Resveratrol

Resveratrol is a polyphenolic compound present in grapes, and is most well known for its presence in red wine. Interestingly, the popularity of resveratrol is due to its discovery in red wine and subsequent hypothesizing that it may be able to explain the 'French paradox' of Heart Disease.^[15] However, resveratrol only contributes slightly to the french paradox, as alcohol (from red wine), diet and lifestyle are also significant factors.^[16][17][18]

It is estimated to have an average intake of 0.2mg daily in a Spanish population,^[19] mostly (98.4%) from wine. North Americans tend to have minute levels in the diet.

Interestingly, it is classified as a phytoalexin (plant toxin). Its reason for existence is to protect grapes from infection, as it is synthesized in response to Bothrytis cinerea infection.^[20][21]

Resveratrol is a polyphenolic compound referred to as a 'stilbene' due to its structure, and it is the most common and well researched stilbene currently known (despite stilbene's per se being a class of molecules). Stilbenes tend to be most well known for occurring in the Vitis family (Grape family) although they extend to many plants

Food products or other common consumables that have a resveratrol content include:

Note: 1umol of Resveratrol is equal to approximately 0.23mg

• The Vitis family of plants,^[22][23] which is usually Wine (0.005-2.861umol/L;^[24] highest in Red Bordeaux) and Grapes (27-35mcg/g trans-resveratrol with 160-460ng/g cis-resveratrol^[25])
• Raspberry, at 38-59ng/g trans-resveratrol ^[25]
• Plums, at 13-20ng/g trans-resveratrol ^[25]
• Grape Tomatoes, at 168-175ng/g trans-resveratrol ^[25]
• Piper Imperiale^[26]
• Açai berries^[27]
• Peanuts and peanut products^[28] at possibly as high as 5mcg/g in boiled peanuts, 0.3mcg/g in peanut butter and 0.05mcg/g in roasted peanuts.

With common nutritional supplements that have a resveratrol content including:

• Japanese Knotweed (Polypodium Cuspidatum), a source of resveratrol and stilbenes in Traditional Chinese Medicine and economical source of resveratrol for industries. Tends to be a common source of supplemental resveratrol
• Grape Seed Extract (about 0.53% dry weight of GSE)
• Cissus Quadrangularis
• Morus Alba (0.2358mg/g of the ethanolic extract of the branch, about 5mg/g of Oxyresveratrol)

Resveratrol can exist in one of two isomers: trans-resveratrol and cis-resveratrol. The configuration greatly changes the structure:

Trans-resveratrol is commonly seen as the active form of resveratrol. As the simple change results in a largely different molecule, many actions seen from trans-resveratrol are not seen with cis-resveratrol. These actions include modulation of the inflammation response,^[29] and a more potent anti-proliferative effect on cancer cells.^[30]

It should be noted that the cis isomer is still bioactive, but most research is focused on trans-resveratrol. Cis-resveratrol is still an anti-oxidant^[31] and can interact with genetic transcription.^[32]

When implemented into a gel base, trans-resveratrol stored at 4C does not convert into its cis isomer over a period of 30 days, suggesting possible usage as a topical ingredient.^[33] Resveratrol is also poorly soluble in water^[34] but many studies that use supplemental resveratrol on an empty stomach note that it can be absorbed without fatty acids.

Sirtuins are a class of 7 protein messengers in mammalian cells with a myriad of effects.^[35] Specifically, they are NAD-dependent deacetylases; they deacetylase a wide variety of other compounds, and they use NAD (Nicotinamide Dinucleatide) to do this.^[36] The enzyme (SIRT1) is induced (activated) in periods of fasting and is inhibited during high nicotinamide concentrations (correlated with feeding)^[37] and, as prior research shows a link between SIRT1 and resveratrol, the above activation pattern of SIRT1 and caloric intake is the most famous hypothesized 'link' between longevity (via caloric restriction) and resveratrol.

The entire sirtuin family is highly involved with regulation of mitochondria^[38] and energy metabolism,^[39] and is a player in musclular energy metabolism in response to exercise.^[40]

The sirtuin system is a group of cytoplasmic and mitochondrial proteins that are involved in energy restriction and metabolism and thought to be related to aging, with activation of this system thought to promote longevity

Resveratrol has been noted to exert many actions in a manner that is dependent on SIRT1^[41][42][43] and it was initially hypothesized that resveratrol allosterically modified SIRT1 to increase its affinity for substrate and NAD+ (directly increasing its activity^[44]), although this is currently thought to not be the case since the fluorescent used in past research (Fluor de Lys-SIRT1 peptide) may have been a research artefact (unintended mistake);^[45][46][47] while a direct interaction isn't fully ruled out yet^[48] it seems more likely that the influence on SIRT1 is indirect.

It seems unlikely that resveratrol directly activates SIRT1, although an activation of SIRT1 does appear to exist with resveratrol and it is currently thought to be downstream of directly influencing other molecular targets

Resveratrol may act on SIRT1 secondary to acting on AMPK in various tissues^{[49][50][51][52]} which is thought to be due to AMPK increasing levels of NAD+ in a cell, which is the cofactor for SIRT1.^[53] The opposite also holds true as abolishing SIRT1 activity prevents resveratrol from acting on AMPK,^[54] and SIRT1 has the ability to deacetylate the AMPK kinase known as LKB1 which would increase AMPK activity^[55][56] while abolishing SIRT1 prevents resveratrol from acting on AMPK.^[54]

It was noted that AMPK was activated in a SIRT1 dependent manner at lower concentrations (25μM) but independent at higher concentrations (50μM) in C2C12 cells,^[54] there is a possible pathway towards AMPK via phosphodiesterase inhibition since resveratrol inhibits PDE4 and PDE3 causing an increase in cAMP levels in the cell; increase cAMP will cause Epac1 to release calcium from the sarcoplasmic reticulum, and said calcium increases AMPK activity via the CamKKβ-AMPK pathway;^[57] this was validated when Rolipram (selective PDE4 inhibitor) mimicked the effects of resveratrol.^[57] Since the influence of SIRT1 on AMPK (deacetylation of LKB1) is that of a positive modulator, it is possible that PDE inhibition explains the direct AMPK activation while the lower concentrations indicate an alterate mechanism of resveratrol on SIRT1, which facilitates normally subactive concentrations of resveratrol on AMPK to then become active.

