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Endothelial progenitor cells (EPCs) and microparticles are emerging as novel markers of cardiovascular disease (CVD) risk, which could potentially be modified by dietary fat. We have previously shown that replacing dietary saturated fatty acids (SFAs) with monounsaturated or n-6 (ω-6) polyunsaturated fatty acids (MUFAs or PUFAs, respectively) improved lipid biomarkers, blood pressure, and markers of endothelial activation, but their effects on circulating EPCs and microparticles are unclear.The Dietary Intervention and VAScular function (DIVAS) Study investigated the replacement of 9.5-9.6% of total energy (%TE) contributed by SFAs with MUFAs or n-6 PUFAs for 16 wk on EPC and microparticle numbers in United Kingdom adults with moderate CVD risk.In this randomized, controlled, single-blind, parallel-group dietary intervention, men and women aged 21-60 y (n = 190) with moderate CVD risk (≥50% above the population mean) consumed 1 of three 16-wk isoenergetic diets. Target compositions for total fat, SFAs, MUFAs, and n-6 PUFAs (%TE) were as follows: SFA-rich diet (36:17:11:4; n = 64), MUFA-rich diet (36:9:19:4; n = 62), and n-6 PUFA-rich diet (36:9:13:10; n = 66). Circulating EPC, endothelial microparticle (EMP), and platelet microparticle (PMP) numbers were analyzed by flow cytometry. Dietary intake, vascular function, and other cardiometabolic risk factors were determined at baseline.Relative to the SFA-rich diet, MUFA- and n-6 PUFA-rich diets decreased EMP (-47.3%, -44.9%) respectively and PMP (-36.8%, -39.1%) numbers (overall diet effects, P 0.01). The MUFA-rich diet increased EPC numbers (+28.4%; P = 0.023). Additional analyses that used stepwise regression models identified the augmentation index (measuring arterial stiffness determined by pulse-wave analysis) as an independent predictor of baseline EPC and microparticle numbers.Replacement of 9.5-9.6%TE dietary SFAs with MUFAs increased EPC numbers, and replacement with either MUFAs or n-6 PUFAs decreased microparticle numbers, suggesting beneficial effects on endothelial repair and maintenance. Further studies are warranted to determine the mechanisms underlying the favorable effects on EPC and microparticle numbers after SFA replacement. This trial was registered at www.clinicaltrials.gov as NCT01478958.

Public health strategies to lower cardiovascular disease (CVD) risk involve reducing dietary saturated fatty acid (SFA) intake to ≤10% of total energy (%TE). However, the optimal type of replacement fat is unclear.We investigated the substitution of 9.5-9.6%TE dietary SFAs with either monounsaturated fatty acids (MUFAs) or n-6 (ω-6) polyunsaturated fatty acids (PUFAs) on vascular function and other CVD risk factors.In a randomized, controlled, single-blind, parallel-group dietary intervention, 195 men and women aged 21-60 y from the United Kingdom with moderate CVD risk (≥50% above the population mean) followed one of three 16-wk isoenergetic diets (%TE target compositions, total fat:SFA:MUFA:n-6 PUFA) that were rich in SFAs (36:17:11:4, n = 65), MUFAs (36:9:19:4, n = 64), or n-6 PUFAs (36:9:13:10, n = 66). The primary outcome measure was flow-mediated dilatation; secondary outcome measures included fasting serum lipids, microvascular reactivity, arterial stiffness, ambulatory blood pressure, and markers of insulin resistance, inflammation, and endothelial activation.Replacing SFAs with MUFAs or n-6 PUFAs did not affect the percentage of flow-mediated dilatation (primary endpoint) or other measures of vascular reactivity. Of the secondary outcome measures, substitution of SFAs with MUFAs attenuated the increase in night systolic blood pressure (-4.9 mm Hg, P = 0.019) and reduced E-selectin (-7.8%, P = 0.012). Replacement with MUFAs or n-6 PUFAs lowered fasting serum total cholesterol (-8.4% and -9.2%, respectively), low-density lipoprotein cholesterol (-11.3% and -13.6%), and total cholesterol to high-density lipoprotein cholesterol ratio (-5.6% and -8.5%) (P ≤ 0.001). These changes in low-density lipoprotein cholesterol equate to an estimated 17-20% reduction in CVD mortality.Substitution of 9.5-9.6%TE dietary SFAs with either MUFAs or n-6 PUFAs did not significantly affect the percentage of flow-mediated dilatation or other measures of vascular function. However, the beneficial effects on serum lipid biomarkers, blood pressure, and E-selectin offer a potential public health strategy for CVD risk reduction. This trial was registered at www.clinicaltrials.gov as NCT01478958.

