Figure 2.

Effect of polarization on membrane expression of FcγRs and CD13. Monocytes from healthy donors were cultured for 6 days in a medium supplemented with macrophage colony-stimulating factor (M-CSF) to differentiate into M0. The resulting human monocyte-derived macrophages were polarized by incubation with IFN-γ (30 ng/mL), IL-4 (50 ng/mL), or IL-10 (20 ng/mL) for 48 h and subsequently analyzed by flow cytometry for the expression of FcγRI, FcγRII, FcγRIII, and CD13. (A) Representative histograms of cells from a single donor. The arrows indicate significant changes in receptor expression induced by polarization. Colored histograms are from cytokine-treated cells. (B) Upper plots show the mean fluorescence intensity (MFI) of FcγRI and FcγRIII in non-polarized and polarized cells from 30 individual donors, and lower plots show the same data plotted as average fold increase in MFI relative to control (non-polarized macrophages or M0). (C) After polarization, cells were lysed, and RNA was isolated for quantification of mRNA for FcγRI, FcγRIIa, FcγRIIb, and FcγRIII by real-time PCR. Average fold increase of mRNA relative to non-polarized macrophages in cells from 10 different donors analyzed in triplicate. Results are expressed as mean + SD of independent experiments. Statistical significance was calculated using one-way ANOVA with Tukey post hoc test. For analysis of the expression of FcγRs using normalization of data [(B), lower plots] ANOVA was used to compare Mϕ-IFN-γ, Mϕ-IL-4, and Mϕ-IL-10, followed by Tukey’s post hoc test for comparisons between treatment groups (*p < 0.05 and ***p < 0.001).