Inorganic Nitrite Supplementation Improves Endothelial Function With Aging

Abstract

Supplemental Digital Content is available in the text. To determine the efficacy of inorganic nitrite supplementation on endothelial function in humans and mechanisms of action, we performed (1) a randomized, placebo-controlled, parallel-group clinical trial with sodium nitrite (80 mg/day, 12 weeks) in older adults (N=49, 68±1 year) and (2) reverse-translation experiments in young (6 months) and old (27 months) c57BL/6 mice. In the clinical trial, sodium nitrite increased plasma nitrite (P<0.05) and was well tolerated. Brachial artery flow-mediated dilation (endothelial function) was increased 28% versus baseline after nitrite supplementation (P<0.05) but unchanged with placebo. Nitrotyrosine, a marker of oxidative stress, was reduced by 45% versus baseline in biopsied endothelial cells after nitrite, but not placebo, treatment. Plasma from nitrite-treated, but not placebo-treated, subjects decreased whole-cell (CellROX) and mitochondria-specific (MitoSOX) reactive oxygen species in cultured human umbilical vein endothelial cells (P<0.05). Old mice (old [27 months] control, n=9) had ≈30% lower ex vivo carotid artery endothelium-dependent dilation (EDD) versus young mice (young [6 months] control, n=9) due to reduced NO bioavailability (P<0.05). Nitrite supplementation (drinking water, 50 mg/L, 8 weeks) restored EDD and NO bioavailability in old mice (n=10) to (6 months) control. Mitochondrial reactive oxygen species suppression of EDD was present in old control (increased EDD with a mitochondrial-targeted antioxidant, P<0.05) but not in young control or old mice supplemented with sodium nitrite. A mitochondrial reactive oxygen species inducer (rotenone) further impaired EDD in old control (P<0.05); young control and old mice supplemented with sodium nitrite were protected. Markers of mitochondrial health were greater in aorta of old mice supplemented with sodium nitrite versus old control (P<0.05). Inorganic nitrite supplementation improves endothelial function with aging by increasing NO, decreasing mitochondrial reactive oxygen species/oxidative stress, and increasing mitochondrial stress resistance. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT02393742.