Endothelial Mineralocorticoid Receptors Contribute to Sex‐Dependent Vascular Inflammation and Plaque Vulnerability in Atherosclerosis

Abstract

BACKGROUNDIschemic complications of atherosclerosis are the leading cause of death in the USA. Ischemic events, including heart attacks and strokes, are predominantly caused by rupture and thrombosis of inflamed or “vulnerable” atherosclerotic plaques. Clinical studies show that excess of the hormone aldosterone predisposes to cardiovascular ischemia and that inhibition of the aldosterone‐binding mineralocorticoid receptor (MR) prevents these adverse outcomes. We previously showed that MR is expressed in human vascular endothelial cells (ECs) and contributes to leukocyte adhesion to ECs in culture.We hypothesize that EC‐specific MR promotes vascular inflammation in hyperlipidemia, thereby contributing to development of atherosclerotic plaques with a vulnerable phenotype.METHODS & RESULTSWe first investigated the effect of whole‐body MR inhibition with spironolactone (Spiro) on vascular inflammation in the ApoE‐KO atherosclerotic mouse model fed high fat diet (HFD) for 8 weeks. Using flow cytometry to quantify specific leukocyte populations in the aortic arches of male ApoE‐KO mice, preliminary results reveal that Spiro reduces total leukocytes, T cells, and monocytes within the aortic arch by up to 80% compared to vehicle. In vitro treatment of human coronary artery ECs with oxidized phospholipids to mimic atherogenic conditions increased leukocyte adhesion by 50% (p<0.01), which was blocked by pretreatment with Spiro. This suggests that MR in ECs contributes to leukocyte adhesion in dyslipidemia. To explore the role of EC‐MR in vascular inflammation due to hyperlipidemia in vivo, transgenic mice with MR specifically deleted from ECs (VE‐Cadherin Cre x floxed MR) were injected with an adeno‐associated virus expressing human PCSK9 (AAV‐PCSK9) and fed HFD for 12 weeks, resulting in severe hyperlipidemia (fasting cholesterol >1000 mg/dL) and atherosclerosis. Cardiovascular risk factors and other potential confounders were measured: there was no difference in serum cholesterol, fasting glucose, body weight gain, blood pressure, or serum aldosterone levels between EC‐MR‐KO and MR‐intact mice of the same sex. Histologic analysis of plaques in the aortic root revealed that EC‐MR‐KO did not affect overall plaque size. However in EC‐MR‐KO mice of both sexes, plaque lipid content tended to be lower (p=0.09) and inflammatory marker staining was reduced (p=0.05), a phenotype consistent with more stable plaques with EC‐MR KO. Quantitative flow cytometry analysis of cells in the aortic arch revealed that EC‐MR‐KO males had a 40% reduction in total leukocytes (p<0.05) and monocytes (p=0.05) compared to MR‐intact males. MR‐intact females had 50–60% fewer intravascular leukocytes (p<0.001), T cells (p<0.01), and monocytes (p<0.01) than MR‐intact males, and this was not further decreased by EC‐MR KO.CONCLUSIONSMR inhibition attenuates atherosclerotic plaque inflammation in male mice. Endothelial MR contributes directly to leukocyte adhesion to ECs in vitro and to plaque inflammation in vivo in dyslipidemic males. Females are protected from vascular inflammation even with intact EC‐MR. Our results suggest a new mechanism for the higher incidence of cardiovascular ischemia and death in people with increased MR activation and for sex differences in the incidence of ischemic cardiovascular events. Further studies are underway to determine the mechanisms for the proinflammatory role of EC‐MR in atherosclerosis and for the observed sex difference.Support or Funding InformationThis work was funded by: NIH/NHLBI HL095590 (to IZJ) and AHA EIA18290005 (to IZJ)