Clinical trial NCT06502535: Inulin and Nitrate's Effects on Vascular Health and Gut Microbiome
Official title: The Effects of Inulin and Inorganic Nitrate on Flow-Mediated Dilation and Gut Microbiome in Adults with Pre-Hypertension and Hypertension
Study type: INTERVENTIONAL
Overall status: COMPLETED
Phase: NA
Conditions: Endothelial Dysfunction
High Blood Pressure
Gut Microbiota Dysbiosis and Nutrition
Lead sponsor: University of Exeter
Sponsor class: OTHER
Collaborators: name: University of East Anglia
First posted: 2024-07-16
Enrollment: 22
Sex: ALL
Minimum age (years): 45
Maximum age (years): 74
Standard ages: ADULT
OLDER_ADULT
Accepts healthy volunteers: no
Allocation: RANDOMIZED
Intervention model: CROSSOVER
Intervention model description: Participants will be assigned randomly to receive either A) a combination of inulin and nitrate, or B) inulin alone. This will be done following a crossover and counterbalanced design, ensuring that all participants will receive both treatments in a randomised order.
Primary purpose: BASIC_SCIENCE

Brief summary: Previous research has revealed that adults with pre-hypertensive and hypertensive blood pressure levels typically exhibit impaired vasodilation, which is a crucial aspect of blood vessel function, due to decreased nitric oxide production. Flow-mediated dilation (FMD) is a widely used technique for assessing vasodilation, which gauges the capacity of blood vessels to widen in response to increased blood flow. Nitrate supplements have been shown to enhance blood vessel function both in the short and long term by elevating nitric oxide (NO) production. Similarly, acetate, a by-product of fibre digestion, has demonstrated anti-inflammatory effects in the gut and may also increase NO production. However, further investigation is needed to determine whether these two supplements work synergistically to improve FMD responses by augmenting NO production in the human body. This study will involve administering inulin plus nitrate supplements to adults aged 45-74 with untreated high blood pressure (120-139/80-89 mmHg and 140/90 mmHg or higher) for a period of 4 weeks, followed by a subsequent 4-week period where only inulin supplements are consumed. The supplements will be provided in the form of powder, to be mixed with water, and will be given in a cross-over and counterbalanced order. Various measurements, including brachial artery function, faecal samples, blood pressure, plasma nitrate and nitrite levels, plasma short-chain fatty acid levels, red blood cell and whole blood S-Nitrosothiols, and salivary nitrate and nitrite levels, will be taken at the baseline and after each supplementation period. Additionally, acute supplementation measurements will be collected on two separate occasions before the start of the longer-term supplement regimen.

Detailed description: Endothelial dysfunction contributes to the increased cardiovascular risk associated with high blood pressure levels (e.g., 120-139/80-89 mmHg and 140/90 mmHg or higher), and it is considered as an early event in the development of atherosclerotic vascular disease. Mechanistically, endothelial dysfunction is characterised by a reduced bioavailability of the endothelium-derived molecule nitric oxide (NO), which is reflected by impaired brachial artery flow-mediated dilation (FMD). However, endothelial dysfunction is largely preventable by its early detection, thus approaches that might restore the loss of NO in the body could have obvious therapeutic potential.

Consumption of the prebiotic fibre inulin has been shown to increase the fermentation of bacterial-derived metabolites such as acetate by our gut microbiome. While increased acetate has local benefits in the gut (i.e., anti-inflammatory, lower luminal pH), it also has the potential to improve endothelial health. Research has shown that acetate can increase NO bioavailability via the activation of the NO synthase pathway by increasing phosphorylation of endothelial NO synthase (eNOS), a key enzyme in the maintenance of endothelial homeostasis, following a 15-day supplementation with inulin. Similarly, NO is produced following the consumption of nitrate, with individual responses being dependent on the basal eNOS activity. Therefore, individuals with compromised eNOS activity (e.g., pre- and hypertensive) might show an augmented response to nitrate supplementation. Since both nitrate and inulin supplementations can increase NO production, it is possible that their combined ingestion might enhance endothelial function to a greater extent. When supplemented in isolation, nitrate proved an effective dietary strategy to ameliorate endothelial dysfunction in older and clinical populations. For example, a 4-week chronic dietary nitrate supplementation (400 mg nitrate/day) was associated with an increased peak brachial artery FMD of 1.0% (95% CI, 0.3-1.5; p \< 0.001), which was not evident in the placebo group. Similarly, following a 4-week dietary nitrate supplementation (400 mg nitrate/day), brachial artery FMD (%) values increased significantly in the nitrate group compared to the placebo group. Unlike nitrate, the effects of inulin on brachial artery FMD in humans are yet to be established. Preliminary findings demonstrated an enhancement of endothelial function after inulin supplementation in mice. These findings were further supported by another study which demonstrated a brachial artery FMD increase of \~1% following a 3-month prebiotic supplementation with 12 g fructo-oligosaccharides in clinical patients with less severe endothelial dysfunction.

