Resveratrol and Alzheimer’s disease: message in a bottle on red wine and cognition

Cognitive impairment is the final outcome of a complex network of molecular mechanisms ultimately leading to dementia. Despite major efforts aimed at unraveling the molecular determinants of dementia of Alzheimer type (DAT), effective disease-modifying approaches are still missing. An interesting and still largely unexplored avenue is offered by nutraceutical intervention. For instance, robust epidemiological data have suggested that moderate intake of red wine may protect against several age-related pathological conditions (i.e., cardiovascular diseases, diabetes, and cancer) as well as DAT-related cognitive decline. Wine is highly enriched in many polyphenols, including resveratrol. Resveratrol is a well recognized antioxidant which may modulate metal ion deregulation outcomes as well as main features of the Alzheimer’s disease (AD) brain. The review will discuss the potentiality of resveratrol as a neuroprotectant in dementia in relation to the oxidative stress produced by amyloid and metal dysmetabolism.


INTRODUCTION
The so-called French paradox arises from the epidemiological fact that French people, despite their indulgence to a high fat diet, show a relative low incidence of cardiovascular diseases (Renaud and De Lorgeril, 1992). Several epidemiological studies have shown that moderate wine consumption can be effective in slowing down age-related cognitive decline (Wang et al., 2006;Panza et al., 2012;Corona et al., 2013). A possible explanation of this phenomenon has been linked to the national high consumption of wine (20-30 g/day) (Renaud and De Lorgeril, 1992). Albeit moderate ethanol intake is, generally speaking, "beneficial", some more specific effects appear to be related to red wine. Red wine consumption seems in fact to promote far more protective effects than consumption of other ethanol containing beverages (Baur and Sinclair, 2006). Resveratrol, a natural polyphenol, is mainly present in red wine and has been suspected to be the major driving force behind the French paradox (Siemann and Creasy, 1992).
AD is one of the most common forms of dementia in the elderly. To date, no disease-modifying therapies are still available for AD.
In the last 20 years, deregulation of Aβ metabolism (amyloid oligomerization, aggregation, and plaques formation) has been considered the main trigger for AD-related synaptic dysfunction. Amyloid has been therefore the major target for therapeutic intervention (Hardy and Higgins, 1992;Mucke and Selkoe, 2012). Unfortunately, most of these attempts have dramatically failed or have produced only marginal effects (Reitz, 2012;Krstic and Knuesel, 2013;Doody et al., 2014).
Better therapeutic strategies are thus needed along with new acknowledgment that AD is a complex multifactorial syndrome.
Aging is the required paramount condition (Herrup, 2010) on which, in addition to Aβ together with tau deregulation, genes, chronic inflammation, mitochondrial, metabolic dysfunctions, impaired insulin signaling, oxidative stress, aberrant cell cycle reentry, cholesterol dysmetabolism as well as metal ion dyshomeostasis must synergistically work to promote AD pathological manifestation (Herrup, 2010;Querfurth and Laferla, 2010;Roberts et al., 2012). While a single-target therapeutic strategy seems to produce only suboptimal results a broader neuroprotective approach, at least theoretically, appears more appealing (Mudher and Lovestone, 2002).
In this review, we are providing some evidence for resveratrol as a broad-spectrum neuroprotective agent in aging and hopefully in AD.
A recent review on aging determinants has proposed nine hallmarks for the process (López-Otín et al., 2013). Not surprisingly, almost all of them are also involved in AD development and progression (Figure 1; Herrup, 2010;Querfurth and Laferla, 2010) and, notably, at least five, are well recognized target for resveratrol modulation.
In the following sections we have outlined potential effects of resveratrol on these aging and/or AD molecular targets.

