Modeling Early Events in Food Sensitization: Complementary Insights from Caco-2 and T84 Epithelial Barriers Exposed to Peanut Allergens

Food allergies are increasing worldwide, yet the early epithelial mechanisms that initiate allergic sensitization remain incompletely defined. As the intestinal epithelium governs both allergen translocation and epithelial-immune crosstalk, it constitutes a critical but underutilized model for predicting allergenicity. In this study, we used Caco-2 and T84 intestinal epithelial monolayers cultured on Transwell^® inserts to compare barrier properties and responses to peanut protein extract. Phenotypic characterization included biomarker profiling, transepithelial electrical resistance (TEER) measurements, tight junction integrity assessment, and analysis of cytokine levels as well as oxidative and nitrosative stress. Peanut exposure caused moderate TEER reductions without overt tight junction disruption while allowing translocation of the major allergen, Arachis hypogaea allergen 1 (Ara h 1), likely via transcellular pathways. Peanut protein extracts also induced epithelial stress responses, characterized by increased reactive oxygen species and nitric oxide production, alongside time-dependent secretion of innate and type 2-associated mediators, including IL-1β, TSLP, IL-25, and IL-33, indicating epithelial activation in the absence of complete barrier breakdown. Notably, basolateral supernatants from peanut-exposed epithelial monolayers activated THP-1-derived macrophages and enhanced IL-6 secretion, demonstrating that limited allergen passage across an otherwise intact epithelial barrier is sufficient to elicit early innate immune responses. Collectively, these findings indicate that peanut extract induce subtle functional perturbations in the intestinal epithelium while promoting downstream immune activation, highlighting Caco-2 and T84 cells as complementary in vitro platforms for studying barrier-dependent mechanisms of allergic sensitization.