The Mineralocorticoid Receptor in Myeloid Cells Promotes Vascular Inflammation and Atherosclerosis via Transcriptional Regulation of Selplg

Abstract

Elevated plasma aldosterone (Aldo) is associated with increased risk of cardiovascular (CV) ischemia, and inhibition of the Aldo-activated mineralocorticoid receptor (MR) reduces CV-related mortality. Ischemic events including heart attack and stroke are caused by rupture of atherosclerotic plaques with abundant leukocytes. In mouse models of atherosclerosis, MR antagonists reduce plaque size and inflammation. MR in myeloid cells (MyMR) regulates macrophage (Mp) polarization in vitro, but its role in vascular inflammation in vivo is unknown. Here we test the hypothesis that MyMR contributes to vascular inflammation by facilitating leukocyte recruitment. A MyMR-knockout (KO) mouse was created (LysM-Cre/MRfl/fl) and crossed with atheroprone apolipoprotein E-KO (ApoE) mice. Eight-week-old male and female MyMR-intact/ApoE-KO and MyMR-KO/ApoE-KO littermates were fed high fat diet for 8 weeks to accelerate atherogenesis. Aortic root plaque area was modestly decreased with MyMR-KO. FACS analysis of aortic arch plaques revealed fewer Mp (37%, p < 0.01) and T cells (26%, p < 0.05) with MyMR-KO among both sexes. Female plaques were larger but had fewer myeloid (34%, p < 0.05) and T cells (30%, p < 0.01) compared to male plaques. Seven-week-old LysM-Cre/MRfl/fl mice were injected with TNFa intraperitoneally, and leukocyte trafficking was quantified in the mesenteric veins by intravital microscopy four hours later. Male and female MyMR-KO mice had 40% fewer slow-rolling cells (p < 0.0001) and 56% fewer adherent cells (p < 0.01) compared to MyMR-intact controls. There were 39% fewer slow-rolling cells in females versus males (p < 0.01). To explore mechanism, expression of leukocyte adhesion ligands was quantified by FACS in circulating leukocytes and RT-PCR in peritoneal Mp. Surface levels of P-selectin glycoprotein ligand-1 (Selplg) was attenuated on monocytes, but not on granulocytes or T cells, from MyMR-KO mice, particularly among males. Selplg mRNA was decreased by 35% in Mp from MyMR-KO mice (p < 0.05). To determine whether Selplg is transcriptionally regulated by the MR, U937 cells were pre-treated with the MR antagonist spironolactone (1 μM, Spiro) and then treated with Aldo (10 nM). Spiro alone decreased Selplg transcription (p < 0.05) compared to vehicle. Aldo increased Selplg transcription (p < 0.05) compared to vehicle, and this effect was blocked by Spiro (p < 0.0001). MyMR-KO decreases plaque size and inflammation, which may be partially due to a role of MyMR in leukocyte recruitment in early atherosclerosis likely by regulation of Selplg. The sex differences in leukocyte trafficking and plaque inflammation are worth further exploration as a mechanism for female protection from CV disease. This study suggests that MyMR regulation of vascular inflammation is a novel mechanism for the association of Aldo with increased ischemic events and supports testing of MR antagonists to prevent CV ischemia.