Fig. 6: Analysis of antigen-specific CD4 T cells from the liver and the spleen of an in vivo non-TCR-transgenic mouse model.
A Experimental design for investigation of immune response against the HA antigen in the liver. HA/iCre mice received either tamoxifen treatment (n = 9), HA immunization treatment (HA i.m; n = 15), or HA immunization followed 2 weeks later by tamoxifen treatment (HA i.m + Tamoxifen; n = 12). Mice were euthanized 2 weeks after the last treatment. B Relative HA mRNA expression in the liver. ACTB was used as loading control. C Analysis of normalized anti-HA antibody rate in serum of mice. D Analysis of HA-tetramer-specific CD4 T cells in the spleen (top) and in the liver (bottom). Contour plot representation of HA tetramer staining and CD44 expression in CD4 T cells from HA i.m and HA i.m + Tamoxifen mice (left). Frequency of HA-tetramer-specific memory (CD44high) CD4 T cells per million cells (right). Dotted lines in the frequency graphs represent threshold of positive detection of HA-specific CD4 T cell population. E Analysis of PD-1 expression in detectable HA-specific memory CD4 T cells from the spleen and the liver of HA i.m and HA i.m + Tamoxifen mice. F Principal component analysis (PCA) of bulk-RNA-seq transcriptome of HA-specific CD4 T cells isolated from the liver or the spleen. G Gene expression heatmap of HA-specific CD4 T cells. H Volcano plot of gene expression change between liver and spleen HA-specific CD4 T cells. I Genes from GO enrichment pathways upregulated and shared between mouse liver HA-specific CD4 T cells and human autoreactive CD4 T cells. Data are presented as mean values ± SD in graphs (B, C, and E). Dunn’s multiple comparisons test was used for (B–D). Two-sided Mann-Whitney test was used for (E). p-values and adjusted p-values (multiple comparisons) are indicated. Source data are provided as a Source Data file.