β-AMYLOID AND HYPERPHOSPHORYLATED TAU MISFOLDING
Blockade of Aβ deposition into SPs and inhibition of hyperphosphorylation of tau into NFTs has been considered mandatory to prevent or, at least, delay AD-related cognitive decline.
Resveratrol has been shown to inhibit Aβ fibrils formation (Porat et al., 2006;Rivière et al., 2007). Moreover, in vitro and in vivo studies have also indicated that resveratrol reduces amyloid toxicity by decreasing Aβ production through sirtuindependent activation of a disintegrin and metalloproteinase domain-containing protein 10 (Donmez et al., 2010). The compound also increases clearance and metabolism via an AMPactivated protein kinase-pathway and can induce autophagic and lysosomal Aβ degradation (Marambaud et al., 2005;Vingtdeux et al., 2010). Resveratrol can effectively interject in the amyloid cascade through its antioxidant and anti-inflammatory activity, thereby reducing Aβ-driven production of reactive oxygen species (ROS) as well as neuroinflammation (Liu and Bitan, 2012).

CELLULAR METABOLISM
Caloric restriction has been proposed to be effective in increasing lifespan in several animal models. Fasting has been observed to promote beneficial effects on preclinical models of AD and aging not only by extending lifespan but also by ameliorating cognitive performances (Halagappa et al., 2007). Caloric restriction can in fact promote release of brain-derived neurotrophic factor (BDNF), a neurotrophin critically involved in counteracting cognitive decline (Weinstein et al., 2014).
In this context, resveratrol efficiently mimics caloric restriction by inducing expression of SIRT1 (a nicotinamide adenine dinucleotide (NAD + ) dependent deacetylase) which in turn sets in motion a cascade of PGC-1α-dependent events that ultimately lead to improved mitochondrial functioning and biogenesis and boost cellular ROS scavenging (Gomes et al., 2013;López-Otín et al., 2013).

INFLAMMATION
Areas of localized inflammation and active microglia contribute to neurodegeneration and cognitive decline in AD brains Resveratrol, compared to many other disease modifying approaches, shows a broader range of beneficial effects targeting many molecular aspects of AD pathogenesis. Thus, resveratrol may represent a promising broad-spectrum neuroprotective agent in AD. (McGeer and McGeer, 2013). Pharmacological and genetic manipulations aimed at reducing brain inflammation appear to be effective in slowing/modifying the disease progression in AD animal models (Heneka et al., 2013;Giuliani et al., 2014).
Mechanisms by which resveratrol attenuate neuroinflammation are still not completely clear. A major pathway seems to involve sirtuin-dependent arrest of nuclear factor kappa-lightchain-enhancer of activated B cells signaling cascades, a step that results in downstream blockade of microglia activation (Capiralla et al., 2012;Donmez, 2012;Ye et al., 2013).

MITOCHONDRIAL DYSFUNCTION AND ROS
Mitochondria play an essential role in the cell wellbeing. The organelles critically control cellular energy and metabolism as well as intracellular signaling (Rizzuto et al., 2012). On the dark side, mitochondria are also key players in modulating cellular death through release of apoptotic factors, blockade of energy supply and generation and release of ROS. Alterations of mitochondrial functioning are known in aging and early stages of AD (Wang et al., 2013).
Mitochondrial electron leakage, followed by ROS production occurs in neurodegenerative conditions paving the way to lipid peroxidation, nucleic acid damage, protein oxidation, and, eventually, neuronal death (Wang et al., 2013).

TELOMERES SHORTENING
Telomeres shortening plays a key role in cellular aging and AD (Cai et al., 2013;Mathur et al., 2014). Short telomeres increase DNA vulnerability to stressful insults (i.e., UV irradiation, ROS production) ultimately leading to aberrant cell functioning and cell death. Polyphenols has a positive impact upon maintenance of telomeres length (Jayasena et al., 2013). In that respect, resveratrol promotes the expression of Werner syndrome ATP-dependent helicase, a telomere maintenance factor (Uchiumi et al., 2011), increases the activity of telomerase via a SIRT1-dependent pathway (Palacios et al., 2010), and spares telomeres and DNA from ROS dependent damages thanks to its intrinsic scavenging properties (Jayasena et al., 2013).

