Fig. 4

UC-MSCs-CM from 2D and 3D cultures navigated the polarization of human macrophages. A The concentration of TNF-α in culture media of MDM. MDM were treated with UC-MSCs-CM from 2D and 3D cultures, macrophages of control group were treated with DMEM complete media for 48 h. All cells were stimulated with LPS (10 ng/ml) and IFN-γ (50 ng/ml). The concentration of TNF-α in UC-MSCs-CM-treated cells is presented as percent of TNF-α concentration in control cells. Statistical significance was determined using a one-way ANOVA with Tukey’s post hoc test; **p < 0.01; ns = p > 0.05; B Surface expression of CD206 in untreated and UC-MSCs-CM-treated M2 macrophages evaluated by flow cytometry. Human macrophages of experimental group were treated with UC-MSCs-CM from 2D and 3D cultures, and macrophages of control group were treated with DMEM complete media for 48 h. All cells were additionally stimulated with IL-4 (20 ng/ml). Statistical significance was determined using Welch’s t test; ns = p > 0.05. Box plot demonstrates the fold change of median fluorescence intensity (MFI) of CD206 expression in UC-MSCs-CM-treated macrophages compared to untreated control which is represented by a dashed line; C Gating strategy used to identify CD206+ -positive macrophages. Forward scattering area (FSC-A) versus side scattering area (SSC-A) density plot was used to identify cells and exclude debris. FSC-Height versus FSC-A density plot was performed to analyze only single cells. Histograms represent fluorescence intensity of antiCD206-PE-Cy7 and allow to distinguish M0 and M2 macrophages; D Scatter plots showing correlation between macrophage markers and concentration of cytokines. Correlation coefficients (ρ) were calculated using Spearman’s rank test; *p < 0.05; **p < 0.01; ***p < 0.001