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. 2016 Dec 8;1(20):e89289. doi: 10.1172/jci.insight.89289

Figure 5. Proliferation of tumor-resident T cells is independent of TCR signaling but partially dependent on IL-15 signaling.

Figure 5

(A) Schematic of experiments to address TCR dependence of the proliferation of tumor-resident T cells. Genetically marked OT-I T cells expressing either a single copy of wild-type ZAP70 or the HXJ42-sensitive mutant ZAP70AS were transferred to PyMT-ChOVA mice. Fourteen days later, tumor slice cultures were prepared from tumors and incubated for 24 hours in the presence of EdU and either HXJ42 inhibitor or DMSO control. (B) Representative analysis of EdU incorporation of inhibitor-resistant OT-I/ZAP70+/– or inhibitor-sensitive OT-I/ZAP70AS T cells in the presence or absence of the inhibitor using flow cytometry. (C) The ratio of proliferation in tumor slices incubated with HXJ42 inhibitor over the proliferation in control slices was analyzed using a Mann-Whitney test. P > 0.05. Data are pooled from 3 experiments. (D) Similar to A–C, tumor slices of PyMT-ChOVA mice treated with OT-I T cells were incubated with EdU in absence or presence of an antibody that blocks the MHCI-presented ovalbumin peptide (anti-SL8). EdU incorporation was compared against the average of the control group using a Mann-Whitney test. P > 0.05. Data are pooled from 3 experiments. (E) Analysis of homeostatic cytokine expression by myeloid cells in the tumor microenvironment. Cytokine RNA expression data from ref. 7 were mined for each indicated cell type and plotted using the indicated color scale and using Gapdh expression for normalization. (F) Schematic of experiments addressing the influence of IL-15 signaling on T cell proliferation in two tumor models. KI67+ OT-I cells served as a readout of cell proliferation. (G) Statistical analysis of KI67 expression of tumor-resident OT-I T cells in B78-ChOVA and PyMT-ChOVA tumors treated with IL-15-blocking antibody or control using a Mann-Whitney test. *P < 0.05. Data are pooled from 2 experiments. (H) Statistical analysis of KI67 expression of tumor-resident OT-I T cells in B78-ChOVA tumors grown in wild-type or IL-15-deficient animals using a Mann-Whitney test. ***P < 0.001. Data are pooled from 2 experiments.