Epigenetic Features of FoxP3 in Children With Cow's Milk Allergy

Epigenetic mechanisms have been implicated in the pathogenesis of food allergy. The investigators previously demonstrated that tolerance acquisition in children with Immunoglobulin E- (IgE) mediated cow's milk allergy (CMA) is driven by epigenetic modulation of the Th1 and Th2 cytokine genes. A regulatory T cell (Treg) suppressive phenotype, characterized by stable expression of the transcription factor "Forkhead box Protein 3" (FoxP3), plays a pivotal role in food tolerance. FoxP3 mRNA expression is lower in children with atopic asthma or IgE-mediated food allergy than in healthy children. FoxP3 stable expression requires full CpG demethylation of its transcriptional regulatory regions, and, moreover, hypermethylation of the FoxP3 gene has been associated with reduced Treg function and allergy.

DNA methylation is a biologically and chemically stable epigenetic modification that locks in long-term gene expression patterns. The demethylation status of FoxP3 at a highly conserved region within the Treg-specific-demethylated-region (TSDR), a CpG-rich, located on the 2nd conserved non-coding sequence of FoxP3 (CNS2), is restricted to Tregs. Transcriptional activity of the TSDR is essentially determined by its methylation status : it is completely inactive in its methylated state, but when the TSDR is demethylated, transcription factors such as Ets-1 and Creb can bind to the TSDR. TSDR demethylated and open chromatin conformation in the Foxp3 locus leads to stable phenotype differentiated Foxp3+ Treg. FoxP3 TSDR demethylation in peripheral blood mononuclear cells (PBMCs) has been associated with reduced atopic sensitization and asthma in children. Epigenetic regulation of antigen-induced T-cell subsets may predict a state of immune tolerance in food allergy. Indeed, DNA methylation of the FoxP3 gene in Tregs decreased during oral tolerance acquisition in patients with peanut allergy undergoing oral immunotherapy. The aim of this study was to evaluate further the epigenetic regulation of FoxP3 gene in children with IgE-mediated CMA.

Study Overview

• Status: Completed
• Conditions: Cow's Milk Allergy
• Intervention / Treatment: Dietary supplement: Extensively hydrolyzed casein formula plus LGG

• Study Type: Observational
• Enrollment (Actual): 40

Contacts and Locations

Study Locations

• Italy
  • Naples, Italy, 80131
    • University of Naples Federico II

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 3 months to 1 year (Child)
• Accepts Healthy Volunteers: No
• Genders Eligible for Study: All

• Sampling Method: Probability Sample

Study Population

IgE-mediated CMA children (aged 3 to 18 months) consecutively referred to the tertiary Pediatric Allergy Center for oral food challenge.

During the same study period, consecutive healthy children, not at risk of atopic disorders (namely, those without a first-degree relative affected by an atopic disorder), attending the Center because of minimal surgical procedures served as a control group.

A venous blood sample (4 ml) was obtained from all patients after oral challenge.

Description

Inclusion Criteria:

• IgE-mediated CMA children (aged 3 to 18 months)

Exclusion Criteria:

• allergic disorders or food allergies other than cow's milk allergy
• eosinophilic disorders of the gastrointestinal tract
• food protein-induced enterocolitis syndrome
• concomitant chronic systemic diseases
• congenital cardiac defects
• active tuberculosis
• autoimmune diseases
• immunodeficiency
• chronic inflammatory bowel diseases
• celiac disease
• cystic fibrosis
• metabolic diseases
• lactose intolerance
• malignancy
• chronic pulmonary diseases
• malformations of the gastrointestinal tract

Study Plan

How is the study designed?

Design Details

• Observational Models: Case-Crossover
• Time Perspectives: Cross-Sectional

Number of groups / cohorts

4

Cohorts and Interventions

Group / Cohort	Intervention / Treatment
Healthy controls
Children at diagnosis of cow's milk allergy
Subjects outgrown cow's milk allergy with formula+probiotic; Tolerant with extensively hydrolyzed casein formula with Lactobacillus rhamnosus GG	Dietary supplement: Extensively hydrolyzed casein formula plus LGG
Subjects outgrown cow's milk allergy assuming other formulas; Subjects tolerant with other formulas

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Time Frame
DNA demethylation (rate, in %) of the Treg-specific-demethylated-region (TSDR) of FoxP3	At enrollment

Collaborators and Investigators

• Sponsor: Federico II University

Publications and helpful links

General Publications

• Paparo L, Nocerino R, Cosenza L, Aitoro R, D'Argenio V, Del Monaco V, Di Scala C, Amoroso A, Di Costanzo M, Salvatore F, Berni Canani R. Epigenetic features of FoxP3 in children with cow's milk allergy. Clin Epigenetics. 2016 Aug 12;8:86. doi: 10.1186/s13148-016-0252-z. eCollection 2016.

Study record dates

Study Major Dates

• Study Start: December 1, 2012
• Primary Completion (Actual): April 1, 2014
• Study Completion (Actual): April 1, 2014

Study Registration Dates

• First Submitted: May 16, 2016
• First Submitted That Met QC Criteria: May 19, 2016
• First Posted (Estimate): May 23, 2016

Study Record Updates

• Last Update Posted (Estimate): May 23, 2016
• Last Update Submitted That Met QC Criteria: May 19, 2016
• Last Verified: May 1, 2016

More Information

Terms related to this study

• Additional Relevant MeSH Terms: Immune System Diseases; Food Hypersensitivity; Hypersensitivity, Immediate; Hypersensitivity; Milk Hypersensitivity; Molecular Mechanisms of Pharmacological Action; Chelating Agents; Sequestering Agents; Caseins
• Other Study ID Numbers: 1-14

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: No