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. 2020 Jan 9;10:2945. doi: 10.3389/fimmu.2019.02945

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Copyright © 2020 Gan, Zhang, Gu, Ju, Wu, Wang, Peng, Qiu, Zhou, Cheng and Lu.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Reversal of acidic microenvironment restores Foxp3 expression and iTreg function. (A) The proportion of CD4⁺CD25⁺Foxp3⁺ iTregs cultured with IL-2 and TGF-β for 3 days. (B,C) The proportion of CD4⁺IL-10⁺ and CD4⁺ TGF-β⁺ T cells cultured with IL-2 and TGF-β for 3 days and stimulated with phorbol 12-myristate 13-acetate, brefeldin A, and ionomycin for 6 h before the assay. (D) Statistical analysis of the suppression assay in vitro. (E) iTregs induced in media of varying pH values for 3 days were co-cultured with carboxyfluorescein succinimidyl ester-stained human naïve CD4⁺ T cells (responders) at the indicated ratio. After 72 h of activation with anti-CD3/CD28-conjugated beads, responder cell proliferation was assessed by flow cytometry. (F–H) Statistical analysis of related results of flow cytometry. Data are presented as the means ± SD from three independent experiments. ***p < 0.001.