Characterization of erythroblastic island and tissue macrophages using a novel CD169-tdTomato-Cre mouse model and scRNA-seq

Macrophages play diverse roles, including supporting erythropoiesis. CD169 is a specific marker for macrophages. However, recent studies showed that anti-CD169 staining detected non-macrophages, raising the concern for using anti-CD169 antibody to correctly assign macrophages. Genetically engineered mouse model provides another means of monitoring gene expression. However, no mouse model is currently available to directly monitor CD169 expression. Erythropoiesis relies on macrophage-mediated iron recycle for iron supply. While it is established that spleen macrophages play critical roles in erythrophagocytosis and iron recycle, whether and how other tissue macrophages are involved remain incompletely understood. To address these issues, we generated CD169-tdTomato-Cre mice. We also compared various tissue macrophages by scRNA-seq, flow cytometry and cytospin analyses. We showed that CD169-tdTomato was detected, and that CD169-Cre mediated recombination in only macrophages. Notably, bone marrow F4/80 + CD169 + macrophages resemble erythroblastic island macrophages as demonstrated by the expression of Epor along with other macrophage feature genes. Comparison of tissue macrophages revealed significant differences including erythrophagocytosis by spleen and bone marrow but not by liver macrophages at steady state, differential expression of macrophage surface markers CD169, Vcam1, Cx3cr1 and Csf1r, and iron recycle genes Hmox1, Spic and CD91, selective expression of globin genes in lung macrophages. Our findings have uncovered previously unknown differences among tissue macrophages and provide novel insights into the mechanisms by which tissue macrophages participate in iron recycle to support erythropoiesis. The novel dual functional CD169-tdTomato-Cre mouse line will have wide applications for studies of gene function in macrophages and their role in modulating erythropoiesis.