Evidence suggests that a combination of oxidative stress, neuroinflammation and the formation of endogenous neurotoxins contribute to the underlying neuronal death associated with neurodegenerative diseases. In this study we have investigated the ability of the berry-derived flavonoids to protect against neuronal damage induced by neuroinflammation and the neurotoxin 5-S-cysteinyl-dopamine. The flavanols (+)-catechin and (-)-epicatechin, but not the anthocyanidin pelargonidin, were observed to attenuate LPS/IFN-?-induced TNF-a production in glial cells and associated neuronal injury. In contrast, pre-treatment of primary cortical neurons with pelargonidin, (+)-catechin and (-)-epicatechin (0.1 and 0.3 µM) resulted in concentration-dependant protection against 5-S-cysteinyl-dopamine-induced neurotoxicity. Together these data suggest that berry-derived flavonoids may offer some protection against the neuronal injury relevant to the aetiology of the Parkinson’s and Alzheimer’s diseases.

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.

Flavonoids exert a multiplicity of neuroprotective actions within the brain, including a potential to protect neurons against injury induced by neurotoxins, an ability to suppress neuroinflammation, and the potential to promote memory, learning and cognitive function. These effects appear to be underpinned by two common processes. Firstly, they interact with critical protein and lipid kinase signalling cascades in the brain leading to an inhibition of apoptosis triggered by neurotoxic species and to a promotion of neuronal survival and synaptic plasticity. Secondly, they induce beneficial effects on the vascular system leading to changes in cerebrovascular blood flow capable of causing angiogenesis, neurogenesis and changes in neuronal morphology. Through these mechanisms, the consumption of flavonoid-rich foods throughout life holds the potential to limit neurodegeneration and to prevent or reverse age-dependent loses in cognitive performance. The intense interest in the development of drugs capable of enhancing brain function means that flavonoids may represent important precursor molecules in the quest to develop of a new generation of brain enhancing drugs.

There is growing interest in the potential beneficial effects of flavonoids in the aging and diseased brain. We have investigated the potential of the flavanone hesperetin and two of its metabolites, hesperetin-7-O-beta-d-glucuronide and 5-nitro-hesperetin, to inhibit oxidative stress-induced neuronal apoptosis. Exposure of cortical neurons to hydrogen peroxide led to the activation of apoptosis signal-regulating kinase 1 via its de-phosphorylation at Ser963, the phosphorylation of c-jun N-terminal kinase and c-Jun (Ser73) and the activation of caspase 3 and caspase 9. Whilst hesperetin glucuronide failed to exert protection, both hesperetin and 5-nitro-hesperetin were effective at preventing neuronal apoptosis via a mechanism involving the activation/phosphorylation of both Akt/protein kinase B and extracellular signal-regulated kinase 1 and 2 (ERK1/2). Protection against oxidative injury and the activation of Akt and ERK1/2 followed a bell-shaped response and was most apparent at 100 nmol/L concentrations. The activation of ERK1/2 and Akt by flavanones led to the inhibition of the pro-apoptotic proteins, apoptosis signal-regulating kinase 1, by phosphorylation at Ser83 and Bad, by phosphorylation at both Ser136 and Ser112 and to the inhibition of peroxide-induced caspase 9 and caspase 3 activation. Thus, flavanones may protect neurons against oxidative insults via the modulation of neuronal apoptotic machinery.

The death of nigral neurons in Parkinson's disease is thought to involve the formation of the endogenous neurotoxin, 5-S-cysteinyl-dopamine. In the present study, we show that the polyphenols, (+)-catechin and caffeic acid, which contain a catechol moiety, inhibit tyrosinase-induced formation of 5-S-cysteinyl-dopamine via their capacity to undergo tyrosinase-induced oxidation to yield cysteinyl-polyphenol adducts. In contrast, the inhibition afforded by the flavanone, hesperetin, was not accompanied by the formation of cysteinyl-hesperetin adducts, indicating that it may inhibit via direct interaction with tyrosinase. Whilst the stilbene resveratrol also inhibited 5-S-cysteinyl-dopamine formation, this was accompanied by the formation of dihydrobenzothiazine, a strong neurotoxin. Our data indicate that the inhibitory effects of polyphenols against 5-S-cysteinyl-dopamine formation are structure-dependent and shed further light on the mechanisms by which polyphenols exert protection against neuronal injury relevant to neurodegenerative diseases.

