Abstract Rationale This study provides single cell characterization of bronchial epithelial cells (BECs) from children with asthma. We hypothesized that there would be differences in BEC composition and gene expression when comparing asthma vs healthy and non-type 2 (non-T2) vs T2 asthma both at baseline and after rhinovirus infection, the most common trigger of viral-induced pediatric asthma exacerbations. Methods BECs were collected from children with (n = 14) and without (n = 5) asthma. They were differentiated to an organotypic state at an air-liquid interface for 21 days, then infected with rhinovirus (RV)-A16 (MOI 0.5). BECs were analyzed via single-cell RNAseq prior to infection and 2 days-post infection. Leiden clustering identified cell clusters that were annotated based on established gene signatures. T2 asthma was defined by accepted clinical parameters of absolute eosinophil count 300cells/mm2 and/or FeNO 35ppb if steroid naïve or 20ppb if non-steroid naïve. Results We identified 11 distinct cell clusters. Further subclustering characterized rare populations including Hillock, ionocytes, pulmonary neuroendocrine, and tuft cells. Comparing healthy and asthma, preliminary analyses did not show differences in the cell composition but identified transcriptomic differences, ranging from 50-2825 DEGs among the distinct cell populations (FDR0.01) both pre- and post-rhinovirus infection. Using clinical criteria to dichotomize type 2 vs non-type 2 asthma, there is again no difference in the cell composition, but there were transcriptomic differences at baseline and post rhinovirus infection ranging from 32-682 DEGs among distinct cell clusters (FDR 0.01). Following rhinovirus infection, non-T2 donor basal and suprabasal BECs were enriched for pathways associated with ER stress (ATF3, HSPA5, PPP1R15A, SELENOK, HERPUD1; FDR 0.01), apoptosis (GADD45A, IL18, BTG2, PMAIP1, IER3; FDR 0.01), and TNF alpha signaling (FOS, IL18, EGR1, SOCS3, DUSP5; FDR 0.01). There was also an upregulation in epithelial-mesenchymal transition related genes in suprabasal, club, and secretory cells (CCN1/2, ITGA2, GADD45A, CXCL1/8; FDR0.01). Conversely, non-T2 asthma ciliated and goblet cells demonstrated a downregulation in genes associated with interferon alpha and gamma signaling (CXCL10, IFIT3, IFI35, MX1, OAS3; FDR0.01) post infection. Conclusions Preliminary results suggest that while there are no differences in cellular composition, there are transcriptomic differences in specific bronchial epithelial cell populations at baseline and post-rhinovirus infection when comparing healthy controls to children with asthma and when comparing between T2 vs non-T2 asthma. This study provides a novel dataset, characterizing, to our knowledge, the largest pediatric asthma single-cell dataset of primary bronchial epithelial cells differentiated at ALI to an organotypic state. This abstract is funded by: NIH
Cruz et al. (Fri,) studied this question.