FIG 4.

Reduced bactericidal activity of Hs2st-deficient neutrophils. Bone marrow neutrophils (A) and LPS-elicited neutrophils (B) were collected from WT or Hs2St-deficient mice. After challenge with GBS, CFU were measured at the indicated time points by serial plating. Data shown are means ± SEM from one of the two independent experiments. (C) Reactive oxygen species production from bone marrow neutrophils after GBS stimulation detected by a luminol-based assay (n = 2 mice). (D) Bone marrow neutrophils were directly lysed or stimulated with GBS for 30 min. Supernatants were collected to measure the total neutrophil elastase (top) or released neutrophil elastase (bottom) by Western blot analysis. (E) Whole blood from WT and Hs2st-deficient mice was incubated with pHrodo red-labeled GBS (MOI of 100) for 1 h at 37°C (phagocytosis) or 4°C (control). Neutrophils were fixed and subjected to flow cytometry to measure internalized GBS. (F) LPS-elicited neutrophils were stimulated with GBS at an MOI of 10 for 2 h, washed, fixed, and stained with H2A/H2B MAb to visualize NET formation. Differences between groups were calculated by an unpaired t test. **, P < 0.01; *, P < 0.05 (A, B, C, and F).