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. 2020 Apr 9;10:6156. doi: 10.1038/s41598-020-62881-4

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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LL-CFA/I maintains suppressive Tregs in the PaLNs at 17 weeks. Four wk-old NOD females were orally dosed with 5 × 10⁷ CFUs of LL-CFA/I, LL vector, or PBS per the treatment regimen described in Fig. 5. (A,B) At 17 wks, one wk after their final dose, purified PaLN lymphocytes were stimulated with anti-CD3 and anti-CD28 mAbs to assess by flow cytometry (A) Foxp3⁺CD4⁺ Tregs and co-expression of (B) IFN-γ and Tbet. Expression of RoRγt and GATA3 by splenic (C,D) PaLN CD4⁺ T cells was also measured. (E) Staining was performed on unstimulated diabetogenic PaLN T cells to detect tetramer-specific GAD:206-220 and InsB:9-23 CD4⁺ T cells, and tetramer-specific InsB:15-23 CD8⁺ T cells. Data are represented as means ± SEM; *p < 0.05 for LL-CFA/I vs PBS; ^#p < 0.05 for LL-CFA/I vs LL vector.