White wines are generally low in polyphenol content as compared to red wines. However, Champagne wines have been shown to contain relatively high amounts of phenolic acids that may exert protective cellular actions in vivo. In this study, we have investigated the potential neuroprotective effects of Champagne wine extracts, and individual phenolics present in these extracts, against peroxynitrite-induced injury. Organic and aqueous Champagne wine extracts exhibited potent neuroprotective activity against peroxynitrite-induced injury at low concentrations (0.1 microg/mL). This protection appeared to be in part due to the cellular actions of individual components found in the organic extracts, notably tyrosol, caffeic acid, and gallic acid. These phenolics were observed to exert potent neuroprotection at concentrations between 0.1 and 10 microM. Together, these data suggest that polyphenols present in Champagne wine may induce a neuroprotective effect against oxidative neuronal injury.

There is much interest in the bioactivity of in vivo flavonoid metabolites. We report for the first time the hierarchy of reactivity of flavonoid metabolites with peroxynitrite and characterise novel reaction products. O-Methylation of the B-ring catechol containing flavonoids epicatechin and quercetin, and O-glucuronidation of all flavonoids reduced their reactivity with peroxynitrite. The reaction of the flavanones hesperetin and naringenin and their glucuronides resulted in the formation of multiple mono-nitrated and nitrosated products. In contrast, the catechol-containing flavonoids epicatechin and quercetin yielded oxidation products which when trapped with glutathione led to the production of glutathionyl-conjugates. However, the O-methylated metabolites of epicatechin yielded both mono- and di-nitrated products and nitrosated metabolites. The 3'-O-methyl metabolite of quercetin also yielded a nitrosated species, although its counterpart 4'-O-methyl quercetin yielded only oxidation products. Such products may represent novel metabolic products in vivo and may also express cellular activity.

The increase in CVD incidence following the menopause is associated with oestrogen loss. Dietary isoflavones are thought to be cardioprotective via their oestrogenic and oestrogen receptor-independent effects, but evidence to support this role is scarce. Individual variation in response to diet may be considerable and can obscure potential beneficial effects in a sample population; in particular, the response to isoflavone treatment may vary according to genotype and equol-production status. The effects of isoflavone supplementation (50 mg/d) on a range of established and novel biomarkers of CVD, including markers of lipid and glucose metabolism and inflammatory biomarkers, have been investigated in a placebo-controlled 2×8-week randomised cross-over study in 117 healthy post-menopausal women. Responsiveness to isoflavone supplementation according to (1) single nucleotide polymorphisms in a range of key CVD genes, including oestrogen receptor (ER) α and β and (2) equol-production status has been examined. Isoflavones supplementation was found to have no effect on markers of lipids and glucose metabolism. Isoflavones improve C-reactive protein concentrations but do not affect other plasma inflammatory markers. There are no differences in response to isoflavones according to equol-production status. However, differences in HDL-cholesterol and vascular cell adhesion molecule 1 response to isoflavones v. placebo are evident with specific ERβ genotypes. In conclusion, isoflavones have beneficial effects on C-reactive protein, but not other cardiovascular risk markers. However, specific ERβ gene polymorphic subgroups may benefit from isoflavone supplementation.

Homocysteine and asymmetric dimethylarginine (ADMA) affect nitric oxide (NO) concentration, thereby contributing to cardiovascular disease (CVD). Both amino acids can be reduced in vivo by estrogen. Variation in the estrogen receptor (ER) may influence homocysteine and ADMA, yet no information is available on associations with single nucleotide polymorphisms in the estrogen receptor genes ERalpha (PvuII and XbaI) and ERbeta (1730G--A and cx + 56 G--A).To find relationships between common polymorphisms associated with cardiovascular disease and cardiovascular risk factors homocysteine and ADMA.In a cross-sectional study with healthy postmenopausal women (n = 89), homocysteine, ADMA, nitric oxide metabolites (NOx), plasma folate and ERalpha and beta polymorphisms ERalpha PvuII, ERalpha XbaI; ERbeta 1730G--A (AluI), ERbeta cx + 56 G--A (Tsp509I) were analyzed.Women who are homozygotic for ERbetacx + 56 G--A A/A exhibited higher homocysteine (p = 0.012) and NOx (p = 0.056) levels than wildtype or heterozygotes. NOx concentration was also significantly affected by ERbeta 1730 G --A polymorphism (p = 0.025). The ERbeta (p0.001) and ERalpha (p0.001) polymorphisms were in linkage disequilibrium.Women who are homozygotic for ERbetacx + 56 G--A A/A may be at increased risk for cardiovascular disease due to higher homocysteine levels.