METAL IMBALANCE IN THE AD BRAIN: A POTENT TRIGGER OF OXIDATIVE STRESS
In the brain, metal ions are involved in many essential processes such as intracellular signaling, modulation of cellular redox and metabolic states, enzymatic activities and channels functioning (Billard, 2006;Sensi et al., 2009;Rizzuto et al., 2012;Sekler and Silverman, 2012;Gaier et al., 2013). Metal homeostasis is strictly controlled by the interplay of transporters, channels, chaperones and metalloregulatory sensors (Finney and O'halloran, 2003). In neurodegenerative conditions and/or aging, this tightly controlled system is lost, thereby leading to disease-promoting metal imbalance Breydo and Uversky, 2011;Jellinger, 2013).
Metal ion dyshomeostasis is in fact involved in several neurological disorders like Parkinson's disease (PD), Amyotrophic Lateral Sclerosis (ALS), Prion Protein disease, Huntington's disease (HD), and AD. All these neurodegenerative conditions share common pathological features that include deposition of misfolded proteins, metal ion deregulation and exposure to oxidative stress (Boillee et al., 2006;Duce and Bush, 2010;Roberts et al., 2012;Gonzalez-Dominguez et al., 2014).
In AD, metal ion dyshomeostasis represents a key, though too often overlooked, pathological step. A metal hypothesis for AD has been proposed by many authors (Bush, 2008). In that respect, copper, iron, zinc and aluminum are the metals found deregulated in AD. All of them are able to alter Aβ metabolism and deposition . SPs but also NFTs are highly enriched of these metals. Moreover, all these ions can promote ROS generation (Sayre et al., 2000;Pithadia and Lim, 2012;Ayton et al., 2013).
Recent findings have shown that low levels of copper are sufficient to dramatically affect Aβ homeostasis by increasing Aβ accumulation and neuroinflammation related to Aβ-deposition (Singh et al., 2013). Compared to nondemented elderly controls, brains of AD patients show an increased presence of labile copper pools, which correlate with oxidative damage in these tissues (James et al., 2012). Resveratrol is a well known copper chelator (Tamboli et al., 2011) and, in theory, of some use in AD (Faux et al., 2010). Unfortunately, the copper-resveratrol complex seems to be more harmful than beneficial in the context of AD. Resveratrol promotes the reduction of copper (II) to copper (I) (de la Lastra and Villegas, 2007) and several studies have indicated a pro-oxidant activity of the compound when bound to copper (Zheng et al., 2006;de la Lastra and Villegas, 2007;Muqbil et al., 2012). Thus, resveratrol activity on copper homeostasis appears more harmful than neuroprotective if used as standalone therapeutic approach. A feasible and, in our opinion, clinically relevant approach might be represented by the administration of resveratrol in association with a higher affinity copper chelator. This would lead, at least in theory, to a dual beneficial effect: reduction of copper dyshomeostasis coupled with decreased ROS production.
Iron deregulation has been linked to AD (Weinreb et al., 2013;Crespo et al., 2014;Gonzalez-Dominguez et al., 2014). Role of iron in AD pathogenesis is substantiated by the effectiveness of metal homeostatic therapies aiming at reducing iron deregulation (Crouch et al., 2007), which results in (1) decreased free iron accumulation and ferroptosis (Dixon et al., 2012); (2) decreased iron-dependent ROS production; and (3) blockade of neurotoxic Aβ-iron conjugates formation . To date, in vivo evidence for iron chelation by resveratrol is missing, however the compound prevents iron-driven mitochondrial dysfunction by inhibiting glycogen synthase kinase-3 beta activity (a mechanism useful also to prevent tau hyperphosphorylation) (Shin et al., 2009), and by reducing peroxidation of lipoproteins and lipids through its activity as scavenger (Belguendouz et al., 1997;Tadolini et al., 2000).
Zinc dyshomeostasis has been proposed as a risk factor for AD. Accumulation of excessive zinc, or its deficiency, are both involved in the neuronal loss which leads to AD and aging related cognitive decline (Brewer, 2012). While zinc deficiency increases neuroinflammation and also affects BDNF maturation and ultimately cognition, aberrant intracellular zinc mobilization or accumulation leads to mitochondrial failure and ROS production. Extracellular zinc overload within SPs also inhibits the iron-export ferroxidase activity further increasing ROS production and ultimately neuronal death . Resveratrol does not directly affect zinc levels however it can be useful in preventing the full development of zincdependent injurious mechanisms. Actually, resveratrol inability to sequester zinc does not represent a limitation, as the compound can exert antioxidant activities without producing zinc deficiency.
Aluminum lacks modulatory functions in biological processes; however, its accumulation in the brain has been demonstrated to be linked to several neuropathological conditions (Zatta et al., 2003;Walton, 2013). To date three are the main mechanisms through which aluminum exerts its neurotoxic effects: (1) production of ROS; (2) induction of neuroinflammation; and (3) formation of toxic aggregates of misfolded proteins (Perl, 2006;Kumar et al., 2009;Wu et al., 2012;Bolognin et al., 2013). In AD, aluminum seems to act as an effective cross-linker between tau phospho-sites, to "freeze" Aβ in its toxic oligomeric state, and to induce exposure of Aβ hydrophobic clusters aggregates, thereby boosting toxic properties of these misfolded proteins Bolognin et al., 2011;Chen et al., 2011;. Aluminum-related oxidative damage occurs through lipid peroxidation, alteration of the activity of antioxidant enzymes, alterations of mitochondrial functioning and biogenesis and promotion of DNA injury (Zatta et al., 2002;Sharma et al., 2013). Resveratrol shows a negligible ability to bind aluminum in vitro Frontiers in Aging Neuroscience www.frontiersin.org May 2014 | Volume 6 | Article 95 | 3 FIGURE 2 | Metal ions dyshomeostasis is closely related to different hallmarks of AD pathogenesis, mainly protein misfolding, ROS production, mitochondrial failure and inflammation. In this figure the mechanisms through which resveratrol exert its neuroprotective role against selected metal ions are reported. Of note, most of the resveratrol beneficial effects against metal ion dyshomeostasis belong to its scavenging properties. , nevertheless, it seems effective in reducing in vivo the downstream events of aluminum overload, namely the aluminum-related ROS production and neuroinflammatory response activation (Zaky et al., 2013).

