Vascular protection by extracts of polyphenols

That polyphenols cause endothelium-dependent relaxations was first observed by Fitzpatrick et al. ⁽Reference Fitzpatrick, Hirschfield and Coffey¹⁸⁾, where some wines, grape juices and grape skin extracts caused endothelium-dependent relaxations in aortic rings. Other studies confirmed that polyphenol-rich sources such as extracts from red wines, green and black tea, and several plants caused endothelium-dependent relaxations in large arteries, arterioles and veins that were prevented by competitive inhibitors of eNOS and guanylyl cyclase⁽Reference Fitzpatrick, Hirschfield and Coffey^18, Reference Schini-Kerth, Auger and Kim¹⁹⁾. Direct proof that polyphenols (1 μg/ml) stimulate endothelial formation of NO was obtained using electron paramagnetic resonance spectroscopy using rat aortic rings and cultured endothelial cells⁽Reference Andriambeloson, Kleschyov and Muller²⁰⁾. However, in porcine coronary arteries, red wine extract-induced relaxation was only partially prevented by a competitive inhibitor of eNOS but abolished by the addition of the combination charybdotoxin plus apamin, two inhibitors of the EDHF-mediated relaxation, indicating the involvement of both NO and EDHF⁽Reference Ndiaye, Chataigneau and Lobysheva²¹⁾. The endothelium-dependent relaxation induced by red wine polyphenols is observed at concentrations of 3 μg/ml (or greater) in porcine coronary artery rings⁽Reference Ndiaye, Chataigneau and Lobysheva²¹⁾. Although the concentration of red wine polyphenols in the blood after the intake of red wine remains unknown, estimates are that an intake of 100 ml of red wine by healthy volunteers increases plasma concentrations of polyphenolic monomers to 2·5 μg/ml⁽Reference Duthie, Pedersen and Gardner²²⁾. Thus, the stimulatory effect of red wine polyphenols on NO levels is observed at plasma concentrations likely to be achieved with the moderate consumption of red wine.

The signal transduction pathway mediating the stimulatory effect of polyphenols on eNOS suggests a key role of an intracellular redox-sensitive mechanism⁽Reference Duarte, Andriambeloson and Diebolt²³⁾. Thus, vasodilation to red wine polyphenols, purple grape juice and grape skin extracts are reduced by membrane-permeant analogues of superoxide dismutase (SOD) and also to some extent by a membrane-permeant analogue of catalase⁽Reference Madeira, Auger and Anselm²⁴⁾. Exposure of cultured endothelial cells to polyphenols increased the intracellular formation of reactive oxygen species (ROS)⁽Reference Ndiaye, Chataigneau and Lobysheva²¹⁾. ROS can trigger the activation of sarcoma oncogene homolog (Src) kinase by phosphorylation, which subsequently leads to a phosphatidylinositol-3-kinase-dependent activation of Akt by phosphorylation, which ultimately causes the phosphorylation of eNOS at Ser1177 to increase its activity in response to polyphenols⁽Reference Ndiaye, Chataigneau and Lobysheva^21, Reference Anselm, Chataigneau and Ndiaye²⁵⁾. Changes in Ca signalling and oestrogen receptor (ER) function also contribute to eNOS activation caused by some polyphenols⁽Reference Chalopin, Tesse and Martinez²⁶⁾.

Polyphenol extracts reduce CVD

Polyphenols prevent and/or improve endothelial dysfunction and reduce blood pressure in spontaneously hypertensive rats (SHR)⁽Reference Peng, Clark and Prasain^27, Reference Machha and Mustafa²⁸⁾, and in deoxycorticosterone acetate salt⁽Reference Jimenez, Lopez-Sepulveda and Kadmiri²⁹⁾, the N^ω-nitro-l-arginine⁽Reference de Moura, Miranda and Pinto³⁰⁾ and angiotensin II⁽Reference Sarr, Chataigneau and Martins³¹⁾ hypertension models. In the latter case, ingestion of 150 mg/kg per d of a red wine polyphenol extract in the drinking-water reduced hypertension induced by angiotensin II in rats (0·4 mg/kg per d for 28 d)⁽Reference Sarr, Chataigneau and Martins³¹⁾. Angiotensin II-induced hypertension was associated with blunted endothelium-dependent vasodilation that was reversed by the ingestion of red wine polyphenols. Moreover, angiotensin II-induced hypertension also increased oxidative stress due to the increased formation of ROS in the arterial wall through the up-regulation of NADPH oxidase via angiotensin type 1 receptors⁽Reference Griendling and FitzGerald^12, Reference Griendling and FitzGerald^13, Reference Sarr, Chataigneau and Martins^31, Reference Harrison, Cai and Landmesser³²⁾. Polyphenol-rich red wine extracts abrogate the angiotensin II-stimulated up-regulation of several NADPH oxidase subunits including Nox 1 and p22phox and the associated oxidative stress⁽Reference Sarr, Chataigneau and Martins³¹⁾, probably due to the inhibition of the angiotensin II-induced expression of NADPH oxidase by preventing angiotensin type 1 receptor expression⁽Reference Miyazaki, Ichiki and Hashimoto³³⁾. Polyphenols also exert antioxidant activities in endothelial cells not only by reducing NADPH oxidase expression but also reducing its activity, and increasing the expression of antioxidant enzymes such as catalase⁽Reference Steffen, Gruber and Schewe³⁴⁾. Angiotensin II-induced endothelial dysfunction includes endothelium-dependent contractile responses to acetylcholine⁽Reference Kane, Etienne-Selloum and Madeira³⁵⁾, which involves COX-dependent formation of endothelium-derived contracting factors that act on thromboxane receptors located on vascular smooth muscle cells. Both the angiotensin II-induced vascular expression of COX and the increased endothelium-derived contracting factors are significantly reduced by red wine polyphenols⁽Reference Kane, Etienne-Selloum and Madeira³⁵⁾. Thus, polyphenols prevent ROS-mediated degradation of NO, and blunt vasoconstrictor and pro-inflammatory responses (Fig. 3).

Fig. 3

Potential mechanism(s) in the cardiovascular and metabolic effects of polyphenols. Polyphenols interact with oestrogen receptor α (ERα) to activate the sirtuin-1 (SIRT1)–AMP-activated protein kinase (AMPK) network. Stimulation of SIRT1 and AMPK results in the activation of PPARγ coactivator 1 α (PGC-1α), placing mitochondria at the epicentre of targets for polyphenols in CVD and metabolic disorders. (A colour version of this figure can be found online at http://www.journals.cambridge.org/bjn)

Polyphenol-rich products increase basal flow-mediated dilation in healthy subjects at relatively low doses such as those achieved after the intake of two glasses of red wine⁽Reference Agewall, Wright and Doughty³⁶⁾ or 2 weeks of daily consumption of flavonoid-rich dark chocolate bars (46 g)⁽Reference Engler, Engler and Chen³⁷⁾. Similar beneficial effects of polyphenol-rich products on flow-mediated dilation occur in patients with coronary artery disease after consumption of black tea⁽Reference Duffy, Vita and Holbrook³⁸⁾, a green tea extract⁽Reference Widlansky, Hamburg and Anter³⁹⁾ or a red grape extract⁽Reference Lekakis, Rallidis and Andreadou⁴⁰⁾. Systolic blood pressure is improved in hypertensive patients by daily ingestion of polyphenol-rich products such as a piece of a sixteen-piece dark chocolate bar⁽Reference Taubert, Roesen and Lehmann⁴¹⁾, two glasses of purple grape juice⁽Reference Park, Kim and Kang⁴²⁾ or 50 ml of pomegranate juice⁽Reference Aviram and Dornfeld⁴³⁾.

