Fig. 5

FL-MSCs are able to induce T regs with more activated phenotype in comparison to BM-MSCs. Activated CD3⁺ CD25⁻ effector T cells were co-cultured with BM- or FL-MSCs in a fixed 1:5 MSC to T cell ratio. After 4 days, T cells were collected and the expression of different activated markers (CD25, GITR, TNFR2, and ICOS) was determined in CD4⁺Foxp3⁺ T regs by flow cytometry measurements. Representative flow cytometry dot plots show the percentage of CD25, GITR, TNFR2, and ICOS, within CD4⁺Foxp3⁺ T regs from the FL-MSC group (left panel). After delimitating the lymphocyte region by a forward-scatter-area (FSC-A) versus side-scatter area (SSC-A) plot, a CD4 versus Foxp3 plot was used to gate on CD4⁺Foxp3⁺ T regs. Frames defined the positive subpopulations for each marker analysis in the CD4⁺Foxp3⁺ population. Statistical summary dot-plot graphs showing the percentage or the MFI value of each marker analyzed in CD4⁺Foxp3⁻⁺ iT regs (right panel). Red dots stand for BM-MSCs iT regs (n = 9) and blues stand for FL-MSCs iT regs conditions (n = 9). All data are collected from 3 different experiments. MFI values have been normalized with BM-MSCs iT regs group. For each group of values, horizontal lines represent mean value ± SEM. Unpaired Student t test analysis was performed to generate P values. ns, non-significant; iTregs, induced regulatory T cells. *P < .05, **P < .01, ***P < .001, ****P < .0001