Investigation of the influence of dietary nitrate on cardiovascular function and on the oral microbiome in mouse models of atherosclerosis

PhD ThesisReduced endothelium-derived nitric oxide (NO) bioavailability is a characteristic of endothelial dysfunction thought to partly account for the enhanced platelet activity in cardiovascular disease (CVD). Supplementation with dietary nitrate (NO3 - ) has been shown to be effective for in-vivo delivery of exogenous NO and this relies on commensal oral bacteria presenting a nitrate reductase enzyme able to reduce NO3 - to nitrite, then reduced, within the mammalian vasculature, in bioavailable NO. Dietary NO3 - supplementation suppresses ex-vivo platelet activation in healthy volunteers and in hypercholesterolemic patients. Whether dietary NO3 - is an effective antiplatelet agent in a scenario of chronic NO deficiency is unknown. Furthermore, the impact of long-term NO3 - supplementation on CV function and on the composition of the oral microbiome population in a scenario of CVD is currently unclear. The aim of this investigation was to test the impact of NO3 - on platelet function in-vivo and ex-vivo in mouse models of endothelial dysfunction (eNOS-/-) and atherosclerosis (ApoE-/-). Secondly, I assessed the effect of a long-term NO3 - supplementation and whether its effects are sustained once supplementation ceases upon cardiovascular function in the ApoE-/- mouse by monitoring blood pressure (BP), tail bleeding time and cardiac function. Simultaneously, the impact of the intervention upon the oral microbiome population was assessed through culture of oral swabs and DNA sequencing of samples collected before, during and following ceasing of the supplementation with NO3 -. Supplementation with dietary NO3 - in the setting of atherosclerosis reduces platelet reactivity and decreases BP suggesting that NO3 - ingestion delivers bioavailable NO within the vasculature in the setting of NO deficiency and these effects are sustained with long-term NO3 - supplementation and for substantial duration following cessation of supplementation. These effects were associated with a shift in the oral microbiome towards a population linked with health state, primarily mediated by a restoration of the abundance of Streptococcus danieliae, indicating a potential for nitrate to prevent dysbiosis associated with CVD and development of periodontitis in previous investigations. Together my findings suggest that dietary NO3 - when incorporated as part of a lifestyle change is likely to affect substantial improvements in the cardiovascular function that are sustained for several weeks in the absence of the dietary intervention. Supplementation with dietary NO3 - in the setting of atherosclerosis reduces platelet reactivity and decreases BP suggesting that NO3 - ingestion delivers bioavailable NO within the vasculature in the setting of NO deficiency and these effects are sustained with long-term NO3 - supplementation and for substantial duration following cessation of supplementation. These effects were associated with a shift in the oral microbiome towards a population linked with health state, primarily mediated by a restoration of the abundance of Streptococcus danieliae, indicating a potential for nitrate to prevent dysbiosis associated with CVD and development of periodontitis in previous investigations. Together my findings suggest that dietary NO3 - when incorporated as part of a lifestyle change is likely to affect substantial improvements in the cardiovascular function that are sustained for several weeks in the absence of the dietary intervention