An advanced in vitro human mucosal immune model to predict food sensitizing allergenicity risk: A proof of concept using ovalbumin as model allergen

Abstract

BackgroundThe global demand of sustainable food sources leads to introduction of novel foods on the market, which may pose a risk of inducing allergic sensitization. Currently there are no validated in vitro assays mimicking the human mucosal immune system to study sensitizing allergenicity risk of novel food proteins. The aim of this study was to introduce a series of sequential human epithelial and immune cell cocultures mimicking key immune events after exposure to the common food allergen ovalbumin from intestinal epithelial cell (IEC) activation up to mast cell degranulation.MethodsThis in vitro human mucosal food sensitizing allergenicity model combines crosstalk between IEC and monocyte-derived dendritic cells (moDC), followed by coculture of the primed moDCs with allogenic naïve CD4+ T cells. During subsequent coculture of primed CD4+ T cells with naïve B cells, IgE isotype-switching was monitored and supernatants were added to primary human mast cells to investigate degranulation upon IgE crosslinking. Mediator secretion and surface marker expression of immune cells were determined.ResultsOvalbumin activates IEC and underlying moDCs, both resulting in downstream IgE isotype-switching. However, only direct exposure of moDCs to ovalbumin drives Th2 polarization and a humoral B cell response allowing for IgE mediated mast cell degranulation, IL13 and IL4 release in this sequential DC-T cell-B cell-mast cell model, indicating also an immunomodulatory role for IEC.ConclusionThis in vitro coculture model combines multiple key events involved in allergic sensitization from epithelial cell to mast cell, which can be applied to study the allergic mechanism and sensitizing capacity of proteins.

Allergenic Shrimp Tropomyosin Distinguishes from a Non-Allergenic Chicken Homolog by Pronounced Intestinal Barrier Disruption and Downstream Th2 Responses in Epithelial and Dendritic Cell (Co)Culture

“…After IEC-DC or moDC were exposed for 48 h to chicken TM or shrimp TM, primed DC were subsequently cocultured with naïve T cells to assess the functional immune response after TM exposure. The effects of OVA exposure in this sequential mucosal immune model were already recently published, revealing type 2 immune polarization when OVA was directly exposed to DC and to a lesser extent this was also the case for IEC-DC [ 19 ]. The current study shows the differential activating capacity of shrimp TM versus chicken TM in IEC and moDC (visually summarized in Supplemental Figure S4B ).…”

“…Hen’s egg-derived OVA was used as a control allergen, as it is among the top three most common food allergy inducers [ 2 ]. Previously, it was shown that OVA promotes a type 2 immune response in vitro, demonstrating its sensitizing capacity [ 19 , 26 ].…”

“…A ‘control’ condition was used in every cell culture experiment; this control medium condition was not exposed to any additional protein component. OVA was used as an allergenic protein model known to induce activation of IEC and/or moDC to drive sequential mucosal type 2 responses [ 19 , 21 ].…”

“…This coculture model to study mucosal immunity was previously described in more detail [ 19 ]. In brief, healthy donor PBMC from volunteers, who had given written informed consent for research purposes, were obtained from the Dutch Blood bank (Amsterdam, The Netherlands).…”

“…The first step in this cascade is the breaching and/or activation of the epithelial barrier followed by activation of DC and a type 2 T cell response [ 18 ]. A previous study has shown this response for ovalbumin (OVA) as model allergen, which was able to activate intestinal epithelial cells (IEC) and/or monocyte-derived dendritic cells (moDC) [ 19 ].…”