Resveratrol and oxidative stress

Resveratrol increases the expression/activity of SOD, catalase and glutathione peroxidase (GPx) in cardiac H9C2 cells⁽Reference Cao and Li⁴⁸⁾ and aortic smooth muscle cells⁽Reference Li, Xia and Forstermann^47, Reference Li, Cao and Zhu⁴⁹⁾. Another study, however, found no changes in the protein levels of SOD1 or SOD2 but an up-regulation of GPx1 and catalase in aortic segments or cultured aortic smooth muscle⁽Reference Ungvari, Orosz and Rivera⁵⁰⁾. In a hamster model of dilated cardiomyopathy, treatment with resveratrol increases SOD2 levels, suppresses fibrosis, preserves cardiac function and significantly improves survival⁽Reference Tanno, Kuno and Yano⁵¹⁾. Treating hypercholesterolaemic, atherosclerosis-prone apoE-knockout (KO) mice (as a model of oxidative stress) with resveratrol (30–100 mg/kg per d for 7 d) leads to the up-regulation of SOD1, SOD2, SOD3, GPx1 and catalase in the heart⁽Reference Tanno, Kuno and Yano⁵¹⁾. The expression of these enzymes is also increased by resveratrol in cultured human endothelial cells⁽Reference Xia, Daiber and Habermeier^52, Reference Spanier, Xu and Xia⁵³⁾.

The antioxidant transcription factor nuclear factor erythroid 2-related factor-2 (Nrf2) is a recently identified target of resveratrol⁽Reference Hasko and Pacher^54, Reference Ungvari, Bagi and Feher⁵⁵⁾. In cultured coronary arterial endothelial cells, resveratrol increases the transcriptional activity of Nrf2 and up-regulates the expression of Nrf2 target genes NAD(P)H:quinone oxidoreductase 1, γ-glutamylcysteine synthetase (glutamate cysteine ligase catalytic subunit, GCLC) and haem oxygenase-1⁽Reference Ungvari, Bagi and Feher⁵⁵⁾. All these enzymes, together with thioredoxin-1⁽Reference Thirunavukkarasu, Penumathsa and Koneru⁵⁶⁾, could well contribute to the antioxidant actions of resveratrol.

NADPH oxidases (NOX) are major sources of ROS in the cardiovascular system⁽Reference Brandes and Kreuzer^57, Reference Griendling⁵⁸⁾. Resveratrol reduces the expression of Nox2 and Nox4 in the heart of apoE-KO mice⁽Reference Xia, Daiber and Habermeier⁵²⁾ and also prevents Nox2 expression in the aorta of diabetic mice⁽Reference Zhang, Zhang and Ungvari⁵⁹⁾. In human umbilical vein endothelial cells (HUVEC) and HUVEC-derived EA.hy 926 endothelial cells, resveratrol decreases the expression of Nox4⁽Reference Spanier, Xu and Xia⁵³⁾, the most predominant Nox isoform in these cell types⁽Reference Xu, Goettsch and Xia⁶⁰⁾. Small-interfering RNA (siRNA)-mediated knockdown of sirtuin 1 (SIRT1) has no effect on the Nox4 down-regulation by resveratrol, indicating that the effect of resveratrol on Nox4 is likely to be SIRT1-independent⁽Reference Xia, Daiber and Habermeier⁵²⁾.

Resveratrol and endothelial NO synthase uncoupling

Uncoupling of eNOS switches it from a NO-producing enzyme to a superoxide-generating molecule. The major cause of eNOS uncoupling under pathological conditions is a deficiency of the eNOS cofactor tetrahydrobiopterin (BH₄)⁽Reference Forstermann and Munzel^61, Reference Li and Forstermann⁶²⁾. Tissue levels of BH₄ are a balance of its biosynthesis and degradation/oxidation: synthesis of BH₄ from GTP via a de novo pathway, with GTP cyclohydrolase 1 as the rate-limiting enzyme, while rapid oxidation by peroxynitrite makes the cofactor unavailable for eNOS generation of NO.

Untreated apoE-KO mice show increased oxidation of BH₄⁽Reference Alp, McAteer and Khoo⁶³⁾ and significant ROS production in their aorta⁽Reference Alp, McAteer and Khoo^63, Reference Wohlfart, Xu and Endlich⁶⁴⁾ and heart⁽Reference Xia, Daiber and Habermeier⁵²⁾. Both aortic⁽Reference Alp, McAteer and Khoo^63, Reference Wohlfart, Xu and Endlich⁶⁴⁾ and cardiac⁽Reference Xia, Daiber and Habermeier⁵²⁾ superoxide production are reduced by the NOS inhibitor l-N^G-nitroarginine methyl ester (l-NAME), indicating that eNOS is in an uncoupled state and that it produces ROS in this pathological model. Resveratrol treatment enhances the expression of GTP cyclohydrolase 1 and BH₄ biosynthesis. In addition, resveratrol decreases the cardiac content of superoxide and peroxynitrite, and thereby decreases BH₄ oxidation⁽Reference Xia, Daiber and Habermeier⁵²⁾. As a result, the cardiac levels of BH₄ are increased by resveratrol. Cardiac superoxide production in resveratrol-treated mice is markedly reduced to a level that cannot be lowered any further by l-NAME⁽Reference Xia, Daiber and Habermeier⁵²⁾, suggesting that eNOS no longer produces superoxide in resveratrol-treated apoE-KO mice, i.e. resveratrol reverses eNOS uncoupling. The expression of GTP cyclohydrolase 1 in cultured human endothelial cells is increased by resveratrol. This up-regulation is reduced by the SIRT1 inhibitor sirtinol or by siRNA-mediated SIRT1 knockdown, indicating SIRT1-dependent mechanisms⁽Reference Xia, Daiber and Habermeier⁵²⁾.