Regardless of the exact mechanisms, AMPK and SIRT1 both beneficially influence mitochondrial biogenesis^[58][59] as since both SIRT1^[60][61] and AMPK^[62] promote the activity of PGC1α, which then promotes the transcription of proteins involved in mitochondrial biogenesis.^[63] This pathway, however, has been noted to be primarily AMPK directly but dependent on SIRT1 (suggesting a positive modulator role).^[59]

There is ultimately one shared result of activate SIRT1 and AMPK in a cell (increase in mitochondrial biogenesis secondary to PGC1α activatioN) although there appear to be two distinct pathways involved; it seems that resveratrol can directly activate AMPK via PDE inhibition at higher concentrations, and another influence on SIRT1 may positively regulate the AMPK pathway allowing it to occur at lower concentrations

Resveratrol has a good absorption but low bioavailability when orally administered to man, as evidence by one study noting an oral dose of 25mg resulting in less than 5ug/mL in the serum while 0.2mg (125-fold lower) dose injected into serum resulted in levels of 16.4-30.7 ng/mL.^[64] This is due to rapid conjugation via sulphation and glucuronidation (P450 enzymes) that reduce the amount of free resveratrol.^[65]

There appears to be variation between individuals between dosages, which is less of a concern with low dosages and more of a concern with superloading. At an oral dose of 25mg there is merely a 2-fold difference in maximal differences^[64] whereas at 5g a range of 52-2834 ng/g when measuring liver levels of resveratrol, and plasma levels varying from 800-5000ng/mL, exists.^[66]

Interestingly, there is a circadian rhythm with resveratrol absorption. It appears the concentration of resveratrol in the blood is more dose-efficient (more bioavailable) in the morning relative to the PM,^[67] which may be due to diurnal variations in one of the main system that metabolizes free resveratrol, P450 glucuronidation.^[68] Additionally, enterohepatic circulation, which resveratrol is subject to^[69] decreases in the morning.^[70]

When consumed with a balanced meal, the overall bioavailability of Resveratrol does not change. However, the time until peak levels in the blood (T_max) and the peak levels (C_max) are delayed and reduced, respectively. Overall exposure (AUC) does not change with the balanced meal,^[71] but may in response to a higher-fat meal (45g relative to 15g in the balanced meal).^[72]

Its well absorbed in the intestines, and food does not affect its overall absorption. Bioavailability is another issue though, as conjugation begins at this stage.

When ingested orally, resveratrol is taken up from the intestines to the liver (like any xenobiotic). It can be sulphated at either location^[73] and it along with glucuronide conjugates account for up to 90% of serum resveratrol levels (the other 10% being free resveratrol)^[74][75][76]. Coingestion with bioflavonoids causing competitive inhibition that may free up Resveratrol by having the other bioflavonoids act in a sacrificial manner,^[77] specifically Quercetin.^[78][65] However, at least one study using 500mg Quercetin and 2000mg trans-resveratrol noted no differences in resveratrol pharmacokinetics after co-ingestion of quercetin.

Dose dependence can be seen with capsules, with C_max values of around 73ng/mL (100mg oral dose) 147ng/mL (1g oral) 268ng/mL (2.5g) and 534ng/mL (5g).^[79] The larger doses gradually shifted the T_max backwards towards 1.5 hours from 1 hour.^[79] These dosages are relatively high, but lower have been studied; C_max values have been noted at 1.48–3.83ng/mL (25mg) 6.59–7.39ng/mL (50mg) 21.4–23.1ng/mL (100mg) and 24.8–63.8ng/mL (150mg).^[67] T_max is harder to get from this study due to multiple dosing throughout the day.

In human trials with only trans-resveratrol, doses of 500mg resulted in a 24hour mean plasma concentration of 8.36ug/L and higher doses of 5g resulted in a 24 hour mean plasma concentration of 51.9ug/L; the C_max was 72.6 μg/L at 50 minutes and 538.8 μg/L at 90 minutes, respectively.^[79] These numbers are approximations, as there appears to be large inter-individual differences in the correlation between oral dose and blood levels.^[80] The C_max appears to follow loading parameters, and repeated dosages appear to induce subsequently higher levels of blood resveratrol.^[81][82]

Micronized Resveratrol (SRT501) with particulate sizes of less than 5μm, the C_max was found to be 8.51nmoL/L (1942 ng/mL) at 2.8 hours after oral ingestion.^[66] A high inter-individual variation exists with this study noting a range of range 52-2834 ng/g when measuring liver levels of resveratrol, and plasma levels varying from 800-5000ng/mL.^[66] These numbers appear to suggest that micronized resveratrol has a 3.6-fold increase in bioavailability^[66] when compared to previous studies with non-micronized.^[79]

Resveratrol's absorption is relatively the same in ethanol as it is in water^[83] and, as stated earlier, other bioflavonoids in wine act to increase the bioavailability of Resveratrol.^[77] Thus, there is some synergism in absorption when consumed in the format of wine but this is not due to the alcohol content.

Consumption of 300mL of white wine can increase plasma levels in the range of 0.72±0.3 to 1.33±0.3umol/L and red wine in the range of 0.71±0.2 to 1.72±0.1 μmol/L.^[84] As compiled in this review^[65] other studies have been conducted on the pharmacokinetics of resveratrol after wine consumption. Three studies looked at 25mg of resveratrol consumption in the format of wine^[85][83][86] and found, on average, a C_max of 1.5-8ug/L (more often at the higher end) at 30 minutes after ingestion.

