Can resveratrol win the equivalent of a 2nd Nobel Prize?

Resveratrol’s Second
Life Extension Mechanism

Resveratrol operates as an mTOR inhibitor,
thus adding another mechanism to its maximum lifespan proficiencies

By Will Block

When mice were administered extra copies of the SIRT1 gene, or fed with the sirtuin activator resveratrol, they had more efficient DNA repair without deterioration in the control by SIRT1 of gene expression; moreover, their mean lifespan was extended by 24% to 46%.
— Durk Pearson & Sandy Shaw [commenting in Life Extension News, Volume 12 No. 1, Feb. 2009 on Oberdoerffer et al. SIRT1 redistribution on chromatin promotes genetic stability but alters gene expression during aging. Cell 135:907-18 (2008).]

P opulation geneticists and molecular anthropologists now use the study of DNA to determine when significant events occurred in human evolution. As an example, it is now possible to determine when humans began to use fully modern language (about 50,000 years ago), or when the world’s ancestral population of about 150 humans left the African continent (also around 50,000 years ago).* Both of these are cheerful events.


* See “Did Shellfish Omega-3s Spur Brain Evolution?” in the April issue.


However, there is a darker side, away from heightened word craft and the diaspora of a remnant population into a more inviting world, and it involves some disturbing ideas of DNA evidence: that our ancestors wiped out the Neanderthal and Homo ergaster populations in genocidal warfare that spanned millennia, and that they practiced cannibalism (also called anthropophagy).1 Yes, cannibalism! It shows up in our genes.

Speaking of which, we still practice a form of cannibalism (aside from the Congressional legislative process), called autophagy and rather than speaking disparagingly of it, we should praise it because it may be a key to another mechanism for maximum lifespan extension.


Rapamycin’s life extending benefits
are, at least in part,
due to autophagy.


Why Autophagy is Good for You

In cell biology, autophagy is the process of self-digestion by a cell through the action of enzymes originating within the same cell. It is the tightly-regulated catabolic breakdown and degradation of a cell’s components through the lysosomal enzymatic machinery. Autophagy is a normal part of cell growth, development, and homeostasis that helps maintain a balance between the production, degradation, and consequent recycling of cellular material. When a cell is starving, this process plays an instrumental role in reallocating nutrients from unnecessary to more-essential processes.

There are different forms of autophagic processes, but all degrade intracellular components via the lysosome. Perhaps the most studied mechanism of autophagy involves membrane formation around a targeted region of the cell, separating the contents from the rest of the cell’s cytoplasm. The vesicle thus formed fuses with a lysosome and then degrades the contents. (See Fig. 1.)


The Four Stages of Autophagy
  1. Induction: Following external/internal stimuli (e.g. nutrient depletion or ischemia), mTOR is inhibited, leading to induction of autophagy.
  2. Autophagosome formation: Cytosolic proteins and organelles are sequestered by a double membrane vesicle, the origin of which is uncertain, but may arise from the endoplasmic reticulum. Formation of this vesicle is co-ordinated by complexes of Atg proteins, which encode for the “start codons,” which encode for the amino acid methionine.
  3. Docking and fusion with the lysosome.
  4. Breakdown of the autophagic vesicle. The molecular mechanism behind the fusion with the lysosome and subsequent breakdown of the autophagic vesicle are poorly understood.
Autophagy was first described in the 1960s, yet many questions remain about the actual processes and mechanisms involved. Its role in disease is still not well categorized. However, it may help to prevent or halt the progression of some diseases, including some types of neurodegeneration and cancer. Also, it may play a protective role against infection by invoking intracellular pathogens. And it may be evocable by various nutrients.

Excitement Caused by Aging Delay

Fireworks went off last July in the scientific world when a paper appeared in Nature finding that rapamycin, an antifungal drug used to prevent organ rejection in transplant patients, could delay aging in laboratory animals, even when given late in life.2

Working in parallel at three institutions, the researchers—David E. Harrison of the Jackson Laboratory, a mouse-breeding center in Bar Harbor, ME; Richard A. Miller of the University of Michigan; and Randy Strong of the University of Texas Health Science Center—knew that inhibiting the TOR (target of rapamycin) signaling pathway by genetic or pharmacological intervention extends lifespan in variety of lower organisms. But no one knew if the same process of mTOR (mammalian target of rapamycin) inhibition could extend lifespan in mammals. This was the basis of their design, and what they found was that rapamycin, an inhibitor of mTOR, extends median and maximal lifespan of both male and female mice, even when started at 600 days of age, the equivalent of a 60 year old human.

Rapamycin led to an increase in mean lifespan between 28–38% from the beginning of treatment, or 9–14% in total increased maximum lifespan (14% for females and 9% for males, on the basis of age at 90% of mortality). In another part of the study, rapamycin begun at 270 days of age also increased survival in both males and females. However, additional data are needed to provide an accurate estimate of the measure of success with midlife mortality, and to evaluate effects on maximal longevity.

