Figure 3. Flow cytometry analysis of spleen and ILN T cells at day 56 post-induction of γHV68-CIA and control CIA mice.

(A–D) Representative flow cytometry plots of spleen samples previously gated on lymphocytes, live cells, singlets, CD45⁺CD3⁺ cells, and (A, D) CD4⁺ cells, (C) CD8⁺ cells, of uninfected mice with collagen-induced arthritis (CIA) (upper plot) and γHV68-CIA mice (lower plot). (A, C) Side-scatter (SSC) plotted on the y-axis. (A–D) Percent of immune subsets (y-axis) in the spleens of uninfected mice with CIA (filled circles) and γHV68-CIA mice (filled squares). (A) %FoxP3⁺ of CD4⁺; (B) %CD3⁺CD8⁺ and %CD3⁺CD4⁺ of CD45⁺; (C) %IFNγ⁺ of CD8⁺; (D) IFNγ⁺ or IL17A⁺ of CD4⁺; (E) RNA extracted from inguinal lymph node (ILN) cells, real-time quantitative PCR (RT-qPCR) performed for Ifng and Il17a, and relative expression plotted for uninfected mice with CIA (filled circles) and γHV68-CIA mice (filled squares). (A) n = 12–17 mice per group, three experiments; (B) n = 14–17 mice per group, three experiments; (C, D) n = 19–22 mice per group, three experiments; (E) n = 4–8 mice per group, one experiment. (A–E) Each data point represents an individual mouse. Data presented as mean ± SEM, analyzed by Mann-Whitney test; ****p<0.0001, ***p<0.001, **p<0.01, *p<0.05.

Figure 3—figure supplement 1. Total numbers of spleen and inguinal lymph node T cell populations at day 56 post-CIA induction.

(A–J) Total numbers of T cell and dendritic cell subsets (y-axis) in the spleen and inguinal lymph nodes (ILNs), as determined by flow cytometry in uninfected mice with collagen-induced arthritis (CIA) (filled circles) and γHV68-CIA mice (filled squares). (A) Total number of cells in the ILN, counted on hemocytometer, (B) total number of CD4⁺FoxP3⁺ cells in spleen, (C) total number of CD3⁺CD8⁺ cells in spleen, and (D) total number of CD3⁺CD4⁺ cells in spleen. (E) Total number of CD4⁺FoxP3⁺ cells in ILN, (F) total number of CD3⁺CD8⁺ cells in ILN, and (G) total number of CD3⁺CD4⁺ cells in ILN. Proportion of CD11c⁺CD8⁺ cells of CD45⁺CD3^- in the (H) spleen and (I) ILN. (J) Proportion of CD11c⁺CD11b⁺ of CD45⁺CD19^-CD3^- in the spleen. (A) n = 4–17 mice per group, three experiments; (B–G) n = 8–9 mice per group, one experiment; (H) n = 16–17 mice per group, three experiments; (I) n = 12–14 mice per group, two experiments; (J) n = 12 mice per group, two experiments. (A–J) Each data point represents an individual mouse. Data presented as mean ± SEM, analyzed by (A) one-way ANOVA, (B–J) Mann-Whitney test; ****p<0.0001, ***p<0.001, **p<0.01, *p<0.05.