To date, no studies that the investigators are aware of have explored the role of nitrate and inulin in combination; however, based on the above, these two nutrients have the potential to work synergistically. Therefore, the primary aim of this study is to assess whether adding nitrate to inulin during a 4-week chronic supplementation might lead to an increase in brachial artery FMD in middle-aged and youngest elderly adults (45-74 years old) with elevated blood pressure and compare this response to inulin supplemented in isolation. The secondary aims of this study include brachial artery FMD % changes at 4 h post-consumption of a single dose of 15 g inulin and 1300 mg potassium nitrate combined and 15 g inulin supplemented in isolation. This study will also explore whether changes in brachial artery FMD % following the two supplementations are mediated by potential changes in the gut microbiome composition and diversity that may occur following prebiotics supplementation. Further secondary outcomes of the study will investigate differences in plasma nitrate, nitrite, and short-chain fatty acids (SCFAs) following both types of supplementation. Additionally, the study will examine changes in the levels of S-nitrosothiols (RSNO) in red blood cells and whole blood. The study will also compare changes in resting blood pressure between the combined inulin and nitrate supplementation and the inulin-only supplementation.

Eligibility criteria: Inclusion Criteria:

* Able to provide informed consent
* Adults aged 45-74
* Adults with untreated elevated blood pressure as SBP 120 to 129 mm Hg and DBP \< 80 mm Hg or untreated stage 1 hypertension (i.e., pre-hypertension) as SBP 130 to 139 mm Hg or DBP 80 to 89 mm Hg or untreated stage 2 hypertension as SBP equal to or higher than 140 mm Hg or DBP equal to or higher than 90 mm Hg (ACC/AHA and ESC/ESH guidelines and Blood Pressure UK)
* Weight-stable in the 3 months prior to enrolment (self-report)

Exclusion Criteria:

* Body mass index \> 40 kg/m\^2 at screening;
* History of symptomatic coronary artery disease, stroke, or other known atherosclerotic disease;
* History of gastrointestinal disease (e.g., irritable bowel syndrome, inflammatory bowel disease, coeliac disease, Chron's disease) or former abdominal surgery, weight loss surgery e.g., gastric by-pass (except for appendectomy);
* History of chronic viral hepatitis (including presence of hepatitis B surface antigen or hepatitis C antibody), or other chronic hepatic disorders;
* History of malignancy within the past 5 years, other than non-melanoma skin cancer;
* History of diabetes and/or any endocrine disorder;
* Current use of nitrate medications, including glyceryl trinitrate (GTN), isosorbide dinitrate and isosorbide mononitrate and blood pressure medications;
* Consumption of a low calorie or other special diet during the last month prior to the study, or if following a slimming or medically prescribed diet, or a vegan or macrobiotic lifestyle;
* Use of antibiotics during the three months prior to screening;
* Consumption of pre- or probiotics during the last month prior to screening and during the study, unless discontinued 3 weeks before the start of the trial. This is referred to the use of supplements only and NOT foods containing probiotics and/or prebiotics foods;
* Consumption of dietary supplements such as vitamins C and E during the last month prior to the screening and during the trial, unless discontinued 3 weeks before the start of the trial;
* Consumption of fish oil 6 weeks prior to the screening and during the trial, unless discontinued 6 weeks before the start of the trial;
* Allergies or intolerance against the substances used in the study and/or food intolerances associated with gastrointestinal upset;
* Any apparent dependence on or abuse of alcohol, tobacco, and cannabis products;
* Pregnancy, breast-feeding, or plans to become pregnant during the duration of the study;
* Current use of nitrate supplements in the form of beetroot juice, beetroot crystals, potassium nitrate and sodium nitrate, including L-arginine or L-citrulline supplements.
* Self-reported use of antimicrobial mouthwash or tongue scrapes, unless discontinued during the trial.

