Generation of Alveolar Assembloids (AlvAssemb) with Functional Alveolar-like Macrophages

Within the human lung, interplay between alveolar epithelial cells and resident macrophages shape lung development and function in both health and disease. To study these processes, we devised a strategy for co-culturing hPSC-derived alveolar epithelial organoids and induced macrophages to form functional environment, which we name alveolar assembloids (AlvAssemb). By scRNAseq and functional analysis of AlvAssemb, we identified alveolar type 2-like cells that produced GM-CSF, a response associated with tissue adaptation of macrophages, and that resident alveolar macrophage-like cells secreted interleukin-1β and interleukin-6, a core immune response function, and expressed genes required for surfactant metabolism. Remarkably, in response to alveolar epithelial injury, we found that macrophage-like cells within AlvAssemb efficiently eliminated damaged cells and absorbed oxidized lipids. Moreover, in response to lipopolysaccharide exposure and Mycobacterium tuberculosis infection, we found that core features of the human respiratory defense response were replicated in AlvAssemb. These findings support that AlvAssemb could serve as a valuable platform to investigate human lung development and pathology. We cultured iMφ with and without GM-CSF (10 ng/ml) for 7 days. The cells were then infected with M.tb (H37Rv) at MOIs of 5 and 10 for 48 hours. Following infection, RNA sequencing libraries were prepared and the results were analyzed using RNA-seq. Please note that GSM4106642 (Ni, huPrimary_0_r1) and GSM4106641 (Moi-20, huPrimary_20) data (in GSE138398) was re-analyzed in the current study.