Resveratrol and vasodilation

Resveratrol causes vasodilation by releasing NO from endothelial cells⁽Reference Cruz, Luksha and Logman⁶⁵⁾ and/or improving NO bioavailability⁽Reference Zhang, Zhang and Ungvari⁵⁹⁾. Resveratrol increases endothelial eNOS mRNA⁽Reference Nicholson, Tucker and Brameld⁶⁶⁾ and protein⁽Reference Appeldoorn, Venema and Peters⁶⁷⁾ expressions, and causes rapid phosphorylation of eNOS at Ser1177 (the activator site of this enzyme), and thereby increasing eNOS enzymatic activity⁽Reference Klinge, Wickramasinghe and Ivanova⁶⁸⁾. In parallel, resveratrol improves NO bioavailability by decreasing oxidative stress per se ⁽Reference Pearson, Baur and Lewis⁶⁹⁾. These actions combine to stimulate cyclic guanylyl monophosphate formation, protein kinase G activation and vasodilation⁽Reference Murad⁷⁰⁾. Voltage-gated K⁺ channels, large Ca²⁺-activated-K⁺ channels or voltage-gated Ca²⁺ channels⁽Reference Gojkovic-Bukarica, Novakovic and Kanjuh⁷¹⁾ mediate the endothelium-independent vasodilatation caused by resveratrol. The vasodilator properties of resveratrol offer cardiovascular and vascular protection in several models of CVD.

Resveratrol and vasoconstriction

The endothelium also releases vasoconstrictor and mitogenic substances such as endothelin-1, which under pathophysiological conditions, counteracts the protective effects of vasodilator products from endothelial cells. Resveratrol is able to reduce endothelial mRNA expression and secretion of endothelin-1⁽Reference Nicholson, Tucker and Brameld⁶⁶⁾, inhibit H₂O₂-induced endothelin-1 expression in human vascular smooth muscle cells⁽Reference Ruef, Moser and Kubler⁷²⁾ and reduce endothelin-1 expression in the ischaemia–reperfused heart⁽Reference Lekli, Szabo and Juhasz⁷³⁾.

The renin–angiotensin system regulates blood pressure via the release of angiotensin II that interacts with angiotensin type 1 receptors to evoke vasoconstriction⁽Reference Touyz and Schiffrin⁷⁴⁾. Resveratrol suppresses the mRNA and protein expressions of angiotensin type 1 receptors in intact mice and also in isolated vascular smooth muscle cells⁽Reference Miyazaki, Ichiki and Hashimoto³³⁾. Resveratrol also possesses a potent in vitro angiotensin-converting enzyme (ACE) inhibitory activity⁽Reference Geng, He and Yang⁷⁵⁾, which can partially account for resveratrol-induced blood pressure-lowering effects in various animal models of hypertension.

Resveratrol and inflammation

Resveratrol has in vitro and in vivo anti-inflammatory effects. Resveratrol treatment decreases the overexpression of adhesion molecules (vascular cell adhesion molecule-1 and intercellular adhesion molecule-1) by inhibiting the NF-κB pathway in TNFα-activated endothelial cells⁽Reference Deng, Alex and Huang⁷⁶⁾. In intact animal studies, resveratrol inhibits the angiotensin II-induced adhesion of leucocytes to arterioles, partially by reducing cellular adhesion molecule expression and circulating levels of monocyte chemoattractant protein-1 and macrophage inflammatory protein-1α⁽Reference Rius, Abu-Taha and Hermenegildo⁷⁷⁾. These effects may partially contribute to the cardiovascular protective activity of resveratrol, especially during the early phase of the atherosclerotic process.

Resveratrol probably improves the pro-inflammatory profile in human obesity by decreasing pro-inflammatory cytokine secretion and increasing adiponectin release from human adipose tissue⁽Reference Olholm, Paulsen and Cullberg⁷⁸⁾. Resveratrol modulates adipokine expression and improves insulin sensitivity in murine adipocytes, where resveratrol treatment reduces the levels of pro-inflammatory cytokines and adipokines (TNFα, IL-6 and resistin), and increases adiponectin and PPARγ expression and the Ser/Thr phosphorylation state of insulin receptor substrate-1⁽Reference Kang, Heng and Yuan⁷⁹⁾. In addition, resveratrol also normalises the levels of pro-inflammatory cytokines (IL-6 and TNF-α) and COX-2 expression by decreasing NF-κB activation in diabetic rats⁽Reference Kumar and Sharma⁸⁰⁾.

Resveratrol and platelet function

Resveratrol alters several functions of platelets: adhesion, activation and aggregation of platelets, and thrombus formation⁽Reference Wang, Huang and Zou⁸¹⁾. Since tissue factor is the major determinant for the extrinsic coagulation pathway, decreases in tissue factor expression can reduce thrombosis risk. Resveratrol attenuates an agonist-induced increase in tissue factor mRNA in endothelial and mononuclear cells, resulting from the inhibition of IκBα (inhibitor of kappa B) degradation, thus decreasing the DNA-binding occupancy by the transcription factor c-Rel/p65⁽Reference Di Santo, Mezzetti and Napoleone⁸²⁾. Additionally, resveratrol inhibits platelet aggregation induced by collagen, thrombin, ADP or arachidonic acid⁽Reference Crescente, Jessen and Momi⁸³⁾. Resveratrol inhibits COX-1⁽Reference Szewczuk, Forti and Stivala⁸⁴⁾ and modifies COX metabolite production to modulate platelet activation, and inhibits the arachidonate-dependent synthesis of inflammatory agents such as thromboxane B2, hydroxyheptadecatrienoate and 12-hydroxyeicosatetraenoate⁽Reference Crescente, Jessen and Momi⁸³⁾. Data from molecular modelling studies performed by in silico docking show that resveratrol forms stable complexes in platelet COX-1 channels⁽Reference Crescente, Jessen and Momi⁸³⁾.

Resveratrol effects and in vivo relevance

Resveratrol at concentrations of up to 100 μm is used in many cell-culture studies⁽Reference Li, Xia and Forstermann⁴⁷⁾; the molecular mechanisms obtained with such concentrations may not easily extend to understanding the effects of dietary resveratrol. It is unlikely that such high plasma concentrations of resveratrol are achieved, either by drinking red wine or by consuming resveratrol-containing food. However, high doses of resveratrol are well tolerated by animals⁽Reference Baur and Sinclair⁴⁴⁾ and by humans⁽Reference Patel, Scott and Brown⁸⁵⁾. The low toxicity of resveratrol favours its use as a nutraceutical (to reach higher in vivo concentrations).

As much as 70 % of orally ingested resveratrol can be absorbed. However, the bioavailability of unchanged resveratrol is very low, due to rapid and extensive metabolism⁽Reference Patel, Scott and Brown⁸⁵⁾. The plasma concentration and the half-life of resveratrol metabolites are much greater than those of resveratrol⁽Reference Baur and Sinclair⁴⁴⁾, indicating higher systemic exposure to the modified form than to unchanged resveratrol. It is possible that part of the in vivo effects of resveratrol can be attributed to its metabolites.

