Long-term culture expanded alveolar macrophages restore full epigenetic identity in vivo

Alveolar macrophages (AM) are tissue resident macrophages of the lung that can be expanded in culture, but it is unknown to what extent culture affects their in vivo identity. Here we show that long-term ex vivo expanded mouse AM (exAM) maintain core AM gene expression but show culture adaptations related to adhesion, metabolism and proliferation. Strikingly, even after several months in culture exAM reacquired full transcriptional and epigenetic identity upon transplantation into the lung and could self-maintain in the natural niche long-term. Changes in open chromatin regions (OCR) observed in culture were fully reversible in transplanted exAM (texAM) and resulted in a gene expression profile indistinguishable from resident AM. Our results demonstrate that long-term proliferation of AM in culture does not compromise cellular identity in vivo. The demonstrated robustness of exAM identity provides new opportunities for mechanistic analysis and highlights the therapeutic potential of ex vivo expanded macrophages.