Multiple Subsets of T Helper 2 Lung-resident Memory Cells Are Established in a Mouse Model of Allergic Inflammation

Asthma is one of the most common chronic respiratory diseases. Clinical studies have shown that asthma represents a spectrum of inflammatory phenotypes, including predominantly eosinophilic, mixed eosinophilic/neutrophilic, and paucigranulocytic signatures. To investigate the mechanism underlying the heterogeneity of inflammatory phenotypes in asthma, we employed a well-established house dust mite (HDM) challenge model in mice and performed Il13-Cre-mediated lineage-tracing of T helper 2 tissue resident memory cells (Th2-TRMs) in the lung. The lineage-labelled Th2-TRMs were HDM-specific based on Derp1 tetramer labeling and their quick activation in response to HDM stimulation in precision-cut lung slices prepared from mice previously challenged with HDM. Since allergen-specific Th2-TRMs are a central mediator of anamnestic inflammation in asthma, we investigated the molecular phenotypes of IL13-lineage labelled Th2-TRMs by Single cell RNA sequencing (scRNAseq). We identified a major TRM2 subset (Gata3+ only) and additional subsets that co-expressed specific markers of Th1 (TRM2+1), Th17 (TRM2+17), and Treg (TRM2+reg) lineages, which reveals significant heterogeneity of allergen-specific, Th2-TRMs in the lung. By assessing the kinetics of marker expression among Il13-lineage labelled, CD4+ T cells following HDM challenges, we found that the Th cells co-expressing Th2 and Th17 lineage markers were generated in the acute phase of inflammation and subsequently established as TRM2+17 subset in the lung over time. Ongoing experiments aim to study how the heterogeneity of Th2-TRMs is regulated and whether different subsets play distinct roles in eosinophilic and neutrophilic inflammation upon HDM re-challenges. Our ultimate goal is to identify novel mechanisms underlying the spectrum of inflammatory phenotypes in asthma so that effective therapeutics targeting each asthma endotype can be developed.