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Background: Evidence suggests that the intake of blueberry (poly)phenols is associated with improvements in vascular function and cognitive performance. Whether these cognitive effects are linked to increases in cerebral and vascular blood flow or changes in the gut microbiota is currently unknown., Methods: A double-blind, parallel randomized controlled trial was conducted in 61 healthy older individuals aged 65-80 y. Participants received either 26 g of freeze-dried wild blueberry (WBB) powder (302 mg anthocyanins) or a matched placebo (0 mg anthocyanins). Endothelial function measured by flow-mediated dilation (FMD), cognitive function, arterial stiffness, blood pressure (BP), cerebral blood flow (CBF), gut microbiome, and blood parameters were measured at baseline and 12 wk following daily consumption. Plasma and urinary (poly)phenol metabolites were analyzed using microelution solid-phase extraction coupled with liquid chromatography-mass spectrometry., Results: A significant increase in FMD and reduction in 24 h ambulatory systolic BP were found in the WBB group compared with the placebo group (0.86%; 95% CI: 0.56, 1.17, P < 0.001; -3.59 mmHg; 95% CI: -6.95, -0.23, P = 0.037; respectively). Enhanced immediate recall on the auditory verbal learning task, alongside better accuracy on a task-switch task was also found following WBB treatment compared with placebo (P < 0.05). Total 24 h urinary (poly)phenol excretion increased significantly in the WBB group compared with placebo. No changes in the CBF or gut microbiota composition were found., Conclusions: Daily intake of WBB powder, equivalent to 178 g fresh weight, improves vascular and cognitive function and decreases 24 h ambulatory systolic BP in healthy older individuals. This suggests that WBB (poly)phenols may reduce future CVD risk in an older population and may improve episodic memory processes and executive functioning in older adults at risk for cognitive decline. Clinical Trial Registration number in clinicaltrials.gov: NCT04084457., (Copyright © 2023 American Society for Nutrition. Published by Elsevier Inc. All rights reserved.)

