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IL-33 induced gene expression in activated Th2 effector cells is dependent on IL-1RL1 haplotype and asthma statusTo the Editor:Asthma is a heterogeneous respiratory disease caused by the interaction between environmental and genetic factors 1.The IL-33 and IL-1RL1 genes are strongly associated with childhood-onset and type-2 high asthma, and the asthma risk alleles amplify interleukin (IL)-33 pathway activity 2.Environmental factors, such as allergens and viral infections, trigger bronchial epithelial cells to release IL-33, which can activate signalling by binding to the IL-1RL1/IL-1RAcP receptor complex 3, and contribute to hyper-responsiveness, remodelling and chronic type 2 inflammation of the airways 4.IL-1RL1 is expressed in immune and structural cells of the airways, such as epithelial cells, mast cells, macrophages, Th2 cells and type 2 innate lymphoid cells.IL-1RL1 encodes two protein isoforms: the transmembrane receptor subunit (IL-1RL1b) and a soluble (IL-1RL1a) isoform that functions as an antagonistic decoy receptor.Human Th2 cells respond to IL-33 by enhancing cytokine production 5.Genetic variation at the IL-1RL1 locus, particularly rs1420101 in intron 5 and a block of four non-synonymous single nucleotide polymorphisms (SNPs) in full linkage disequilibrium in exon 11, alter IL-1RL1 expression levels and IL-33 induced signalling activity 2,6.However, it is not known whether genetic variation at the IL-1RL1 locus affects the response of Th2 cells to IL-33.Therefore, we tested whether IL-1RL1 haplotype altered the IL-33 induced response of Th2 cells from healthy controls and patients with asthma.Moreover, we explored whether IL-33-induced gene signatures from Th2 cells could identify subgroups of asthma patients in transcriptomic datasets.We selected peripheral blood mononuclear cells (PBMCs) from asthma patients and controls (figure 1a andb) based on the genotype of asthma-associated IL-1RL1 SNPs and grouped them into carriers of the high risk haplotype (rs1420101-AA, rs4988956-GG) or low risk haplotype (rs1420101-GG, rs4988956-AA/AG).CD4 + CD25 -T cells were isolated from PBMCs and differentiated into Th2 cells (CellXVivo, #CDK002).Th2 cells were re-stimulated through CD3 and CD28 (555725, BD Pharmingen) in the presence/absence of 100 ng•mL -1 IL-33 BioTech).RNA was extracted and sequenced using NextSeq500 (Illumina, San Diego, CA, USA), processed data can be found at GEO. Univariant (paired) analysis of differential gene expression induced by CD3/CD28 crosslinking or IL-33 was performed using Limma-voom in R, stratifying by haplotype or disease status of the donor.We generated an IL-33 response signature in Th2 cells by selecting those differentially expressed genes (DEGs) specifically expressed in T cells (using the Human Lung Cell Atlas (HLCA)) by removing genes also expressed in mast cells, macrophages and eosinophils 7.We subsequently selected the genes with top-25% baseline expression to allow detection in single-cell RNA-sequencing data and therein the genes with the top-5% largest LogFoldChange after IL-33 stimulation.Enrichment of this Th2-IL33 gene signature was analysed in single-cell RNA-sequencing data 7 and in bulk RNA-sequencing 8 data from asthma patients and controls in the INDURAIN 9 (bronchial biopsies) and U-BIOPRED 10 (induced sputum) studies using gene set variation analysis 11.Statistical tests included t-test, Wilcoxon rank-sum test, and Fisher's exact test.CD3/CD28 restimulation of Th2 cells resulted in 9299 significantly differentially expressed genes compared to unstimulated Th2 cells (figure 1c), mapping to pathways related to T cell receptor signalling, proliferation and cytokine production (not shown).No differences were observed between IL-1RL1 haplotype groups or disease categories for this response (not shown).Presence of IL-33 during CD3/CD28
Taneja et al. (Sat,) studied this question.