These numbers should be taken with a grain of salt, as in addition to the high inter-individual variation trans-resveratrol also varies in content due to growing conditions for grapes.^[87]

There appears to be no difference between the resveratrol in food and wine compared to capsule form. However, resveratrol has low bioavailability anyways at around 10% of ingested dose being bioactive. Food can have an advantage due to ingestion of compounds that increase bioavailability (like Quercetin) but capsules can be micronized to increased bioavailability 3.6-fold. Additionally, it would be hard to get very large dosages through wine without saying farewell to your liver (in regards to Alcohol)

Resveratrol appears to be readily taken up into cells, as evidenced by rapid serum depletion of resveratrol when injected into human subjects^[64] and a robust accumulation into isolated cells (reponsive to resveratrol) within 10 minutes (the rate becoming more steady after 10 minutes).^[88] The subcellular distribution of resveratrol after uptake appears to favor the membrane/organelle, with less concentration in the nucleus and less in the cytoskeleton and cytosol (the latter being equivalent).^[88]

To travel through the blood, it may bind itself to albumin due to its structure^[89] and incubation of free resveratrol with albumin in vitro causes a decrease in free resveratrol, from binding.^[90] This binding affinity to albumin has been noted in other studies,^[91] and appears to be enhanced when in the presence of fatty acids;^[90] most likely due to structural changes of albumin when bound to fatty acids.^[92]

Studies on HepG2 cells (liver cells) note that cellular uptake is mediated in part by passive diffusion and by carrier-mediated processes^[93]

It has been quantified in vivo in gerbil brains^[94] and human liver tissue^[66] as well, demonstrating that it can diffuse into a cell. It has also been found in colon, lung, and heart tissue after oral administration.^[95][96]

After ingestion of resveratrol, it can be conjugated by liver P450 enzymes. The results are resveratrol sulphate (via sulphation) and two glucuronides, resveratrol-O-glucuronide and resveratrol-C-glucuronide.^[65]

Resveratrol has anti-oxidant properties, and when it sequesters oxidants (typically hydroxyl groups via dismutation) it can turn into one of four metabolites: piceatannol (PCT), 3,5-dihydroxybenzoic acid (3,5-DHBA), 3,5-dihydroxybenzaldehyde (3,5-DHB) and para-hydroxybenzaldehyde (PHB).^[97]

Resveratrol is excreted in both the urine and the feces.

Half-life of the resveratrol molecule appears to be in the range of 1-3 hours, and can be extended slightly (2-5 hours) with multiple doses.^[67]

Elimination half-life is estimated at being around 7-14 hours.^[64]

In the end, resveratrol is readily absorbed and distributed to the body. It is mostly excreted within a day, and has a half-life of a few (1-3) hours. Blood levels are relatively dose-dependent, and the 'peak' level of blood concentration is slowly pushed backwards (up to 90 minutes, from 30) with the more resveratrol you consume. It can get into cells as well, when should preclude its metabolic actions. Its only real downfall is that it is heavily conjugated by the liver (P450)

In Drosophilia and C.Elegans (two research creatures), the mechanism of longevity appears to be activation of Sirt2; a NAD+ dependent histone deacetylase protein.^[98] In Drosophilia, there appears to be interaction with both gender and diet with females on low-carbohydrate diets having benefit at the lowest dose resveratrol.^[99]

As mentioned in the section on Sirtuins, these beneficially influences on the Sirtuin system may be vicariously through activation of AMPK.

Downregulation of p53 expression in neurons has also been shown to dose-dependently be associated with increasing lifespan in Drosophilia,^[100][101] which is hypothesized to be downstream of the junction of interest.^[102]

In a drosophilia genomic-wide analysis,^[102] cross-referencing caloric restriction, Sirt2 long-living flies and p53 long-living flies, there came to be 21 genes that were shared between the three long-living creatures. These genes include takeout (expression related to food intake), which is related to Juvenile Hormone (insect-exclusive) and life-extension.^[102][103] Of interest to humans is that the 21 overlapping genes had "relationships to chromatin structure, circadian rhythm, neural activity, detoxification/chaperone activity, muscle maintenance, immune function, growth factor activity and feeding behavior/response to starvation".^[102]

Hypothesized mechanisms in mammals are various and were outlined in the Sirtuin section on 'Direct/Indirect influences'; less promosing than Drosophilia and C.Elegans mechanisms.

It appears that resveratrol reliably increases lifespan in these two non-mammalian models,^{[104][104][102][105][98]} which may be related to acting on Juvenile Hormone; a longevity promoting yet insect exclusive hormone.^[102][103]

In insect models (commonly used in longevity research as they have short lifespans and a study spanning an insects lifetime is actually practical to conduct and publish), resveratrol may promote lifespan by a mechanism that does not exist in humans

In mammalian models of premature aging, resveratrol does not seem to be indicative of enhanced lifespan per se.^{[106][107][108][98]} However, resveratrol is effective in inhibiting or reversing salient effects of aging (osteoporosis, sarcopenia, cognitive decline, etc.) and may give the illlusion of longer life or otherwise give life to your years rather than add years to your life.^[76] In rats, these beneficial effects (parameters of aging) have been treated with doses as low as 4.9mg/kg bodyweight a day.^[109]

It should be noted that this is not the consensus, and some studies do note increase longevity in lab animals.^[110] However, it is currently up for debate whether the effects seen are due to a legitimate life extension mechanism or whether they protect from causes of death acutely (such as heart attacks) and push the median lifespan past statistical significance.

At this moment in time, it appears that resveratrol 'adds life to years' rather than 'adds years to life'. It may protect against common causes of death (covered in heart health and cancer metabolism) as well as metabolic syndrome, which could then push the median lifespan higher and exert a 'pseudo-life extension' appearance; however, a novel life extension mechanism independent of lifestyle that can be attributed to resveratrol appears to be lacking.

Resveratrol is able to cross the mammalian blood brain barrier, and incorporate itself into brain tissue.^[94]

Resveratrol, at doses between 250-500mg of the trans-resveratrol isomer, have been noted to increase cerebral blood flow in healthy human subects.^[111] This was not accompanied by an increase in cognitive performance.

Resveratrol may suppress glutamate release from neurons^[112] although other evidence suggests that this does not occur.^[113] Regardless of its effects presynaptically, resveratrol appears to attenuate glutamatergic activation of activated neurons^{[112][113][114]} with an IC₅₀ of 53.3+/-9.4µM which differntially affected glutamate receptors. 100µM of resveratrol inhibited AMPA currents (19.4+/-6.3%) less than NMDA (49.8+/-8.9%) and kainate (74.1+/-4.5%) with this inhibition was reversible.^[113] Although it is not known why this occurs, it is thought that REDOX modulation may occur similar to pyrroloquinoline quinone (since it affects the NMDA but not AMPA receptors^[115][116]) or that the known inhibition of L-type calcium channels.^[117]

Other studies have noted that glutamate-induced changes (associated with excitotoxicity) are attenuated with resveratrol,^[118][119] lending support for the suppressive effects on glutamatergic signalling.