The importance of chronic low-grade inflammation in the pathology of numerous age-related chronic conditions is now clear. An unresolved inflammatory response is likely to be involved from the early stages of disease development. The present position paper is the most recent in a series produced by the International Life Sciences Institute's European Branch (ILSI Europe). It is co-authored by the speakers from a 2013 workshop led by the Obesity and Diabetes Task Force entitled 'Low-grade inflammation, a high-grade challenge: biomarkers and modulation by dietary strategies'. The latest research in the areas of acute and chronic inflammation and cardiometabolic, gut and cognitive health is presented along with the cellular and molecular mechanisms underlying inflammation-health/disease associations. The evidence relating diet composition and early-life nutrition to inflammatory status is reviewed. Human epidemiological and intervention data are thus far heavily reliant on the measurement of inflammatory markers in the circulation, and in particular cytokines in the fasting state, which are recognised as an insensitive and highly variable index of tissue inflammation. Potential novel kinetic and integrated approaches to capture inflammatory status in humans are discussed. Such approaches are likely to provide a more discriminating means of quantifying inflammation-health/disease associations, and the ability of diet to positively modulate inflammation and provide the much needed evidence to develop research portfolios that will inform new product development and associated health claims. Copyright © 2015 ILSI Europe. Chemicals/CAS: C reactive protein, 9007-41-4

The recommendation to reduce saturated fatty acid (SFA) consumption to ≤10% of total energy (%TE) is a key public health target aimed at lowering cardiovascular disease (CVD) risk. Replacement of SFA with unsaturated fats may provide greater benefit than replacement with carbohydrates, yet the optimal type of fat is unclear. The aim of the DIVAS (Dietary Intervention and Vascular Function) study was to develop a flexible food-exchange model to investigate the effects of substituting SFAs with monounsaturated fatty acids (MUFAs) or n-6 (ω-6) polyunsaturated fatty acids (PUFAs) on CVD risk factors. In this parallel study, UK adults aged 21-60 y with moderate CVD risk (50% greater than the population mean) were identified using a risk assessment tool (n = 195; 56% females). Three 16-wk isoenergetic diets of specific fatty acid (FA) composition (%TE SFA:%TE MUFA:%TE n-6 PUFA) were designed using spreads, oils, dairy products, and snacks as follows: 1) SFA-rich diet (17:11:4; n = 65); 2) MUFA-rich diet (9:19:4; n = 64); and 3) n-6 PUFA-rich diet (9:13:10; n = 66). Each diet provided 36%TE total fat. Dietary targets were broadly met for all intervention groups, reaching 17.6 ± 0.4%TE SFA, 18.5 ± 0.3%TE MUFA, and 10.4 ± 0.3%TE n-6 PUFA in the respective diets, with significant overall diet effects for the changes in SFAs, MUFAs, and n-6 PUFAs between groups (P < 0.001). There were no differences in the changes of total fat, protein, carbohydrate, and alcohol intake or anthropometric measures between groups. Plasma phospholipid FA composition showed changes from baseline in the proportions of total SFAs, MUFAs, and n-6 PUFAs for each diet group, with the changes in SFAs and MUFAs differing between the groups (P < 0.001). In conclusion, successful implementation of the food-exchange model broadly achieved the dietary target intakes for the exchange of SFAs with MUFAs or n-6 PUFAs with minimal disruption to the overall diet in a free-living population. This trial was registered at clinicaltrials.gov as NCT01478958.

