The small intestine epithelium exempts Foxp3+ Tregs from their IL-2 requirement for homeostasis and effector function
Praveen Prakhar, Jaylene Alvarez-DelValle, Hilary Keller, Assiatu Crossman, Xuguang Tai, Yoo Kyoung Park, Jung-Hyun Park
Praveen Prakhar, Jaylene Alvarez-DelValle, Hilary Keller, Assiatu Crossman, Xuguang Tai, Yoo Kyoung Park, Jung-Hyun Park
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Research Article Immunology

The small intestine epithelium exempts Foxp3+ Tregs from their IL-2 requirement for homeostasis and effector function

Abstract

Foxp3+ Tregs are potent immunosuppressive CD4+ T cells that are critical to maintain immune quiescence and prevent autoimmunity. Both the generation and maintenance of Foxp3+ Tregs depend on the cytokine IL-2. Hence, the expression of the IL-2 receptor α-chain (CD25) is not only considered a specific marker, but also a nonredundant requirement for Tregs. Here, we report that Foxp3+ Tregs in the small intestine (SI) epithelium, a critical barrier tissue, are exempt from such an IL-2 requirement, since they had dramatically downregulated CD25 expression, showed minimal STAT5 phosphorylation ex vivo, and were unable to respond to IL-2 in vitro. Nonetheless, SI epithelial Tregs survived and were present at the same frequency as in other lymphoid organs, and they retained potent suppressor function that was associated with high levels of CTLA-4 expression and the production of copious amounts of IL-10. Moreover, adoptive transfer experiments of Foxp3+ Tregs revealed that such IL-2–independent survival and effector functions were imposed by the SI epithelial tissue, suggesting that tissue adaptation is a mechanism that tailors the effector function and survival requirements of Foxp3+ Tregs specific to the tissue environment.

Authors

Praveen Prakhar, Jaylene Alvarez-DelValle, Hilary Keller, Assiatu Crossman, Xuguang Tai, Yoo Kyoung Park, Jung-Hyun Park

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Figure 2

Phenotypic and functional characterization of SI IEL Foxp3+ Tregs.

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Phenotypic and functional characterization of SI IEL Foxp3+ Tregs. (A) C...
(A) Cell numbers of SI IEL Foxp3+ Tregs isolated from Foxp3-GFP reporter mice by Percoll enrichment or by EpCAM+ cell depletion. The graph shows the summary of 2 independent experiments with a total of 3 Foxp3-GFP reporter mice. (B) Surface expression (left) and ΔMFI (right) of CD25 on Foxp3-GFP+CD4+TCRβ+ cells from LN and IELs. The histogram is representative, and the graph shows the summary of 2 independent experiments with a total of 4 Foxp3-GFP reporter mice. (C) Intracellular pSTAT5 in freshly isolated Foxp3+ Tregs of LN and SI IELs. The histogram is representative (left), and the graph is a summary (right), of 3 independent experiments with a total of 4 WT mice. (D) IL-2–induced STAT5 phosphorylation in Foxp3+ Tregs of LN and SI IELs. Freshly isolated lymphocytes of LN and SI IELs were stimulated with recombinant IL-2 (1 ng/mL) for 30 minutes and then assessed for pSTAT5 contents gated on Foxp3+ Tregs. Histogram is representative (top), and the fold increase in pSTAT5 (bottom graph) is the summary, of 2 independent experiments with a total of 4 WT mice. (E) Surface expression (top) and ΔMFI (bottom) of CD39 and CD73 on Foxp3-GFP+CD4+ T cells from LN and IELs. Histograms are representative (top), and the graphs show the summary (bottom), of 2 independent experiments with a total of 6 Foxp3-GFP reporter mice. (F) Intracellular staining for CTLA-4 in Foxp3-GFP+CD4+ T cells from LN and IELs. Histograms are representative (top), and the ΔMFI graphs show the summary (bottom), from 2 independent experiments with a total of 6 Foxp3-GFP reporter mice. (G) Surface expression of CTLA-4 on Foxp3-GFP+CD4+ T cells from LN and IELs. Histogram is representative (top), and the ΔMFI graph shows the summary (bottom), from 2 independent experiments with a total of 6 Foxp3-GFP reporter mice. (H) Surface expression of PD-1 and LAG-3 on Foxp3-GFP+CD4+ T cells from LN and IELs. Histograms are representative (top), and the ΔMFI graphs show the summary (bottom), from 2 independent experiments with a total of 6 Foxp3-GFP reporter mice. (I) Surface expression (top) and ΔMFI (bottom) of GITR and Neuropilin-1 on Foxp3-GFP+CD4+ T cells from LN and IELs. Histograms are representative (top), and the graph show the summary (bottom), of 2 independent experiments with a total of 6 Foxp3-GFP reporter mice. The data are represented as the mean ± SEM. P values were determined by paired Student’s t test. *P < 0.05, **P < 0.01, and ***P < 0.001.