Beta-Casein(94-123)-derived peptides differently modulate production of mucins in intestinal goblet cells

We recently reported the identification of a peptide from yoghurts with promising potential for intestinal health: the sequence (94-123) of bovine β-casein. This peptide, composed of 30 amino acid residues, maintains intestinal homoeostasis through production of the secreted mucin MUC2 and of the transmembrane-associated mucin MUC4. Our study aimed to search for the minimal sequence responsible for the biological activity of β-CN(94-123) by using several strategies based on (i) known bioactive peptides encrypted in β-CN(94-123), (ii) in silico prediction of peptides reactivity and (iii) digestion of β-CN(94-123) by enzymes of intestinal brush border membranes. The revealed sequences were tested in vitro on human intestinal mucus-producing HT29-MTX cells. We demonstrated that β-CN(108-113) (an ACE-inhibitory peptide) and β-CN(114-119) (an opioid peptide named neocasomorphin-6) up-regulated MUC4 expression whereas levels of the secreted mucins MUC2 and MUC5AC remained unchanged. The digestion of β-CN(94-123) by intestinal enzymes showed that the peptides β-CN(94-108) and β-CN(117-123) were present throughout 1·5 to 3 h of digestion, respectively. These two peptides raised MUC5AC expression while β-CN(117-123) also induced a decrease in the level of MUC2 mRNA and protein. In addition, this inhibitory effect was reproduced in airway epithelial cells. In conclusion, β-CN(94-123) is a multifunctional molecule but only the sequence of 30 amino acids has a stimulating effect on the production of MUC2, a crucial factor of intestinal protection

Le peptide β-CN(94-123), un peptide bioactif des laits fermentés, comme modulateur de la protection intestinale

Deux populations de cellules épithéliales jouent un rôle crucial dans la défense intestinale : les cellules à mucus qui produisent la mucine MUC2 à l’origine de la formation du gel de mucus et les cellules de Paneth qui libèrent dans la lumière intestinale des molécules à action antimicrobienne ( -défensines, lysozyme, ..). Lors de travaux antérieurs, nous avons montré que la -casomorphine-7 (peptide opioïde issu de la -caséine bovine) est un puissant sécrétagogue du mucus intestinal, suggérant ainsi que des peptides bioactifs du lait pourraient renforcer l’arsenal défensif de l’intestin. Cependant pour être active par voie luminale, la -casomorphine-7 devait être administrée à des concentrations élevées (100 M ou plus). Parallèlement à ces résultats, des données de la littérature dévoilaient la présence de très nombreux peptides bioactifs dans les laits fermentés. Le premier objectif de notre étude était de déterminer si le pool peptidique total (PPT) d’un yaourt pouvait moduler la production de la mucine MUC2 in vitro sur la lignée mucipare intestinale humaine (HT29-MTX). Notre 2ème objectif a ensuite été d'identifier le peptide portant l'activité biologique et d'étudier son impact in vivo sur des facteurs impliqués dans la protection intestinale après administration par voie orale à des rats (une fois par jour durant 9 jours consécutifs). Les résultats obtenus ont montré que le PPT augmente l'expression des mucines MUC2 et MUC4 ainsi que la sécrétion de mucus par les cellules HT29-MTX. Parmi les quatre fractions peptidiques obtenues à partir du PPT par RP-HPLC préparative, seule la fraction C2 était capable de reproduire l'effet in vitro du PPT. La séquence [94-123] de la β-caséine, présente uniquement dans cette fraction C2, régulait également la production de MUC2 et MUC4 dans les cellules HT29-MTX. L’étude menée chez le rat a montré clairement que l'administration orale de ce peptide à des concentrations comparables à celles détectées dans un yaourt (0.01 à 1 M) induit l’expansion des cellules à mucus et de Paneth le long de l'intestin grêle. Ces effets étaient associés à une augmentation de l’expression des mucines MUC2 et MUC4 et de facteurs antibactériens (lysozyme, rdefa5). En conclusion : le peptide -CN(94-123), identifié dans les yaourts, est un nouveau peptide à effet santé ciblant le tractus intestinal. Grâce à son action sur les cellules à mucus et de Paneth, il pourrait maintenir ou restaurer l'homéostasie intestinale et jouer un rôle important dans la protection contre les agents délétères présents dans la lumière