Background and Aims: Berry (poly)phenol consumption has been associated with cardioprotective benefits, however little is known on the role the gut microbiome may play on such health benefits. Our objective was to investigate the effects of aronia berry (poly)phenol consumption on cardiometabolic health and gut microbiome richness and composition in prehypertensive middle-aged men and women., Methods: A total of 102 prehypertensive participants were included in a parallel 12-week randomized double-blind placebo-controlled trial. Volunteers were randomly allocated to daily consume an encapsulated (poly)phenol-rich aronia berry extract (Aronia, n = 51) or a matched maltodextrin placebo (Control, n = 51). Blood pressure (BP) and arterial function (office and 24 h), endothelial function (measured as flow-mediated dilation), serum biochemistry (including blood lipids), plasma and urine (poly)phenol metabolites as well as gut microbiome composition through shotgun metagenomic sequencing were monitored over the study period. Relationships between vascular outcomes, (poly)phenol metabolites and gut microbiome were investigated using an integrated multi-levels approach., Results: A significant improvement in arterial indices measured as augmentation index (AIx) and pulse wave velocity (PWV) was found in the Aronia compared to Control group (awake Δ PWV = -0.24 m/s; 95% CI: -0.79, -0.01 m/s, P < 0.05; 24 h peripheral Δ AIx = -6.8; -11.2, -2.3, %, P = 0.003; 24 h central Δ AIx = -3.3; -5.5, -1.0, %, P = 0.006). No changes in BP, endothelial function or blood lipids were found following the intervention. Consumption of aronia (poly)phenols led to a significant increase in gut microbiome gene richness and in the abundance of butyrate-producing species such as Lawsonibacter asaccharolyticus and Intestinimonas butyriciproducens species, compared to Control group. Results from an approach including metabolomic, metagenomic and clinical outcomes highlighted associations between aronia-derived phenolic metabolites, arterial stiffness, and gut microbiome., Conclusions: Aronia berry (poly)phenol consumption improved arterial function in prehypertensive middle-aged individuals, possibly via modulation of gut microbiome richness and composition based on the associations observed between these parameters., Clinical Trial Registry: The National Institutes of Health (NIH)-randomized trial records held on the NIH ClinicalTrials.gov website (NCT03434574). Aronia Berry Consumption on Blood Pressure., Competing Interests: Conflict of interest EF, MLecomte, PFB are employees of Naturex SA, part of Givaudan. MLS, YX, FAG, DK, CD, UK, SA, JKC and ARM were supported by a collaborative research agreement with Naturex SA, part of Givaudan. FPO and MLaiola were supported by a service agreement with Naturex SA, part of Givaudan., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Betalains are natural red color pigments abundant in red-fleshed dragon fruit (Hylocereus polyrhizus). Recent research has shown that dragon fruit consumption may help improve blood glucose and lipid profile. However, investigations of its cardioprotective properties in human trials, especially in nutritionally achievable amounts, remain nonexistent., Objectives: The aim of this study was to investigate the effects of acute and short-term consumption of dragon fruit on vascular function in a healthy population., Methods: A randomized, double-blind, placebo-controlled, crossover trial was conducted in 19 young, healthy, nonsmoking men and women assigned to consume 24 g whole dragon fruit powder (33 mg betalains) or a nutrient-matched placebo, daily for 14 d. Flow-mediated dilation (FMD), arterial stiffness, and blood pressure (BP) were measured at 0 h, 1 h, 2 h, 3 h, and 4 h and finally at 14 d after daily consumption., Results: A total of 18 participants completed the trial. Dragon fruit consumption significantly improved acute FMD at 2 h (+0.8 ± 0.3%, P = 0.01), 3 h (+1.0 ± 0.3%, P = 0.001), and 4 h (+1.3 ± 0.4%, P < 0.001) postconsumption compared with placebo. This effect was sustained up until 14 d (+1.3 ± 0.2%, P < 0.001). Pulse-wave velocity was acutely significantly reduced at 3 h (-0.5 ± 0.2 m/s, P = 0.003), whereas augmentation index (AIx) also improved after 14 d (-7.0 ± 3.3%, P = 0.02) when compared with placebo. No differences were found in either peripheral or central BP across all time points., Conclusions: Acute and short-term consumption of dragon fruit in dietary achievable amounts improved endothelial function and arterial stiffness in healthy individuals. This implies that regular dragon fruit consumption may have a meaningful impact on cardiovascular disease risk likely due to the high betalain content. This trial was registered at ClinicalTrials.gov as NCT03995602., (© The Author(s) 2022. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Nutritional epidemiological studies have frequently reported associations between higher (poly)phenol intake and a decrease in the risk or incidence of noncommunicable diseases. However, the assessment methods that have been used to quantify the intakes of these compounds in large-population samples are highly variable. This systematic review aims to characterize the methods used to assess dietary (poly)phenol intake in observational studies, report the validation status of the methods, and give recommendations on method selection and data reporting. Three databases were searched for publications that have used dietary assessment methods to measure (poly)phenol intake and 549 eligible full texts were identified. Food-frequency questionnaires were found to be the most commonly used tool to assess dietary (poly)phenol intake (73%). Published data from peer-reviewed journals were the major source of (poly)phenol content data (25%). An increasing number of studies used open-access databases such as Phenol-Explorer and USDA databases on flavonoid content since their inception, which accounted for 11% and 23% of the data sources, respectively. Only 16% of the studies reported a method that had been validated for measuring the target (poly)phenols. For future research we recommend: 1) selecting a validated dietary assessment tool according to the target compounds and target period of measurement; 2) applying and combining comprehensive (poly)phenol content databases such as USDA and Phenol-Explorer; 3) detailing the methods used to assess (poly)phenol intake, including dietary assessment method, (poly)phenol content data source; 4) follow the Strengthening the Reporting of Observational Studies in Epidemiology-Nutritional Epidemiology (STROBE-nut) framework; and 5) complementing dietary intake assessment based on questionnaires with measurement of (poly)phenols in biofluids using appropriate and validated analytical methods., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Society for Nutrition.)

