Differences in the Uptake of Ara h 3 from Raw and Roasted Peanut by Monocyte-Derived Dendritic Cells

Abstract

Roasting has been implicated in the increase of peanut allergenicity due to the chemical reactions that occur during the process. However, this increase is not fully understood, and little information is available regarding the role of roasted peanut allergens in the initial phase of allergy, where dendritic cells (DCs) play a key role. We sought to analyze differences in the internalization of Ara h 3 from raw and roasted peanut by immature monocyte-derived DCs (MDDCs) and the implication of the mannose receptor in the uptake. Ara h 3 was purified from raw and roasted peanut (Ara h 3-raw and Ara h 3-roas) and labeled with a fluorescent dye. The labeled allergens were added to MDDCs obtained from 7 donors and internalization was analyzed after 10, 30, and 120 min by flow cytometry. In parallel, mannan, which blocks the mannose receptor, was added 30 min before adding the labeled allergens. Results showed that the internalization of Ara h 3-roas by MDDCs was significantly increased at every time point. However, the increase in the internalization of Ara h 3-raw was only significant after 2 h of incubation. Ara h 3-roas had an enhanced capacity to be internalized by MDDCs in comparison with Ara h 3-raw at every time point. Blocking the mannose receptor decreased the internalization of Ara h 3-roas but not Ara h 3-raw. In conclusion, the internalization of Ara h 3-roas by the MDDCs is enhanced when compared to Ara h 3-raw, and the mannose receptor might be implicated in this enhancement.

The Effect of Digestion and Digestibility on Allergenicity of Food

“…[ 67 ]) was shown [ 68 ]. Binding of roasted Ara h 1 was shown to occur to scavenger receptor CD36 and receptor for advanced glycated end products RAGE [ 69 , 70 ], and for roasted Ara h 3 to mannose receptor [ 71 ]. Most importantly, this engagement leads to cellular signaling resulting in pro-inflammatory responses and enhanced allergic sensitization, as was shown in a mouse model comparing raw vs. roasted peanut [ 69 ].…”

Interaction of Monocyte-Derived Dendritic Cells with Ara h 2 from Raw and Roasted Peanuts

“…This could be due to the number of samples used, and a more thorough analysis will be necessary in the future. In our previous study with Ara h 3, we found a statistically significant increase in the internalization of purified Ara h 3 from roasted peanuts compared to Ara h 3 purified from raw peanuts [21]. These apparent differences between Ara h 2 and Ara h 3 might be explained by the different effects that the roasting process has on both allergens.…”

“…After these analyses, the purities of Ara h 2 from raw and roasted peanuts were found to be greater than 95%. Ara h 2 purified from raw and roasted peanuts, raw-Ara h 2, and roast-Ara h 2, respectively, were labeled with the fluorescent dye DyLight amine-reactive dye 488 NHS (Thermo Fisher Scientific, Bremen, Germany) as previously described [21]. The raw-Ara h 2 and roast-Ara h 2 labeled with the fluorescent dye were stored protected from light at −20 • C until use.…”

“…DCs with a plethora of receptors and mechanisms involved in the recognition and internalization of antigens are the key cells implicated in the higher or lower allergic sensitization to specific proteins [17][18][19][20]. In that sense, previous studies have shown that monocyte-derived dendritic cells (MDDCs) uptake and internalize Ara h 3 from roasted peanuts more efficiently than Ara h 3 purified from raw peanuts, which could explain the higher allergic sensitization potential of roasted Ara h 3 [21]. However, the interaction of DCs with highly relevant peanut allergens such as Ara h 2 and its allergenic sensitization potential has not yet been analyzed.…”

Contribution of Chemical Modifications and Conformational Epitopes to IgE Binding by Ara h 3

“…From the linear sequence of Ara h 3, 3D models have been developed [ 28 ] and the crystal structure has been solved [ 29 ]. It is important to note that it has been shown that monocyte-derived dendritic cells, which are antigen presenting cells, display differential uptake of Ara h 3 purified from raw (R), light roast (LR) and dark roast (DR) peanuts [ 30 ]. In this study, we investigated differently processed Ara h 3 at the molecular level.…”