Biomedical subjects
Cathie Connaughton
Publications and source records attributed to Cathie Connaughton.
A neonatal swine model for peanut allergy.
BACKGROUND: Peanut allergy represents a significant health threat in the United States. The factors contributing to the severity of the allergic response and the immunopathogenic mechanisms underlying peanut allergy remain to be completely characterized. As yet, no animal model has been developed that will completely mimic the physical, immunologic, and histologic features of food allergy. OBJECTIVE: The purpose of this investigation was to develop a neonatal pig model of peanut allergy that would mimic the allergic symptoms and the immunologic and histologic profile of human peanut allergy. METHODS: Newborn piglets sensitized intraperitoneally with peanut extract and cholera toxin were orally challenged repeatedly with peanut meal. Physical symptoms, including emesis, lethargy, diarrhea, and respiratory distress, were monitored to determine the allergic response. Immunologic assessment was conducted through use of skin testing and the antigenic response to peanut proteins. Histologically, tissues derived from the esophagus, stomach, small intestine, and colon were assessed for morphologic changes after the oral challenge. RESULTS: Peanut-sensitized pigs responded with physical symptoms that mimicked those seen in double-blinded, placebo-controlled oral food challenges to peanuts in children and adults. Skin testing suggested an IgE-mediated response; this was confirmed by a negative passive cutaneous anaphylaxis response of heat-treated sera obtained from peanut-sensitized animals. Damage to villi of the small intestine was similar to that seen in endoscopically obtained tissue specimens from certain food-allergic individuals. CONCLUSION: The neonatal pig model of peanut allergy mimics the physical and immunologic characteristics of peanut allergy in human beings. The model will be useful for determining IgE-mediated mechanisms and conducting endoscopic histologic assessment of tissues and immunotherapeutic intervention strategies with repeated allergen challenges.
Modification of peanut allergen Ara h 3: effects on IgE binding and T cell stimulation.
BACKGROUND: Peanut allergy is a major health concern due to the increased prevalence, potential severity, and chronicity of the reaction. The cDNA encoding a third peanut allergen, Ara h 3, has been previously cloned and characterized. Mutational analysis of the Ara h 3 IgE-binding epitopes with synthetic peptides revealed that single amino acid changes at critical residues could diminish IgE binding. METHODS: Specific oligonucleotides were used in polymerase chain reactions to modify the cDNA encoding Ara h 3 at critical IgE binding sites. Four point mutations were introduced into the Ara h 3 cDNA at codons encoding critical amino acids in epitopes 1, 2, 3 and 4. Recombinant modified proteins were used in SDS-PAGE/Western IgE immunoblot, SDS-PAGE/Western IgE immunoblot inhibition and T cell proliferation assays to determine the effects of these changes on in vitro clinical indicators of peanut hypersensitivity. RESULTS: Higher amounts of modified Ara h 3 were required to compete with the wild-type allergen for peanut-specific serum IgE. Immunoblot analysis with individual serum IgE from Ara-h-3-allergic patients showed that IgE binding to the modified protein decreased approximately 35-85% in comparison to IgE binding to wild-type Ara h 3. Also, the modified Ara h 3 retained the ability to stimulate T cell activation in PBMCs donated by Ara-h-3-allergic patients. CONCLUSIONS: The engineered hypoallergenic Ara h 3 variant displays two characteristics essential for recombinant allergen immunotherapy; it has a reduced binding capacity for serum IgE from peanut-hypersensitive patients and it can stimulate T-cell proliferation and activation.