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. 2015 Oct 20;125(11):4212–4222. doi: 10.1172/JCI81151

Figure 2. High-salt conditions directly influence Treg function.

Figure 2

(A) Tregs were activated with plate-bound αCD3 and soluble αCD28 at 1 μg/ml either in normal media (media, white) or media enriched with 40 mM NaCl (gray) for 72 hours with IL-2 (25 U/ml). Following incubation, Tregs were washed, counted, and plated in a suppression assay in normal media with freshly sorted allogeneic CD4 effector cells stained with CFSE. CFSE dilution was measured via flow cytometry after 5 days. (B) Line graph depicts a summary of experiments (n = 5). At onset of preactivation and following 72-hour preactivation, an aliquot of Tregs was analyzed. (C and D) We confirmed that Foxp3 expression (C) (n = 5) and percent demethylation of the TSDR (D) (n = 3) did not change with activation and that these are human Tregs. Statistical analyses were performed using paired Student’s t test.