There is a lack of focus on the protective health effects of phytochemicals in dietary guidelines. Although a number of chemical libraries and databases contain dietary phytochemicals belonging to the plant metabolome, they are not entirely relevant to human health because many constituents are extensively metabolized within the body following ingestion. This is especially apparent for the highly abundant dietary (poly)phenols, for which the situation is compounded by confusion regarding their bioavailability and metabolism, partially because of the variety of nomenclatures and trivial names used to describe compounds arising from microbial catabolism in the gastrointestinal tract. This confusion, which is perpetuated in online chemical/metabolite databases, will hinder future discovery of bioactivities and affect the establishment of future dietary guidelines if steps are not taken to overcome these issues. In order to resolve this situation, a nomenclature system for phenolic catabolites and their human phase II metabolites is proposed in this article and the basis of its format outlined. Previous names used in the literature are cited along with the recommended nomenclature, International Union of Pure and Applied Chemistry terminology, and, where appropriate, Chemical Abstracts Service numbers, InChIKey, and accurate mass., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Background: Aronia melanocarpa is a rich source of (poly)phenols. Previous research has demonstrated that these berries may provide cardiovascular health benefits in high-risk populations. However, very few studies have investigated the effects of daily consumption of dietary achievable amounts of the berries in healthy subjects., Objectives: The aim of this study was to investigate the effects of aronia berries on vascular function and gut microbiota composition in a healthy population., Methods: A double-blind, placebo-controlled, parallel designed study was conducted in 66 healthy men randomly allocated to consume a (poly)phenol-rich extract (116 mg, 75 g berries), a whole fruit powder (12 mg, 10 g berries), or placebo (maltodextrin) for 12 wk. Flow-mediated dilation (FMD), arterial stiffness, blood pressure, heart rate, and serum biochemistry were assessed. Plasma (poly)phenol metabolites were analyzed by LC-MS. Gut microbiota composition was determined via 16S rRNA sequencing in stool samples., Results: Consumption of aronia whole fruit and extract powder for 12 wk led to a significant increase in FMD over control of 0.9% ± 0.4% (95% CI: 0.13%, 1.72%) and 1.2% ± 0.4% (95% CI: 0.36%, 1.97%), respectively. Acute improvements in FMD were also observed 2 h after consumption of aronia extract on day 1 (1.1% ± 0.3%, P = 0.003) and 12 wk later (1.5% ± 0.4%, P = 0.0001). Circulating plasma phenolic metabolites increased upon consumption of the aronia treatments. Although no changes were found in gut microbiota diversity, consumption of aronia extract increased the growth of Anaerostipes (+10.6%, P = 0.01), whereas aronia whole fruit showed significant increases in Bacteroides (+193%, P = 0.01). Correlation analysis identified significant associations between changes in FMD, aronia-derived phenolic metabolites, and specific gut microbial genera., Conclusions: In healthy men, consumption of aronia berry (poly)phenols improved endothelial function and modulated gut microbiota composition, indicating that regular aronia consumption has the potential to maintain cardiovascular health in individuals at low risk of cardiovascular disease. This trial was registered at CLINICALTRIALs.gov as NCT03041961., (Copyright © American Society for Nutrition 2019. All rights reserved.)

Objective: This work provides a detailed description and differential diagnosis of a Pleistocene cave bear (Ursus spelaeus)., Materials: The specimen was recovered at the Cueva de Guantes archaeo- paleontological site, located in the North of the Iberian Peninsula and dated to more than 30k yr BP., Methods: The study was carried out by macroscopic and radiological analysis., Results: The specimen has unusual morphology, with two vertebrae (C6-C7) connected in the ventrodorsal projection by osseous tissue, without a space or disruption between them. However, a separation is visible in the dorsoventral projection. Moreover, C7 shows a "wedge-shape" conformation., Conclusions: The lack of clear radiological and macroscopic evidence of degenerative processes and trauma suggests a congenital anomaly or pathology. The short height of the ventral margin of the block and evidence of a radiological 'waist' lead us to propose congenital block vertebra (CBV) as the most likely diagnosis., Significance: The Cueva de Guantes specimen would be the first reported evidence of CBV in a Pleistocene cave bear (Ursus spelaeus)., Limitations: All diagnosis of archaeological animal remains should be undertaken with caution, especially when based on partial remains, as in this case. Moreover, this specimen lacks the neural arches of C6 and C7, preventing evaluation of the vertebral foramina., Suggestions for Further Research: Intensive review of cave bear skeletal collections is advised to find new cases and perform an epidemiological approach to the palaeopathology of cave bears., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: Flavanols are an important class of food bioactives that can improve vascular function even in healthy subjects. Cocoa flavanols (CFs) are composed principally of the monomer (-)-epicatechin (∼20%), with a degree of polymerisation (DP) of 1 (DP1), and oligomeric procyanidins (∼80%, DP2-10)., Objective: Our objective was to investigate the relative contribution of procyanidins and (-)-epicatechin to CF intake-related improvements in vascular function in healthy volunteers., Design: In a randomized, controlled, double-masked, parallel-group dietary intervention trial, 45 healthy men (aged 18-35 y) consumed the following once daily for 1 mo: 1) a DP1-10 cocoa extract containing 130 mg (-)-epicatechin and 560 mg procyanidins, 2) a DP2-10 cocoa extract containing 20 mg (-)-epicatechin and 540 mg procyanidins, or 3) a control capsule, which was flavanol-free but had identical micro- and macronutrient composition., Results: Consumption of DP1-10, but not of either DP2-10 or the control capsule, significantly increased flow-mediated vasodilation (primary endpoint) and the concentration of structurally related (-)-epicatechin metabolites (SREMs) in the circulatory system while decreasing pulse wave velocity and blood pressure. Total cholesterol significantly decreased after daily intake of both DP1-10 and DP2-10 as compared with the control., Conclusions: CF-related improvements in vascular function are predominantly related to the intake of flavanol monomers and circulating SREMs in healthy humans but not to the more abundant procyanidins and gut microbiome-derived CF catabolites. Reduction in total cholesterol was linked to consumption of procyanidins but not necessarily to that of (-)-epicatechin. This trial was registered at clinicaltrials.gov as NCT02728466.

