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. 2022 Feb 18;10(4):437–452. doi: 10.1158/2326-6066.CIR-20-0973

Figure 2.

Figure 2. Cbl-b KO CD8+ T cells display resistance to Treg-mediated suppression. A, Proliferation of WT and Cbl-b KO CD8+ T cells in the Treg-suppression assay, analyzed on day 3 after stimulation (n = 3). B and C, The percentage of suppression (B) and CD25 expression (C) of WT and Cbl-b KO CD8+ at different Teff to Treg ratios (n = 3). D, Intracellular expression of IFNγ and IL2 by WT and Cbl-b KO CD8+ effector T cells in the presence of WT Tregs. After 24 hours of coculture, T cells were restimulated using PMA/ionomycin and Golgi block and were stained for CD8, IFNγ, and IL2 for flow cytometry. E, Cytokine secretion by WT and Cbl-b KO CD8+ T cells in the presence of Tregs. Supernatants from stimulated WT and Cbl-b KO CD8+ T cells in the presence (or absence) of Tregs were collected on days 1 and 3 after stimulation (n = 3). ELISAs were performed to assess secretion of IFNγ, IL2, and TNFα by CD8+ T cells. Statistical analyses were performed using repeated measure (B and C) or nonrepeated (E) two-way ANOVA with Holm–Sidak test (*, P < 0. 05; **, P < 0.01; ns, not significant).

Cbl-b KO CD8+ T cells display resistance to Treg-mediated suppression. A, Proliferation of WT and Cbl-b KO CD8+ T cells in the Treg-suppression assay, analyzed on day 3 after stimulation (n = 3). B and C, The percentage of suppression (B) and CD25 expression (C) of WT and Cbl-b KO CD8+ at different Teff to Treg ratios (n = 3). D, Intracellular expression of IFNγ and IL2 by WT and Cbl-b KO CD8+ effector T cells in the presence of WT Tregs. After 24 hours of coculture, T cells were restimulated using PMA/ionomycin and Golgi block and were stained for CD8, IFNγ, and IL2 for flow cytometry. E, Cytokine secretion by WT and Cbl-b KO CD8+ T cells in the presence of Tregs. Supernatants from stimulated WT and Cbl-b KO CD8+ T cells in the presence (or absence) of Tregs were collected on days 1 and 3 after stimulation (n = 3). ELISAs were performed to assess secretion of IFNγ, IL2, and TNFα by CD8+ T cells. Statistical analyses were performed using repeated measure (B and C) or nonrepeated (E) two-way ANOVA with Holm–Sidak test (*, P < 0. 05; **, P < 0.01; ns, not significant).