The endogenous neurotoxin 5-S-cysteinyl-dopamine (CysDA) has been linked to the degeneration of dopaminergic neurons in the Substantia nigra. Sulforaphane (SFN) is an isothiocyanate derived from cruciferous vegetables that has been proposed as a potential chemopreventive agent, although its effects on other cells are less clear. This study was undertaken to investigate the protective effects of SFN against CysDA-induced injury in primary cortical neurons. The results indicate that SFN protects primary cortical neurons against CysDA-induced injury by a mechanism involving the increased nuclear translocation of Nrf2 and the increased activity of phase II enzymes such as glutathione-S-transferase, glutathione reductase, and thioredoxine reductase. Moreover, the protection exerted by SFN appears to be mediated by the activation of the extracellular signal-regulated kinase 1 and 2 (ERK1/2) and Akt/protein kinase B (PKB) pathways.

Neuroinflammatory processes are known to contribute to the cascade of events culminating in the neuronal damage that underpins neurodegenerative disorders such as Parkinson's and Alzheimer's disease. Recently, there has been much interest in the potential neuroprotective effects of flavonoids, a group of plant secondary metabolites known to have diverse biological activity in vivo. With respect to the brain, flavonoids, such as those found in cocoa, tea, berries and citrus, have been shown to be highly effective in preventing age-related cognitive decline and neurodegeneration in both animals and humans. Evidence suggests that flavonoids may express such ability through a multitude of physiological functions, including an ability to modulate the brains immune system. This review will highlight the evidence for their potential to inhibit neuroinflammation through an attenuation of microglial activation and associated cytokine release, iNOS expression, nitric oxide production and NADPH oxidase activity. We will also detail the current evidence indicting that their regulation of these immune events appear to be mediated by their actions on intracellular signaling pathways, including the nuclear factor-κB (NF-κB) cascade and mitogen-activated protein kinase (MAPK) pathway. As such, flavonoids represent important precursor molecules in the quest to develop of a new generation of drugs capable of counteracting neuroinflammation and neurodegenerative disease.

The degeneration of dopaminergic neurons in the substantia nigra has been linked to the formation of the endogenous neurotoxin 5-S-cysteinyl-dopamine. Sulforaphane (SFN), an isothiocyanate derived from the corresponding precursor glucosinolate found in cruciferous vegetables has been observed to exert a range of biological activities in various cell populations. In this study, we show that SFN protects primary cortical neurons against 5-S-cysteinyl-dopamine induced neuronal injury. Pre-treatment of cortical neurons with SFN (0.01-1 microM) resulted in protection against 5-S-cysteinyl-dopamine-induced neurotoxicity, which peaked at 100 nM. This protection was observed to be mediated by the ability of SFN to modulate the extracellular signal-regulated kinase 1 and 2 and the activation of Kelch-like ECH-associated protein 1/NF-E2-related factor-2 leading to the increased expression and activity of glutathione-S-transferase (M1, M3 and M5), glutathione reductase, thioredoxin reductase and NAD(P)H oxidoreductase 1. These data suggest that SFN stimulates the NF-E2-related factor-2 pathway of antioxidant gene expression in neurons and may protect against neuronal injury relevant to the aetiology of Parkinson's disease.

The olive oil polyphenol, hydroxytyrosol (HT), is believed to be capable of exerting protection against oxidative kidney injury. In this study we have investigated the ability of HT and its O-methylated metabolite, homovanillic alcohol (HVA) to protect renal cells against oxidative damage induced by hydrogen peroxide. We show that both compounds were capable of inhibiting hydrogen peroxide-induced kidney cell injury via an ability to interact with both MAP kinase and PI3 kinase signalling pathways, albeit at different concentrations. HT strongly inhibited death and prevented peroxide-induced increases in ERK1/2 and JNK1/2/3 phosphorylation at 0.3 microM, whilst HVA was effective at 10 microM. At similar concentrations, both compounds also prevented peroxide-induced reductions in Akt phosphorylation. We suggest that one potential protective effect exerted by olive oil polyphenols against oxidative kidney cell injury may be attributed to the interactions of HT and HVA with these important intracellular signalling pathways.

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.