At least one study noted that neuroprotection from resveratrol was abolished with an NMDA receptor antagonist,^[120] which the authors thought reflected a preconditioning effect (as despite NMDA overactivation contributing to toxicity^[121] a low level of activation preconditions the neurons and is neuroprotective^[122]).

There are unclear effects of resveratrol on the glutaminergic receptors. While in general it appears to suppress overactivation of glutaminergic signalling (all three major receptor subsets), some neuroprotective effects appear to be prevented by blocking the receptor. It is possible that resveratrol is a weak agonist of sorts while outcompeting stronger agonists

In regards to seizures induced by kainic acid (acting via kainate receptors, one of the glutamate receptor isoforms), resveratrol appears to be able to reduce seizures and hippocampal neurotoxicity^{[123][124][125][126]} although it appears ineffective in younger still developing rats^[127] who are known to be more sensitive to kainate-induced seizures.^[128]

May suppress the signalling through kainate receptors, and reduces the seizure potential of kainic acid. This appears to be well replicated in rat models, although it failed in the one more sensitive to kainic acid

In morphine tolerant mice (experiencing upregulations in NR1 and NR2B subunits of NMDA receptors which contributes to morphine tolerance^[129][130]), resveratrol administration into the brain (7.5-30µg) appears to downregulate these two subunits via preventing an increase in PSD-95 expression with no inhernet suppressive effect without morphine.^[131] PSD-95 provides a physical anchorage for NMDA receptors at the membrane^[132] and this was thought to mediate the suppression in NMDA receptor increases.^[131]

With agents that predispose the neurons to excitotoxicity secondary to upregulating NMDA receptors, resveratrol has been implicated in preventing this upregulation. Although the mechanism is not known, it appears to be through preventing a scaffolding protein from supporting NMDA receptors in the membrane

Glial cells are cells used to support neurons, and are highly involved in neurological systems being associated with their own form of plasticity and metabolic coupling^[133] and influencing synaptic function;^[134][135] some nutritional supplements such as D-Serine are also known as gliotransmitters as they are borne from astroglia, and glial cells are involved with neurons (presynaptically and postsynaptically) in a sort of tripartite synapse.^[136]

Astrocytes are also involved in glutamatergic neurotransmission as they can convert glutamate into glutamine (via glutamine synthetase)^[137] and the released glutamine is taken up by neurons for converion into glutamate via a glutamate-glutamine cycle^[138][139] to then be released into the synapse.

Glial cells, particularly the astrocytes, are intimately involved in neurotransmission between neurons and are further implicated in glutaminergic neurotransmission

It is hypothesized^[140] that resveratrol acts on glial cells as it has been noted to increase glutamate uptake into glial cells in the dosage range of 0.1-250μM^[141] which also appears to increase glutathione content of these cells.^[142] Resveratrol has also been noted to protect astroglia from neurotoxicity from ammonia^[143] which is used in the process of converting glutamate into glutamine via glutamine synthetase. It seems that under periods of ammonia toxicity that this enzyme is downregulated due to increased oxidative stress^{[144][145][146]} and as such the upregulation of this enzyme seen with resveratrol^[141] may underlie the protective effects.^[143]

Resveratrol appears to increase glial cell uptake of glutamate, and encourage its production into glutamine. This mechanisms can be used to explain neuroprotection from ammonia, and it may also contribute to the observed anti-glutaminergic effects of resveratrol

Resveratrol has also been shown in some studies to be synergistic with Melatonin supplementation in preventing beta-amyloid induced neurotoxicity^[147] and potentiates Resveratrol's induction of the anti-oxidant enzyme, heme-oxygenase 1, which is linked to neuroprotection.^[148]

Additionally, dosages of 30mg/kg bodyweight in gerbils (RP injection) have been linked to significant protective effects following ischemia, protecting neurons from delayed cell death.^[94]

In age-accelerated mice (SAMP8), it appears that lifelong supplementation of resveratrol may increase lifespan and delay biomarkers of Alzheimer's (beta-amyloid and tau protein aggregation).^[149]

When investigating SIRT1 levels of the endothelium of persons with coronary artery bypass, they were found to be expressed at lower levels in artherosclerotic arteries relative to normal vessels (about 60% of normal level for tissue, 20% for the endothelium itself).^[150]

The typical target of resveratrol, SIRT1, appears to be reduced in artherosclerotic arteries

In regards to oxidized LDL (oLDL; a form of LDL with is seen as more artherogenic), resveratrol is able to attenuate the rate of oLDL formation form LDL secondary to its direct anti-oxidative effects against both metal ions and hydrogen peroxide.^{[151][152][34]}

When looking at the endothelial nitric oxide synthase enzyme (eNOS), resveratrol can upregulate eNOS mRNA in isolated endothelial cells (HUVEC and EA.hy 926) at 1-100μM associated with increased activity of the promoter (2-fold at 10μM) secondary to increasing its stability and half-life;^[153] the effect was not associated with the estrogen receptors,^[153] which is a trait of estrogen receptor activation.^[154]

Uncoupling of eNOS (which is caused by deficiency of the tetrahydrobiopterin subunit aka. BH₄^[155][156]) is when the eNOS enzyme no longer couples exclusively to its substrate L-arginine and can begin producing superoxide (O₂^-) radicals;^[157][156] such uncoupling can be treated by either NADPH oxidase inhibition^[158] or via increasing BH₄ synthesis. Resveratrol possesses both of these properties, being able to reduce the expression of the NOX4 subunit of NADPH oxidase^[159] and able to increase the activity of GTP cyclohydrolase 1 (GCH1), the rate limiting enzyme of BH₄ synthesis (30-100mg/kg resveratrol in mice).^[160]

Resveratrol appears to stabilize the mRNA which signals the genome to produce the eNOS enzyme (thereby increasing overall production of eNOS), which is not related to the estrogenic effects of resveratrol. There is also a recoupling effect on eNOS (by mitigating some of the pathological changes seen in unhealthy states)

Resveratrol is being investigated for its benefits to the endothelium as it is a direct free radical scavenger (specifically ROS) at 10-100µM^[150][161] may inhibit NADPH,^[162] and at least in vitro appears to induce eNOS;^[163] these effects of resveratrol suggest it can be beneficial in preserving endothelial responsiveness to agents that induce vasorelaxation (such as nitric oxide or acetylcholine), which are commonly hindered by alterations of the above three mechanisms.^[164][165]