Quercetin and oxidative stress

Quercetin scavenges free radicals in vitro and has epidemiological correlates. Quercetin is a potent scavenger of superoxide anion and peroxynitrite, inhibits superoxide anion generation by suppressing xanthine oxidase activity⁽Reference Hanasaki, Ogawa and Fukui⁸⁷⁾ and inhibits the mitochondrial NADH/NAD⁺ system⁽Reference Wilms, Kleinjans and Moonen⁸⁸⁾. Important is the finding that the hydroxyl groups of quercetin contribute to the generation of intracellular superoxide, leading to the inhibition of cell proliferation and the induction of apoptosis in leukaemia cells⁽Reference Sakao, Fujii and Hou⁸⁹⁾.

Quercetin and vasodilation

Quercetin causes endothelium-dependent vasodilation through the production of NO⁽Reference Perez-Vizcaino, Duarte and Jimenez^86, Reference Perez-Vizcaino, Duarte and Andriantsitohaina⁹⁰⁾ probably by increasing eNOS phosphorylation at 5 μm⁽Reference Khoo, White and Pozzo-Miller⁹¹⁾. Additionally, 50 μm-quercetin is proposed to also increase NO release by causing a hyperpolarisation-dependent capacitative Ca²⁺ entry in isolated cultured endothelial cells⁽Reference Kuhlmann, Schaefer and Kosok⁹²⁾. These effects result in endothelium-dependent vasodilatation that is inhibited by eNOS inhibitors and charybdotoxin, thus demonstrating that the quercetin effect is dependent on both the NO/cyclic guanylyl monophosphate pathway and EDHF⁽Reference Khoo, White and Pozzo-Miller⁹¹⁾. Similar to the effects of resveratrol, quercetin, at a physiologically relevant concentration of 0·1 μm, also increases eNOS mRNA expression in HUVEC⁽Reference Nicholson, Tucker and Brameld⁶⁶⁾. Additionally, quercetin enhancement of cyclic guanylyl monophosphate-dependent relaxation in porcine isolated coronary arteries is insensitive to phosphodiesterase 5 inhibition. Quercetin reduces the development of glyceryl trinitrate-induced tolerance in vitro in porcine arteries; these findings can benefit patients with angina pectoris and await confirmation in humans⁽Reference Suri, Liu and Rayment⁹³⁾.

Other studies report that quercetin treatment (100 μm) suppresses eNOS activity in bovine aortic endothelial cells as a result of decreased eNOS phosphorylation⁽Reference Jackson and Venema⁹⁴⁾. This effect is associated with an in vitro disruption of mitotic microtubule polymerisation and an in vivo inhibition of angiogenesis, as quercetin inhibits vascular endothelial growth factor-induced endothelial cell function and angiogenesis through the inhibition of ERK1/2 (extracellular signal-regulated kinase 1/2) phosphorylation⁽Reference Donnini, Finetti and Lusini⁹⁵⁾.

Quercetin and vasoconstriction

As with resveratrol, quercetin decreases H₂O₂-induced endothelin-1 mRNA expression and reduces endothelin-1 release in HUVEC⁽Reference Nicholson, Tucker and Brameld⁶⁶⁾. Moreover, quercetin, at 1 μm and more so at 10 μm, prevents endothelin-1-induced endothelial dysfunction and NADPH oxidase subunit p47phox overexpression by inhibiting protein kinase C⁽Reference Romero, Jimenez and Sanchez⁹⁶⁾.

The detailed effects of quercetin on the renin–angiotensin system are not known. Treatment of Dahl salt-sensitive hypertensive rats with quercetin (10 mg/kg) for 4 weeks reduces blood pressure along with decreases in angiotensin II type 1 receptor mRNA, suggesting modulation of renal function by quercetin⁽Reference Mackraj, Govender and Ramesar⁹⁷⁾. It should be noted that quercetin fails to modify ACE activity either in vitro using rat kidney membranes⁽Reference Actis-Goretta, Ottaviani and Fraga⁹⁸⁾ or in vivo after administration to rats⁽Reference Neto-Neves, Montenegro and Dias-Junior⁹⁹⁾, suggesting that the antihypertensive effect of quercetin may be unrelated to actions on the renin–angiotensin system.

Quercetin and inflammation

Although various mechanisms are involved in the anti-inflammatory properties of quercetin, it mainly targets signalling pathways related to NF-κB activation. Thus, quercetin (10 μm) decreases mRNA and protein levels of TNFα, IL-1β, IL-6, macrophage inflammatory protein-1α and inducible NO synthase in several in vitro and in vivo studies⁽Reference Boesch-Saadatmandi, Loboda and Wagner¹⁰⁰⁾. Quercetin has pleiotropic effects in apoE-KO mice related to the reduction of pro-inflammatory markers (isoprostane, leukotriene B4 and P-selectin) and the enhancement of anti-inflammatory indicators (eNOS and haem oxygenase-1 expression)⁽Reference Loke, Proudfoot and Hodgson¹⁰¹⁾, suggesting that quercetin at a dose of 1·3 mg/d could delay the atherosclerotic process through its anti-inflammatory properties. Also, adiponectin mRNA levels are enhanced in adipose tissue from rats receiving quercetin fed high-fat diets⁽Reference Wein, Behm and Petersen¹⁰²⁾. High concentrations of quercetin (40 μm) suppress the Akt phosphorylation and transactivation of nuclear factor activator protein-1 and NF-κB, resulting in an inhibition of the TNF-α-induced up-regulation of cell migration⁽Reference Hwang, Song and Kang¹⁰³⁾. In addition, by reducing the production of pro-inflammatory cytokines and enzymes, quercetin (50 μm) inhibits mouse dendritic cell activation, suggesting that quercetin could be a potent immunosuppressive agent⁽Reference Huang, Yu and Cheng¹⁰⁴⁾. However, contradictory results have been described in human subjects as no effects on the inflammatory profile were detected in females receiving a 12-week supplementation with quercetin (0·5–1 g/d)⁽Reference Heinz, Henson and Nieman¹⁰⁵⁾. Dietary supplementation of quercetin in combination with vitamin C for 4 weeks does not change plasma biomarkers of inflammation (TNF-α, IL-1β, IL-6 and C-reactive protein) and the disease severity of rheumatoid arthritis patients⁽Reference Bae, Jung and Lee¹⁰⁶⁾.

Quercetin and platelet function

Platelet aggregation contributes to both the development of atherosclerosis and to acute platelet thrombus formation, followed by embolisation of stenosed arteries. Quercetin impairs in vitro platelet aggregation induced by thrombin by interfering with Ca²⁺ mobilisation and serotonin secretion⁽Reference Navarro-Nunez, Rivera and Guerrero¹⁰⁷⁾, and inhibiting platelet kinases such as phosphatidylinositol-3-kinsase and Src kinases⁽Reference Navarro-Nunez, Lozano and Martinez¹⁰⁸⁾. These results were obtained with concentrations that exceed those attained after standard consumption of flavonoid-rich foods. Quercetin inhibits platelet aggregation independently of the agonist used (arachidonic acid or ADP)⁽Reference Crescente, Jessen and Momi⁸³⁾. Quercetin inhibits platelet activation through the blockade of activity of the proto-oncogene tyrosine-protein kinase Fyn and the tyrosine phosphorylation of spleen tyrosine kinase (Syk) and phospholipase C gamma 2 (PLCγ2) following quercetin internalisation in platelets⁽Reference Wright, Moraes and Kemp¹⁰⁹⁾.

