FcεRI Cluster Size Determines Effective Mast Cell Desensitization without Effector Responses in vitro

Abstract

Background: Allergen-specific desensitization of mast cell (MC) IgE receptors (FcεRI) is an important mechanism of allergen-specific immunotherapy that enables tolerance induction via systemic desensitization. Experimental in vitro IgE-mediated MC desensitization is a potential tool to understand the molecular mechanisms underlying this therapy. Desensitized MCs exhibit internalized IgE and its FcεRI receptors in response to suboptimal doses of allergen without provoking activation. The ovalbumin (OVA) allergen exhibits altered allergenicity upon heat treatment. MC reactions are fundamentally regulated by allergen features (i.e., allergenicity); however, the effects of allergenicity on desensitization remain unclear. Objectives: This study aimed to examine the impact of allergenicity on the establishment of in vitro MC desensitization using naive OVA (nOVA) and heated OVA (hOVA), which could induce varying MC effector responses. Method: Bone marrow-derived MCs (BMMCs) were sensitized with OVA-specific IgE, desensitized with sequentially increasing doses of nOVA or hOVA at 10-min intervals, and challenged with nOVA. To evaluate desensitization, the cell surface expression level and subcellular localization of FcεRI-bound IgE were analyzed before and after the final nOVA challenge. MC activation was determined by measuring the release of β-hexosaminidase into supernatants. Results: Desensitized cells exhibited impaired activation following OVA challenge. Both nOVA and hOVA induced BMMC desensitization under different conditions. Formation of small IgE-FcεRI cluster BMMCs, which adequately represent the desensitized state, was significant. The size of the internalized IgE-FcεRI clusters might be correlated with the desensitized state of MCs. Conclusions: We demonstrate that the optimal size of IgE-FcεRI clusters for in vitro BMMC desensitization differed significantly depending on allergenicity, and the efficacy of desensitization was reflected by IgE-FcεRI cluster formation. Our study provides information on the characteristics of IgE-FcεRI internalization for successful desensitization in vitro.