Pulmonary macrophage subsets display distinct metabolic responses to polarising stimuli in vivo

Macrophage activation is underpinned by metabolic changes required to fight infection, resolve inflammation and enable effective wound healing. While metabolic control of macrophage activation is increasingly understood in culture systems in vitro, it remains poorly understood in more complex in vivo settings, like the lung. Here we applied novel flow cytometry based immunometabolic techniques to profile immune cell metabolism in the murine lung. We revealed a surprising role for glucose in naive alveolar macrophages (AMs) that was retained by AMs polarised in vivo with either interleukin-4 (IL-4) or lipopolysaccharide (LPS). We identified that naive interstitial macrophages (IMs) were dependent on mitochondrial and glucose metabolism, IMs polarised with IL-4 failed to induce metabolic alterations but displayed a glycolytic phenotype following LPS exposure as they adopted an M1 like metabolic profile. We also demonstrated that AMs were metabolically less responsive than IMs to intranasal delivery of LPS, but upregulated glycolysis and metabolic features of M2 polarisation (defined in vitro) in response to intranasal IL-4, including oxidative phosphorylation (OXPHOS), fatty acid oxidation (FAO) and arginine metabolism. Finally, we identified AM M2 polarisation as highly sensitive to glucose inhibition ex vivo. Thus, lung macrophage subsets display distinct metabolic responses to polarising stimuli in vivo. HighlightsO_LINaive alveolar macrophages require glucose metabolism despite residing in a low glucose environment. C_LIO_LIAlveolar macrophages are more responsive to IL-4 in vivo than LPS upregulating oxidative metabolism, lipid metabolism and glycolysis. C_LIO_LIInterstitial macrophages adopt a glycolytic phenotype characteristic of M1 BMDMs in vitro following in vivo LPS administration. C_LIO_LIAlternatively activated alveolar macrophages are extremely sensitive to glucose inhibition ex vivo. C_LI