Marco-deficient mice have elevated aldosterone and reduced serum cholesterol. (A) Schematic depicting the murine adrenal corticosteroid biosynthetic pathway. Plasma aldosterone and corticosterone concentrations from wild type (WT) and Marco-deficient male (B, C) and female (D, E) mice as measured by ELISA. (F) Weights of both adrenal glands from WT or Marco-deficient male mice. Serum total cholesterol levels (G) and relative intra-adrenal cholesterol levels, normalised to adrenal weight, in WT and Marco-deficient male mice (H). Data in B-H were analysed by two-tailed unpaired Student’s t-test and are shown as average ± s.e.m. *P < 0.05, **P < 0.01, ***P < 0.001, **** P < 0.0001.

Marco is expressed in the lung and not adrenal glands. (A) Single cell RNA seq data plots from PMID 33571131 representing mRNA expression of Ptprc (CD45), Adgre1 (F4/80), Cd68, and Marco, in male murine adrenal glands. (B) Representative image showing a central cryosection of the male murine adrenal gland from a C57bl/6 mouse stained against CD68 (green) Marco (magenta), and DAPI (cyan). (C) Single cell RNA seq data plots from PMID 30283141 representing mRNA expression of Ptprc (CD45), Adgre1 (F4/80), Cd68, and Marco, in the male murine lung. (D) Representative image showing a cryosection of the male murine lung from a hCD68-GFP reporter mouse stained against GFP (green) Marco (magenta), and DAPI (cyan). (E) qPCR data showing the relative gene expression data for Marco in the indicated tissues. M = medulla, ZF = zona fasciculata, ZG = zona glomerulosa.

Marco-mediated elevation of aldosterone is not explained by upregulation of adrenal biosynthetic enzymes, the zona fasciculata, or renin. (A-E) qPCR data reporting the expression of adrenal corticosteroid biosynthetic enzymes between wild type (WT) and Marco-deficient male mice. (F) A schematic illustrating that pituitary-derived adrenocorticotropic hormone (ACTH) stimulates the upregulation of Cyp11b1 from the zona fasciculata (ZF) and the dexamethasone-mediated suppression of this effect. (G) A bar graph showing the plasma aldosterone concentrations of Marco-deficient mice treated with vehicle or dexamethasone- supplemented drinking water for 14 days. (H) A bar graph showing the plasma renin activity of wild type (WT) and Marco-deficient mice at steady state. All data were analysed by two- tailed unpaired Student’s t-test and are shown as average ± s.e.m. ns P>0.05. * P < 0.05.

Marco-deficient mice have enhanced expression of Ace in the lung. (A) qPCR data reporting the expression of angiotensin converting enzyme (Ace) in the lungs of wild type (WT) and Marco-deficient male mice. (B) Representative images showing a cryosection of wild-type and Marco-deficient male murine lungs stained against ACE (green) CD68 (magenta), MARCO (yellow) and DAPI (blue). Quantitation of DAPI-normalised ACE median fluorescence intensity (MFI) (C) and CD68+ myeloid cell presence (D) in the lungs of wild type and Marco-deficient male mice. (E) Single cell RNA seq UMAP plot depicting the cell types present in the male murine lung. (F) Marker heatmap showing the top three gene markers for each cell cluster in (E). (G) A dot plot showing the gene expression levels of Marco and Ace in the different cell clusters of the male murine lung. (H) A schematic, generated using BioRender, showing the working model by which Marco+ alveolar macrophages regulate aldosterone output from the adrenal gland. Data in A, C, and D, were analysed by two-tailed unpaired Student’s t-test and are shown as average ± s.e.m. *P < 0.05, **P < 0.01, ***P < 0.001, **** P < 0.0001.