Chronic inorganic nitrate supplementation does not improve metabolic health and worsens disease progression in mice with diet-induced obesity.

Inorganic nitrate (NO(3)(-)) has been proposed to be of therapeutic use as a dietary supplement in obesity and related conditions including the metabolic syndrome (MetS), type II diabetes, and metabolic dysfunction-associated steatotic liver disease (MASLD). Administration of NO(3)(-) to endothelial nitric oxide synthase-deficient mice reversed aspects of MetS; however, the impact of NO(3)(-) supplementation in diet-induced obesity is not well understood. Here we investigated the whole body metabolic phenotype and cardiac and hepatic metabolism in mice fed a high-fat, high-sucrose (HFHS) diet for up to 12 mo of age, supplemented with 1 mM NaNO(3) (or NaCl) in their drinking water. HFHS feeding was associated with a progressive obesogenic and diabetogenic phenotype, which was not ameliorated by NO(3)(-). Furthermore, HFHS-fed mice supplemented with NO(3)(-) showed elevated levels of cardiac fibrosis and accelerated progression of MASLD including development of hepatocellular carcinoma in comparison with NaCl-supplemented mice. NO(3)(-) did not enhance mitochondrial β-oxidation capacity in any tissue assayed and did not suppress hepatic lipid accumulation, suggesting it does not prevent lipotoxicity. We conclude that NO(3)(-) is ineffective in preventing the metabolic consequences of an obesogenic diet and may instead be detrimental to metabolic health against the background of HFHS feeding. This is the first report of an unfavorable effect of long-term nitrate supplementation in the context of the metabolic challenges of overfeeding, warranting urgent further investigation into the mechanism of this interaction.NEW & NOTEWORTHY Inorganic nitrate has been suggested to be of therapeutic benefit in obesity-related conditions, as it increases nitric oxide bioavailability, enhances mitochondrial β-oxidation, and reverses metabolic syndrome in eNOS(-/-) mice. However, we here show that over 12 months nitrate was ineffective in preventing metabolic consequences in high fat, high sucrose-fed mice and worsened aspects of metabolic health, impairing cholesterol handling, increasing cardiac fibrosis, and exacerbating steatotic liver disease progression, with acceleration to hepatocellular carcinoma.