An integrated framework using high-dimensional mass cytometry and fluorescent flow cytometry identifies discrete B cell subsets in patients with red meat allergy.

Abstract

B cells play a critical role in the development and maintenance of food allergy by producing allergen-specific IgE. Despite the importance of B cells in IgE-mediated food allergy, the identity of sIgE-producing human B cells and how IgE is regulated are poorly understood. To identify the immunophenotypes of circulating B cells associated with the production of galactose-alpha-1,3-galactose-specific IgE production in patients with red meat allergy. B cells in PBMC samples obtained from 19 adults with physician-diagnosed red meat allergy and 20 non-meat allergic healthy controls were assessed by mass cytometry along with a bioinformatics analysis pipeline to identify discrete B cell phenotypes that associated with serum sIgE. Fluorescent flow cytometry was then applied to sort purify discrete B cell subsets, and B cells were functionally evaluated on an individual cell level for the production of sIgE by ELISPOT. Discrete B cell phenotypes abundant in meat allergic subjects compared to non-meat allergic controls were found in peripheral blood that do not share typical characteristics of classical isotype-switched memory B cells that express high levels of CD27. These B cell subsets shared higher IgD and lower IgM expression levels coupled with CXCR4, CCR6 and CD25 expression. In vitro polyclonal stimulation of purified B cell subsets from meat allergic subjects demonstrated that these subsets were enriched for cells induced to secrete sIgE. Circulating B cells display increased abundance of discrete B cell subsets in meat allergic subjects. This observation, coupled with the capacity of individual B cell subsets to produce sIgE following activation, implicates these novel B cell phenotypes in promoting IgE in meat allergy.