FC FELLOWSHIP - CALL FOR PROPOSALS 2018

Triennial Programmes granted in the frame of the FC Fellowships - Call for Proposals 2018 

 

Triennial Fellowship Programme 001_FC_2018 

MOLECULAR AND CELLULAR EFFECTS OF GLUTEN-EXORPHINS ON TUMORIGENESIS: AN IN VITRO PILOT STUDY.

 

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Federico Manai, University of Pavia
Topic: Celiac Disease – Area: Oncology

 

Background

Gluten-exorphins (GEs) are opioid-peptides digestion products of wheat-based food, showing exceptional resistance to enzymatic hydrolysis and potent bioactivity. To exert the opioid-like effect, GEs enter the bloodstream through the intestinal epithelium, whose permeability has been altered in pathological conditions. Then, GEs interact with opioid receptors (ORs) activating cell signalling pathways. There is emerging evidence that opioid-mediated signalling play a role in cancer progression by interacting with MAPK, ERK and other signalling pathways, controlling cell proliferation and apoptosis. It has been reported that the incidence of some neoplastic disorders, particularly malignant lymphoma and small intestinal adenocarcinoma, are increased in celiac disease (CD). In particular, the main cause of mortality in CD, within 5 years of diagnosis, is malignancy and CD survival is improved by early diagnosis and strict adherence to a gluten-free diet.

Hypothesis

Enteric glial cells (EGCs) are potent regulators of intestinal epithelium functions, but their impact on cancer cell progression in CD has not been deeply investigated.

In this study, we hypothesize that EGCs can bind GEs to their ORs, thereby influencing molecular and paracellular signaling cascades that finally modulate cancer cells functions and associated tumorigenesis.

Aims

The goal of the proposal is to identify pro-tumorigenic molecular signalling differences between pediatric CD/controls, modulated after GEs administration. These signalling alteration might potentially drive to specific target genes that will be interfered using antisense or siRNAs strategies.

Research Plan 

In this proposal, the effect of synthetic GEs on EGCs isolated from a cohort of pediatric CD patients and controls will be evaluated in their cellular uptake, intracellular traffic, influence on molecular and paracellular signaling toward human duodenal adenocarcinoma or lymphoma established cells, to finally evaluate the GEs induced effects on tumorigenicy (viability, proliferation and cell death modulation).

Expected results and Impact on Celiac disease/Dermatitis Herpetiformis 

The possible final outcome on CD could be to early understand the effects of by-products of gluten digestion in promoting tumorigenesis, evaluating the differences between pediatric CD and control subjects, and to develop preclinical strategies to counteract tumor cell proliferation.



Triennial Fellowship Programme 008_FC_2018

EXPLOITING THE MAST CELLS PLASTICITY FOR PREDICTIVE SIGNATURES OF CANDIDA ALBICANS COMMENSALISM VS PATHOGENICITY IN CELIAC DISEASE

 

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Giorgia Renga, University of Perugia
Topic: Celiac Disease – Area: Infiammation

 

Background

Celiac disease (CD) is an immune-mediated disorder resulting from an interaction between diet, genome, and immunity, and more frequent and severe in female vs male. Whilst many patients respond to a gluten free diet with the resolution of symptoms, a substantive number of individuals have continuous problems with potentially serious consequences. Candida albicans is a fungal commensal of human gut often implicated in inflammatory intestinal diseases and it has been reported that Candida may trigger CD development.

Hypothesis

Mast cells (MCs) have recently gained importance for their ability to modulate both innate and adaptive immune responses in the gut. Indeed, we have recently identified MCs and IL-9 as key players of Candida commensalism and pathogenicity. Specifically, we have unveiled two molecular pathways that link MC/IL-9 axis to Candida albicans. The inflammatory pathway, upon which Candida promotes inflammation and intestinal permeability, acts through mucosal mast cells (MMCs) and IL-9. Instead, the tolerogenic pathway occurs through the connective tissue mast cells (CTMCs) and IL-9 via indoleamine 2,3-dioxygenase (IDO), an enzyme required for long-term tolerance in the gut. We have also shown that the MC-IL-9 inflammatory pathway seems to play a major role in experimental leaky gut models, such as celiac disease. In agreement with recent literature, we could confirm MCs expansion in CD patients, and additionally could show that IL-9 positivity increases, while IDO expression decreases, with the severity of the disease. Based on these findings, our working hypothesis is that MCs contributes to Candida overgrowth and onset and development of CD by integrating multiple signals from the environment and different mechanisms in the gastrointestinal tract. 