Limitations of the use of quercetin

Quercetin is absorbed through the gastrointestinal tract and rapidly metabolised by methylation and conjugation with glucuronic acid and/or sulphate in enterocytes and in the liver⁽Reference Manach, Morand and Crespy^110, Reference Morand, Crespy and Manach¹¹¹⁾. Once conjugated, quercetin is present in plasma after repeated daily dosage⁽Reference Hollman, vd Gaag and Mengelers¹¹²⁾, and, paradoxically, although the plasma concentrations of free quercetin are very low, it can occur in relatively high concentrations in several tissues indicating that in situ deconjugation of quercetin can occur⁽Reference Galindo, Rodriguez-Gomez and Gonzalez-Manzano¹¹³⁾.

Delphinidin and oxidative stress

Delphinidin possesses antioxidant effects in a wide range of chemical oxidation systems by virtue of two hydroxyl groups on the phenyl ring⁽Reference Azuma, Ohyama and Ippoushi¹¹⁶⁾, and among the anthocyanins, delphinidin has the greatest in vitro potency against superoxide anions and peroxynitrite⁽Reference Rahman, Ichiyanagi and Komiyama¹¹⁷⁾. Since this study was performed at neutral pH, it is not clear whether this potency is maintained in vivo. Its ability to scavenge ROS protects endothelial cells from LDL-induced lipid oxidation, although it is not clear whether the effects of delphinidin in quenching ROS are by direct actions on LDL. Nevertheless, delphinidin (25–200 μm) restores SOD activity to a similar extent to that produced by vitamin C, suggesting that delphinidin maintains endothelial cell function by preserving endogenous antioxidants and by attenuating lipid peroxidation⁽Reference Chen, Yi and Jin¹¹⁸⁾. Treatment of CCl₄-intoxicated mice with delphinidin (25 mg/kg, once daily for 2 weeks) decreases oxidative stress in the liver as reflected by the recovery of GPx activity and the ratio reduced glutathione:oxidised glutathione. These antioxidant effects of delphinidin are associated with antifibrotic activity, indicating that delphinidin possesses a tissue-regenerative capability⁽Reference Domitrovic and Jakovac¹¹⁹⁾. Cytotoxic effects of delphinidin (100 μm for 24 h) in metastatic cells (but not in cells originating from a primary tumour site) are related to cellular free radical accumulation, inhibition of glutathione reductase and depletion of glutathione, suggesting that delphinidin could be used as a sensitising agent in metastatic therapy⁽Reference Cvorovic, Tramer and Granzotto¹²⁰⁾.

Delphinidin and vasodilation

Delphinidin stimulates NO production independently of its antioxidant property⁽Reference Andriambeloson, Kleschyov and Muller²⁰⁾. Delphinidin activates NO release by increasing intracellular Ca²⁺ concentrations through the release from intracellular stores and the entry from the extracellular space. In bovine aortic endothelial cells, delphinidin-induced increases in intracellular Ca²⁺ are accompanied by tyrosine phosphorylation of several intracellular proteins⁽Reference Martin, Andriambeloson and Takeda¹²¹⁾. Acute treatment with delphinidin (10 min) enhances NO release and eNOS phosphorylation at Ser1177⁽Reference Chalopin, Tesse and Martinez²⁶⁾.

The only study of the angiogenic properties of low doses of delphinidin (0·06 mg/kg per d) reports no effects on the recovery of blood flow in ischaemic hindlimbs, while higher doses of delphinidin (0·6 mg/kg per d) have anti-angiogenic effects as characterised by impaired blood flow and decreased vascular density in the ischaemic leg of rats⁽Reference Baron-Menguy, Bocquet and Guihot¹²²⁾. These results are similar to those obtained with the whole extracts from red wine, suggesting that delphinidin could play an important role in the anti-angiogenic effect of red wine.

By targeting STAT1 (a nuclear transcriptional factor of the signal transducers and activators family and which has a critical role in cardiomyocyte apoptosis), delphinidin (more potently than quercetin) provides protection against ischaemia–reperfusion injury in isolated cardiomyocytes and in the Langerdoff-perfused rat heart when used at 10 μm 2 h before the onset of the ischaemic insult⁽Reference Scarabelli, Mariotto and Abdel-Azeim¹²³⁾.

Delphinidin and vasoconstriction

Delphinidin reduces both mRNA and protein levels of endothelin-1 in cultured HUVEC⁽Reference Lazze, Pizzala and Perucca¹²⁴⁾. While resveratrol and quercetin (30 μm) reduce endothelin-1 production by only 20 %, similar concentrations of delphinidin lower endothelin-1 production by approximately 75 %⁽Reference Khan, Lees and Douthwaite¹²⁵⁾. Although the inhibition of purified ACE by oligomeric procyanidins (mainly oligomeric epicatechins) is well established⁽Reference Actis-Goretta, Ottaviani and Keen¹²⁶⁾, the effects of delphinidin-3-O-sambubioside on ACE have only been recently reported. This compound inhibits ACE by competing with the active site of the enzyme, with an half-maximal inhibitory concentration value similar to that obtained with quercetin⁽Reference Ojeda, Jimenez-Ferrer and Zamilpa¹²⁷⁾.

Delphinidin and inflammation

Several in vitro studies report that delphinidin interacts directly with kinases; however, it is not established whether delphinidin also has similar effects in vivo. Delphinidin (5–20 μm) suppresses COX-2 promoter activity and COX-2 expression in mouse epidermal cells by inhibiting activator protein-1 and NF-κB pathways; these effects result from the direct binding of delphinidin to the ATP-binding site in the kinase domain of mitogen-activated protein kinase kinase 4 and to the ATP-binding site of the catalytic domain of phosphatidylinositol-3-kinsase⁽Reference Kwon, Lee and Kim¹²⁸⁾. Delphinidin (10–40 μm) inhibits phosphorylations of c-Jun N-terminal kinases, p38 mitogen-activated protein kinase, Akt and ERK as well as Fyn kinase in mouse epidermal cells, and directly binds with Fyn kinase in a non-competitive manner with ATP⁽Reference Hwang, Kang and Heo¹²⁹⁾. Additionally, delphinidin also inhibits a broad spectrum of receptor tyrosine kinases of the epidermal growth factor receptor B (ErbB) and vascular endothelial growth factor receptor families in both cell-free assays and intact cell systems⁽Reference Teller, Thiele and Boettler¹³⁰⁾. Other enzymes potentially playing a role in inflammation are also inhibited by delphinidin: for example, a mixed competitive and non-competitive phospholipase A2 inhibition has been described for delphinidin in a cell-free assay⁽Reference Dreiseitel, Korte and Schreier¹³¹⁾, while delphinidin also weakly inhibits proteasome activity⁽Reference Dreiseitel, Schreier and Oehme¹³²⁾. These data highlight the ability of delphinidin to interfere with pro-inflammatory pathways, although no evidence of in vivo effects on these enzyme systems is available.

