Fig. 3

Evaluation of polyfunctional CD4⁺/CD8⁺ T cells and depression effects on BCG immunomodulation. A-D The mRNA expression of IFN-γ, IL12p40, IL1β and TNF-α cytokines was calculated in the spleen after euthanization in all four4 groups of mice. The significance is calculated compared with the control group. E Demonstrative gating of CD4⁺ and CD8 ⁺ cells of the indicated groups generated with CytExpert flow cytometry software. F-G Quantification of CD4⁺ and CD8⁺ frequencies in the spleens of all four groups of mice. H-I Gating images for phenotypic characterization of CD4⁺IFNγ⁺ and CD8⁺ IFNγ⁺ T cells. J-K The percentage of IFNγ⁺ cells in both CD4⁺ and CD8⁺ T cells was analyzed and depicted for all four groups of mice. Their significance was evaluated by comparison with the control group. L Descriptive images for gating both CD4⁺ central and effector memory immune cells. M–N Similarly, the relative percentage of CD4⁺ cells, including the phenotype of effector memory (CD25⁻CD44⁺CD62L⁻) and central memory (CD25⁻CD44⁺CD62L⁺) T cells, in the spleens of all four groups of mice was determined *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001; ****, p ≤ 0.0001 (one-way ANOVA). Data are representative of three independent experiments with three biological replicates (mean ± SEM)