Aims

We will resort to wild-type, MC-deficient (KitW/W-v) and tolerogenic MC-deficient (Tph1—/—) mice, infected or not with C. albicans after induction of gluten sensitivity, to achieve the following aims:

1) Define the inflammatory vs tolerogenic role of MCs;

2) Elucidate the contribution of IL-9 and IDO1/IDO2;

3) Explain the potential contribution of MCs in modifying the microbiome by metagenomics and metabolomics analysis;

4) Define the impact of antifungals and post-biotics;

5) Identify gender-specific effects

Research Plan 

The experimental design is as follows:

Induction of gluten sensibilization in different mouse models, infected or not with Candida and allocations in the following groups:

- anti-IL-9 mAb vs isotype control

- antifungals or post-biotics vs vehicle

- male vs female

The following endpoints will be evaluated:

- inflammatory status and levels of selected markers by histology, ELISA, qPCR, Wester blotting

- fungal burden by CFU counting

- microbiome and metabolome changes by Next-Generation Sequencing and mass spectrometry, respectively

Expected results and Impact on Celiac disease/Dermatitis Herpetiformis 

The present project is expected to elucidate the role of MCs in CD and the specific mechanisms employed in disease pathogenesis. The specific aims should unravel the plasticity of MCs and the reciprocal interactions with the surrounding environment and the presence of gender effects. These results are expected to be rapidly translated to the human setting and should allow to stratify and treat the patients according to their risk to develop a pathological Candida infection.

 

 

Triennial Fellowship Programme 011_FC_2018

DEFINITION AND VALIDATION OF BIOMARKERS TO EVALUATE GLUTEN FREE DIET (GFD) COMPLIANCE IN CELIAC PATIENTS

 

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Albino Coglianese, University of Salerno
Topic: Celiac Disease – Area: Epidemiology

 

Background

Celiac disease (CD) is an autoimmune disorder triggered by gluten ingestion in genetically susceptible individuals. A scrupulous and permanent gluten-free diet (GFD) is the only treatment available for CD, and strict adherence to GFD is essential. However, according to various reports, the transgression in the diet is relatively frequent (32.6-55.4%) in the celiac patient. Currently, available methods for assessing GFD compliance are not enough sensitive to detect occasional diet transgressions.

Hypothesis

Currently, no validated methods to monitor the adherence to GFD are available; this aspect is often evaluated through questionnaires, which have severe limits in terms of reliability. It is therefore necessary to identify and validate suitable biomarkers to reveal possible distraction from GFD. The peptide “33-mer” and some alkyl-resorcinols are molecules that have provided encouraging results in that aims; their presence in human biological matrices is indeed related to gluten intake.

Aims

This project is focused on the development of an analytical protocol allowing the identification of non-adherence of celiac patients to GFD through the qualitative-quantitative measurement in human biological matrices, such as feces and urine, of biomarkers specific for gluten exposure. Specifically, the correlation between the dietary intake of gluten-containing products and the presence of the "33 mer" peptide and specific alkyl-resorcinols in those matrices will be evaluated.

Research Plan

In the first phase of the project, an analytical method for the simultaneous detection of the two biomarkers of interest in human feces and urine will be the developed and validated. Individuals belonging to three groups of healthy volunteers will be involved: the first not subjected to any controlled diet, the second following a diet - designed in collaboration with a dietician - with a controlled content of gluten, and the third subjected to a GFD. The second phase will consist in the application of the aforementioned analytical method for the monitoring of the two biomarkers in a small group of celiac subjects treated with a GFD. In the third phase, the study will be extended to a larger group of celiac patients undergoing a GFD - randomized for different degrees of severity of CD- and healthy subjects; biomarkers levels detected in these individuals will then be related to the possible manifestation of CD typical symptoms. The analytical technique chosen for the quantization of biomarkers is high performance liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). The whole process will be validated as required by the European Medicines Agency (EMA) guidelines for the validation of analytical methods.

Expected results and Impact on Celiac disease/Dermatitis Herpetiformis 

This project will finally provide diagnostic an easily applicable procedure aimed at maximizing the adherence to the GFD. This procedure will also allow detecting any distractions from the diet or involuntary intake of food contaminated by gluten after appearance of acute symptoms in CD patients. Moreover, a quick and correct evaluation of the proposed biomarkers in patients showing CD symptoms could favor an early diagnosis of the pathology, also in patients with refractory CD or with IgA deficiency

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