Fibroblasts Impact Goblet Cell Responses to Lactic Acid Bacteria After Exposure to Inflammatory Cytokines and Mucus Disruptors.

SCOPE: Mucus produced by goblet cells contributes to gut barrier function. Lactic acid bacteria (LAB) have been shown to impact mucus production. It is not completely known whether mucus production is influenced by the abundantly present fibroblasts in the intestine. METHODS AND RESULTS: The influence of fibroblasts on mucus-related genes including mucin-2 (MUC2), trefoil factor 3 (TFF3), resistin-like molecule ß (RETNLB), carbohydrate sulfotransferase 5 (CHST5), and galactose-3-O-sulfotransferase 2 (GAL3ST2) is examined after co-culture of LS174T-goblet cells and CCD-18Co colonic fibroblasts in the presence and absence of LAB-strains known to impact mucus function. This is also tested after exposure to TNF-α, IL-13, or the mucin synthesis inhibitor tunicamycin (Tm). Effects of fibroblasts are treatment duration- and bacterial species-dependent under homeostatic conditions. During TNF-α challenge, fibroblasts reverse Lactobacillus (L.) rhamnosus CCFM237-elicited declined TFF3 expression. After IL-13 exposure, L. rhamnosus CCFM237 and L. fermentum CCFM787 attenuate enhanced TFF3 and RETNLB expression, respectively, only in the presence of fibroblasts. LAB has no effects on Tm-induced decreased expression of goblet cell-related genes regardless of the presence of fibroblasts. CONCLUSION: It is demonstrated that goblet cell-fibroblast crosstalk impacts mucus synthesis and influences the effects of LAB on goblet cell-related genes. Effects are LAB-species and stressor dependent.

Lactic Acid Bacteria May Impact Intestinal Barrier Function by Modulating Goblet Cells.

SCOPE: Lactic acid bacteria (LAB) are recognized to promote gastrointestinal health by mechanisms that are not fully understood. LABs might modulate the mucus and thereby enhance intestinal barrier function. Herein, we investigate effects of different LAB strains and species on goblet cell genes involved in mucus synthesis. METHODS AND RESULTS: Gene expression profiles of goblet-cell-associated products (mucin MUC2, trefoil factor 3, resistin-like molecule ß, carbohydrate sulfotransferase 5, and galactose-3-O-sulfotransferase 2) induced by LAB or their derived conditioned medium in human goblet cell line LS174T are studied. Effects of LAB on gene transcription are assessed with or without exposure to TNF-α, IL-13, or the mucus damaging agent tunicamycin. LAB do impact the related genes in a species- and strain-specific fashion and their effects are different in the presence of the cytokines and tunicamycin. Bioactive factors secreted by some strains are also found to regulate goblet cell-related genes. CONCLUSION: Our findings provide novel insights in differences in modulatory efficacy on mucus genes between LAB species and strains. This study further unravels direct interactions between LAB and intestinal goblet cells, and highlights the importance of rationally selecting appropriate LAB candidates to achieve specific benefits in the gut.

Writing of H3K4Me3 overcomes epigenetic silencing in a sustained but context-dependent manner.

Histone modifications reflect gene activity, but the relationship between cause and consequence of transcriptional control is heavily debated. Recent developments in rewriting local histone codes of endogenous genes elucidated instructiveness of certain marks in regulating gene expression. Maintenance of such repressive epigenome editing is controversial, while stable reactivation is still largely unexplored. Here we demonstrate sustained gene re-expression using two types of engineered DNA-binding domains fused to a H3K4 methyltransferase. Local induction of H3K4me3 is sufficient to allow re-expression of silenced target genes in various cell types. Maintenance of the re-expression is achieved, but strongly depends on the chromatin microenvironment (that is, DNA methylation status). We further identify H3K79me to be essential in allowing stable gene re-expression, confirming its role in epigenetic crosstalk for stable reactivation. Our approach uncovers potent epigenetic modifications to be directly written onto genomic loci to stably activate any given gene.

Particulate ß-glucans synergistically activate TLR4 and Dectin-1 in human dendritic cells.

SCOPE: The major receptor for ß(1-3)-glucans on immune cells is considered to be Dectin-1 receptor. Particulate ß-glucans induce stronger immune responses than soluble ß-glucans by clustering of Dectin-1 receptors. Here, it was hypothesized that activation of other pattern recognition receptors such as Toll-like receptor 4 (TLR4) can also contribute to enhanced activity of immune cells after exposure to particulate ß-glucans. METHODS AND RESULTS: To test this hypothesis, reporter cell lines were designed expressing TLR4 with either Dectin-1A or Dectin-1B, that is, one of the two transcript variants of human Dectin-1 receptors. Enhanced NF-κB activation was observed after stimulation with particulate ß-glucans in both Dectin-1A-TLR4 and the Dectin-1B-TLR4 cell lines. This was different with soluble ß-glucans, which enhanced activation in Dectin-1A-TLR4 cell lines but not in Dectin-1B-TLR4 cells. The synergistic activation of TLR4 and Dectin-1 by particulate ß-glucans was confirmed in human dendritic cells. The effects of particulate ß-glucan induced TLR4 binding were regulatory as blocking TLR4 enhanced pro-inflammatory cytokine IL-23, IL-4, IL-6, and TNF-α production. CONCLUSION: These results suggest that TLR4 and Dectin-1 are synergistically activated by particulate ß-glucans, wherein TLR4 activates an immune regulatory pathway in human dendritic cells. Our data suggest that ß-glucan is an immune regulatory ligand for TLR4.