Various epidemiological investigations, as well as dietary supplementation studies in humans and animals, have established a positive correlation between the intake of flavonoid-rich foods or beverages and the attenuation or delayed onset of cardiovascular disease, cancer and neurodegeneration. Flavonoids are founds ubiquitously in the human diet, although some foods and beverages, such as tea, wine, cocoa, berries and citrus fruits are particularly rich sources of these plant phytochemicals. They are extensively metabolised at various sites within the human body, most notably the small intestine, large intestine and liver, where they are converted into a variety of metabolic forms. Due to this metabolism and the low extent of their overall absorption, it is likely that their beneficial actions are mediated by their abilities to interact with both protein and lipid kinase signalling cascades, rather than via their potential to act as classical antioxidants. The concentrations of flavonoids encountered in vivo are sufficiently high to exert pharmacological activity at receptors, and on kinases and transcription factors. Presently the precise sites of action are unknown, although it is likely that their activity depends on their ability to: 1) bind to ATP sites on enzymes and receptors; 2) modulate the activity of kinases directly, i.e. MAPKKK, MAPKK or MAPK; 3) affect the function of important phosphatases, which act in opposition to kinases; 4) preserve Ca2+ homeostasis, thereby preventing Ca2+-dependent activation of kinases in neurons; and 5) modulate signalling cascades lying downstream of kinases, i.e. transcription factor activation and binding to promoter sequences. Such interactions will be discussed in relation to the precise site(s) of action of flavonoids within signalling pathways and the sequence of events that allow them to regulate cellular function at various sites within the body.

There is increasing evidence to suggest that neuroinflammatory processes contribute to the cascade of events that lead to the progressive neuronal damage observed in neurodegenerative disorders such as Parkinson's disease and Alzheimer's disease. Therefore, treatment regimes aimed at modulating neuroinflammatory processes may act to slow the progression of these debilitating brain disorders. Recently, a group of dietary polyphenols known as flavonoids have been shown to exert neuroprotective effects in vivo and in neuronal cell models. In this review we discuss the evidence relating to the modulation of neuroinflammation by flavonoids. We highlight the evidence which suggests their mechanism of action involves: 1) attenuation of the release of cytokines, such as interleukin-1beta (IL-1beta and tumor necrosis factor-alpha (TNF-alpha); 2) an inhibitory action against inducible nitric oxide synthase (iNOS) induction and subsequent nitric oxide (NO(*)) production; 3) inhibition of the activation of NADPH oxidase and subsequent reactive oxygen species generation; 4) a capacity to down-regulate the activity of pro-inflammatory transcription factors such as nuclear factor-kappaB (NF-kappaB); and 5) the potential to modulate signalling pathways such as mitogen-activated protein kinase (MAPK) cascade. We also consider the potential of these dietary compounds to represent novel therapeutic agents by considering their metabolism in the body and their ability to access the brain via the blood brain barrier. Finally, we discuss future areas of study which are necessary before dietary flavonoids can be established as therapeutic agents against neuroinflammation.

The cellular actions of genistein, and its in vivo metabolites, are believed to mediate the decreased risk of breast cancer associated with high soy consumption. The genistein metabolite, 5,7,3',4'-tetrahydroxyisoflavone (THIF), induced G2-M cell cycle arrest in T47D tumorigenic breast epithelial cells via a mechanism involving the activation of ataxia telangiectasia and Rad3-related kinase (ATR) via its phosphorylation at Ser428. This activation of ATR appeared to result from THIF-induced increases in intracellular oxidative stress, a depletion of cellular GSH and an increase in DNA strand breakage. THIF treatment also led to an inhibition of cdc2, which was accompanied by the phosphorylation of both p53 (Ser15) and Chk1 (Ser296) and the de-activation of cdc25C phosphatase. We suggest the anti-proliferative actions of THIF may be mediated by initial oxidative DNA damage, activation of ATR and downstream regulation of the p53 and Chk1 pathways leading to cell cycle arrest in G2-M. This may represent one mechanism by which genistein exerts its cellular activity in vivo.

Background: The incidence of cardiovascular diseases increases after menopause, and soy consumption is suggested to inhibit disease development. Objective: The objective was to identify biomarkers of response to a dietary supplementation with an isoflavone extract in postmenopausal women by proteome analysis of peripheral blood mononuclear cells. Design: The study with healthy postmenopausal woman was performed in a placebo-controlled sequential design. Peripheral mononuclear blood cells were collected from 10 volunteers after 8 wk of receiving daily 2 placebo cereal bars and after a subsequent 8 wk of intervention with 2 cereal bars each providing 25 mg of isoflavones. The proteome of the cells was visualized after 2-dimensional gel electrophoresis, and peptide mass fingerprinting served to identify proteins that by the intervention displayed altered protein concentrations. Results: Twenty-nine proteins were identified that showed significantly altered expression in the mononuclear blood cells under the soy-isoflavone intervention, including a variety of proteins involved in an antiinflammatory response. Heat shock protein 70 or a lymphocyte-specific protein phosphatase and proteins that promote increased fibrinolysis, such as a-enolase, were found at increased intensities, whereas those that mediate adhesion, migration, and proliferation of vascular smooth muscle cells, such as galectin-1, were found at reduced intensities after soy extract consumption. Conclusion: Protcome analysis identified in vivo markers that respond to a dietary intervention with isoflavone-enriched soy extract in postmenopausal women. The nature of the proteins identified suggests that soy isoflavones may increase the anti inflammatory response in blood mononuclear cells that might contribute to the atherosclerosis-preventive activities of a soy-rich diet.

