Increased frequencies of ROS+ inflammatory monocytes and neutrophils that generate higher levels of ROS during the secondary infection.
(A) Spleen cells from wt mice (3/group) were independently analyzed by FACS for surface expression of F4/80, CD11b, Ly-6C and Ly-6G. Data show representative FACS profiles in a representative (out of 2) experiment. Red gate: inflammatory Ly6C+ monocytes, black gate: neutrophils, blue gate: macrophages. (B) Mice (9–10 per group) primarily injected with PBS (closed circles, primary) or 0.1xLD50 (3×103) wt Lm (open circles, secondary) were challenged 30 days later with 10xLD50 (3×105) wt Lm. At the indicated times after challenge, spleen cells were restimulated with Heat Killed Lm (HKLM) in the presence of hydroethidine and analyzed by FACS for CD11b and Ly-6C expression. Data show the number (mean +/− SE) (left panels) and the MFI (mean +/− SE) (right panels) of ROS-producing inflammatory monocytes (upper panel), neutrophils (middle panel) and macrophages (bottom panel) per total gated inflammatory monocytes, neutrophils or macrophages and are representative of a pool of 2–3 replicate experiments. (C) Primary and memory mice (10/group) were treated 30 days later with a control isotype or serum (black bars), anti-CD8 or anti-CCL3 (white bars) and challenged with 3×105 wt Lm. At 10 hrs after the infection, the frequencies of ROS-producing cells were analyzed as in (B). Data result from the pool of 2 independent experiments. P (*<0.05) values were calculated in (B–C) with n = 9–10.
