was used to determine the percent PMNs with bound/ingested USA300. (F) Bactericidal activity of PMNs toward USA300 following priming by 1 nM PVL or 100 ng/mL LPS, as described for E. Results for E and F are the mean ± se of four separate experiments as indicated. *P < 0.05 for the indicated comparisons using a one-way ANOVA and Tukey's post-test. S.a., S. aureus.