Background: Dietary isoflavones are thought to be cardioprotective because of their structural similarity to estrogen. The reduction of concentrations of circulating inflammatory markers by estrogen may be one of the mechanisms by which premenopausal women are protected against cardiovascular disease. Objective: Our aim was to investigate the effects of isolated soy isoflavones on inflammatory biomarkers [von Willebrand factor, intracellular adhesion molecule 1, vascular cell adhesion molecule 1 (VCAM-1), E-selectin, monocyte chemoattractant protein 1, C-reactive protein (CRP), and endothelin 1 concentrations]. Differences with respect to single-nucleotide polymorphisms in selected genes [estrogen receptor alpha (XbaI and PvuII), estrogen receptor beta [ER beta (AluI) and ER beta[cx] (Tsp5091), endothelial nitric oxide synthase (Glu298Asp), apolipoprotein E (Apo E2, E3, and E4), and cholesteryl ester transfer protein (TaqIB)] and equol production were investigated. Design: One hundred seventeen healthy European postmenopausal women participated in this randomized, double-blind, placebo-controlled, crossover dietary intervention trial. Isoflavone-enriched (genistein-to-daidzein ratio of 2:1;50 mg/d) or placebo cereal bars were consumed for 8 wk, with a washout period of 8 wk between the crossover. Plasma inflammatory factors were measured at 0 and 8 wk of each study arm. Results: Isoflavones improved CRP concentrations [odds ratio (95% Cl) for CRP values >1 mg/L for isoflavone compared with placebo: 0.43 (0.27, 0.69)]; no significant effects of isoflavone treatment on other plasma inflammatory markers were observed. No significant differences in the response to isoflavones were observed according to subgroups of equol production. Differences in the VCAM-1 response to isoflavones and to placebo were found with ER beta AluI genotypes. Conclusion: Isoflavones have beneficial effects on CRP concentrations, but not on other inflammatory biomarkers of cardiovascular disease risk in postmenopausal women, and may improve VCAM-1 in an ER beta gene polymorphic subgroup.

Postmenopausal women are at increased risk for cardiovascular disease because many risk factors are aggravated by menopause. Phytoestrogens may modulate risk factors favorably, involving mechanisms similar to estrogen. The effect of phytoestrogens on the atherogenic amino acids homocysteine and asymmetric dimethylarginine (ADMA) was investigated in a controlled intervention study in healthy postmenopausal women. A multicenter, double-blind, crossover intervention trial in 89 postmenopausal women from Denmark, Germany, and the UK was performed. Subjects consumed fruit cereal bars with or without soy isoflavones (50 mg/d) for 8 wk each with an 8-wk washout period in between. Urinary phytoestrogens increased significantly after isoflavone intervention (P < 0.001). Isoflavone supplementation did not affect plasma total homocysteine or ADMA. For homocysteine, changes from baseline were 0.32 micromol/L (range: -0.31-0.92; 95% CI 0.13-0.72), and 0.29 micromol/L (range: -0.45-1.09; 95% CI 0.01-0.63, P = 0.286) for isoflavone treatment and placebo, respectively. For ADMA concentrations, changes from baseline were -0.02 micromol/L (range: -0.08-0.03; 95% CI -0.04-0.01, and 0.00 micromol/L (range: -0.05-0.03; 95% CI -0.03-0.01, P = 0.397) for isoflavone treatment and placebo, respectively. There was no association between plasma total homocysteine and ADMA. Changes from baseline in plasma ADMA and folate were negatively correlated (r = -0.18, P = 0.017). These results challenge the overall health effect of isoflavone supplementation in healthy postmenopausal women.