Fig. 5
MenSCs pre-treated with IFN-γ- and IL-1β-augmented functional Treg generation. Purified CD4 + T cells were co-cultured with MenSCs at different ratios and activated with T cell activation beads. After five days, percent of CD4 + CD25highFOXP3 + Tregs was quantified by flow cytometry. Gating strategy for Tregs is shown in panel (A). MenSCs were either remained untreated (B) or primed with IFN-γ and IL-1β (C). T cells cultured in the absence of MenSCs served as biological control (BC). The data obtained from each group were compared with biological control, and the data were expressed as mean ± SEM of 12 experiments. IL-1β/IFN-γ-treated MenSCs were co-cultured with CD4 + T cells for five days and induced Tregs were then isolated by MACS. Tregs were added at 1:5 and 1:10 ratio to the CFSC-labeled autologous PBMCs cultures activated with activation beads and proliferation of PBMCs were measured after 4 days by flow cytometry (D). Wells containing single PBMCs, Tregs (generated in the absence of MenSCs, BC-Treg), or MenSCs-induced Tregs (Coculture-Treg) served as negative controls. Co-cultures containing Tregs (generated in the absence of MenSCs) and PBMCs at ratio of 1:5 and 1:10 were considered as positive control. Results were displayed as Mean ± SEM of 6 different experiments. The signs (**) and (***) stand for p < 0.0021 and p < 0.0002, respectively