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. 2018 Dec 17;9:2953. doi: 10.3389/fimmu.2018.02953

Figure 7.

Figure 7

Functional improvement of CMV-specific T cells treated with sirolimus strongly depends on IL-2R-driven STAT-5 signaling. (A) Percentages (%) of CMV-specific multimer+CD8+ T cells after 7 days of aAPC stimulation with or without sirolimus in the presence of the following cytokines: IL-7, IL-12, IL-15, IL-21 (each 10 ng/ml) which were added independently as different sets of cell cultures. Results for supplemental IL-2 (50 U/ml) without sirolimus (black dashed lines) and with sirolimus (gray dashed lines) are shown (n = 4). (B) Percentages of IFN-γ following 5 h peptide re-stimulation, as determined by intracellular cytokine staining on CMV-specific multimer+CD8+ T cells (n ≥ 3). (C) Percentage of expanded CMV-specific multimer+CD8+ T cells following overnight STAT-5 inhibition (STAT-5i) on day 7. (D) Percentages of pS6 and pSTAT-5, as determined by phospho-flow cytometry or (E) fluorescence microscopy on CMV-specific CD8+ T cells after 1 h of peptide re-stimulation. (F) Intracellular expression of IFN-γ (%) following 5 h of peptide re-stimulation, as measured by multicolor flow cytometry. Values are displayed as mean (±) SD. Statistical analysis: (A,B) Student's t-test and (C,D,F) two-way analysis of variance (*p < 0.05, **p < 0.01, ***p < 0.001).