Fig. 6
CD26high T cells have stemness and increased migratory capacity. a, b Tumors from treated PANC1-bearing mice (from Fig. 5) were harvested and then frozen in cryomatrix. Tumor samples were then sliced and used for immunohistochemistry analysis (purple = H&E, brown = CD45; N = 5–9 tumors per group). Magnification = ×10. In b, the integrated optical density (IOD) of CD45 in CD26-sorted groups was quantified with ImageJ software and graphed (N = 5–8 per group). c Sorted T cells were activated with CD3/ICOS beads and expanded in 100 IU/ml IL-2 for 10 days prior to testing cell migration via a transwell assay. 0.75e6 sorted cells were re-suspended and assessed for percent cell migration towards M108 or PANC1 supernatant in a 2 h time period (N = 5). d, e Sorted T cells were analyzed for chemokine receptor expression by flow cytometry (d N = 12–15) and gene array (e N = 3–5) prior to bead stimulation. In e, data shown in heat map as (+/−) log2-fold change in chemokine receptor expression compared to bulk CD4+. f Viability of T cells that migrated towards PANC1 was determined by live/dead staining (N = 3). g Anti-apoptotic and stemness genes were analyzed and displayed as a (+/−) log2-fold change compared to bulk CD4+ (N = 3–5). h–j Protein from pre-activation (day 0) and post-activation (day 10) cells was isolated and used for western blot analysis (N = 2–3). In j, the fold change of β-catenin, BCL2 and cleaved Caspase-3 for each subset compared to bulk CD4+ T cells was graphed (N = 2–3). P values were calculated using a One-way ANOVA with a Kruskal-Wallis comparison. Data with error bars represent mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001
