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. 2021 Oct 26;12:746469. doi: 10.3389/fimmu.2021.746469

Figure 5.

Figure 5

All 3 DC subsets can present Class I, and II MHC restricted OVA peptide antigens, but only DC1b cells present glycolipid antigens. (A) Mean concentrations of IL-2 (Right), IL-13 (Right Middle), IL-4 (Left Middle), and IFNγ (Left) in culture supernatants of FACS-sorted DC subsets (DC1a, DC1b and DC2) (1x104 per well) from wild type untreated C57BL/6 mice that were cultured with OT-I restricted OVA257-264 peptide (1g/mL) and with CFSE-labelled OT-I CD8+ T-cells (1x105 cells per well). In control experiments, CFSE-labelled OT-I CD8+ T-cells (1x105 cells per well) were incubated with OVA257-264 peptide (1g/mL) but without DCs for 5 days. The concentrations of IL-4, IL-13, IL-2, IFN in culture supernatants was measured by Luminex. (B) Mean concentrations of IL-2 (Right), IL-13 (Right Middle), IL-4 (Left Middle), and IFNγ (Lef) in culture supernatants of FACS-sorted DC subsets (DC1a, DC1b and DC2) (1x104 per well) from wild type untreated C57BL/6 mice that were cultured with OT-II restricted OVA323-339 peptide (10g/mL) with CFSE-labelled OT-II CD4+ T-cells (1x105 cells per well). In control experiments, CFSE-labelled OT-II CD4+ T-cells (1x105 cells per well) were incubated with OVA323-339 peptide (10g/mL) without DCs for 5 days. The concentrations of IL-4, IL-13, IL-2, IFN in culture supernatants was measured by Luminex. All data are representative of three independent experiments (6 mice per experiment). (C, D) Mean concentrations of cytokines in cultures of sorted DC1a (Pop 1), DC1b (Pop 2) and DC2 (Pop 3) (15x103 cells/well) subsets incubated with sorted NKT cells (5x105 cells/well) from untreated wild type BALB/c spleen cells. AlphaGalcer (α-galactosylceramide) (100ng/mL) was added to all cultures in (C) but not in (D). Control cultures contained NKT cells with (C) or without (D) glycolipid but without DCs. Supernatants were harvested after 5 days, and concentrations were measured by Luminex. Data pooled from 2 or more independent experiments for a total of at least 10-12 mice per group, represented as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, NS, not significant (p > 0.05). One-way ANOVA (Holm-Sidak’s multiple comparison test) was used for statistical analysis.