Primary outcomes:
measure: Flow-mediated dilation (FMD) with ultrasound to detect a change in brachial artery FMD
timeFrame: Day 0, Month 3
measure: Plasma nitrite and nitrate levels
timeFrame: Day 0, Month 3
measure: Plasma Short-Chain Fatty Acid (SCFA) levels
timeFrame: Day 0, Month 3
measure: Gut microbial composition using 16S sequencing
timeFrame: Day 0, Month 3
measure: Gut microbial composition using 16S sequencing
timeFrame: Day 0, Month 3
measure: Gut microbial composition using 16S sequencing
timeFrame: Day 0, Month 3
Secondary outcomes:
measure: Flow-mediated dilation (FMD) with ultrasound to detect a change in brachial artery FMD
timeFrame: Baseline, 4 hour post supplement consumption
measure: Plasma nitrite and nitrate levels
timeFrame: Baseline, 4 hour post supplement consumption
measure: Plasma Short-Chain Fatty Acid (SCFA) levels
timeFrame: Baseline, 4 hour post supplement consumption
measure: S-nitrosothiols (RSNO) in red blood cells and whole blood
timeFrame: Baseline, 4 hour post supplement consumption
measure: S-nitrosothiols (RSNO) in red blood cells and whole blood
timeFrame: Day 0, Month 3
measure: Resting blood pressure
timeFrame: Baseline, 4 hour post supplement consumption
measure: Resting blood pressure
timeFrame: Day 0, Month 3
measure: Tolerability of inulin defined by gastrointestinal wellbeing
timeFrame: Day 0, Month 3
measure: Rate of habitual fibre intake before and during the intervention
timeFrame: Pre intervention, at baseline
measure: Dietary intake
timeFrame: Day 0, Month 3
measure: Body weight
timeFrame: Day 0, Month 3
measure: Body fat percentage
timeFrame: Day 0, Month 3
measure: Hip circumference
timeFrame: Day 0, Month 3
measure: Waist circumference
timeFrame: Day 0, Month 3
Other outcomes:
Locations:
city: Exeter
country: United Kingdom
facility: University of Exeter
state: Devon
Study officials:
affiliation: University of Exeter
name: Luciana Torquati, PhD
role: PRINCIPAL_INVESTIGATOR
affiliation: University of Exeter
name: Jessica Virgili, MSc
role: PRINCIPAL_INVESTIGATOR
References:
citation: Armani RG, Carvalho AB, Ramos CI, Hong V, Bortolotto LA, Cassiolato JL, Oliveira NF, Cieslarova Z, do Lago CL, Klassen A, Cuppari L, Raj DS, Canziani MEF. Effect of fructooligosaccharide on endothelial function in CKD patients: a randomized controlled trial. Nephrol Dial Transplant. 2021 Dec 31;37(1):85-91. doi: 10.1093/ndt/gfaa335.
citation: Bhardwaj R, Dod H, Sandhu MS, Bedi R, Dod S, Konat G, Chopra HK, Sharma R, Jain AC, Nanda N. Acute effects of diets rich in almonds and walnuts on endothelial function. Indian Heart J. 2018 Jul-Aug;70(4):497-501. doi: 10.1016/j.ihj.2018.01.030. Epub 2018 Feb 1.
citation: Boets E, Deroover L, Houben E, Vermeulen K, Gomand SV, Delcour JA, Verbeke K. Quantification of in Vivo Colonic Short Chain Fatty Acid Production from Inulin. Nutrients. 2015 Oct 28;7(11):8916-29. doi: 10.3390/nu7115440.
citation: Carlstrom M, Liu M, Yang T, Zollbrecht C, Huang L, Peleli M, Borniquel S, Kishikawa H, Hezel M, Persson AE, Weitzberg E, Lundberg JO. Cross-talk Between Nitrate-Nitrite-NO and NO Synthase Pathways in Control of Vascular NO Homeostasis. Antioxid Redox Signal. 2015 Aug 1;23(4):295-306. doi: 10.1089/ars.2013.5481. Epub 2014 Feb 6.
citation: Catry E, Bindels LB, Tailleux A, Lestavel S, Neyrinck AM, Goossens JF, Lobysheva I, Plovier H, Essaghir A, Demoulin JB, Bouzin C, Pachikian BD, Cani PD, Staels B, Dessy C, Delzenne NM. Targeting the gut microbiota with inulin-type fructans: preclinical demonstration of a novel approach in the management of endothelial dysfunction. Gut. 2018 Feb;67(2):271-283. doi: 10.1136/gutjnl-2016-313316. Epub 2017 Apr 4.