Delphinidin and platelet function

Although aqueous residues containing the anthocyanic fraction from red wine suppressed ADP-induced platelet aggregation⁽Reference Garcia-Alonso, Rimbach and Rivas-Gonzalo¹³³⁾, delphinidin was unable to inhibit collagen-induced platelet aggregation in vitro ⁽Reference Garcia-Alonso, Rimbach and Rivas-Gonzalo¹³³⁾. Delphinidin containing fractions from purple grapes inhibits whole-blood aggregation, suggesting a potential mechanism for the beneficial effects of polyphenols on the suppression of platelet-mediated thrombosis⁽Reference Freedman, Parker and Li¹³⁴⁾.

Limitations on the use of delphinidin

Although abundant in the diet, anthocyanins, in general, and delphinidin in particular, are either poorly absorbed or not absorbed at all. One consequence of the poor bioavailability of anthocyanins is that many effects observed in vitro (e.g. inhibition of COX-2) are unlikely to occur in vivo. The measurement in plasma or urine of the original anthocyanins and their conjugated metabolites (glucuronidated and sulphated anthocyanins) indicates their very low bioavailability⁽Reference Manach, Scalbert and Morand^7, Reference Manach, Williamson and Morand¹³⁵⁾. In addition, intestinal microflora play an important role in the metabolism of anthocyanins⁽Reference Avila, Ojcius and Yilmaz¹³⁶⁾. Clearly, additional in vivo studies on the effects of delphinidin are needed to establish the beneficial effects of concentrations used in in vitro studies, which generally tend to be higher than those attained physiologically.

AMP-activated protein kinase

AMP-activated protein kinase (AMPK) is a Ser/Thr protein kinase involved in ATP production in mammalian cells⁽Reference Hwang, Kwon and Yoon¹³⁷⁾. The AMPK cascade may have an important role in preventing diseases since AMPK inhibits fat accumulation, reduces cholesterol synthesis and modulates inflammatory cytokines. Polyphenols found in natural products can target and activate AMPK leading to numerous beneficial effects in cardiovascular and metabolic diseases, as shown by the finding that activation of AMPK by resveratrol is SIRT1-independent⁽Reference Dasgupta and Milbrandt¹³⁸⁾ (see below). By increasing AMPK phosphorylation, resveratrol prevents the development of hyperlipidaemia and atherosclerosis in diabetic mice⁽Reference Zang, Xu and Maitland-Toolan¹³⁹⁾. These effects may be related to reduced fat accumulation⁽Reference Um, Park and Kang¹⁴⁰⁾, enhanced glucose transporter GLUT4 translocation and increases in glucose uptake by diabetic rat cardiomyocytes⁽Reference Penumathsa, Thirunavukkarasu and Zhan¹⁴¹⁾. Resveratrol increases physical endurance and mitochondrial biogenesis as revealed by increases in the expressions of PPARγ coactivator (PGC)-1α, PGC-1β, oestrogen-related receptor α and nuclear respiratory factor (NRF) in AMPK-deficient mice, leading to improved glucose homeostasis through mechanisms dependent on AMPK activation⁽Reference Um, Park and Kang¹⁴⁰⁾.

Quercetin also activates the AMP–AMPK pathway via down-regulation of protein phosphatase 2C in the brains of old mice fed a cholesterol-rich diet, indicating that quercetin enhances the resistance of neurons to age-related diseases via AMPK pathway activation⁽Reference Lu, Wu and Zheng¹⁴²⁾. Furthermore, quercetin inhibits adipocyte 3T3-L1 differentiation by decreasing adipogenic transcription factors such as PPARγ and CCAAT/enhancer-binding protein via the phosphorylation of mitogen-activated protein kinase, suggesting that quercetin can regulate the adipocyte life cycle⁽Reference Ahn, Lee and Kim¹⁴³⁾. Dietary bilberry extracts rich in anthocyanidins ameliorate hyperglycaemia and insulin sensitivity in diabetic mice by activating AMPK in the adipose tissue, skeletal muscle and liver⁽Reference Takikawa, Inoue and Horio¹⁴⁴⁾.

Sirtuin 1

Polyphenols such as resveratrol activate a NAD⁺-dependent protein deacetylase, silent information regulator orthologue 1 (SIRT1)⁽Reference Milne, Lambert and Schenk¹⁴⁵⁾, which regulates a variety of cellular functions such as genome maintenance, longevity and metabolism⁽Reference Michan and Sinclair^146, Reference Milne and Denu¹⁴⁷⁾. Resveratrol increases the lifespan in animals partially via the stimulation of SIRT1, in a manner similar to energy restriction⁽Reference Guarente¹⁴⁸⁾. Resveratrol augments exercise endurance in mice through the deacetylation of PGC-1α (a mitochondrial biogenesis factor) by SIRT1 to stimulate mitochondrial function in muscle and brown adipose tissue⁽Reference Lagouge, Argmann and Gerhart-Hines¹⁴⁹⁾. The pleiotropic effects of resveratrol, which occur by the activation of SIRT1, could protect animals from obesity and diabetes by shifting the energy balance towards energy consumption rather than storage⁽Reference Lagouge, Argmann and Gerhart-Hines¹⁴⁹⁾.

Small-interfering RNA against SOD2 or SIRT1 reduce the cell-protective effects of resveratrol⁽Reference Tanno, Kuno and Yano⁵¹⁾. Although a recent study using cell-free assays questions the ability of resveratrol to activate SIRT1 directly⁽Reference Pacholec, Bleasdale and Chrunyk¹⁵⁰⁾, it is highly likely that resveratrol (or its metabolites) can promote SIRT1 activation in vivo. Moreover, resveratrol also enhances the expression levels of SIRT1. In an attempt to address this, Li's group reported that inhibition of SIRT1 activity with sirtinol or knockdown of SIRT1 expression with siRNA both reduced the effects of resveratrol on SOD1, SOD2 and GPx1, but not those on SOD3 and catalase⁽Reference Xia, Daiber and Habermeier⁵²⁾. This finding is consistent with the findings that resveratrol up-regulates SOD2 in C2C12 myoblasts in a SIRT1-dependent manner⁽Reference Tanno, Kuno and Yano⁵¹⁾.

Of note is the report that SIRT1 also activates the transcriptional activity of PGC-1α, and subsequently induces mitochondrial biogenesis and lipolysis, and so inhibits the generation of ROS from the mitochondria⁽Reference Liang, Kume and Koya¹⁵¹⁾. The activation of SIRT1 is related to both lipid and glucose homeostasis; thus, SIRT1 inhibits adipogenesis, reduces fat storage in adipose tissue⁽Reference Picard, Kurtev and Chung¹⁵²⁾ and increases insulin secretion and sensitivity⁽Reference Bordone, Motta and Picard¹⁵³⁾.

