Soluble Non-Starch Polysaccharides From Plantain (Musa x paradisiaca L.) Diminish Epithelial Impact of Clostridioides difficile

Clostridioides difficile infection (CDI) is a leading cause of antibiotic-associated diarrhoea. Adhesion of this Gram-positive pathogen to the intestinal epithelium is a crucial step in CDI, with recurrence and relapse of disease dependent on epithelial interaction of its endospores. Close proximity, or adhesion of, hypervirulent strains to the intestinal mucosa are also likely to be necessary for the release of C. difficile toxins, which when internalized, result in intestinal epithelial cell rounding, damage, inflammation, loss of barrier function and diarrhoea. Interrupting these C. difficile-epithelium interactions could therefore represent a promising therapeutic strategy to prevent and treat CDI. Intake of dietary fibre is widely recognised as being beneficial for intestinal health, and we have previously shown that soluble non-starch polysaccharides (NSP) from plantain banana (Musa spp.), can block epithelial adhesion and invasion of a number of gut pathogens, such as E. coli and Salmonellae. Here, we assessed the action of plantain NSP, and a range of alternative soluble plant fibres, for inhibitory action on epithelial interactions of C. difficile clinical isolates, purified endospore preparations and toxins. We found that plantain NSP possessed ability to disrupt epithelial adhesion of C. difficile vegetative cells and spores, with inhibitory activity against C. difficile found within the acidic (pectin-rich) polysaccharide component, through interaction with the intestinal epithelium. Similar activity was found with NSP purified from broccoli and leek, although seen to be less potent than NSP from plantain. Whilst plantain NSP could not block the interaction and intracellular action of purified C. difficile toxins, it significantly diminished the epithelial impact of C. difficile, reducing both bacteria and toxin induced inflammation, activation of caspase 3/7 and cytotoxicity in human intestinal cell-line and murine intestinal organoid cultures. Dietary supplementation with soluble NSP from plantain may therefore confer a protective effect in CDI patients by preventing adhesion of C. difficile to the mucosa, i.e. a “contrabiotic” effect, and diminishing its epithelial impact. This suggests that plantain soluble dietary fibre may be a therapeutically effective nutritional product for use in the prevention or treatment of CDI and antibiotic-associated diarrhoea.

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Characterization of the human peptide transporter PEPT1 promoter: Sp1 functions as a basal transcriptional regulator of human PEPT1

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00025.2005 ◽

2005 ◽

Vol 289 (3) ◽

pp. G471-G477 ◽

Cited By ~ 27

Author(s):

Jin Shimakura ◽

Tomohiro Terada ◽

Toshiya Katsura ◽

Ken-Ichi Inui

Keyword(s):

Transcriptional Regulation ◽

Transcriptional Activity ◽

Mutational Analysis ◽

Peptide Transporter ◽

Intestinal Cell ◽

Dependent Manner ◽

Intestinal Epithelial ◽

Intestinal Cell Line ◽

Human Intestinal Cell Line

H+-coupled peptide transporter 1 (PEPT1, SLC15A1) localized at the brush-border membranes of intestinal epithelial cells plays an important role in the intestinal absorption of small peptides and a variety of peptidemimetic drugs. PEPT1 is regulated by various factors, including hormones, dietary conditions, some pharmaceutics, and diurnal rhythm. But there is little information about the transcriptional regulation of PEPT1. In the present study, therefore, we cloned the human (h)PEPT1 promoter region and examined its promoter activity using a human intestinal cell line, Caco-2. Deletion analysis of the hPEPT1 promoter suggested that the region spanning −172 to −35 bp was essential for basal transcriptional activity. This region lacked a TATA-box but contained some GC-rich sites that supposedly bind with the transcription factor Sp1. Mutational analysis revealed that three of these putative Sp1 sites contributed to the transcriptional activity. EMSA showed that Sp1 bound to two GC-rich sites. Furthermore, inhibition of Sp1 binding by mithramycin A treatment significantly reduced the transcriptional activity. Finally, overexpression of Sp1 increased the transcriptional activity in a dose-dependent manner. This study reports the first characterization of the hPEPT1 promoter and shows the significant role of Sp1 in the basal transcriptional regulation of hPEPT1.

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Dipeptide transporters in apical and basolateral membranes of the human intestinal cell line Caco-2

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1993.265.2.g289 ◽

1993 ◽

Vol 265 (2) ◽

pp. G289-G294 ◽

Cited By ~ 13

Author(s):

H. Saito ◽

K. Inui

Keyword(s):

Cell Line ◽

Inhibitory Effect ◽

Intestinal Cell ◽

Epithelial Cell Line ◽

Apical Surface ◽

Intestinal Epithelial ◽

Basolateral Membranes ◽

Intestinal Cell Line ◽

Basolateral Side ◽

Human Intestinal Cell Line

The localization and transport characteristics of dipeptide transporters of the intestinal epithelial cell line Caco-2 were examined by measuring the intracellular accumulation and transcellular flux of Bestatin, a dipeptide-like anticancer agent. When added to the apical surface of Caco-2 monolayers grown on microporous membrane filters, Bestatin was accumulated in the cells and was transported unidirectionally to the basolateral side. The cellular uptake of Bestatin from the basolateral as well as from the apical surface was inhibited by excess dipeptides. Bestatin accumulation from the apical surface was dependent on the pH of the incubation medium with an optimal pH of 6.0, whereas uptake from the basolateral surface was insensitive to the medium pH. Kinetic parameters for Bestatin uptake also indicated that the basolateral and apical dipeptide transporters could be distinguished from each other. A sulfhydryl reagent, p-chloromercuribenzene sulfonate, inhibited Bestatin accumulation from both surfaces, although the inhibitory effect on the basolateral transport was greater than that on the apical transport. These findings suggest not only that dipeptide transporters exist on both the apical and basolateral membranes of Caco-2 cells but also that the basolateral dipeptide transporter is distinct from the apical H(+)-dipeptide cotransporter.

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