citation: Craighead DH, Freeberg KA, Seals DR. Vascular Endothelial Function in Midlife/Older Adults Classified According to 2017 American College of Cardiology/American Heart Association Blood Pressure Guidelines. J Am Heart Assoc. 2020 Sep;9(17):e016625. doi: 10.1161/JAHA.120.016625. Epub 2020 Aug 20.
citation: Heiss C, Rodriguez-Mateos A, Kelm M. Central role of eNOS in the maintenance of endothelial homeostasis. Antioxid Redox Signal. 2015 May 10;22(14):1230-42. doi: 10.1089/ars.2014.6158. Epub 2014 Dec 10.
citation: Inaba Y, Chen JA, Bergmann SR. Prediction of future cardiovascular outcomes by flow-mediated vasodilatation of brachial artery: a meta-analysis. Int J Cardiovasc Imaging. 2010 Aug;26(6):631-40. doi: 10.1007/s10554-010-9616-1. Epub 2010 Mar 26.
citation: Jones T, Dunn EL, Macdonald JH, Kubis HP, McMahon N, Sandoo A. The Effects of Beetroot Juice on Blood Pressure, Microvascular Function and Large-Vessel Endothelial Function: A Randomized, Double-Blind, Placebo-Controlled Pilot Study in Healthy Older Adults. Nutrients. 2019 Aug 2;11(8):1792. doi: 10.3390/nu11081792.
citation: Kapil V, Khambata RS, Robertson A, Caulfield MJ, Ahluwalia A. Dietary nitrate provides sustained blood pressure lowering in hypertensive patients: a randomized, phase 2, double-blind, placebo-controlled study. Hypertension. 2015 Feb;65(2):320-7. doi: 10.1161/HYPERTENSIONAHA.114.04675. Epub 2014 Nov 24.
citation: Kleinbongard P, Dejam A, Lauer T, Jax T, Kerber S, Gharini P, Balzer J, Zotz RB, Scharf RE, Willers R, Schechter AN, Feelisch M, Kelm M. Plasma nitrite concentrations reflect the degree of endothelial dysfunction in humans. Free Radic Biol Med. 2006 Jan 15;40(2):295-302. doi: 10.1016/j.freeradbiomed.2005.08.025. Epub 2005 Nov 10.
citation: Koch CD, Gladwin MT, Freeman BA, Lundberg JO, Weitzberg E, Morris A. Enterosalivary nitrate metabolism and the microbiome: Intersection of microbial metabolism, nitric oxide and diet in cardiac and pulmonary vascular health. Free Radic Biol Med. 2017 Apr;105:48-67. doi: 10.1016/j.freeradbiomed.2016.12.015. Epub 2016 Dec 16.
citation: Medina-Leyte DJ, Zepeda-Garcia O, Dominguez-Perez M, Gonzalez-Garrido A, Villarreal-Molina T, Jacobo-Albavera L. Endothelial Dysfunction, Inflammation and Coronary Artery Disease: Potential Biomarkers and Promising Therapeutical Approaches. Int J Mol Sci. 2021 Apr 8;22(8):3850. doi: 10.3390/ijms22083850.
citation: Park KH, Park WJ. Endothelial Dysfunction: Clinical Implications in Cardiovascular Disease and Therapeutic Approaches. J Korean Med Sci. 2015 Sep;30(9):1213-25. doi: 10.3346/jkms.2015.30.9.1213. Epub 2015 Aug 13.
citation: Rios-Covian D, Ruas-Madiedo P, Margolles A, Gueimonde M, de Los Reyes-Gavilan CG, Salazar N. Intestinal Short Chain Fatty Acids and their Link with Diet and Human Health. Front Microbiol. 2016 Feb 17;7:185. doi: 10.3389/fmicb.2016.00185. eCollection 2016.
citation: Velmurugan S, Gan JM, Rathod KS, Khambata RS, Ghosh SM, Hartley A, Van Eijl S, Sagi-Kiss V, Chowdhury TA, Curtis M, Kuhnle GG, Wade WG, Ahluwalia A. Dietary nitrate improves vascular function in patients with hypercholesterolemia: a randomized, double-blind, placebo-controlled study. Am J Clin Nutr. 2016 Jan;103(1):25-38. doi: 10.3945/ajcn.115.116244. Epub 2015 Nov 25.