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. 2010 Jan 5;107(1):228-33.
doi: 10.1073/pnas.0906112107. Epub 2009 Dec 4.

The bacterial signal indole increases epithelial-cell tight-junction resistance and attenuates indicators of inflammation

Tarun Bansal  1 , Robert C Alaniz, Thomas K Wood, Arul Jayaraman
Affiliations

The bacterial signal indole increases epithelial-cell tight-junction resistance and attenuates indicators of inflammation

Tarun Bansal et al. Proc Natl Acad Sci U S A. .

Abstract

Interkingdom signaling is established in the gastrointestinal tract in that human hormones trigger responses in bacteria; here, we show that the corollary is true, that a specific bacterial signal, indole, is recognized as a beneficial signal in intestinal epithelial cells. Our prior work has shown that indole, secreted by commensal Escherichia coli and detected in human feces, reduces pathogenic E. coli chemotaxis, motility, and attachment to epithelial cells. However, the effect of indole on intestinal epithelial cells is not known. Because intestinal epithelial cells are likely to be exposed continuously to indole, we hypothesized that indole may be beneficial for these cells, and investigated changes in gene expression with the human enterocyte cell line HCT-8 upon exposure to indole. Exposure to physiologically relevant amounts of indole increased expression of genes involved in strengthening the mucosal barrier and mucin production, which were consistent with an increase in the transepithelial resistance of HCT-8 cells. Indole also decreased TNF-alpha-mediated activation of NF-kappaB, expression of the proinflammatory chemokine IL-8, and the attachment of pathogenic E. coli to HCT-8 cells, as well as increased expression of the antiinflammatory cytokine IL-10. The changes in transepithelial resistance and NF-kappaB activation were specific to indole: other indole-like molecules did not elicit a similar response. Our results are similar to those observed with probiotic strains and suggest that indole could be important in the intestinal epithelial cells response to gastrointestinal tract pathogens.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Changes in tight junction proteins and TER. (A) Pathways containing differentially expressed genes involved in tight junction formation were adapted from Pathway Express using KEGG classifications. Red, up-regulation; blue, down-regulation; gray, no change in expression; arrow, molecular interaction leading to activation; blunt line without an arrowhead, molecular interaction leading to inhibition. The pathway scheme shown is based on microarray data and does not include posttranscriptional regulation. (B) Changes in the TER of polarized HCT-8 cells exposed to either solvent (filled circles) or indole (open circles) for 24 h. Data represent mean ± SD from seven measurements at each time point and two independent experiments. (C) Effect of indole pretreatment on adherence of EHEC to HCT-8 cells in the presence of 500 μM norepinephrine. Data represent mean ± SD from three wells per experiment and three independent experiments. *, statistical significance, using Student's t test, at P < 0.05; **, significance at P < 0.005.
Fig. 2.
Fig. 2.
Changes in Toll-like receptor, IL-8, and IL-10 signaling. (A) Pathways containing differentially expressed genes involved in tight junction formation were adapted from Pathway Express using KEGG classifications. Colors and symbols as in Fig. 1. The pathway scheme shown is based on microarray data and does not include posttranscriptional regulation. (B) Secretion of IL-8 in HCT-8 cells exposed to indole for 4 h or 24 h, in the presence or absence of 40 ng/mL TNF-α. Data represent mean ± SD from three independent experiments. (C) Intracellular staining and flow cytometry of IL-10 in HCT-8 cells exposed to indole for 24 h. Two-color dot plots of number of PE-stained cells and forward scatter for solvent control, indole-treated, and unstained cells are shown. Percentages in the boxed region indicate the proportion of HCT-8 cells producing IL-10 in one representative experiment. (D) Average increase in IL-10 expression data from three independent experiments. *, statistical significance, using Student’s t test, at P < 0.05; **, significance at P < 0.005.
Fig. 3.
Fig. 3.
Modulation of NF-κB activity by indole. (A) Effect of indole on NF-κB activation by TNF-α in HCT-8 NF-κB-GFP reporter cells and the resultant GFP expression. (B) Quantification of NF-κB activity in control cells (filled circles), cells treated with TNF-α to induce NF-κB activation (open circles), and cells treated with indole and TNF-α (filled inverted triangles). Data shown are mean ± SD from images obtained at four locations in three independent experiments.
Fig. 4.
Fig. 4.
Changes in HCT-8 cell phenotypes with aromatic bicyclic molecules. (A) NF-κB activation in HCT-8 NF-κB-GFP reporter cells and the resultant GFP expression. Images shown are after 18 h of exposure to TNF-α and are representative of data from three independent experiments. (B) Changes in TER after exposure to indole-like molecules for 24 h. Data represent mean ± SD from three independent experiments. **, statistical significance, using Student's t test, at P < 0.005.
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