Raspberries are a rich source of ellagitannins and anthocyanins. The aim of this work was to investigate whether raspberry consumption can improve vascular function and to understand which phenolic metabolites may be responsible for the effects. A 3 arm double-blind randomized controlled crossover human intervention trial was conducted in 10 healthy males. Flow-mediated dilation (FMD) was measured at baseline, 2 h, and 24 h post-consumption of 200 g and 400 g of red raspberries containing 201 or 403 mg of total (poly)phenols, or a matched control drink. Raspberry (poly)phenol metabolites were analyzed in plasma and urine by UPLC-QTOF mass spectrometry using authentic standards. Significant improvements in FMD were observed at 2 h (1.6% (95%CI 1.2, 1.9) and 1.2% (95% CI 0.8, 1.5)) and 24 h (1.0% (95% CI 0.6, 1.2) and 0.7% (95%CI 0.2, 0.9)) post-consumption of the 200 and 400 g raspberry drinks as compared to control, respectively. Plasma ellagic acid, urolithin A-3-glucuronide and urolithin A-sulfate correlated with the improvements in FMD at 2 and 24 h post consumption, respectively. Consumption of dietary achievable amounts of red raspberries acutely improves endothelial function up to 24 h and ellagitannins may be responsible for the observed effect., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Cardiometabolic disease, comprising cardiovascular diseases, type 2 diabetes, and their associated risk factors including metabolic syndrome and obesity, is the leading cause of death worldwide. Plant foods are rich sources of different groups of bioactive compounds, which might not be essential throughout life but promote health and well-being by reducing the risk of age-related chronic diseases. However, heterogeneity in the responsiveness to bioactive compounds can obscure associations between their intakes and health outcomes, resulting in the hiding of health benefits for specific population groups and thereby limiting our knowledge of the exact role of the different bioactive compounds for health. The heterogeneity in response suggests that some individuals may benefit more than others from the health effects of these bioactive compounds. However, to date, this interindividual variation after habitual intake of plant bioactive compounds has been little explored. The aim of this review is to provide an overview of the existing research that has revealed interindividual variability in the responsiveness to plant-food bioactive compound consumption regarding cardiometabolic outcomes, focusing on polyphenols, caffeine and plant sterols, and the identified potential determinants involved., Competing Interests: Author disclosures: DM, CM, AC, AK-R, FT-B, JMO, PK, RDC, and AR-M, no conflicts of interest., (© 2017 American Society for Nutrition.)