Rapa Nui from Easter Island

It is curious to note that rapamycin was first discovered as a product of the bacterium Streptomyces hygroscopicus in a soil sample from Easter Island, also known as “Rapa Nui,” and hence the name. In summary, Harrison et al. concluded that rapamycin may extend lifespan either by postponing death due to cancer or by retarding aging mechanisms, or both. But they could not say with certainty exactly how rapamycin locks in its anti-aging effects. This paper is the first to show a role for mTOR in the regulation of mammalian lifespan, and the extension of lifespan in both genders by a drug. Why couldn’t rapamycin do the same for humans? To throw down a damp towel, a lot is not known, and any drug that inhibits the immune system is not to be fooled around with. However, there may be other alternatives, not in the world of Pharma, and not subject to the regulatory system, where things tend to get done a lot faster. And fortunately, the findings of this paper have implications for these other interventions targeting mTOR for the treatment and prevention of age-related diseases.

Rapamycin is an Autophagy Inducer

What has not been widely reported, especially in light of all the publicity surrounding the Nature study, is that rapamycin has been found to induce autophagy.3 One study has even found that rapamycin’s life extending benefits are, at least in part, due to autophagy.4 The researchers in this study fed the antifungal drug to adult Drosophila (fruit flies) and found that it could produce the life span extension seen in some TOR mutants. This was associated with increased resistance to both starvation and paraquat (a noxious herbicide that the US has used to defoliate marijuana fields in Mexico).


A plaque commemorating the discovery of rapamycin (sirolimus) on Rapa Nui (Easter Island)
In this study, several mechanisms were revealed for increased longevity involving the TORC1 protein complex branch of the TOR pathway, which operated through alterations to both autophagy and translation machinery. Elevated levels of autophagy are generally thought to be beneficial for aging prevention, owing to increased rates of removal of damaged molecules and organelles. Other mechanisms identified for lifespan increase involve rapamycin’s ability to amplify the insulin/insulin-like growth factor signaling pathway activity and that of dietary restriction.

Other mTOR Inhibitors

Among nutrients that have been found to target mTOR are curcumin (one of the active compounds in turmeric) as well as resveratrol (see Durk Pearson & Sandy Shaw’s “Emerging Science: Life Extension by Inhibiting Growth” in their Life Extension News, Volume 13 No. 1, February 2010 in the April issue of Life Enhancement). Rapamycin has been shown to induce autophagy, which through programmed self-digestion helps to clear aggregated proteins in neurodegenerative diseases such as Alzheimer’s disease. Could autophagy be involved in mTOR inhibition?

Resveratrol is Autophagic

In a new study, conducted at the University of Connecticut School of Medicine’s Cardiovascular Research Center, Farmington, CT, Dr. Narasimman Gurusamy, et al. have found that resveratrol given in lower doses—0.1 and 1 µM in cardiac myoblast cells (a type of progenitor cell that gives rise to muscle cells) and 2.5 mg/kg/day in rats—induced cardiac autophagy enhanced formation of autophagosomes and component markers after hypoxia/reoxygenation or ischemia/reperfusion.5 The induction of autophagy was correlated with enhanced cell survival and decreased apoptosis. However, with the higher dose of resveratrol, autophagy was not increased but reduced.

Remarkably, treatment with rapamycin (100 nM), a known inducer of autophagy, did not further increase autophagy compared with resveratrol alone. The activation of mTOR was differentially regulated by low-dose resveratrol, i.e. the phosphorylation of mTOR was inhibited at one site of interactivity but increased at another; but these results were lessened with a higher dose of resveratrol. This and other results suggest that the entire process of autophagy is upregulated after low-dose resveratrol treatment.

Trehalose, an mTOR-Independent
Inducer of Autophagy

Trehalose is an unusual disaccharide present in many plants and non-mammalian species that protects against a wide variety of environmental stresses (see “Maintain Your Youthful Edge” in the April issue). While many of its protective benefits are due to its chemical chaperone properties, much is still unknown. In a recent scientific paper, trehalose is report to possess a novel function as an mTOR-independent autophagy activator.1 This is the first time that trehalose has been shown to induce autophagy.

Trehalose-induced autophagy enhanced the clearance of autophagy substrates like mutant huntingtin and two mutants of α-synuclein (a protein expressed in neural tissue) that are associated with Huntington and Parkinson diseases, respectively. Furthermore, trehalose and mTOR inhibition by rapamycin together exerted an additive effect on the clearance of these aggregate-prone proteins because of increased autophagic activity. By inducing autophagy, the researchers showed that trehalose also protects cells against subsequent pro-apoptotic insults via the mitochondrial pathway.

This is great news! We know that trehalose is an osmolyte and that it can help to prevent protein misfolding that is a principle cause of protein aggregation, and at the root of many degenerative diseases. When you combine these protective properties of trehalose (as an inducer of autophagy and chemical chaperone), with other known autophagy inducers, you have a powerful nutrient formulation. And should you need to be treated for Huntington or Parkinson disease, the combinatorial strategy of trehalose along with rapamycin may be relevant for these and other related diseases, where the mutant proteins are autophagy substrates.

