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. 2018 May 31;9:1235. doi: 10.3389/fimmu.2018.01235

Figure 4.

Figure 4

RhoA expression is important for pan T-cell responses. (A) Representative histogram of the proliferation of RhoA+/+ and RhoA−/− T-cells in vitro based on their CFSE labeling. (B) Proliferation index of RhoA+/+ (filled bars) and RhoA−/− (striped bars) T-cells upon pan-activation with anti-CD3/CD28 antibodies for 72 h. Data represent the mean of two independent experiments, n = 3 mice per group. (C) Representative gating strategy for the analysis of cell activation. (D) Quantification of the percentage of CD4+CD25+RhoA+/+ and RhoA−/− T-cells in the presence or absence of pan-activation. Data represent the mean of two independent experiments, n = 3 mice per group. (E) Representative gating strategy for the analysis of cell viability. (F) Quantification of the percentage of necrotic (N), apoptotic (EA + LA) and living (L) cells in RhoA+/+ and RhoA−/− T-cells after 24 h of pan-activation in vitro. Data represent the mean of three independent experiments, n = 4 mice per group. (G–L) Amount of IFN-γ, IL-6, TNF-α, IL-4, IL-5, and IL-17A present in the supernatant of RhoA+/+ and RhoA−/− T-cells after 24 h stimulation with anti-CD3/CD28. Data shown as mean ± SEM *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 [(B) Two-way ANOVA, followed by Tukey’s test. (D) One-way ANOVA, followed by Tukey’s test. (F) Multiple t-test, followed by FDR correction. (G–K) Unpaired t-test]. Abbreviation: FSC, forward scatter; SSC, side scatter; PI, propidium iodide; FLICA, fluorescent inhibitor of caspases; L, living; N, necrotic; EA, early apoptotic; LA, late apoptotic; FDR, false discovery rate.