Background: Cocoa flavanol intake, especially that of (-)-epicatechin, has been linked to beneficial effects on human cardiovascular function. However, cocoa also contains the methylxanthines theobromine and caffeine, which may also affect vascular function., Objective: We sought to determine whether an interaction between cocoa flavanols and methylxanthines exists that influences cocoa flavanol-dependent vascular effects., Design: Test drinks that contained various amounts of cocoa flavanols (0-820 mg) and methylxanthines (0-220 mg), either together or individually, were consumed by healthy volunteers (n = 47) in 4 different clinical studies-3 with a randomized, double-masked crossover design and 1 with 4 parallel crossover studies. Vascular status was assessed by measuring flow-mediated vasodilation (FMD), brachial pulse wave velocity (bPWV), circulating angiogenic cells (CACs), and blood pressure before and 2 h after the ingestion of test drinks., Results: Although cocoa flavanol intake increased FMD 2 h after intake, the consumption of cocoa flavanols with methylxanthines resulted in a greater enhancement of FMD. Methylxanthine intake alone did not result in statistically significant changes in FMD. Cocoa flavanol ingestion alone decreased bPWV and diastolic blood pressure and increased CACs. Each of these changes was more pronounced when cocoa flavanols and methylxanthines were ingested together. It is important to note that the area under the curve of the plasma concentration of (-)-epicatechin metabolites over time was higher after the co-ingestion of cocoa flavanols and methylxanthines than after the intake of cocoa flavanols alone. Similar results were obtained when pure (-)-epicatechin and the methylxanthines theobromine and caffeine were consumed together., Conclusion: A substantial interaction between cocoa flavanols and methylxanthines exists at the level of absorption, in which the methylxanthines mediate an increased plasma concentration of (-)-epicatechin metabolites that coincides with enhanced vascular effects commonly ascribed to cocoa flavanol intake. This trial was registered at clinicaltrials.gov as NCT02149238., (© 2017 American Society for Nutrition.)

A rapid and high-throughput micro-solid phase extraction (μ-SPE) method coupled with ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC Q-TOF MS) analysis was optimized and validated for the quantification of 67 (poly)phenol metabolites in human plasma and urine using authentic standards. The method was fully validated in terms of specificity, linearity, method detection limit (MDL), method quantification limit (MQL), repeatability, intra- and inter-day precision, accuracy and matrix effects. The method proved to be specific and results showed linearity of responses for all compounds, with MDL ranging between 0.04nM and 86nM in plasma and between 0.01nM and 136nM in urine. MQL ranged between 0.14nM and 286nM in plasma and between 0.03nM and 465nM in urine. Repeatability varied between 1.7 and 9.2% in plasma and between 2.2% and 10.4% in urine. Median precision values of 8.7 and 11.5% (intra-day), and 10.8% and 10.0% (inter-day) were obtained in plasma and urine, respectively. The median recovery was 89% in both biological matrices. Matrix effects were determined and median values of -1.2% and -6.8% in plasma and urine were obtained. After method validation, 49 and 57 compounds, including phase II and gut microbial metabolites, were quantified in plasma and urine, respectively, following cranberry juice consumption. This methodology can be applied to large-scale human dietary intervention trials allowing for high sample throughput., (Copyright © 2016. Published by Elsevier B.V.)

Recent advances in cranberry research have expanded the evidence for the role of this Vaccinium berry fruit in modulating gut microbiota function and cardiometabolic risk factors. The A-type structure of cranberry proanthocyanidins seems to be responsible for much of this fruit's efficacy as a natural antimicrobial. Cranberry proanthocyanidins interfere with colonization of the gut by extraintestinal pathogenic Escherichia coli in vitro and attenuate gut barrier dysfunction caused by dietary insults in vivo. Furthermore, new studies indicate synergy between these proanthocyanidins, other cranberry components such as isoprenoids and xyloglucans, and gut microbiota. Together, cranberry constituents and their bioactive catabolites have been found to contribute to mechanisms affecting bacterial adhesion, coaggregation, and biofilm formation that may underlie potential clinical benefits on gastrointestinal and urinary tract infections, as well as on systemic anti-inflammatory actions mediated via the gut microbiome. A limited but growing body of evidence from randomized clinical trials reveals favorable effects of cranberry consumption on measures of cardiometabolic health, including serum lipid profiles, blood pressure, endothelial function, glucoregulation, and a variety of biomarkers of inflammation and oxidative stress. These results warrant further research, particularly studies dedicated to the elucidation of dose-response relations, pharmacokinetic/metabolomics profiles, and relevant biomarkers of action with the use of fully characterized cranberry products. Freeze-dried whole cranberry powder and a matched placebo were recently made available to investigators to facilitate such work, including interlaboratory comparability., (© 2016 American Society for Nutrition.)