Resveratrol is thought to preserve the response of the endothelium to endogenous (occurring within the body) agents that induce relaxation. Relaxation of the vessel wall by these agents tends to be impaired in chronic disease, and resveratrol may reverse or attenuate this impairment

28 days of resveratrol supplementation in spontaneously hypertensive rats at 0.448-4.48mg/L (the lower dose to mimic moderate wine consumption; 0.05-0.5mg/kg in rats and equivalent to 3.3-33mg resveratrol in humans) was able to increase acetylcholine-induced maximal vasorelaxation from 60.7+/-1.4% in control to 80.8% (no dose dependence noted) without influencing KCl or phenylephrine induced contraction nor the EC₅₀ of acetylcholine induced contraction.^[166] Enhancement of vasorelaxation has been noted elsewhere with a higher dose of 5mg/kg to a similar degree of efficacy.^[167]

One study in spontaneously hypertensive rats noted that endothelial nitric oxide synthase (eNOS) was not significantly altered despite enhanced acetylcholine responsiveness of the endothelium,^[166] although in vitro studies note increases in eNOS at physiologically relevant concentrations.^{[153][163][168]} This was thought to explain enhanced endothelial-dependent vasodilation noted in rats given 5mg/kg resveratrol (a supplemental dose),^[167] and noted that it can occur at lower doses occurring in wine (human equivalent of 3.3mg) to a similar degree as higher doses.^[166]

Appears to enhance acetylcholine induced relaxation of the arteries in rats with hypertension, which is thought to be related to the endothelial-dependent vasorelaxation noted in rat models; this may be independent of changes in blood pressure.

Several rat studies using resveratrol that note benefits to the endothelium and vasorelaxation have noted that reductions in resting blood pressure may not occur.^[166]

Resveratrol and related grape phenolics are investigated for their effects on blood pressure since dealcoholized wine has been demonstrated to reduce blood pressure in persons at risk for heart disease.^[169]

Mechanistically, the protein quinone reductase NQO2 appears to have remarkably high affinity for resveratrol with a K_M of less than 50nM;^[170][171] complexing can be read here.^[172] This protein does not appear to be highly expressed in the aorta of rats, but is highly expressed in the heart tissue (as well as liver and kidneys) and appears to decline during the rat aging process.^[88]

Resveratrol has been investigated for its contribution to heart health after a meta-analysis first found a significant risk reduction associated with 1-2 glasses (150-300mL) of wine daily.^[173] There was a hormetic curve (J-curve) with the peak of preventative actions at 300mL resulting in a Risk Ratio of 0.61 (approximately 61% of the risk of vascular health complications in those consuming 150-300mL wine daily); this prompted research into both resveratrol and alcohol and heart health.^[173]

Resveratrol, per se, has been shown to increase insulin sensitivity when supplemented obese persons at 150mg daily as measured by HOMA index and measured at 13.3% improvement over 30 days alongside reductions in glucose (4.2%) and insulin (13.7% reduction).^[174] It has been suggested to do most of its mechanisms of action on the level of the cell, as it increases Akt phosphorylation (Type II diabetics at 5-10mg daily)^[175] and activates AMPK (150mg in non-diabetic obese persons).^[174] This mechanism of insulin sensitization, AMPK, is not activated in healthy non-obese persons nor are there apparent benefits to insulin sensitivity of muscle, fat, or the liver in this population.^[176] This study, however, used 75mg, a dose higher than the one associated with insulin sensitization in type II diabetics^[175] and half of that which activated AMPK in healthy obese adults.^[174] Additionally, these benefits appear to fade after 6 months cessation of supplementation.^[177]

Resveratrol appears to benefit glucose metabolism, with lower doses needed for those in worse metabolic condition (insulin resistant, diabetic) and higher doses needed for those in pre-clinical disease states; may not be effective in healthy persons at increasing insulin sensitivity, and appears to exert temporary benefit

On the level of the pancreas, resveratrol can reduce the degree of pancreatic beta-cell death in rats fed a high dose (70-400mg/kg bodyweight).^[178][179] It has been seen to have protective effects against oxidation at a low dose of 0.04% dietary intake in diabetic mice.^[180] These interactions with the pancrease do not appear to influence insulin secretion, as evidenced by a prolonged study in lemurs in which resveratrol (and caloric restriction) were both implicated in improved glycemic control independent of insulin secretion.^[181]

When investigated the changes seen in a group subject to an obesogenic diet compared to one with the same diet but 100mg/kg resveratrol daily, pigs fed resveratrol^[182] had increased phosphorylated Akt, GLUT4 expression, and PGC-1a levels.

Fat cells are borne from mesenchymal cells, which are pluripotent stem cells that can turn into muscle cells (myocytes), fat cells (adipocytes), bone cells (osteoblasts) or cartilage (chondroblasts). A general overview of how resveratrol affects fat metabolism is that it hinders mesenchymal cells from turning into adipocytes and thus indirectly favors the other pathways. Thus, it indirectly benefits bone health and muscle health over a long period of time (theoretically).^[43]

In preadipocytes (the stage between mesenchymal cell and adipocyte), resveratrol intervention can cause a reduced viability of preadipocytes and reduced differentiation via increasing SIRT1 which suppresses the transcriptional factor PPARy and CCAAT^[183] which are two proteins required for differentiation of preadipocytes into mature adipoctes.^[184] This can reduce cell viability and at concentrations of 25-50uM can reduce lipid accumulation into preadipocytes.^[185]

In mature adipocytes, resveratrol can induce apoptosis (possibly via non-SIRT1 mediated synergysm with TNF-alpha mediated cellular apoptosis)^[186] and increased ephedrine-induced lipolysis while decreasing insulin-induced lipogenesis^[187][188] and increasing insulin and basal mediated glucose uptake into adipocytes.^[189] In essence, resveratrol seems to have non-significant benefits towards fat metabolism in adult cells as well as preadipocytes.

These mechanisms suggest resveratrol may be a great long-term anti-obesity agent, but it is unlikely that the above will cause fat loss over a short period of time independent of an increase in metabolic rate.