Resveratrol stimulates eNOS activity by SIRT1 activation and eNOS deacetylation⁽Reference Mattagajasingh, Kim and Naqvi¹⁵⁴⁾. For example, resveratrol increases mitochondrial mass and up-regulates eNOS by activating SIRT1 in human coronary arterial endothelial cells, where the ability of resveratrol to induce mitochondrial biogenesis is NO-dependent⁽Reference Csiszar, Labinskyy and Pinto¹⁵⁵⁾. Likewise, SIRT1 activation by other stimuli such as laminar flow and statin treatment also increases eNOS activity and NO production⁽Reference Chen, Peng and Cui¹⁵⁶⁾. Thus, the interaction between SIRT1 and eNOS contributes to the cardiovascular beneficial effects of resveratrol. The multifaceted molecular mechanisms for the cardiovascular benefits of resveratrol are summarised in Fig. 4.

Fig. 4

Resveratrol reduces oxidative stress by decreasing reactive oxygen species (ROS) production from NADPH oxidases (NOX), uncoupled endothelial NO synthase (eNOS, by up-regulating GTP cyclohydrolase 1, GCH1) and mitochondria (by stimulating mitochondrial (Mito.) biogenesis). In addition, resveratrol enhances the expression of antioxidant enzymes, such as superoxide dismutases (SOD1–3), catalase (Cat.), glutathione peroxidase 1 (GPx1), NAD(P)H:quinone oxidoreductase 1 (NQO1), γ-glutamylcysteine synthetase (glutamate cysteine ligase catalytic subunit, GCLC), haem oxygenase-1 (HO-1) and thioredoxin-1 (Trx-1). SIRT1, sirtuin 1; PGC-1α, PPARγ coactivator 1-α; Nrf2, nuclear factor E₂-related factor-2. (A colour version of this figure can be found online at http://www.journals.cambridge.org/bjn)

In a similar manner, treating mice with quercetin enhances mRNA expression of PGC-1α and SIRT1 to increase both maximal endurance capacity and running activity⁽Reference Davis, Murphy and Carmichael¹⁵⁷⁾. The possibility of targeting SIRT1 by polyphenols, and thereby co-affecting PGC-1α signalling, makes endothelial mitochondria important in CVD and metabolic disorders.

Oestrogen receptor α

Due to the structural similarities with diethylstilbestrol (a synthetic oestrogen), resveratrol has been proposed to activate the ER. Resveratrol binds to and activates gene transcription via the ER in oestrogen-sensitive tissues and cell lines⁽Reference Gehm, McAndrews and Chien¹⁵⁸⁾. Of interest, resveratrol binds ERβ with a lower affinity than at ERα⁽Reference Kuiper, Carlsson and Grandien¹⁵⁹⁾. Owing to its properties as an agonist for the ER, resveratrol is able to regulate the transcription of oestrogen-responsive target genes, and possibly has cancer chemopreventive effects⁽Reference Le Corre, Chalabi and Delort¹⁶⁰⁾. Activation of ER is a key step in the effects of resveratrol on glucose uptake by muscles⁽Reference Deng, Hsieh and Huang¹⁶¹⁾. At the molecular level, resveratrol rapidly activates ERα in caveolae, leading to eNOS activation by the stimulation of G-protein Ga, caveolin-1 (Cav-1), Src and ERK1/2K; siRNA knockdown of ERα, but not ERβ, or the presence of ER antagonists inhibits the rapid eNOS activation by resveratrol⁽Reference Klinge, Wickramasinghe and Ivanova⁶⁸⁾.

As described for resveratrol, quercetin is also able to reduce oestrogen-sensitive tumour growth in mouse models by directly acting on ER⁽Reference Schlachterman, Valle and Wall¹⁶²⁾ and by down-regulating cytoplasmic ER levels and promotion of a tighter nuclear association of the ER⁽Reference Miodini, Fioravanti and Di Fronzo¹⁶³⁾. Quercetin exhibits a similar potency of both ER subtypes⁽Reference Kuiper, Lemmen and Carlsson¹⁶⁴⁾ and stimulates the expression of the proto-oncogene c-fos through ERα activation⁽Reference Maggiolini, Vivacqua and Fasanella¹⁶⁵⁾.

Recent data suggest that delphinidin interacts directly with the activator site of ERα, leading to the activation of eNOS. Thus, the ability of delphinidin (and of total polyphenolic extract from red wine) to induce NO production and endothelium-dependent vasorelaxation data is lost in ERα-deficient mice or after using siRNA for this receptor⁽Reference Chalopin, Tesse and Martinez²⁶⁾. Silencing the effects of ERα completely prevents delphinidin activation of Src, ERK1/2 and eNOS, while binding assay and docking experiments indicate a direct interaction between delphinidin and the ERα activator site. Oral administration of total polyphenolic extracts from red wine increases the sensitivity of endothelium-dependent relaxation to acetylcholine and is associated with increased NO production and decreased superoxide anions in control mice but absent in ERα-deficient mice⁽Reference Chalopin, Tesse and Martinez²⁶⁾.

Interaction between molecular targets of polyphenols

It is likely that there is an intracellular crosstalk of signalling cascades activated by molecular targets of polyphenols. For example, resveratrol modulates tumour cell proliferation and protein translation via SIRT1-dependent AMPK activation in ER-positive breast cancer cells, highlighting the interactions of ER, SIRT1 and AMPK⁽Reference Lin, Lin and Rau¹⁶⁶⁾. Resveratrol induces deacetylation of PGC-1α mediated by SIRT1 and phosphorylation of AMPK in the liver to promote fatty acid oxidation and inhibit lipogenesis⁽Reference Baur, Pearson and Price¹⁶⁷⁾. It is likely that SIRT1 may be upstream of AMPK, since SIRT1 activation increases AMPK activity⁽Reference Hou, Xu and Maitland-Toolan¹⁶⁸⁾, probably by SIRT1 deacetylation/activation of the upstream AMPK kinase liver kinase B1 (LKB1)⁽Reference Lan, Cacicedo, Ruderman and Ido¹⁶⁹⁾. Finally, eNOS acetylation is higher in AMPKα2-deficient mice, suggesting that AMPK phosphorylation of eNOS is required for SIRT1 deacetylation of eNOS⁽Reference Chen, Peng and Cui¹⁵⁶⁾. These findings suggest that the improvement of cell function produced by the polyphenols resveratrol, quercetin and delphinidin occurs by the activation of several signalling mechanisms in addition to the transcriptional and post-translational effects.

