Transcriptomic analysis of airway macrophages from a murine experimental allergic airway disease model

Airway macrophages (AMs) are the most abundant leukocytes in the healthy airway lumen and have a highly specialised but plastic phenotype that is governed by the local microenvironment. AMs are thought to maintain immunological homeostasis in the steady state, but have also been implicated in the pathogenesis of allergic airway disease (AAD). To better understand these potentially contrasting AM functions, bulk RNA sequencing was performed on murine AMs obtained during experimental AAD driven by repeated house dust mite inhalation (AM[HDM]s), comparing to control AMs from non-allergic mice. AM[HDM]s showed increased expression of genes associated with antigen presentation, inflammatory cell recruitment and tissue repair, including several chemokines and matrix metalloproteinases. This was accompanied by increased expression of mitochondrial electron transport chain subunit genes and the retinoic acid biosynthetic enzyme gene Raldh2. Conversely, AM[HDM]s displayed decreased expression of a number of cell cycle genes, genes related to cytoskeletal functions and a subset of genes implicated in antimicrobial innate immunity, such as Tlr5, Il18 and Tnf. Differential gene expression in AM[HDM]s was consistent with upstream effects of the cytokines IL-4 and IFN-γ, both of which were at increased concentrations in lung tissue after HDM treatment. These data highlight diverse gene expression changes in the total AM population in a clinically relevant mouse model of AAD, overall suggestive of contributions to inflammation and tissue repair/remodelling, but decreases in certain steady state cellular and immunological functions. CD11c+ CD64+ Siglec F+ AMs were flow cytometry sorted from bronchoalveolar lavage of adult female C57BL/6J mice after 3 weeks of intranasal exposures to either house dust mite (HDM) allergen (AM[HDM]s, N=4) or PBS control (control AMs, N=5). RNA was extracted and converted to mRNA libraries before performing Illumina paired end sequencing. Reads were aligned to the Mm10 genome using COBWeb. Transcripts were quantified from aligned reads using DESeq. Low abundance transcripts were filtered (normalised expression <8 in >25 % of samples in both groups) and differential expression analysis performed by moderated t-test with Benjamini-Hochberg false discovery rate correction. Differentially expressed genes were hierarchically clustered using Ward's method and pathway analysis performed to predict changes in AM function during HDM-driven allergic airway disease.