Exploring the Effect of Inorganic Nitrates on the Human Microvascular Physiology - a Pilot Study.
Condition: Aging
Study type: Interventional

Summary: 
      We conducted a randomised, placebo-controlled, double blind, cross-over, trial to examine the
      short-term effect of concentrated beetroot juice on vascular outcomes in a young (18-35
      years) and older group (55-75 years). The study included a run-in period of one day preceding
      the test day. A wash-out period of at least 7 days was implemented to ensure to avoid a
      carry-over effect of the interventions.
    

Detailed description: 
      Cardiovascular disease (CVD) is the largest cause of global mortality accounting for over
      twenty-nine percent of all deaths with coronary heart disease and stroke projected to become
      the global leading causes of mortality by 2020. Increased mortality represents only one side
      of the problem since the clinical care of CVD is costly and prolonged and potentially diverts
      family and societal resources towards life-saving intensive medical treatments. Therefore, it
      is crucial to develop preventative and therapeutic strategies that will reduce the financial
      and social burden of CVD.

      Ageing is a major risk factor for chronic diseases such as cancer, diabetes neurodegeneration
      and CVD. For example, the prevalence and incidence of Chronic Venous Insufficiency (CVI) is
      closely linked to an age-related dysfunction of micro-circulation control. Similarly, ageing
      is associated with increased risk of atherosclerosis and formation of macroscopic lesions in
      large arteries leading to obstruction of blood flow and onset of ischaemic events . The rapid
      changes in worldwide trends of age-demographics are therefore closely correlated with the
      rising burden of CVD.

      Hypertension is an important risk factor for CVD. Raised blood pressure (BP) is primarily
      caused by increased vascular resistance, mainly due to arterial wall thickening, autonomic
      dysregulation and endothelial dysfunction . Endothelium-derived nitric oxide (NO) is a potent
      vasodilator that plays a pivotal role in the moment-to-moment control of vascular tone .
      Therefore, it is no surprise that a key aspect of endothelial dysfunction is a reduced NO
      production via the L-arginine-dependent enzymatic pathway. NO also decreases platelet
      aggregation, improves mitochondrial function, and diminishes cellular vascular adhesion .
      Hence a reduction in NO bioavailability is causally related to an increase in BP and
      atherosclerotic risk.

      Until recently it was believed that dietary nitrate (NO3-) and nitrite (NO2-) offered limited
      physiological benefits, despite them being produced endogenously as part of the NO metabolic
      pathway . The discovery of the entero-salivary circulation and non-enzymatic reduction of
      NO3- into NO2- and NO provides an important alternative source of NO, particularly in
      environment with reduced pH and oxygen tension . The major sources of dietary NO3- are green
      leafy vegetables such as rocket or spinach or tuberous product such as beetroot. The typical
      nitrate intake of a western diet provides between 50-100mg/NO3-/day whereas diets rich in
      fruit- and vegetables such as the Mediterranean or Japanese diets could provide up to
      1000mg/NO3-/day. Increasing evidence seems to support the association between the high NO3-
      content of these dietary patterns with protective cardiovascular effects.

      The cardiovascular benefits of NO3- are thought to be derived by an improvement of
      endothelial function achieved by improved NO bioavailability. Other mechanisms may include a
      direct effect of NO2- on calcium-channels in vascular smooth muscle cells (SMVCs) in
      resistance vessels . The conversion of NO3- into NO2- by the oral micro-environment is a key
      mechanism of the entero-salivary NO3- circulation and non-enzymatic synthesis of NO. The oral
      microbiota is critically involved in the first reducing step (NO3- into NO2-) and an
      alteration of the reducing capacity of the oral microbiota (i.e., spitting, antiseptic
      mouthwash) interrupts the beneficial effects of inorganic NO3- on vascular function. Siervo
      et al has recently demonstrated a reduced efficacy of concentrated beetroot juice
      supplementation in older overweight and obese subjects. Ageing appears to be linked to
      changes in the oral micro-environment as well as a reduced responsiveness of the endothelium
      and SMVCs to NO.

      However, there is limited knowledge on the influence of the ageing process on factors that
      affect NO3- conversion starting from the NO3- -reducing potential in the oral environment.
      Therefore, a better understanding of the age-related mechanisms linked to endothelial
      dysfunction could lead to the development of effective and targeted dietary strategies for
      the early prevention of hypertension, arterial stiffness and CVD across the life-course.
    

Intervention name(s): Crossover intervention
Intervention type(s): Other
Intervention description(s): Drink consumption was followed by regular BP measurements and collection of saliva samples every 30 minutes for four hours. Microcirculatory measurements were then repeated and saliva and urine samples were collected again.
After the last measurement subjects were provided with a standardized meal with a very low nitrate intake to eat within 30minutes. Participant then left the research centre and they were asked to collect a saliva sample after two hours from the completion of the meal. This was immediately followed a dose of labelled nitrate (4mg in 100ml of distilled water) to drink within one minute. Subjects were then asked to collect six additional saliva samples until the next morning (four before going to bed and two in the morning), accompanied by measurements of resting BP.
Cross-over assessments were then repeated in the same order at Visit 3, followed a wash-out period of at least one week before entering the second arm of the study.
Allocation: Randomized
Intervention model: Crossover Assignment
Masking: Triple (Participant, Investigator, Outcomes Assessor)
Primary purpose: Prevention
Keywords: nitrates; beetroot; nutrition; microcirculation; nitrites