IL35 regulation of tumor growth is accompanied by suppression of CD4+ effector T-cell activity and expansion of Tregs. IL35 regulation of tumor growth.

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IL35 regulation of tumor growth is accompanied by suppression of CD4+ effector T-cell activity and expansion of Tregs. IL35 regulation of tumor growth is accompanied by suppression of CD4+ effector T-cell activity and expansion of Tregs. A, Representative images of pancreatic tumors from WT, Il10−/−, p35−/−, and Ebi3−/− mice orthotopically injected with KPC 4662 cells and collected at 3 weeks after injection of tumor cells. B, Quantification of tumor weights from WT, Il10−/−, p35−/−, and Ebi3−/− (N = 12; n = 3/group) mice. C, Quantification of tumor weights from WT (n = 9), Il10−/− (n = 9), and p35−/− (n = 9) mice orthotopically injected with KPC 2173 cells and collected 3 weeks after tumor cell injection. D, Representative flow cytometry plots of gating strategy for identifying CD4+ T cells and frequency of intratumoral CD4+CD25− effector T cells isolated from WT, Il10−/−, p35−/−, and Ebi3−/− mice orthotopically injected with KPC 4662 cells and collected 3 weeks after injection of tumor cells. Percentage of CD45+CD4+ cells is indicated. E, Quantification of the frequency of CD45+CD4+ T cells from WT, Il10−/−, p35−/−, and Ebi3−/− (n = 3/group) mice. F, Representative flow-cytometric plots of CD4+IFNγ+ and CD4+TNFα+ T cells isolated from WT, Il10−/−, p35−/−, and Ebi3−/− mice orthotopically injected with KPC 4662 cells and collected 3 weeks after injection of tumor cells. G, Quantification of the frequency of CD4+IFNγ+ T cells (left graph, % of CD4+ T cells) and CD4+TNFα+ T cells (right graph, % of CD4+ T cells) from WT, Il10−/−, p35−/−, and Ebi3−/− (n = 3/group) mice. H, Representative flow-cytometric plots of intratumoral CD4+Foxp3+ Tregs isolated from WT, Il10−/−, p35−/−, and Ebi3−/− mice. I, Quantification of the frequency of CD4+Foxp3+ (% of CD4+ T cells) from WT, Il10−/−, p35−/−, and Ebi3−/− (n = 3/group) mice. J, Mean CD4+ effector T cell to Treg ratio was calculated based on the percentage of positive lymphocyte population determined by flow cytometry. Error bars, SEM. P values were calculated using Student t test (unpaired, two-tailed); NS, not significant; *, P < 0.05; **, P < 0.01; ***, P < 0.001. Data represent 3–4 independent experiments. Bhalchandra Mirlekar et al. Cancer Immunol Res 2018;6:1014-1024 ©2018 by American Association for Cancer Research