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. 2012 Sep 12;7(9):e43609. doi: 10.1371/journal.pone.0043609

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This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration, which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose.

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Panels A–F show Treg suppression of IL-2-secreting cells in primary MLR cultures. MLR cultures of DO11.10 CD4⁺CD25⁻ cells and allogeneic DCs from WT or IL-2^−/− KO littermates were incubated overnight with or without DO11.10 Tregs, washed, and transferred to IL-2-ELISPOT plates to measure IL-2-secreting cells as per materials and methods. A&B: Tregs fail to suppress MLR-IL-2 response in cultures containing IL-2^−/− KO DCs. Bars are mean ± SE IL-2 producing cells/culture from N = 5 independent experiments with splenic DCs (A) and N = 3–4 independent experiments with BMDCs (B). * indicates p<.001 (A) and p<.005 (B) compared with all other conditions, Holm-Sidak method of multiple pairwise comparison. C&D: Anti-CD25 blocks Treg suppression of MLR-IL-2 response. MLR cultures of DO11.10 CD4⁺CD25⁻ cells and allogeneic WT splenic DCs (C) or BMDCs (D), with or without Treg, were treated with anti-CD25 or isotype control antibody, washed, then transferred to IL-2-ELISPOT plates as in material and methods. C: Symbols are mean ± SE IL-2 producing cells/culture from N = 6 independent experiments. * indicates p<.001, anti-CD25 versus isotype control, Holm-Sidak method of multiple pairwise comparison. D: Symbols are mean ± SE IL-2-producing cells of triplicate cultures from one of four representative experiments. * indicates p<.001, Treg treatment groups anti-CD25 versus isotype control, Tukey's test for all pairwise multiple comparison. E&F: Exogenous IL-2 restores Treg suppression of MLR cultures containing IL-2KO DCs. MLR cultures of DO11.10 CD4⁺CD25⁻ cells and allogeneic IL-2^−/− KO splenic DCs (E) or BMDCs (F), with or without Treg, were treated with exogenous IL-2, washed, then transferred to IL-2 ELISPOT plates as in material and methods. Symbols are mean ± SE IL-2 producing cells of triplicate cultures from one of three representative experiments. * indicates p<.001, IL-2 treatment DC/CD4eff + Treg versus DC/CD4eff control, Tukey's test for all pairwise multiple comparison.