Fig. 5
From: Tumor-released autophagosomes induces CD4+ T cell-mediated immunosuppression via a TLR2–IL-6 cascade
IL-6/IL-10 from TTRAP is responsible for tumor growth and metastasis. a Flow cytometric and statistical analyses of the percentage of IFN-γ+ CD4+ and CD8+ T cells treated with the supernatants from TTRAP (SN/TTRAP) or control CD4+ T cells for 3 d in the presence of anti-CD3/CD28. b SN/TTRAP was pretreated with anti-IL-6, IL-10, IL-21 neutralizing antibodies respectively, and then co-cultured with the purified CD4+ T cells and CD8+ T cells in the presence of anti-CD3/CD28 for 3 d. The percentages of IFN-γ+ CD4+ T cells and CD8+ T cells were evaluated by flow cytometry. c C57BL/6 mice were adoptively transferred i.v. with OT-I spleen cells and then vaccinated s.c. with OVA-loaded DC on day 1, 4 and 7, following injection i.v. with TTRAP or control CD4+ T cells on day 2, 5 and 8. On the 15th day, the frequencies and the number of Vβ5.1+CD8+ T cells in spleen were analyzed by flow cytometry. d C57BL/6 mice were vaccinated with OVA-loaded DC and following adoptively transferred with TTRAP or CD4+ T cells. On the 15th day, the splenocytes were re-stimulated with OVA-protein for 24 h, and the frequencies of IFN-γ+ T cells were determined by flow cytometry. e B16F10 tumor cells were mixed with TTRAP or control CD4+ T cells and injected s.c. into C57BL/6 mice (n = 6 per group). The growth of the tumor was monitored. f B16F10 tumor cells were intravenously injected into C57BL/6 mice (n = 4 to 6 per group) to establish a lung metastasis model. Subsequently, TTRAP or control CD4+ T cells were adoptively transferred i.v. 3 times with 1 d of interval. Three weeks later, the tumor nodules in the lungs were examined. Data (mean ± SEM) represent 3 independent experiments. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, not significant, by 1-way ANOVA with the Tukey-Kramer multiple test, 2-tailed unpaired t-test or Mann-Whitney U test