Reactive Nitrogen Signaling through Fatty Acid Nitration and Sugar Nitrosation: A New Paradigm in Nutrient-Physiology Interactions - SUMMARY Nitrogen is essential to life and exists in many oxidation states. In its reduced form, it is a key component of amino acids and nucleotides—the building blocks of proteins and DNA. In the environment, nitrogen is present in its oxidized forms (nitrite and nitrate), which participate in microbial metabolism and the global nitrogen cycle. Mammals evolved to use intermediate oxidation states of nitrogen (nitric oxide (•NO) and nitrogen dioxide (•NO2)) as mediators of vasodilation, neurotransmission, and inflammation. In humans, dietary nitrate is concentrated in saliva and converted to nitrite by oral microbes. In the acidic stomach environment, nitrite undergoes chemical reactions that modify dietary components, particularly unsaturated fatty acids. This leads to the formation of nitrated fatty acids (NO2-FAs), electrophilic signaling molecules with potent anti-inflammatory, antimicrobial, and tissue-protective effects. NO₂-FAs act by covalently modifying regulatory proteins through nitroalkylation, altering their structure, function, and activity, and thereby influencing key signaling pathways and gene expression programs. Mechanistic studies in human cells and animal models revealed that NO2-FAs suppress inflammation through modulation of cytokine expression, inhibition of Nf-kB, STING, and STAT3 pathways, and activation of AMPK. The protective and beneficial effects of NO2-FAs have been demonstrated in several preclinical animal models of inflammation, fibrosis, hypertension, ischemia/reperfusion injury, and nephropathy. Clinical studies demonstrated that dietary CLA and nitrate/nitrite supplementation elevate circulating NO2-CLA to levels comparable with those achieved in Phase I trials of NO2-oleic acid (NO2-OA), which is in clinical development. These findings highlight a dietary-gastric axis for forming bioactive nitrogen oxides and lipid mediators with therapeutic potential. This R35 proposal will advance the understanding of endogenous nitration/nitrosation processes and support the translation of NO2-FA biology into disease-modifying strategies. It investigates the mechanisms and physiological responses to the nitrite-dependent reaction products. In addition to conjugated fatty acids, we recently learned that the more abundant hydroxy groups present in carbohydrates effectively and dose-dependently quench the reaction, forming novel bioactive products. Building on our R01-funded discoveries, the overarching goal of this R35 project is to investigate the signaling mechanisms and biological impact of lipid and carbohydrate modification during gastric digestion. These efforts are aimed at 1) consolidating our research on nitrated fatty acids and 2) opening a new line of investigation into the novel modifications of carbohydrates.
