Fig. 6: T cell-specific loss of Chat causes alterations to Treg cells that are linked to compromised antitumor immunity.
From: Tumor-specific cholinergic CD4+ T lymphocytes guide immunosurveillance of hepatocellular carcinoma
a–c, Representative flow cytometry plots (a), quantification of the percentages of Foxp3+CD4+ T cells expressing CD25 (b) and the CD25 mean fluorescent intensity (MFI; c) in HCC-bearing livers from Chatfl/fl and Chatfl/fl; Cd4-cre mice on day 35 of HCC induction (n = 9 mice per group). P values were determined by unpaired, two-tailed t-test; data are representative of two independent experiments. d, Experimental protocol used to deplete CD25-expressing Treg cells during HCC induction. e,f, Representative low-magnification histological images of H&E-stained sections (e) and quantification of numbers of tumor nodules per mm2 in these sections (f) of livers from Chatfl/fl and Chatfl/fl; Cd4-cre mice treated as in d. In f, n = 7 mice in the Chatfl/fl + IgG, Chatfl/fl + anti-CD25 and Chatfl/fl; Cd4-cre + anti-CD25 groups, and n = 9 mice in the Chatfl/fl + anti-CD25 group. P values were determined by two-way ANOVA with Tukey’s multiple comparisons tests. The ‘X’ symbols indicate animals that reached the humane endpoint before day 20. g, Experimental protocol used to deplete CD4+ T cells and NK cells during HCC induction. Each mouse received 100 μg of depleting antibody per injection. Mice in the control group received either 100 μg of rat IgG or PBS. h,i, Quantification of numbers of tumor nodules per mm2 in H&E-stained sections (h) and ratios of liver weight to body weight (i) of Chatfl/fl and Chatfl/fl; Cd4-cre mice treated as in g. The ‘X’ symbols indicate animals that reached the humane endpoint before day 20. In h, n = 8 mice in the Chatfl/fl + IgG/PBS and anti-CD4 groups; n = 7 mice in the Chatfl/fl; Cd4-cre + IgG/PBS group; n = 9 mice in the Chatfl/fl + anti-NK1.1 group; and n = 5 mice in the Chatfl/fl; Cd4-cre + anti-NK1.1 group. In i, n = 8 mice in the IgG/PBS and anti-CD4 groups; n = 9 mice in the Chatfl/fl + anti-NK1.1 group; and n = 5 mice in the Chatfl/fl; Cd4-cre + anti-NK1.1 group. In b, c, f, h and i, each dot represents an individual mouse. Data are shown as means ± s.e.m. P values in h were determined by paired, two-tailed t-tests. P values in i were determined by two-tailed Mann–Whitney test because the Chatfl/fl group did not pass the normality test; NS, not significant.