Cranberries are a rich source of (poly)phenols, in particular proanthocyanidins, anthocyanins, flavonols, and phenolic acids. However, little is known about their bioavailability in humans. We investigated the absorption, metabolism, and excretion of cranberry (poly)phenols in plasma and urine of healthy young men after consumption of a cranberry juice (787 mg (poly)phenols). A total of 60 cranberry-derived phenolic metabolites were identified using UPLC-Q-TOF-MS analysis with authentic standards. These included sulfates of pyrogallol, valerolactone, benzoic acids, phenylacetic acids, glucuronides of flavonols, as well as sulfates and glucuronides of cinnamic acids. The most abundant plasma metabolites were small phenolic compounds, in particular hippuric acid, catechol-O-sulfate, 2,3-dihydroxybenzoic acid, phenylacetic acid, isoferulic acid, 4-methylcatechol-O-sulfate, α-hydroxyhippuric acid, ferulic acid 4-O-sulfate, benzoic acid, 4-hydroxyphenyl acetic acid, dihydrocaffeic acid 3-O-sulfate, and vanillic acid-4-O-sulfate. Some benzoic acids, cinnamic acids, and flavonol metabolites appeared in plasma early, at 1-2 h post-consumption. Others such as phenylacetic acids, benzaldehydes, pyrogallols, catechols, hippuric and dihydrocinnamic acid derivatives appear in plasma later (Tmax 4-22 h). The 24 h urinary recovery with respect to the amount of (poly)phenols consumed was 6.2%. Our extensive description of the bioavailability of cranberry (poly)phenols lays important groundwork necessary to start understanding the fate of these compounds in humans., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Skeletal muscle wasting is likely to play an important role in the Huntington's disease (HD) pathogenesis. Our aim was to analyze the body composition, and specifically fat-free mass (FFM), as an indirect marker of skeletal muscle in patients with HD, and its association with HD severity and energy balance., Methods: Cross-sectional, case-control study. Body composition was analyzed using bioelectrical impedance. Information was collected as regards of the anthropometrics, disease severity [Unified Huntington Disease Rating (UHDRS) and Total functional capacity (TFC) scores], CAG repeats, protein catabolism, energy intake and energy expenditure., Results: Twenty two patients with HD [mean age 50.3±15.6, mean UHDRS of 27.9±23.7, median TFC of 11 (IQR: 7; 13); median body mass index 23.6 (IQR: 26.8; 22.5)], and 18 controls were included. Both groups were similar in terms of age, gender, body mass index, body composition, physical activity level, and protein catabolism. FFM was correlated with energy intake (r=0.73, p<0.001), resting energy expenditure (r=0.64, p=0.001) and physical activity (r=0.54, p=0.003), but not with CAG repeats, or HD severity., Conclusions: Our results do not support the presence of significant muscle wasting in patients with early-moderate Huntington's disease. However, to prevent muscle wasting in HD, dietary strategies, in addition to physical exercise, should be further investigated., (Copyright © 2015. Published by Elsevier B.V.)

Accumulating evidence suggest that diets rich in cocoa flavanols may have beneficial effects on cardiovascular health. The major cocoa flavanol monomer, (-)-epicatechin (EC), is readily absorbed and circulates primarily as glucuronidated, sulfated, and O-methylated metabolites in human plasma. However, cellular metabolism, for example in endothelial cells, is less well defined. In the present study we detail the uptake and cellular metabolism of EC and its major in vivo metabolites, (-)-epicatechin-3'-β-D-glucuronide (E3G), (-)-epicatechin-3'-sulfate (E3S), 3'-O-methyl-(-)-epicatechin-5-sulfate (3ME5S), and 3'-O-methyl-(-)-epicatechin-7-sulfate (3ME7S) in human endothelial (HUVEC), liver (HepG2) and intestinal epithelial cells (Caco-2 monolayer). Our results indicate that EC associates with HUVECs, leading to its intracellular metabolism to 3ME7G and 3ME7S. In contrast, none of the metabolites were taken up by the cells. The metabolic rate and pattern of metabolism in HUVECs was similar to that observed in HepG2 cells, whilst in Caco-2 cells EC was metabolized to E3G, 3ME5G, 3ME7G, 4ME5G, 4ME7G and 3ME7S. Our data support the notion that endothelial cells may contribute significantly to EC metabolism. However, major human circulating metabolites are not accounted for in these model systems underscoring that caution should be taken when drawing conclusions on in vivo flavanol metabolism from in vitro experiments., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: There are very limited data regarding the effects of blueberry flavonoid intake on vascular function in healthy humans., Objectives: We investigated the impact of blueberry flavonoid intake on endothelial function in healthy men and assessed potential mechanisms of action by the assessment of circulating metabolites and neutrophil NADPH oxidase activity., Design: Two randomized, controlled, double-blind, crossover human-intervention trials were conducted with 21 healthy men. Initially, the impact of blueberry flavonoid intake on flow-mediated dilation (FMD) and polyphenol absorption and metabolism was assessed at baseline and 1, 2, 4, and 6 h after consumption of blueberry containing 766, 1278, and 1791 mg total blueberry polyphenols or a macronutrient- and micronutrient-matched control drink (0 mg total blueberry polyphenols). Second, an intake-dependence study was conducted (from baseline to 1 h) with 319, 637, 766, 1278, and 1791 mg total blueberry polyphenols and a control., Results: We observed a biphasic time-dependent increase in FMD, with significant increases at 1-2 and 6 h after consumption of blueberry polyphenols. No significant intake-dependence was observed between 766 and 1791 mg. However, at 1 h after consumption, FMD increased dose dependently to ≤766 mg total blueberry polyphenol intake, after which FMD plateaued. Increases in FMD were closely linked to increases in circulating metabolites and by decreases in neutrophil NADPH oxidase activity at 1-2 and 6 h., Conclusions: Blueberry intake acutely improves vascular function in healthy men in a time- and intake-dependent manner. These benefits may be mechanistically linked to the actions of circulating phenolic metabolites on neutrophil NADPH oxidase activity. This trial was registered at clinicaltrials.gov as NCT01292954 and NCT01829542.

