Fig. 3
α-GalCer increases the frequency of NKT cells, IFN-γ secretion, and inhibits tumor growth. Naïve C57BL6 mice were given s.c. injection of B16F10 cells (1 X 106 cells/mouse), and animals were also given i.v. injection of NK1.1 mAb (PK136; 100 μg/mouse/injection) on day − 3, + 1, + 5, + 10 and + 15 (day with respect to tumor cell injection). α-GalCer (2 μg/mouse/i.p injection) was given on day + 1, + 5, + 10, + 15 and + 20. a The tumor area was calculated and plotted. n = 6 mice/group. The data shown are representative of two independent experiments. b On day 13, spleen and tumor tissues from α-GalCer-treated mice were stained with TCR-β, α-GalCer-loaded CD1d-tetramer and nuclear stain DAPI. Images were acquired using a fluorescent microscope, and the representative images of spleen and tumor from α-GalCer treated mice are shown (magnification 200X). c On day 13, NKT cells in the spleen and tumor were analyzed using flow cytometry. Representative contour plots are shown (left panel) and mean percentages of NKT cells are plotted (middle panel). The absolute number of cells were calculated and plotted (right panel) n = 5 mice/group. d On day 13, IFN-γ, IL-4 and IL-17A expression in the NKT cells in the spleen were analyzed and plotted. n = 5–6 mice/group. e On day 13, based on CD62L and CD44 expression, memory NKT cell subsets were analyzed and plotted. n = 5–6 mice/group. f On day 23, the mean percentages of NKT cells and IFN-γ+-producing NKT cells in the spleen were analyzed. The bar represents mean, and each dot represents an individual mouse. n = 4–5 mice/group. (b-e). One-way ANOVA (a), Student’s t-test (c-e), *p < 0.05; ** p < 0.01; ns, not significant