Hypertension

Hypertension causes modifications of the vascular walls that lead to hypertensive cardiomyopathy and heart failure⁽Reference Lan, Cacicedo, Ruderman and Ido¹⁷⁰⁾. Changes in the mechanical properties of arteries affect vascular resistance by altering the pressure–lumen diameter relationship of small arteries⁽Reference Touyz and Schiffrin⁷⁴⁾. Part of the cardioprotective actions of polyphenols is by lowering blood pressure. However, contradictory data are available: for instance, an antihypertensive effect of resveratrol was reported in partially nephrectomised rats⁽Reference Liu, Song and Zhang¹⁷¹⁾, while in double transgenic rats harbouring human renin and angiotensinogen genes, resveratrol reduces blood pressure, ameliorates cardiac hypertrophy and prevents angiotensin II-induced mortality, probably by increasing mitochondrial biogenesis and SIRT1 activity⁽Reference Biala, Tauriainen and Siltanen¹⁷²⁾. Resveratrol probably suppresses angiotensin II type 1 receptor expression through SIRT1 activation, suggesting that the inhibition of the renin–angiotensin system may contribute, at least in part, to the resveratrol-induced cardiprotective effects⁽Reference Miyazaki, Ichiki and Hashimoto³³⁾. Other studies report that resveratrol does not affect established hypertension in SHR⁽Reference Rush, Quadrilatero and Levy¹⁷³⁾, although it attenuates the compliance of arteries from SHR without changes in wall stiffness by reducing eutrophic remodelling⁽Reference Behbahani, Thandapilly and Louis¹⁷⁴⁾.

Chronic treatment with quercetin (10 mg/kg) reduces systolic blood pressure and significantly reduces left ventricular and renal hypertrophy in SHR⁽Reference Romero, Jimenez and Hurtado¹⁷⁵⁾, hypertension induced by the inhibition of NOS⁽Reference Perez-Vizcaino, Duarte and Jimenez⁸⁶⁾ and in deoxycorticosterone acetate-salt hypertensive rats⁽Reference Galisteo, Garcia-Saura and Jimenez¹⁷⁶⁾. It appears that quercetin is effective in all animal models of hypertension studied, and acts independently of the status of renin–angiotensin system, oxidative stress, NO, etc.⁽Reference Perez-Vizcaino, Duarte and Jimenez⁸⁶⁾.

Short-term oral administration of polyphenols from red wine (a rich source of delphinidin) decreases blood pressure in normotensive rats. This haemodynamic effect was associated with an enhanced endothelium-dependent relaxation and an induction of gene expression within the arterial wall, which together maintain unchanged agonist-induced contractility⁽Reference Diebolt, Bucher and Andriantsitohaina¹⁷⁷⁾. Polyphenols from red wine reduce blood pressure elevations caused by chronic inhibition of NOS, attenuate end-organ damage such as myocardial fibrosis and aortic thickening, and decrease protein synthesis in the heart and aorta⁽Reference Bernatova, Pechanova and Babal^178, Reference Pechanova, Bernatova and Babal¹⁷⁹⁾. Polyphenols also prevent endothelium-dysfunction by increasing eNOS activity, moderately enhancing eNOS expression and reducing oxidative stress in the left ventricle and aorta. Endothelial dysfunction associated with excessive NADPH oxidase-dependent vascular formation of ROS in angiotensin II-induced hypertension is prevented by polyphenols⁽Reference Sarr, Chataigneau and Martins³¹⁾. Thus, polyphenols from red wine reduce hypertension by modulating the NO and ROS balance in the cardiovascular system.

Stroke

Cerebral ischaemia is caused by reduced cerebral blood flow. Stroke involves the interaction of neurons, glia, vascular cells and matrix components, all of which participate in the mechanisms of tissue injury and repair. The severe reduction of cerebral blood flow initiates a series of pathophysiological mechanisms such as impaired energy metabolism, loss of ionic homeostasis, excessive release of excitatory amino acids (mainly aspartate and glutamate) and increased oxidative stress. All these processes lead to brain tissue damage and cell death.

Resveratrol reduces infarct volume in various experimental models of stroke⁽Reference Simonyi, Wang and Miller¹⁸⁰⁾. The mechanisms involved in neuroprotection are largely by the inhibition of lipid oxidation processes. More recent data indicate that resveratrol significantly restores ATP content and the activity of mitochondrial respiratory complexes in a model of transient rat middle cerebral artery occlusion by decreasing apoptosis, mitochondrial lipid peroxidation, brain infarct volume and oedema⁽Reference Yousuf, Atif and Ahmad¹⁸¹⁾. In the stroke model, resveratrol improves neurological function by reducing the release of excitatory neurotransmitters (glutamate and aspartate), and increases inhibitory neurotransmitter release (γ-amino-n-butyric acid and glycine)⁽Reference Li, Yan and Yang¹⁸²⁾. It is likely that these effects are mediated through the activation of oestrogen and N-methyl-d-aspartate receptors⁽Reference Saleh, Connell and Saleh¹⁸³⁾ or the SIRT1 pathway⁽Reference Della-Morte, Dave and DeFazio¹⁸⁴⁾. Resveratrol administration also induces angiogenesis in the cortical area of mice exposed to middle cerebral artery ischaemia⁽Reference Dong, Li and Gao¹⁸⁵⁾. These findings highlight the ability of resveratrol to preserve ischaemic neurovascular units in the treatment of ischaemic stroke.

Liposomal preparations of quercetin that enhance neuroprotective capacity reduce cerebral damage provoked by cerebral ischaemia⁽Reference Rivera, Costa and Abin¹⁸⁶⁾. Repeated treatment with quercetin for 15 d before ischaemic surgery in gerbils reduces lipid peroxidation, suggesting that early administration of quercetin could offer protection of neuronal units during cerebral ischaemia⁽Reference Hwang, Lee and Yoo¹⁸⁷⁾.

Feeding rats with diets enriched in anthocyanins from blueberries provides neuroprotection after stroke induced by ligation of the left common carotid artery independently of their ability to scavenge oxygen radicals⁽Reference Sweeney, Kalt and MacKinnon¹⁸⁸⁾. An anthocyanin-rich extract from red wine reduces injury induced by cerebral ischaemia in rats, and protects from ischaemia-induced excitotoxicity (by reducing the release of the excitatory neurotransmitters glutamate and aspartate), energy failure (by increasing glucose concentrations) and oxidative stress (by increasing levels of ascorbic and uric acids)⁽Reference Ritz, Curin and Mendelowitsch¹⁸⁹⁾. Long-term administration of polyphenols partially restores cerebral blood flow during cerebral artery occlusion and improves flow during reperfusion in the cortex, as measured by increased diameters of the arteries of the cerebral tree, while also causing differential expression of proteins involved in neuroprotection, maintenance of neuronal integrity, oxidative stress, energy metabolism and inflammation (such as neurofilament medium polypeptide (NF-M) or TOAD-64)⁽Reference Ritz, Ratajczak and Curin¹⁹⁰⁾. These experimental data indicate the beneficial effects of polyphenols in stroke protection, or in treatment during different phases of the disease.