A nitric oxide synthase (NOS)-like activity has been demonstrated in human red blood cells (RBCs), but doubts about its functional significance, isoform identity and disease relevance remain. Using flow cytometry in combination with the nitric oxide (NO)-imaging probe DAF-FM we find that all blood cells form NO intracellularly, with a rank order of monocytes > neutrophils > lymphocytes > RBCs > platelets. The observation of a NO-related fluorescence within RBCs was unexpected given the abundance of the NO-scavenger oxyhemoglobin. Constitutive normoxic NO formation was abolished by NOS inhibition and intracellular NO scavenging, confirmed by laser-scanning microscopy and unequivocally validated by detection of the DAF-FM reaction product with NO using HPLC and LC-MS/MS. Using immunoprecipitation, ESI-MS/MS-based peptide sequencing and enzymatic assay we further demonstrate that human RBCs contain an endothelial NOS (eNOS) that converts L-(3)H-arginine to L-(3)H-citrulline in a Ca(2+)/calmodulin-dependent fashion. Moreover, in patients with coronary artery disease, red cell eNOS expression and activity are both lower than in age-matched healthy individuals and correlate with the degree of endothelial dysfunction. Thus, human RBCs constitutively produce NO under normoxic conditions via an active eNOS isoform, the activity of which is compromised in patients with coronary artery disease.

Background: The absorption of cocoa flavanols in the small intestine is limited, and the majority of the flavanols reach the large intestine where they may be metabolized by resident microbiota., Objective: We assessed the prebiotic potential of cocoa flavanols in a randomized, double-blind, crossover, controlled intervention study., Design: Twenty-two healthy human volunteers were randomly assigned to either a high-cocoa flavanol (HCF) group (494 mg cocoa flavanols/d) or a low-cocoa flavanol (LCF) group (23 mg cocoa flavanols/d) for 4 wk. This was followed by a 4-wk washout period before volunteers crossed to the alternant arm. Fecal samples were recovered before and after each intervention, and bacterial numbers were measured by fluorescence in situ hybridization. A number of other biochemical and physiologic markers were measured., Results: Compared with the consumption of the LCF drink, the daily consumption of the HCF drink for 4 wk significantly increased the bifidobacterial (P < 0.01) and lactobacilli (P < 0.001) populations but significantly decreased clostridia counts (P < 0.001). These microbial changes were paralleled by significant reductions in plasma triacylglycerol (P < 0.05) and C-reactive protein (P < 0.05) concentrations. Furthermore, changes in C-reactive protein concentrations were linked to changes in lactobacilli counts (P < 0.05, R(2) = -0.33 for the model). These in vivo changes were closely paralleled by cocoa flavanol-induced bacterial changes in mixed-batch culture experiments., Conclusion: This study shows, for the first time to our knowledge, that consumption of cocoa flavanols can significantly affect the growth of select gut microflora in humans, which suggests the potential prebiotic benefits associated with the dietary inclusion of flavanol-rich foods. This trial was registered at clinicaltrials.gov as NCT01091922.

