Basic Study
Copyright ©The Author(s) 2016. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Nov 28, 2016; 22(44): 9734-9743
Published online Nov 28, 2016. doi: 10.3748/wjg.v22.i44.9734
Ex vivo response to mucosal bacteria and muramyl dipeptide in inflammatory bowel disease
Claudia Loganes, Erica Valencic, Alessia Pin, Elisa Marini, Stefano Martelossi, Samuele Naviglio, Luigina De Leo, Tarcisio Not, Lorenzo Monasta, Alberto Tommasini, Annalisa Marcuzzi
Claudia Loganes, Elisa Marini, Samuele Naviglio, Tarcisio Not, Annalisa Marcuzzi, Department of Medicine, Surgery, and Health Sciences, University of Trieste, Strada di Fiume, 447 Trieste, Italy
Erica Valencic, Alessia Pin, Stefano Martelossi, Luigina De Leo, Tarcisio Not, Lorenzo Monasta, Alberto Tommasini, Institute for Maternal and Child Health - IRCCS “Burlo Garofolo”, I-34137 Trieste, Italy
Author contributions: Loganes C, Valencic E and Tommasini A designed the experimental plan and conceived and designed the research; Pin A, Marini E, De Leo L performed all experiments; Loganes C, Monasta L performed the statistical analyses; Martelossi S, Naviglio S and Not T enrolled the patients and provided patient samples; Loganes C, Tommasini A, Marcuzzi A wrote the paper; Martelossi S and Marcuzzi A revised the manuscript critically for important intellectual content; all authors read and approved the final version of this manuscript.
Supported by Institute for Maternal and Child Health - IRCCS “ Burlo Garofolo”, No. RC 03/09.
Institutional review board statement: The study was reviewed and approved by the Institutional Review Board of the Institute for Maternal and Child Health - IRCCS “Burlo Garofolo”, Trieste, Italy (RC 03/2009).
Conflict-of-interest statement: All the authors declare no conflict of interest.
Data sharing statement: Dataset available from the corresponding author at
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See:
Correspondence to: Alberto Tommasini, MD, PhD, Institute for Maternal and Child Health - IRCCS “Burlo Garofolo”, Via dell’Istria 65/1, I-34137 Trieste, Italy.
Telephone: +39-40-3785422
Received: June 28, 2016
Peer-review started: June 29, 2016
First decision: July 29, 2016
Revised: August 12, 2016
Accepted: September 6, 2016
Article in press: September 6, 2016
Published online: November 28, 2016

To evaluate how mucosal bacteria impact on the spontaneous and muramyl dipeptide (MDP)-induced inflammation in Crohn’s disease (CD) and ulcerative colitis (UC).


Colonic mucosal biopsies were collected from children with active or remissive CD, UC and controls. Two tissue samples were taken from inflamed mucosal segments (in patients with active disease) or from non-inflamed mucosa [in patients in remission or in healthy controls (HC)]. Experiments were performed in the presence or absence of antibiotics, to assess whether the disease-associated microbiota can modulate the cytokine response ex vivo. For this purpose, each specimen was half-cut to compare spontaneous and MDP-induced inflammation in the presence of live bacteria (LB) or antibiotics. After 24 h of culture, an array of 17 cytokines was assessed in supernatants. Statistical analyses were performed to find significant differences in single cytokines or in patterns of cytokine response in the different groups.


We demonstrated that subjects with CD display a spontaneous production of inflammatory cytokines including granulocyte-colony stimulating factor (G-CSF), interleukin (IL) 6, IL8, IL10 and IL12, that was not significantly influenced by the addition of antibiotics. UC specimens also displayed a trend of increased spontaneous secretion of several cytokines, which however was not significant due to broader variability among patients. After the addition of antibiotics, spontaneous IL8 secretion was significantly higher in UC than in controls. In HC, a trend towards the weakening of spontaneous IL8 production was observed in the presence of live mucosal bacteria with respect to the presence of antibiotics. In contrast, in the presence of LB UC showed an increasing trend of spontaneous IL8 production, while MDP stimulation resulted in lower IL8 production in the presence of antibiotics. We also showed that subjects with CD seem to have a lowered production of IL8 in response to MDP in the presence of LB. Only with the addition of antibiotics, likely reducing the contribution of LB, multivariate statistical analysis could identify the combination of measures of G-CSF, tumor necrosis factor alpha, IL4 and IL17 as a good discriminator between CD and UC.


We showed that the presence of LB or antibiotics can significantly influence the inflammatory response ex vivo in inflammatory bowel diseases.

Keywords: Gut-microbiota, Crohn’s disease, Ulcerative colitis, Cytokines, Inflammation

Core tip: Even though previous studies have already considered cytokine secretions as marker of an inflammatory condition and mucosal imbalance in Crohn’s disease (CD) subjects, they did not take into account the autochthonous colonization of the intestinal mucosa by the disease-associated microbiota. In this work we investigated whether the microbiota can modulate the ex vivo cytokine response, in the presence or absence of antibiotics, or if a selection of cytokines could discriminate different inflammatory bowel diseases types. Through a multivariate logistic model we identified, only in specimens treated with antibiotics, a specific cytokine profile able to discriminate CD from ulcerative colitis.