Panel A: Flow cytometry plots show embryonic and postnatal pre-alveolar macrophage (preAM) and alveolar macrophage (AM) populations at different developmental stages including embryonic day 17.5 (E17.5), embryonic day 18.5 (E18.5), day after birth 1 (DAB1), day after birth 5 (DAB5), and day after birth 10 (DAB10) in Rxrabfl/fl and VaviCre+Rxrabfl/fl mice. The plots are gated on DAPI- CD45+ CD11c+ F4/80int cells and DAPI- CD45+ Ly6G- single cells. Different marker combinations including CD11b, SiglecF, Ly6C, and F4/80 distinguish preAM and AM populations and demonstrate altered maturation in VaviCre+Rxrabfl/fl mice. Panel B: A bar graph quantifies the total cell numbers of embryonic and postnatal macrophage populations shown in panel A for Rxrabfl/fl and VaviCre+Rxrabfl/fl mice. Significant reductions or increases between genotypes are indicated by asterisks representing statistical significance levels. Panel C: Flow cytometry plots on the left show mature alveolar macrophage (mAM) and immature alveolar macrophage (ImmAM) subpopulations in 9-week-old Rxrabfl/fl and VaviCre+Rxrabfl/fl mice. The accompanying bar graph on the right quantifies the proportions or total numbers of these AM subpopulations and highlights significant genotype-dependent differences. Panel D: A schematic diagram illustrates the competitive bone marrow (BM) transplantation procedure in which donor bone marrow cells from different genotypes are transplanted into recipient mice to evaluate alveolar macrophage reconstitution and competitive fitness. Panel E: A bar graph quantifies peripheral blood (PB) chimerism among CD11b-positive cells following bone marrow transplantation, demonstrating the contribution of donor-derived hematopoietic cells in recipient mice. Panel F: Flow cytometry plots show mature alveolar macrophages (mAMs) and immature alveolar macrophages (ImmAMs) in lung tissue following bone marrow transplantation for different donor genotypes, illustrating altered macrophage differentiation capacity. Panel G: A bar graph quantifies the proportion or number of Rxrabfl/fl-derived versus Cd11cCre positive Rxrabfl/fl-derived mature and immature alveolar macrophages after bone marrow transplantation, with significant differences marked by asterisks.
RXR signaling is necessary for correct AM differentiation and maturation. (A) Representative flow cytometry plots showing embryonic and postnatal preAM and AM populations. (B) Quantification of A, showing total cell numbers of the selected populations in Rxrabfl/fl and VaviCre+Rxrabfl/fl mice. n = 7–10 per genotype from two independent experiments. (C) (Left) Representative flow cytometry plots showing AM subpopulations and (right) quantification in 9-wk-old Rxrabfl/fl and VaviCre+Rxrabfl/fl mice. n = 4–5 per genotype; data representative from two independent experiments. (D) Competitive BM transplant procedure. (E) Quantification of PB chimerism in CD11b+ cells. (F) Representative flow cytometry plots showing mAMs and ImmAMs in lungs after BM transplant. (G) Quantification of Rxrabfl/fl versus Cd11cCre+Rxrabfl/fl-derived mAMs and ImmAMs after BM transplant. n = 9–10 per genotype; data are from two pooled independent experiments. All data are shown as means ± SEM; *P ≤ 0.05; **P ≤ 0.01; ****P ≤ 0.0001; unpaired Student T test.