CD163 and Tim-4 identify resident intestinal macrophages across sub-tissular regions that are spatially regulated by TGF-β

Abstract In bodily organs, macrophages are localised in poorly understood tissular and sub-tissular niches associated with defined macrophage ontogeny and activity. In the intestine, a paradigm is emerging that long-lived macrophages are dominantly present in the muscular layer, while highly monocyte-replenished populations are found in the lamina propria beneath the epithelial barrier. Whether longevity is restricted in such a simplified manner has not been well explored. Moreover, the impact of specific gut-associated factors on long-lived macrophage functionality and niche occupancy is unknown. We generated sc-RNA-Seq data from wild-type and Ccr2 −/− mice to identify phenotypic features of long-lived macrophage populations in distinct intestinal niches and identified CD163 as a useful marker to distinguish submucosal/muscularis (S/M) from lamina propria (LP) macrophages. Challenging the emerging paradigm, long-lived macrophages, identified by Tim-4 expression, were found in the LP and S/M. Long-lived LP macrophages are restrained in their response to proinflammatory stimulation compared to short-lived populations in the same location, and to the long-lived population within the S/M. Employing a novel Timd4 cre Tgfbr2 fl/fl mouse line we demonstrate distinct functions of TGF-β on long-lived macrophages in these two compartments. Importantly, in Timd4 cre Tgfbr2 fl/fl mice, zonation of CD163 + macrophages in the S/M was lost, suggesting TGF-β plays an unappreciated role in positioning of macrophages in the tissue. These data highlight the importance of considering ontogeny and niche when assessing the action of key intestinal regulatory signals.