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. 2020 Oct 5;130(11):5893–5908. doi: 10.1172/JCI136466

Figure 8. ROS is important for Treg aging and functional deterioration.

Figure 8

(A and B) Flow cytometry of ROS levels in aged Tregs (A) and Dcaf1-deficient (CD4-Cre Dcaf1fl/fl) Tregs (B) in the absence (–) or presence (+) of NAC (20 mM) or GSH (10 mM) (blank, no DCFDA; n = 3 mice of 3 experiments; representative results are shown; means ± SD, ****P < 0.0001, by 1-way ANOVA followed by Tukey’s multiple-comparisons test). (C) Proliferation assayed by BrdU incorporation in aged Tregs in the absence (–) or presence (+) of NAC (20 mM) or GSH (10 mM) (n = 4 mice of 4 experiments; representative results are shown; means ± SD, ****P < 0.0001, by 1-way ANOVA followed by Tukey’s multiple-comparisons test). (D) Proliferation assayed by BrdU incorporation in Dcaf1-deficient (CD4-Cre Dcaf1fl/fl) Tregs in the absence (–) or presence (+) of NAC (20 mM) or GSH (10 mM) (n = 4 mice of 4 experiments; representative results are shown; means ± SD, ****P < 0.0001, by 1-way ANOVA followed by Tukey’s multiple-comparisons test). (E) Suppressive activity of aged Tregs without (–) or with (+) pretreatment of NAC (20 mM) or GSH (10 mM), assessed by in vitro suppression assay (n = 3 mice of 3 experiments; representative results are shown; means ± SD, ****P < 0.0001, by 1-way ANOVA followed by Tukey’s multiple-comparisons test). (F) Suppressive activity of Dcaf1-deficient (CD4-Cre Dcaf1fl/fl) Tregs without (–) or with (+) pretreatment of NAC (20 mM) or GSH (10 mM), assessed by in vitro suppression assays (n = 3 mice of 3 experiments; representative results are shown; means ± SD, ***P < 0.001, by 1-way ANOVA followed by Tukey’s multiple-comparisons test).