Resveratrol, like many flavonoid-like compounds, possesses the ability to inhibit fatty acid synthase. Resveratrol can also inhibit lipoprotein lipase and hormone sensitive lipase in addition to the differentiation factors C/EBP-alpha and SREBP-1c.^[186] These effects can, at concentrations of around 25-50uM, reduce fat accumulation into adipocytes.^[189][43]

In addition to numerous 'inhibition' effects of resveratrol on fat metabolism, resveratrol can also induce activity of the mitochondria vicariously through SIRT1 activation of PGC1-alpha (which activates more genetic transcription conducive to fat metabolism)^[60] and by increasing expression of UCP1 (Thermogenin) and SIRT3 which has the ability to reduce mitochondrial membrane potential.^[186][190] These downstream effects on the mitochondria can increase thermogenesis.

Resveratrol has also recently been shown to inhibit phospdiesterase enzymes, which is notable for fat metabolism as this mechanism (which increases cAMP in cells) is the mechanism by which caffeine is synergistic with other fat loss agents such as green tea catechins.^[57]

The combination of decrease fatty acid accumulation and increased fatty acid oxidation shows a promising trend towards acute fat loss mechanisms, and one in vivo human study did note that the thermic effect of food increase (albeit in a statistically insignificant manner) in obese persons.^[174] However, this increase in metabolism was enough to negate an observed decrease in sleeping metabolic rate, and thus 150mg daily may not influence fat mass in either direction over 30 days.

Supplementation with a high dose resveratrol at 200-400mg/kg bodyweight a day resulted in significant resistance to weight gain and increased thermogenesis, mitochondrial biogenesis, and aerobic capacity in administered rats^[191] In this study, rats were immune to weight gain and better tolerated a cold-stress test, indicating that thermogenesis increased; however, the dose was very high and mice have a higher amount of brown fat relative to humans.

Since AMPK deficient mice do not respond to resveratrol well, fat loss effects and increased thermogenesis may act vicariously through AMPK influencing SIRT1.^[192] This activation of AMPK is consistent with the hypothesis that AMPK activation then induces PGC-1a (through SIRT1 or directly) and thus mitochondrial biogenesis, and increased AMPK and PCG-1a protein content has been seen in humans after supplementation of 150mg resveratrol for 30 days.^[174] Although it is unlikely to have as much dramatic effects as seen in rats due to differences in brown fat stores, the mechanisms in white adipose exist.

Accordingly, 150mg resveratrol for 30 days does not result in fat loss in humans. Additionally, a suppression of sleeping metabolic rate was found to be significant although a slight increase in waking metabolic rate (via non-significant increases in diet-induced thermogenesis) made the whole-day metabolic rate not significantly different between groups.^[174]

Practically, resveratrol may cause some fat loss although this is most likely insignificant. Probably only enough to negate its suppression of sleeping metabolic rate and overall have no effect on fat loss in a short period of time.

When ingested at a dose of 4g/kg bodyweight in rats (a very high dose), resveratrol is able to augment force generation by 1.2-1.8 fold and increase exercise tolerance by 21%.^[193] Lower doses have not yet been investigated.

One study on voluntary runners noted that, after running, there was an increase in the biomarker of DNA damage called 8-OH-deoxyguanosine.^[194] Incubation of cells after running with resveratrol showed a _pro_oxidant effect at 100uM; which is the first example of resveratrol acting as a pro-oxidant in a pseudo in vivo model.^[195] It should be noted that there was high individual variation between runners in regards to how much DNA damage (assessed by double strand breaks) existed, and the more damage there was prior to resveratrol the more resveratrol augmented the damage. At low levels of damage, resveratrol was protective and ameliorated further damage.^[194]

Whether this synergistic effect on apoptosis via oxidative stress would exert a harmful or a anti-cancer effect is currently not known.

trans-Resveratrol, at 0.04% of the diet in rats, can increase glucose uptake into L6 myotubes via both insulin stimulated uptake and AMPK.^[180] This has been noted in humans recieving 150mg resveratrol daily.^[174] In addition to glucose uptake, trans-resveratrol supplementation at 150mg daily has been shown in humans to increase myocellular lipid stores.^[174] These muscular changes appear to be similar to caloric restriction.

Resveratrol, like its related compound Rapamycin, is able to inhibit both mTOR^[196][197] and S6K1^[198] by possibly both SIRT1-mediated^[199] and independent mechanisms.^[196] It also has the ability to prevent angiotension-II induced Akt phosphrylation, albeit in smooth muscle cells.^[200][201] If the above mechanisms in vivo are the same as rapamycin, they may then inhibit exercise induced muscle protein synthesis when taken before resistance training (mediated through mTOR and Akt).^[202]

However, at rest Akt1 and phospho Akt levels do not appear to be influenced by even 5g.^[66]

Although Resveratrol seems to possess the ability to preserve fast-twitch muscle function in vivo, it did not (in this study) appear to protect from age-related muscle wasting at 0.05% trans-resveratrol in the feed.^[203] It has been implicated in preserving lean mass at a dose of 400mg/kg in other studies of shorter duration though^[204] and thus it may be an issue of either dose or time.

Indirectly, the mechanism of PCG-1a overexpression is being investigated as a method to alleviate sarcopenia risk^[205] which resveratrol has been found to elevate downstream of SIRT1.^[206]

Nothing overly significant in regards to muscular health, although a concerning (albeit unproven) possibility with regards to mTOR; the Rapamycin study.

One study has noted that supplementation of 250mg resveratrol taken concomitantly with resistance training over the course of 8 weeks in otherwise healthy older men was able to prevent the reductions in blood pressure and improvements in oxygen uptake associated with exercise, which was seen in placebo given exercise alone.^[207] A commentary on this study^[208] has noted that many of the benefits in placebo that were statistically significant that were not significant in the resveratrol condition were of similar practical magnitude (ex. the statistical decrease of LDL-C by 0.3+/-0.2mM in placebo was significant while the 0.2+/-0.2mM in resveratrol was not) and it was claimed the harm of resveratrol was overstated. The defense from the initial authors^[209] of the clinical trial^[207] agreed that while the interpretation could be seen as excessive it was not the study's aim to assess practical relevance.

