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. Author manuscript; available in PMC: 2017 Mar 1.
Published in final edited form as: J Immunol. 2016 Jan 25;196(5):2167–2180. doi: 10.4049/jimmunol.1501853

Figure 2. Particulate β-glucan treatment in vitro subverts splenic MDSC-mediated T cell suppression.

Figure 2

(A) CFSE-labeled OT-II splenocytes co-cultured with sorted M-MDSC or PMN-MDSC from the spleens of LLC-bearing mice in the presence or absence of particulate β-glucan (100 µg/ml in the M-MDSC cultures and 50 µg/ml in the PMN-MDSC cultures) and OVA (100 µg/ml) for 3 days at 1:1 ratio. Data represent the percentage of CFSE diluted cells gated on CD4+ T cells. The experiment was repeated two times with similar results. (B) CFSE-labeled OT-I splenocytes were co-cultured with sorted M-MDSC or PMN-MDSC from the spleens of LLC-bearing mice in the presence of OVA (50 µg/ml) and particulate β-glucan (50 µg/ml) for 3 days at 1:1 ratio. Data represent the percentage of CFSE diluted cells gated on CD8+ T cells. Data is representative of three independent experiments. (C) OT-II splenocytes co-cultured with sorted M-MDSC or PMN-MDSC from the spleens of LLC-bearing mice in the presence of OVA (100 µg/ml) with or without particulate β-glucan (50 µg/ml) for 3–4 days at 1:1 ratio and stimulated with PMA/Ionomycin for intracellular IFN-γ staining. Data represent the percentage of IFN-γ+ cells gated on CD4+ T cells. The experiment was repeated 3 times with similar results. (D) CFSE-labeled OT-I splenocytes co-cultured with sorted M-MDSC or PMN-MDSC from the spleens of LLC-bearing mice in the presence of OVA (50 µg/ml in M-MDSC cultures and 10 µg/ml in PMN-MDSC cultures) and particulate β-glucan (50 µg/ml) for 3 days at 1:1 ratio and then stimulated with PMA/Ionomycin for intracellular IFN-γ staining. Data represent the percentage of IFN-γ+ CFSE diluted cells gated on CD8+ T cells. Results are representative of three independent experiments. (E) CFSE-labeled splenocytes from C57BL/6 mice stimulated with plate-bound anti-CD3 (5 µg/ml) and soluble anti-CD28 (2 µg/ml) (white bar) and co-cultured with sorted splenic M-MDSC or PMN-MDSC from LLC-bearing mice for 3 days with (black bar) or without particulate β-glucan (50 µg/ml) (grey bar). Data represent the frequency of CFSE diluted cells gated on CD4+ or CD8+ T cells. The experiment was repeated twice with similar results. * p<0.05, **p<0.01, ***p<0.001.

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