Reference

  1. Sarkar S, Davies JE, Huang Z, Tunnacliffe A, Rubinsztein DC. Trehalose, a novel mTOR-independent autophagy enhancer, accelerates the clearance of mutant huntingtin and alpha-synuclein. J Biol Chem 2007 Feb 23;282(8):5641-52.

Autophagy Only a Partial Contributor

Although high dose resveratrol decreased autophagy, it did not alter the cell survival as observed with inhibitors of autophagy, suggesting that resveratrol-mediated autophagy is only a partial contributor to cell survival under the given set of experimental conditions.


Inhibition of mTOR with rapamycin
induces autophagy, but in
the current study it did not induce
any more autophagy than induced by
low-dose resveratrol alone.


Though rapamycin-mediated inhibition of mTOR induces autophagy in many cell types, in the current study it did not induce further autophagy than that induced by low dose of resveratrol alone. Thus resveratrol-mediated autophagy could also track the same pathway as rapamycin in the inhibition of mTOR. Nevertheless, mTOR was differentially regulated when autophagy was induced by low dose of resveratrol, i.e. phosphorylation of mTOR varied with interactivity sites, down at one and up at another.

These results compare favorably with a recent study, cited by Gugusamy, et al., showing that resveratrol inhibits oxidized LDL-induced phosphorylation of mTOR along with a downstream molecule in rabbit femoral smooth muscle cells. However, as another cited study has shown, this downstream molecule, a serine/threonine kinase, is needed for the entire process of autophagy, and it must be activated first for the maximal activation of autophagy.

As the paper also showed, there was a positive correlation between the induction of Rictor [a component of mTOR complex 2, (mTORC2)], autophagy, and cell survival with low dose resveratrol. Speculating whether mTORC2, could play any role in the induction of autophagy, the scientists suppressed the expression of Rictor, and this resulted in the suppression of autophagy induced by low dose of resveratrol followed by hypoxia/reoxygenation.

A TOR de Force

Taken together, the Gurusamy et al. results indicate that low-dose resveratrol mediates cell survival through induction of autophagy. To the best of our knowledge, this may be the first study to show that resveratrol induces autophagy in the myocardium and in cardiac myoblast cells. Inhibition of autophagy with 3-methyladenine and wortmannin, known mTORC1 and mTORC2 inhibitors, inhibited resveratrol-mediated autophagy and abolished the cardioprotective abilities of resveratrol. Though the complex interaction and the positive and negative feedback regulation among the various factors are known to some extent, the exact mechanism through which 3-methyladenine and wortmannin causes the inactivation of mTOR and p70s6k is unclear.


The results indicate for the first time
that low dose of resveratrol-induced
cell survival is, at least in part,
mediated through
the induction of autophagy.


In conclusion, the results indicate for the first time that low dose resveratrol-induced cell survival is, at least in part, mediated through the induction of autophagy. So, whereas there has been some concern about the amount of resveratrol needed to grant an mTOR benefit, at last we have an indication that it may not require toxic levels of resveratrol. From the 2.5 mg/kg/day dose given to rats, we can calculate through body surface ratios that the amount would be a mere 35 mg/day for an 85 kg (187 lb) person. High doses (25 mg/kg/day and 100 mg/kg/day) did not produce autophagy. That’s the equivalent of 344 mg/day for an 85 kg (187 lb) person and 1377 mg/day. What we still need to know is the optimal levels and hopefully, future studies will help us to zero in on these. In France, an exceptional creative achievement, a particularly adroit maneuver, or the accomplishment of a difficult feat is called a tour de force. In this vein, the idea of natural mTOR inhibitors might be called a TOR de force.

References

  1. Brookfield J. Human evolution: a legacy of cannibalism in our genes? Current Biology, Volume 13, Issue 15, Pages R592-3
  2. Harrison DE, Strong R, et al. Rapamycin fed late in life extends lifespan in genetically heterogeneous mice. Nature 2009 Jul 16;460(7253):392-5.
  3. Carloni S, Girelli S, Scopa C, Buonocore G, Longini M, Balduini W. Activation of autophagy and Akt/CREB signaling play an equivalent role in the neuroprotective effect of rapamycin in neonatal hypoxia-ischemia. Autophagy 2010 Apr 20;6(3).
  4. Bjedov I, Toivonen JM, Kerr F, Slack C, Jacobson J, Foley A, Partridge L. Mechanisms of life span extension by rapamycin in the fruit fly Drosophila melanogaster. Cell Metab 2010 Jan;11(1):35-46
  5. Gurusamy N, Lekli I, Mukherjee S, Ray D, Ahsan MK, Gherghiceanu M, Popescu LM, Das DK. Cardioprotection by resveratrol: a novel mechanism via autophagy involving the mTORC2 pathway. Cardiovasc Res 2010 Apr 1;86(1):103-12.


Will Block is the publisher and editorial director of Life Enhancement magazine.

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