High dose resveratrol, due to it being an antioxidant and the process of oxidation being required for some adaptive responses to exercise, may have some hindering effects on optimal exercise adaptations. The magnitude of this 'blunting' effect appears to be minor and it may not have much practical relevance

A rat study using 146mg/kg bodyweight resveratrol for 12 weeks in conjunction with an exercise regimen (5 days a week, 60 minutes running until fatigue daily) increased time to exhaustion and performance by about 20%, theoretically secondary to increased fat oxidation and less glucose oxidation, relative to exercise alone.^[193] This improved performance affected sedentary mice as well, who were better able to run (+25%) than sedentary mice given control diets. This dose is approximately 14-23mg/kg bodyweight in humans, based on extrapolation from previous studies.^[210][211]

In human subjects supplementing resveratrol (150mg taken 15 minutes after exercise) over the course of four weeks alongside high intensity interval training (HIIT) in active adults noted that supplementation blunted the increase in VO₂ max seen in placebo and had lesser increases in power output on a wingate test;^[212] the increase in expression of a few genes induced by exercise (PGC-1α, SIRT1, and SOD2) was actually lesser with resveratrol supplementation compared to placebo with no differences in GPx1.^[212] This observation was not thought to be wholly due to the antioxidant effect of resveratrol alone, as Vitamin C and Vitamin E have similar effects on genetic expression due to their antioxidant actions they do not appear to hinder VO₂ max increases.^[213][214]

One rat study noting improvements in aerobic performance over 12 weeks, it was found that the tibialus anterior muscle had 18% greater twitch force (no different in tetanic force) production, and greater twitch and tetanic force production (58 and 22%, respectively) when rats were fed 146mg/kg resveratrol and given exercise relative to exercise alone.^[193]

Resveratrol can influence bone metabolism by directly influencing osteogenesis (as has been reported in vitro) by direct influence on the differentiation of cells as well as redirected the birth of stem (mesenchymal) cells from becoming fat cells into bone cells.^[215][216]

Although potent on it's own, resveratrol (and other bioflavonoids) show synergism with Vitamin D.^[217]

Resveratrol has been found in one in vitro study to increase protein content and induce the activity of steroidogenesis acute regulatory protein (StAR), the rate limiting step in steroid synthesis. This study was conducted in ovarian cells.^[218] One other study conducted in transfected Leydig cells noted a decrease in activity and mRNA content of StAR,^[219] although this was only significant at concentrations of 25-50uM rather than 1-5uM, which saw a slight non-significant increase.

Resveratrol has a sort of similar structure to estrogen, not as similar as many bioflavonoids, but enough to interact with estrogen metabolism.

In breast cancer cells, resveratrol can inhibit aromatase with an IC(50) value of 25microM by both competitive and non-competitive means.^[220] The IC(50) value is slightly higher in placental (JEG-3) cells.^[221]

Testosterone conversion to estrogen (and subsequent proliferation of the cell line) was reduced with resveratrol at 10uM in breast cells,^[220] and 25-50uM is associated with reduced transcription rates in breast and placental cells.^[220][221]

In the liver, doses of 1g daily for 4 weeks has been shown to induce Aromatase (CYP1A2) and inhibit CYP3A4 along two other CYP enzymes.^[80] It did not have a significant affect on Glutathione conjugation (GST) rates nor glucuronidation (UGT1A1), although it seemed to increase activity only in those with low baseline activity.^[80] The induction of aromatase seen in this study is actually pro-estrogenic (induction means to make more proteins), and may be due to the dose and time, as high acute dosages of resveratrol (25mg/kg bodyweight injection) seem to still suppress aromatase in 1-7 days of treatment.^[222]

After resveratrol supplementation, the highest amount of circulating resveratrol seems to have affinity for liver cells^[96][95] where is is taken up by HepG2 cells by both passive diffusion and carrier-mediated processes.^[93] This uptake is fairly rapid, being less than 2 minutes after incubation^[93] and was dose dependent (although no cancer cell death was seen at doses below 30uM).^[223]

In the liver, resveratrol seems to be cancer-preventative by acting against hepatocellular carcinoma proliferation in vitro^[224][223] and, due to these protective effects, resveratrol is currently being investigated for usage in hepatic metastasis prevention.^[66]

One study in rats noted that, relative to untreated control rats, that resveratrol is able to reduce noise-induced inflammatory and oxidative change (COX-2 and ROS, respectively) in the cochlea of rats; this is thought to be a possible mechanism to attenuate hearing loss with aging.^[225]

Resveratrol appears to be a regulator of the topoisomerase II enzyme (dose-dependently, measured at 20,40,80uM) and can induce genomic damage at high does in vitro.^[226] This effect is not due to resveratrol per se in mammalian cells, but appears to be through interactions with other agents that can act on the cell's nucleus,^[227][226] as shown by Cu2+ ions being able to damage the DNA for effectively when incubated with resveratrol.^[228]

Transfection of the p53 protein in cells which do not normally express it appears to mediate apoptosis induced by resveratrol, suggesting this is a key lever point.^[229]

Fas redistribution (also known as the CD65 pathway of apoptosis), which induces cell death, has been noted with concentrations of 10-100uM in colon tumor cells.^[230] Fas works by forming a 'Death Induced Signalling Cascade' (DISC) when activated by a ligand (as Fas is a cytoplasmic receptor). Resveratrol seems to modulate the levels of Fas and FasL, and thus modify the apoptotic response.^[231][230] The CD65 pathway appears to be involed in colon cancer,^[232] breast cancer,^[233] and in lymphocytes.^[234][43]

The NF-kB pathway, related to inflammation, has also been hypothesized to play a role in cancer progression and resveratrol, particularily in regard to skin, prostate, and lung carcinogenesis.^[43] NF-kB is a regulatory gene that is stimulated by stress and inflammation, and induces cell proliferation and survival; it is frequently misregulated in cancers.^[235][236] Resveratrol is able to suppress genes that are induced by NF-kB in response to inflammation and alleviate some cancer progression in some experimental models.^[237][238]

Resveratrol may also downregulate cell-cycle related proteins such as Cyclin D1, Cyclin E and Cyclin-dependent kinase which can block the Akt pathway in rat smooth muscle cells,^[239] bladder,^[240] and liver cancer cells.^[241] The PI3K/Akt pathway is related to some cancer progressions, and in general is related to cell survival, proliferation, and differentiation.