Neuroinflammation plays an integral role in the progression of neurodegeneration. In this study we investigated the anti-inflammatory effects of different classes of flavonoids (flavanones, flavanols and anthocyanidins) in primary mixed glial cells. We found that the flavanones naringenin and hesperetin and the flavanols (+)-catechin and (-)-epicatechin, but not the anthocyanidins cyanidin and pelargonidin, attenuated LPS/IFN-gamma-induced TNF-alpha production in glial cells. Naringenin also inhibited LPS/IFN-gamma-induced iNOS expression and nitric oxide production in glial cells, thus showing the strongest anti-inflammatory activity among all flavonoids tested. Moreover, naringenin protected against inflammatory-induced neuronal death in a primary neuronal-glial co-culture system. Naringenin also inhibited LPS/IFN-gamma-induced p38 mitogen-activated protein kinase (MAPK) phosphorylation and downstream signal transducer and activator of transcription-1 (STAT-1) in LPS/IFN-gamma stimulated primary mixed glial cells. Taken together, our results suggest that naringenin may produce an anti-inflammatory effect in LPS/IFN-gamma stimulated glial cells that may be due to its interaction with p38 signalling cascades and the STAT-1 transcription factor.

Regular consumption of green tea polyphenols (GTP) is thought to reduce the risk of cardiovascular disease (CVD) but has also been associated with liver toxicity. The present trial aimed to assess the safety and potential CVD health beneficial effects of daily GTP consumption. We conducted a placebo-controlled parallel study to evaluate the chronic effects of GTP on liver function and CVD risk biomarkers in healthy men. Volunteers (treatment: n = 17, BMI 26.7 +/- 3.3 kg/m(2), age 41 +/- 9 y; placebo, n = 16, BMI 25.4 +/- 3.3 kg/m(2), age 40 +/- 10 y) consumed for 3 wk 6 capsules per day (2 before each principal meal) containing green tea extracts (equivalent to 714 mg/d GTP) or placebo. At the beginning and end of the intervention period, we collected blood samples from fasting subjects and measured vascular tone using Laser Doppler Iontophoresis. Biomarkers of liver function and CVD risk (including blood pressure, plasma lipids, and asymmetric dimethylarginine) were unaffected by GTP consumption. After treatment, the ratio of total:HDL cholesterol was significantly reduced in participants taking GTP capsules compared with baseline. Endothelial-dependent and -independent vascular reactivity did not significantly differ between treatments. In conclusion, the present data suggests that the daily consumption of high doses of GTP by healthy men for 3 wk is safe but without effects on CVD risk biomarkers other than the total:HDL cholesterol ratio.

The neuroprotective effects of flavonoids will ultimately depend on their interaction with both neuronal and glial cells. In this study, we show that the potential neurotoxic effects of quercetin are modified by glial cell interactions. Specifically, quercetin is rapidly conjugated to glutathione within glial cells to yield 2'-glutathionyl-quercetin, which is exported from cells but has significantly reduced neurotoxicity. In addition, quercetin underwent intracellular O-methylation to yield 3'-O-methyl-quercetin and 4'-O-methyl-quercetin, although these were not exported from glia at the same rate as the glutathionyl adduct. The neurotoxic potential of both quercetin and 2'-glutathionyl-quercetin paralleled their ability to modulate the pro-survival Akt/PKB and extracellular signal-regulated kinase (ERK) signalling pathways. These data were supported by co-culture investigation, where the neurotoxic effects of quercetin were significantly reduced when they were cultured alongside glial cells. We propose that glial cells act to protect neurons against the neurotoxic effects of quercetin and that 2'-glutathionyl-quercetin represents a novel quercetin metabolite.

Mechanisms of nigral cell injury in Parkinson's disease remain unclear, although a combination of increased oxidative stress, the formation of catecholamine-quinones and the subsequent formation of neurotoxic cysteinyl-catecholamine conjugates may contribute. In the present study, peroxynitrite was observed to generate both 2-S- and 5-S-cysteinyl-dopamine and a dihydrobenzothiazine species, DHBT-1, following the reaction of dopamine with l-cysteine. The formation of 5-S-cysteinyl-dopamine and DHBT-1 in the presence of peroxynitrite induced significant neuronal injury. Pre-treatment of cortical neurons with pelargonidin, quercetin, hesperetin, caffeic acid, the 4'-O-Me derivatives of catechin and epicatechin (0.1-3.0 microM) resulted in concentration dependant protection against 5-S-cysteinyl-dopamine-induced neurotoxicity. These data suggest that polyphenols may protect against neuronal injury induced by endogenous neurotoxins relevant to the aetiology of the Parkinson disease.