Amiloride reduces ENaC content and enhances endothelial dependent dilation in mesenteric arteries of female mice fed a Western diet
Authors: Marshall Kurt Daniel , Aroor Annayya R , Ramirez‐Perez Francisco I , Habibi Javad , DeMarco Vincent G , Jia Guanghong , Garro Mona , Whaley‐Connell Adam , Nistala Ravi , Sowers James R , Martinez‐Lemus Luis A 
Published: November 2023

Consumption of a Western diet (WD), high in fat and refined sugars is associated with increased arterial stiffness and endothelial dysfunction, which have been shown to reduce the cardiovascular disease (CVD) protection naturally afforded to premenopausal women. We have also previously shown that WD increases endothelial content and activity of the epithelial sodium channel (ENaC), and that this can be attenuated with the ENaC inhibitor, amiloride. Therefore, we hypothesized that a low dose, blood pressure‐ineffective amiloride treatment would blunt WD associated increases in arterial stiffness in female mice, and that this reduction in stiffness would be accompanied by alleviated deficits in endothelial function. Female mice (n=6/group) were fed a WD (46% fat, 17.5% sucrose and 17.5% high fructose corn syrup) beginning at 4 weeks of age and continuing for 20 weeks with half of the mice receiving amiloride (1 mg/kg/day) in drinking water at the initiation of WD feeding. Amiloride treatment did not alter body fat content, insulin sensitivity or blood pressure, but reduced proteinuria, likely due to changes in arterial stiffness. Additionally, amiloride reduced WD‐induced aortic stiffness by 20% as assessed by pulse wave velocity, endothelial cell stiffness by 47% as assessed by atomic force microscopy and enhanced maximal acetylcholine‐induced endothelial‐dependent dilation by 50% (P<0.05). Structurally, amiloride reduced collagen and nitrotyrosine staining in the aorta (P<0.05), without altering endothelial ENaC content. In mesenteric arteries, amiloride decreased endothelial ENaC content and induced the arterialization of vessels characterized by increased presence of fibrillar actin, collagen and elastin (P<0.05). This was associated with a 70% enhanced endothelial‐dependent dilation (P<0.05), without affecting endothelial‐independent dilation. Amiloride treatment resulted in WD‐fed mesenteric arteries having a left shifted strain‐stress relationship, which was caused by a diminished capacity of vessels to achieve passive diameters at low intravascular pressures similar to those of mice not treated with amiloride. However, amiloride treatment did not alter the incremental modulus of elasticity or overall compliance of mesenteric arteries, and changes in maximal passive diameter or wall cross sectional area were not detected. These data indicate that low dose amiloride treatment reduces WD‐induced aortic stiffening and enhances overall conduit and resistance vessel endothelial function in female mice. The findings presented herein are highly translatable and suggest that treatment with a blood pressure ineffective dose of amiloride may be used to ameliorate CVD in obese females.Support or Funding InformationThis work was supported by grants from the National Institutes of Health (HL‐088105 to L.A.M‐L.; HL‐073101 and HL‐107910 to J.R.S.) and the Department of Veterans Affairs Biomedical Laboratory Research and Development (0018 to J.R.S.).