Epithelial TNF controls cell differentiation and CFTR activity to maintain intestinal mucin homeostasis
Efren A. Reyes, … , Zev J. Gartner, Ophir D. Klein
Efren A. Reyes, … , Zev J. Gartner, Ophir D. Klein
Published August 29, 2023
Citation Information: J Clin Invest. 2023;133(20):e163591. https://doi.org/10.1172/JCI163591.
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Epithelial TNF controls cell differentiation and CFTR activity to maintain intestinal mucin homeostasis

Abstract

The gastrointestinal tract relies on the production, maturation, and transit of mucin to protect against pathogens and to lubricate the epithelial lining. Although the molecular and cellular mechanisms that regulate mucin production and movement are beginning to be understood, the upstream epithelial signals that contribute to mucin regulation remain unclear. Here, we report that the inflammatory cytokine tumor necrosis factor (TNF), generated by the epithelium, contributes to mucin homeostasis by regulating both cell differentiation and cystic fibrosis transmembrane conductance regulator (CFTR) activity. We used genetic mouse models and noninflamed samples from patients with inflammatory bowel disease (IBD) undergoing anti-TNF therapy to assess the effect of in vivo perturbation of TNF. We found that inhibition of epithelial TNF promotes the differentiation of secretory progenitor cells into mucus-producing goblet cells. Furthermore, TNF treatment and CFTR inhibition in intestinal organoids demonstrated that TNF promotes ion transport and luminal flow via CFTR. The absence of TNF led to slower gut transit times, which we propose results from increased mucus accumulation coupled with decreased luminal fluid pumping. These findings point to a TNF/CFTR signaling axis in the adult intestine and identify epithelial cell–derived TNF as an upstream regulator of mucin homeostasis.

Authors

Efren A. Reyes, David Castillo-Azofeifa, Jérémie Rispal, Tomas Wald, Rachel K. Zwick, Brisa Palikuqi, Angela Mujukian, Shervin Rabizadeh, Alexander R. Gupta, James M. Gardner, Dario Boffelli, Zev J. Gartner, Ophir D. Klein

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Figure 2

TNF-null intestines have increased luminal mucin, goblet cell numbers, intestinal transit time, and bacterial load.

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TNF-null intestines have increased luminal mucin, goblet cell numbers, i...
(A) Mucin fluorescence by UEA-1 lectin staining in adult intestine. Scale bar: 200 μm. (B) Quantification of mean fluorescence intensity of intervillus regions (n = 6 mice per group, 15 regions of interest per mouse). (C) Quantification of MUC2 fluorescence intensity per goblet cell (n = 3 mice per group). Mean represents all goblet cells in the distal 10 cm segment of a sectioned intestine. Values are normalized to mean of control. (D) Mucin granule size calculated as the mean fluorescence area of MUC2 signal in cells of the same regions of interest marked in panel C. (E) Immunostaining of MUC2+ goblet cells. Yellow brackets indicate MUC2+ goblet cells above Paneth cell zone in crypts. Scale bar: 50 μm. (F and G) Quantification of goblet cells in crypts and villi of mice (n = 3 mice per group, 30 full profile crypt-villus units counted per mouse). (H) Micrographs of full-length intestines showing FITC displacement after 1 hour. Arrows indicate the most distal FITC signal. Dashed lines outline tissue landmarks. Scale bar: 5 mm. (I) Quantification of FITC displacement represented as percentage of gut length, i.e., FITC signal over total gut length (n = 6 control, 7 Tnf–/– mice). (J) Total gut length was quantified as the distance from the stomach to the rectum (n = 5 control, 8 Tnf–/– mice). (K) Relative bacterial load in feces collected from the ileum of mice represented by the fold change in 16S rRNA gene copies compared with control (n = 8 mice per group). P values calculated by unpaired, 2-tailed t test (B–D, F, G, I, and J) or by unpaired, 1-tailed t test (K).