Single cell gene expression profile of bronchoalveolar lavage fluid (BALF) cells from Apoa1-/- and Apoa1+/+ mice challenged with LPS and house dust mite
Neutrophils play important roles in inflammatory airway diseases.  Here, we assessed whether apolipoprotein A-I (apoA-I) modifies neutrophil heterogeneity as part of the mechanism by which it attenuates acute airway inflammation.  Neutrophilic airway inflammation was induced by daily intranasal administration of LPS plus house dust mite (LPS+HDM) to Apoa1-/- and Apoa1+/+ mice for 3 days. Single cell RNA sequencing was performed on cells recovered in bronchoalveolar lavage fluid (BALF) on day 4.  Unsupervised profiling identified 10 clusters of neutrophils in BALF from Apoa1-/- and Apoa1+/+ mice.  LPS+HDM-challenged Apoa1-/- mice had an increased proportion of the Neu4 neutrophil cluster that expressed S100a8, S100a9, and Mmp8, and had high maturation, aggregation, and TLR4 binding scores.  There was also an increase in the Neu6 cluster of immature neutrophils, whereas neutrophil clusters expressing interferon-stimulated genes were decreased.  An unsupervised trajectory analysis showed that Neu4 represented a distinct lineage in Apoa1-/- mice.  LPS+HDM-challenged Apoa1-/- mice also had an increased proportion of recruited airspace macrophages, which was associated with a reciprocal reduction in resident airspace macrophages.  Increased expression of a common set of pro-inflammatory genes, S100a8, S100a9, and Lcn2, was present in all neutrophils and airspace macrophages from LPS+HDM-challenged Apoa1-/- mice.  Apoa1-/- mice have increases in specific neutrophil and macrophage clusters in the lung during acute inflammation mediated by LPS+HDM, as well as enhanced expression of a common set of pro-inflammatory genes.  This suggests that modifications in neutrophil and macrophage heterogeneity contribute to the mechanism by which apoA-I attenuates acute airway inflammation. Overall design: Apoa1-/- and Apoa1+/+ mice (n = 3) were challenged daily with LPS (1 ug) plus house dust mite (HDM) (25 ug) in a volume of 20 ul for 3 days, and bronchoalveolar lavage was performed on day 4. The single cell gene expression experiment was performed on BALF cells that were pooled from three Apoa1-/- and three Apoa1+/+ mice using the Chromium Next GEM Single Cell 3' Reagent Kit v3.1 (catalogue number: PN-1000121) (10x Genomics, Pleasanton, CA).