Transcriptional analysis of B cells from patients with alpha-gal allergy

Abstract

Patients who develop alpha-gal allergy have tolerated mammalian meat and products for years. Understanding the shift in transcriptional programming of alpha-gal IgE-producing B cells is critical to elucidating the switch from immune tolerance to allergen reactivity. B cells were enriched from alpha-gal allergic and control subjects by negative selection and sorted for CD27highCD38highIgE+aGal+CD138- plasmabast, one cell/well into a 96-well BD precise plate. Target genes were amplified, sequenced and data were analyzed using BD genomic data view software. In conjunction, additional enriched B cell preparations from control and alpha-gal-allergic subjects were analyzed for targeted gene expression using digital barcoded platform. We detected AG+IgE+ plasmablast in the blood of recent tick bitten subject with median percentage of 0.054 (25% Percentile 0.013, 75th percentile 0.105, N=13). Further a positive correlation was observed between alpha-gal sIgE and alpha-gal+IgE+ plasmablast. Projection of data with tSNE plot suggested that genes from subjects with high sIgE annotated together. An increase in gene expression of transcription factors and pseudogenes involved in transcriptional regulation were observed in subjects with high sIgE. Upregulation of TNF gene expression as well as other inflammation-related products was found in alpha-gal allergic subjects without influence of alpha-gal sIgE titer. Subjects with alpha-gal allergy appear to have a strikingly higher percentage of circulating plasmablasts than control subjects. Moreover, these plasmablasts express a distinct transcriptional repertoire consistent with a robust inflammatory stimulus which likely explains the shift from immune tolerance of red meat to clinical food allergy.