CONCLUSIONS
Resveratrol is a multi target compound and may represent an effective therapeutic tool in aging-related neurodegenerative processes. Consistently, several clinical trials are ongoing to test its effectiveness as dietary supplement to slow dementia progression (ClinicalTrials.gov, 2014). In summary, major effects are associated with its scavenging activity as well as in the activation of SIRT1 (see Bordone and Guarente, 2005;Herskovits and Guarente, 2014 for extensive reviews on the topic). The presence of non-SIRT1 neuronal targets of resveratrol is debated, suggesting that resveratrol in vivo may act on other uninvestigated biological targets (Herskovits and Guarente, 2014). The complementary role of modulator of metal dependent oxidative injury (Figure 2) represents a still largely unexplored field in resveratrol biochemistry.
Resveratrol is a multi-target, simple, safe, and cost-effective dietary supplement. Nevertheless, it should be reminded that its role as therapeutic agent is not devoid of potential problems. The pro-oxidant activity in presence of labile copper, the poor bioavailability and ease degradation all represent major issue that require new sophisticated efforts (Goldberg et al., 2003). Synthesis of novel resveratrol analogs is ongoing and improvement of drug delivery might represent in that regard the major targets to be considered in order to overcome current resveratrol limitations. In agreement, pterostilbene, a resveratrol derivative, has shown promise in preclinical models of neurodegeneration, resulting more efficient than resveratrol itself in modifying AD-and aging-related cognitive decline (Joseph et al., 2008;Chang et al., 2012). These results leave the door open for the use of newly synthesized resveratrol analogs in aging-related disorders (Bourzac, 2012;Ogas et al., 2013;Pezzuto et al., 2013).

ACKNOWLEDGMENTS
Authors are grateful to Prof. Stefano L. Sensi for fruitful discussions during manuscript preparation. Alberto Granzotto is supported by "Marisa Trampuz von Langendorf " Fellowship for Neuroscience.