NFAT5 dictates crosstalk between intestinal epithelial regenerative capacity and microbiota in murine colitis models
Se-Hyeon Park, Dae Hee Cheon, Yu-Mi Kim, Yeji Choi, Yong-Joon Cho, Bong-Ki Hong, Sang-Hyun Cho, Mi-Na Kweon, Hyug Moo Kwon, Eugene B. Chang, Donghyun Kim, Wan-Uk Kim
Se-Hyeon Park, Dae Hee Cheon, Yu-Mi Kim, Yeji Choi, Yong-Joon Cho, Bong-Ki Hong, Sang-Hyun Cho, Mi-Na Kweon, Hyug Moo Kwon, Eugene B. Chang, Donghyun Kim, Wan-Uk Kim
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Research Article Gastroenterology Immunology

NFAT5 dictates crosstalk between intestinal epithelial regenerative capacity and microbiota in murine colitis models

Abstract

Hypertonic and hyperosmolar stimuli frequently pose challenges to the intestinal tract. Therefore, a resilient epithelial barrier is essential for maintaining gut homeostasis in the presence of osmotic perturbations. Nuclear factor of activated T cells 5 (NFAT5), an osmosensitive transcription factor, primarily maintains cellular homeostasis under hypertonic conditions. However, the osmoprotective role of NFAT5 in enterocyte homeostasis is poorly understood. Here, we demonstrate that NFAT5 was critical for the survival and proliferation of intestinal epithelial cells (IECs) and that its deficiency accelerated chemically induced or spontaneous colitis in mice. Mechanistically, NFAT5 promoted the survival of IECs and the renewal of intestinal stem cells, thereby regulating the production of mucus and antimicrobial compounds, including RegIII and lysozyme, which consequently shape the gut microbial composition to prevent colitis. Transcriptome analysis identified HSP70 as a key downstream target of NFAT5 in epithelial regeneration. Loss- and gain-of-function experiments involving HSP70 revealed that NFAT5 mitigated experimental colitis through IEC Hsp70, which protected stem cells from inflammation-induced injury and maintained barrier function. In conclusion, our study demonstrates what we believe to be a previously unknown role for NFAT5 in dictating the crosstalk between intestinal stem cells and the microbiota, underscoring the importance of the NFAT5/HSP70 axis in maintaining epithelial regeneration related to gut barrier function, balancing microbial composition, and subsequently preventing colitis progression.

Authors

Se-Hyeon Park, Dae Hee Cheon, Yu-Mi Kim, Yeji Choi, Yong-Joon Cho, Bong-Ki Hong, Sang-Hyun Cho, Mi-Na Kweon, Hyug Moo Kwon, Eugene B. Chang, Donghyun Kim, Wan-Uk Kim

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Figure 6

NFAT5 facilitates mucin and antimicrobial compound production in goblet and Paneth cells by regulating epithelial regenerative capacity.

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NFAT5 facilitates mucin and antimicrobial compound production in goblet ...
(A) Mucin 2–expressing cells in the ileal tissues of separately housed Nfat5+/+ and Nfat5+/– mice were assessed by IHC staining. Representative images and the corresponding graph are presented. Scale bars: 50 μm. (B and C) Muc 2 mRNA expression was analyzed using qPCR in large IECs (L-IECs) and small IECs (S-IECs) from separately housed Nfat5+/+ and Nfat5+/– mice (B). Additionally, mRNA expression levels of Reg3g, Reg3b, Defa5, and Lyz1 were assessed in S-IECs from mice of both groups (C), with Gapdh mRNA serving as the internal control. (D) Lysozyme-expressing cells in the ileal tissues of separately housed Nfat5+/+ and Nfat5+/– mice were assessed. Representative images and their corresponding graph are presented. Scale bars: 100 μm. (E–J) Expression levels of mucin 2, lysozyme, LGR5, and OLFM were determined by qRT-PCR and immunostaining. Muc2 (E), Lyz1 (G), Lgr5, and Olfm4 (I) mRNA expression levels were measured in S-IECs of separately housed Nfat5fl/fl and Nfat5IEC-KO mice by qRT-PCR; Gapdh was used as an internal control for normalization. Mucin 2+ (F), lysozyme+ (H), and OLFM4+ (J) cells were analyzed in the ileal tissues of separately housed Nfat5fl/fl and Nfat5IEC-KO mice to assess their expression patterns. Representative staining images and the corresponding graphs are shown. Scale bars: 100 μm. (K and L) Organoids derived from small intestinal crypts from either separately housed (K) or cohoused (L) Nfat5fl/fl and Nfat5IEC-TG mice. Crypts isolated from a single mouse were seeded and cultured in Matrigel for 5 days. Representative images of organoid culture wells and the corresponding graphs are shown. Scale bars: 200 μm. Each dot in A–J represents an individual mouse, and the means are displayed as lines. Data in K and L are presented as box-and-whisker plots (minimum-to-maximum, line at the median). Data shown in A–L are representative of at least 2 independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001, by Mann-Whitney U test (A–J) and unpaired, 2-tailed Student’s t test (K and L).