A novel bioactive peptide from yoghurts modulates expression of the gel-forming MUC2 mucin as well as population of goblet cells and Paneth cells along the small intestine

Several studies demonstrated that fermented milks may provide a large number of bioactive peptides into the gastrointestinal tract. We previously showed that beta-casomorphin-7, an opioid-like peptide produced from bovine β-casein, strongly stimulates intestinal mucin production in ex vivo and in vitro models, suggesting the potential benefit of milk bioactive peptides on intestinal protection. In the present study, we tested the hypothesis that the total peptide pool (TPP) from a fermented milk (yoghurt) may act on human intestinal mucus-producing cells (HT29-MTX) to induce mucin expression. Our aim was then to identify the peptide(s) carrying the biological activity and to study its impact in vivo on factors involved in gut protection after oral administration to rat pups (once a day, 9 consecutive days). TPP stimulated MUC2 and MUC4 gene expression as well as mucin secretion in HT29-MTX cells. Among the four peptide fractions that were separated by preparative reversed-phase high-performance liquid chromatography, only the C2 fraction was able to mimic the in vitro effect of TPP. Interestingly, the sequence [94-123] of β-casein, present only in C2 fraction, also regulated mucin production in HT29-MTX cells. Oral administration of this peptide to rat pups enhanced the number of goblet cells and Paneth cells along the small intestine. These effects were associated with a higher expression of intestinal mucins (Muc2 and Muc4) and of antibacterial factors (lysozyme, rdefa5). We conclude that the peptide β-CN(94-123) present in yoghurts may maintain or restore intestinal homeostasis and could play an important role in protection against damaging agents of the intestinal lumen

Peptides augmentant la secrétion et/ou l'expression d'au moins une mucine gastrointestinale et/ou induisant l'augmentation de la population de cellules à mucus ou de cellules de paneth.

Date de publication et mention de la délivrance du brevet: 06.03.2019 Bulletin 2019/10The invention relates to a polypeptide including or consisting of SEQ ID NO: 1, or a sequence at least 80% identical to SEQ ID NO: 1, or a sequence of at least 4 consecutive amino acids included in SEQ ID NO: 1, wherein said polypeptide has at least one effect selected from: inducing the expression and/or the secretion of at least one gastrointestinal mucin; and inducing the expression and/or the secretion of at least one intestinal defense molecule expressed by Paneth cells; and inducing an increase in the population of mucus cells and/or in the population of Paneth cells, with the proviso that the polypeptide does not include the SEQ ID NO: 2 sequence

Peptides augmentant la secrétion et/ou l'expression d'au moins une mucine gastrointestinale et/ou induisant l'augmentation de la population de cellules à mucus ou de cellules de paneth.

The invention relates to a polypeptide including or consisting of SEQ ID NO: 1, or a sequence at least 80% identical to SEQ ID NO: 1, or a sequence of at least 4 consecutive amino acids included in SEQ ID NO: 1, wherein said polypeptide has at least one effect selected from: inducing the expression and/or the secretion of at least one gastrointestinal mucin; and inducing the expression and/or the secretion of at least one intestinal defense molecule expressed by Paneth cells; and inducing an increase in the population of mucus cells and/or in the population of Paneth cells, with the proviso that the polypeptide does not include the SEQ ID NO: 2 sequence

DCC constrains tumour progression via its dependence receptor activity.

International audienceThe role of deleted in colorectal carcinoma (DCC) as a tumour suppressor has been a matter of debate for the past 15 years. DCC gene expression is lost or markedly reduced in the majority of advanced colorectal cancers and, by functioning as a dependence receptor, DCC has been shown to induce apoptosis unless engaged by its ligand, netrin-1 (ref. 2). However, so far no animal model has supported the view that the DCC loss-of-function is causally implicated as predisposing to aggressive cancer development. To investigate the role of DCC-induced apoptosis in the control of tumour progression, here we created a mouse model in which the pro-apoptotic activity of DCC is genetically silenced. Although the loss of DCC-induced apoptosis in this mouse model is not associated with a major disorganization of the intestines, it leads to spontaneous intestinal neoplasia at a relatively low frequency. Loss of DCC-induced apoptosis is also associated with an increase in the number and aggressiveness of intestinal tumours in a predisposing APC mutant context, resulting in the development of highly invasive adenocarcinomas. These results demonstrate that DCC functions as a tumour suppressor via its ability to trigger tumour cell apoptosis