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. 2016 Feb 1;6:20250. doi: 10.1038/srep20250

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Copyright © 2016, Macmillan Publishers Limited

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MDSCs induced from bone marrow cells after 4 days of culture in the presence of 40 ng/ml recombinant mouse GM-CSF with or without 1 μM RPM. (A) The macroscopic pictures of cell culture. (B) Lactate production and glucose uptake of M-MDSCs. (C) Glucose uptake of M-MDSCs isolated from spleens of control mice, RPM-treated mice, alloskin-grafted mice and RPM-treated alloskin-grafted mice at day 7 post transplantation. (D) Glucose uptake of M-MDSCs isolated from spleens of alloskin-grafted WT and lyzs-mTOR KO mice at day 7 after skin grafting. (E) Glucose uptake of Lin⁻ cells isolated from bone marrows of alloskin-grafted mice and RPM-treated alloskin-grafted mice at day 7 after skin grafting. (F) Schematic representation of the important genes (red) involved in the glycolysis pathway. (G) Some key genes involved in the glycolysis were detected in GM-CSF-induced M-MDSCs (white bars) and GM-CSF + RPM-induced M-MDSCs (yellow bars) by real-time PCR. (H) Some genes involved in the glycolysis process were detected in M-MDSCs isolated from spleens of control mice, RPM-treated mice, alloskin-grafted mice and RPM-treated alloskin-grafted mice at day 7 after skin grafting. MDSCs were induced from bone marrow cells after 4 days of culture in the presence of 40 ng/ml recombinant mouse GM-CSF, GM-CSF + 1 μM RPM or GM-CSF + 1.5 mM 2-DG. (I) Percentages and cell numbers of CD11b⁺ Ly6C^highLy6G⁻ M-MDSCs in these three groups were analyzed. The numbers in FACS plots represent the percentages of cells in the gates. (J) iNOS mRNA expressions of CD11b⁺ Ly6C^highLy6G⁻ M-MDSCs were determined by real-time PCR. (K) Sorted CD11b⁺ Ly6C^highLy6G⁻ M-MDSCs from different induction groups were assayed for their immunosuppressive ability. Results are expressed as mean ± SD (N = 4). More than two independent experiments showing similar results were done. **p < 0.01, and ***p < 0.001 for comparison between the indicated groups.