Low-dose resveratrol has been hypothesized to inhibit Wnt signaling, preventing nuclear localization of β-catenin.^[242] This is implicated in colon carcinogenesis to a large degree.^[43]

Alteration of the Bax:Bcl2 ratio has also been noted via an increase in Bax.^[243]

There does seem to be some increaes in apoptosis after ingestion, as increases in caspase-3 (marker of apoptosis) have been noted to be increased by 39% after ingestion of 5g micronized resveratrol (large dose) in liver tissue in humans.^[66]

Due to the low systemic bioavailability of free resveratrol, and the relatively high concentrations seen in vitro to achieve anti-cancer effects, the cancers that resveratrol seems to affect most significantly are those that it can come into contact with without being absorbed.^[43] It has shown promise on skin cancers when used topically^[233] and shows efficacy against esophageal cancer when ingested orally in rats.^[244]

Resveratrol is being investigated in reducing the risk of breast cancers.

It has been seen to inhibit a positive feedback loop in breast cells via aromatase promotor regions;^[245] as mentioned in the section on hormones, this occurs at around 25-50uM concentration.

Lots of possible mechanisms and promise, but human evidence is just starting to surface. It is too early to make a conclusion about resveratrol's interaction with cancer metabolism although it looks promising in most cases.

Resveratrol's effects on fat metabolism (inhibiting adipogenesis) are synergistic with the phytonutrient Genistein, of which the effects of synergism were roughly double the sum of the parts.^[246] At 50umol/L, Genistein increased apoptosis of preadipocytes and mature adipocytes by 46±9.2% and Resveratrol at 100umol/L by 46±7.9%, whereas the combination was measured at 242 ± 8.7%.^[246] Similar synergism was seen in decreasing lipid accumulation, and the decreases in adipogenesis may have been through downregulation of PPARy.

The combination is also able to increase Jun-N-terminal phosporylation when no compound in isolation was able to, and increased fat lipolysis by 25.5±4.6% when no compound in isolation did.^[246]

Quercetin was also synergistic with Resveratrol in protection of blood vessels^[247] and inhibition of adipogenesis and slightly more potent than Genistein in the overall percent synergism.^[248]

Combining all three bioflavonoids showed further synergism^[249] and low-dosing the three can provide cumulatively similar benefits at cheaper costs.^[250]

Leucine is the amino acid that appears to regulate muscle protein synthesis, and its metabolite HMB appears to directly activate SIRT1 in cell-free culture;^[251] Leucine's other metabolite (KIC) and leucine itself also had this effect, and it was as potent as 2-10uM resveratrol with KIC being most potent and HMB being least.^[251] Later, a study incubated leucine or HMB alongside resveratrol and noted synergism; modest fatty acid oxidation (+18%) was seen when no isolated compound induced it, and SIRT1 and SIRT3 activity was increased synergistically in adipocytes and skeletal muscle with the combination.^[252]

Outside of biofalvonoids, Resveratrols anti-cancer effects can be augmented with coingestion of Indole-3-Carbinol^[253][254]

Resveratrol is synergistic with curcumin in an animal model of lung cancer.^[255] The mechanism may be related to anti-inflammatory effects, which have been found to be synergystic in a model of osteoarthritis.^[237]

D-Glucaro-1,4-Lactone is produced during metabolism of Calcium-D-glucarate in humans^[256] at about 30% of an oral dose of D-glucarate.^[257]

This metabolite of D-glucarate at 0.5mM was able to cause a very low dose of resveratrol (0.1uM) to potently inhibit thrombin-induced aggregation and increase anti-oxidant potential; both compounds were ineffective at these dosages in isolation.^[258] Synergism between these two molecules has also been noted in preventing skin cancer occurrence when ingested orally.^[259]

Potentially synergistic for reducing blood clotting, currently the studies are not in living systems

β-1,3-Glucans are a class of polysaccharide that have traditionally been known to have immunostimulating effects.^[260] In female BALB/c mice, a resveratrol complex sourced from Japanese Knotweed containing a small emodin and piceid content injected alongside a β-1,3-Glucan complex showed synergistic reactions in increasing CD4 and CD19 positive splenic cells and on splenic cell recovery after experimentally induced leucopenia.^[260] β-1,3-Glucan, despite having no effect on IL-1 or IL-6, increased resveratrol's induction of these proteins synergistically; the combination was able to increase TNF-a levels when neither alone could.^[260]

Resveratrol's ability to induce Heme-Oxygenase 1 (HO-1) is enhanced during incubation with Melatonin in neurons; this synergism was not accompanied by increased mRNA of HO-1, but inhibition of the ubiquitin-proteasome pathway.^[148] Both molecules also exert neuroprotection and anti-oxidative properties, which are enhanced when together.^[148] In addition to HO-1, these two molecules are synergistic via AMPK^[147] and the Sirtuin system.^[261]

Melatonin is also able to work synergistically with resveratrol at low doses in cardioprotection, as 2.3mg/L resveratrol and 75ng/mL melatonin in a model of animal myocardial infarction.^[262] These doses parallel that found in red wine.

Melatonin and Resveratrol are also synergistic in a model of toxin-induced breast cancer progression in mice.^[263]

Grape seed extract is a blend of molecules, mostly proanthocyanidins, found in concord grapes; resveratrol may be a component of Grape Seed Extract.

In the presence of resvertrol, Grape Seed Extract has significantly more potency in destroying cancer cells via the p53 pathway.^[264]

This synergism has also been noted in regards to preventing skin cancer in mice.^[259]

Resveratrol appears to be well-tolerated by rats continuously at dosages up to 100mg/kg bodyweight,^[265] 400mg/kg bodyweight, and no adverse effects have been noted at 750mg/kg bodyweight trans-resveratrol.^[211] Some adverse effects were noted in animals at 300mg/kg bodyweight, but may have been reflected of increased absorption kinetics by gavage feeding.^[266] This may be of a concern to micronized resveratrol (with enhanced absorption) if taken in similar dosages.

The No Observable Adverse Effect Limit (NOAEL) of resveratrol appears to be 200mg/kg bodyweight in rats and 600mg/kg bodyweight in beagle dogs.^[267]

In humans, up to 5g have been taken with no side effects outside of some intestinal upset^[268] and nausea.^[66] Micronization of resveratrol at this dosage showed the severity of symptoms decrease, indicating that nausea and intestinal upset are caused by resveratrol's poor bioavailability.^[66]

A large amount of in vitro (in glass; not living bodies) evidence suggests that resveratrol can harbor toxic effects, but these studies are typically conducted at concentrations that are well beyond feasible with supplementation.