Alternative activation of MΦs decreases N meningitidis uptake. (A) Flow cytometric analysis of ThioMΦ ingestion of Rhodamine Green labeled Neisseria (RdGnX-N.m.). ThioMΦs were stimulated with IL-4 for 48 hours and challenged with ethanol-fixed RdGnX-N.m. (100 bacteria/MΦ) for 2 hours at 37°C. The mean fluorescence for each treatment was determined by flow cytometry. The histogram shows the effect of IL-4 treatment on uptake of N meningitidis. (B) IL-4 induces a dose-dependent inhibition of N meningitidis uptake. ThioMΦs were stimulated with different concentrations of IL-4 for 48 hours and challenged as described. The mean fluorescence for each population was determined by flow cytometry. The average mean fluorescence for each condition is shown. Error bars indicate SDs. (C) IL-4–inhibited RdGnX-N.m. uptake by MΦs in a time-dependent manner. ThioMΦs were stimulated for different periods with IL-4 and challenged as above. Unchallenged cells served as a negative control. The mean fluorescence for each population was determined by flow cytometry. The average mean fluorescence for each condition is shown. Error bars indicate SDs. (D) The inhibition of N meningitidis uptake is specific for alternative activation of MΦs. (Left) ThioMΦs were stimulated with IL-13 (10 ng/mL) for 48 hours and challenged with ethanol-fixed RdGnX-N.m. The mean fluorescence for each treatment was determined by flow cytometry. The histogram shows the effect of IL-13 treatment on uptake of N meningitidis. (Right) IL-4 inhibited N meningitidis uptake by ThioMΦs via IL-4Rα. WT or IL-4Rα^−/− ThioMΦs were incubated for 48 hours with IL-4 (5 ng/mL) and challenged with RdGnX-N.m. as described. After fixation with 2% paraformaldehyde, the mean fluorescence for each population was determined by flow cytometry. The figure showed the average mean fluorescence intensity of 3 independent experiments. Error bars indicate SDs.