Neutrophils condition lung lipid homeostasis in a spatially dependent manner 3172

Description We recently reported dysregulation of lung lipid homeostasis linked to neutrophil immunometabolism during bacterial infection. We sought to extend these findings to understand neutrophil conditioning of the lung niche from the lipid perspective. We mapped lipid populations in resting (untreated), neutropenic (cyclophosphamide), and stimulated (LPS/fMLP) lungs from adult, female C57BL/6 mice. Briefly, triplicate whole lungs were prepared by gelatin inflation, snap frozen, thin sectioned, thaw-mounted, and prepared for mass spectrometry imaging (MSI) of lipids and metabolites by our previously published methods. Spatial lipidomic data was collected at both 10um and 50um spatial resolution to produce near-single cell and cell-neighborhood lipid maps. In all conditions, the lungs had an overall normal appearance with no notable gross histological changes. Resting lungs showed distinct ion patterns in the parenchyma compared to neutropenic lungs and were characterized by mono- and poly-unsaturated fatty acid-containing phospholipids. Stimulated lungs revealed a lipid signature driven by phosphatidylserine and phosphatidylinositol (PI) lipids. These results further validate the observation that neutrophil migration into the lung is coincident with an increase in PI 18:0/20:4 during bacterial infection and that this increase is spatially distinct in the parenchyma compared to the airways. These results highlight the spatial dependence of the lung lipid landscape.