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Is Nrf2 a Master Regulator of Longevity and Healthspan?
This is what is suggested by the vast array of chronic age-related diseases, as shown in Table 1, that are prevented and/or treated by raising Nrf2. This is what is suggested by the large number of health-promoting factors that act by raising Nrf2. This is what is suggested by the ability to Nrf2 to raise antioxidant, anti-inflammatory, mitochondrial function, and autophagy activities, given the established role of oxidative stress, inflammation, mitochondrial dysfunction, and accumulation of damaged proteins, protein aggregates, and organelles. Each of these last four mechanisms is implicated in the aging process.
This was suggested by Lewis et al. in their paper "Nrf2, A Guardian of Healthspan and Gatekeeper of Species Longevity."49 They state, "There is mounting evidence across evolutionarily distant species that Nrf2-ARE-dependent components are associated with both longevity and extension of healthspan." These studies include a number of genetic studies in the mouse and in several other species that raising Nrf2 activity produces prolonged lifespans and healthspans and that lowering Nrf2 produces shorter lifespans and healthspans.
How Is Nrf2 Regulated by the Health-Promoting Factors Listed in Table 2?
Each source has reviewed the mechanisms by which Nrf2 is regulated, together with at least a bit of information on how various factors raise Nrf2.1-22 Their discussions on mechanisms are generally much more detailed than is the discussion here. Consequently, the reader is suggested to go to them and particularly to Hayes & Dinkova-Kostova, Kumar et al., and Baird & Dinkova-Kostova for more detailed information than is provided here.1-3
Nrf2 protein, under what have been called noninduced situations, is mostly contained in an inactive complex with another protein known as Keap1. Keap1 has five reactive cysteine residues, in each of which reaction of inducing chemicals with the cysteine thiol can start a process leading to release of Nrf2 from Keap1. Following release, Nrf2 can move into the nucleus, complex with other proteins called Maf, bind to ARE sequences on DNA, and stimulate transcription of adjacent genes. The agents that react with these thiols are electrophilic and/or oxidative, and the reaction with these thiols is thought to be the most important mechanism of regulation of Nrf2. The five different cysteine thiols differ from one another in what compounds they react with.
However, there are many other mechanisms that come into play, making the Nrf2 control system very complex. There are several protein kinases that have roles in regulating Nrf2, including the ERK/JNK pathway, PI3K/Akt/ GSK-3b pathway, protein kinase C, and protein kinase G. In addition, when Nrf2 is bound to Keap1, it tends to be targeted to proteasomal degradation, so that its levels are kept low. Release from Keap1 increases the stability of Nrf2 roughly 7-fold, leading to substantially increased levels. Furthermore, Nrf2 stimulates the transcription of its own gene and also the MAFG gene, thus further stimulating Nrf2-dependent transcription. But there is also a downregulation mechanism – Nrf2 also stimulates transcription of the Keap1 gene, lowering Nrf2 elevation. Another complication is that agents that stimulate the aryl hydrocarbon receptor (AhR) increase Nrf2 transcription, leading to increases in Nrf2 activity, a subject that has only fairly recently attracted much attention.50
How then do the agents listed in Table 2 stimulate Nrf2 activity? Isothiocyanates, H2O2 and other oxidants, phenolic antioxidants, long-chain omega-3 fatty acids, and carotenoids act by reaction with Keap1 reactive thiols with the last three of these acting through their oxidation products. Allium sulfur compounds, isothiocyanates, and carotenoids act via ERK stimulation, with the latter two acting via two distinct mechanisms to raise Nrf2. Some flavonoids and other phenolics, including some inactive in the Keap1 reactions, act as AhR agonists, as do some terpenoids. Some terpenoids act by raising PI3K and some act directly in Keap1.
It follows from all this that phytochemicals and other agents can increase Nrf2 activity by reacting either directly or through their products with different cysteine residues on Keap1, by regulating the activity of numerous different protein kinases or by stimulating the AhR receptors. It follows from this that phytochemicals and other agents that act in different ways to raise Nrf2 may be expected to act synergistically together.
Consequently, phytochemically rich diets such as the traditional Mediterranean or Okinawan diet may be more active in Nrf2 activation because of possible synergism than may be suggested from just looking at the activities of their individual phytochemicals.
Can Too Much Nrf2 for Too Long Be Toxic?
In general, as indicated in Lewis et al., raising Nrf2 produces prolonged lifespans in animal studies.49 In addition, human diets rich in nutrients that raise Nrf2, including the traditional Mediterranean and Okinawan diets, produce longer lifespans and lowered disease incidences. However, there are situations where chronic high-level Nrf2 stimulation produces pathophysiological responses in the body. Perhaps the clearest, well-documented example of this is where high-level chronic raising of Nrf2 levels by TCDD (dioxin) leads to chloracne.51,52 TCDD also has other, Nrf2-independent toxic effects, but these acne-like changes in skin properties are clearly caused by excessive, long-term levels of Nrf2, such that chloracne may serve as a marker for excessive Nrf2 stimulation. Arsenite and other arsenicals can also produce similar skin responses, acting via excessive Nrf2 activity, but again arsenite has other Nrf2-independent toxic effects.52 Both the TCDD and the arsenite effects act through AhR stimulation to produce elevated Nrf2 activity. These skin effects of excessive Nrf2 appear to be caused in part by the elevated sensitivity of keratinocytes to Nrf2. Chloracne may be useful as a clinical marker of excessive Nrf2 activity in patients using agents known to raise Nrf2.
This keratinocyte role also shows up in perhaps the most dramatic effect of excessive Nrf2. It was shown that Keap1 transgenic mouse knockout mutants developed hyperkeratosis in the esophagus and forestomach during gestation, which led to death from malnutrition after birth. This was shown to be caused by excessive Nrf2 activity.53
In conclusion, it may be expected that levels of Nrf2-raising nutrients that occur in the Mediterranean or Okinawan diets will produce predominantly health-promoting effects. Nevertheless, very high chronic, long-term Nrf2 elevation can produce pathophysiological effects like almost any regulatory effect taken to extreme. Therefore, one needs to take care not to raise Nrf2 levels too high for too long. Given the great amount of genetic heterogeneity in the human population, some individuals may also be much more susceptible to such pathophysiological effects.
The list of diseases in Table 1 wherein raising Nrf2 acts to prevent and/or treat the disease, at least in animal models, is truly stunning. But it should not be surprising, given the known ability of Nrf2 to produce a whole battery of antioxidant effects, lower a whole battery of inflammatory effects, and improve mitochondrial function in a series of different ways. Additional roles in raising whole batteries of detoxification responses and autophagic removal of destructive protein aggregates and dysfunctional organelles may also be useful in disease prevention and treatment. Each of these diseases is well established to have oxidative stress and inflammatory causal mechanisms, and most of them also are known to have mitochondrial dysfunction mechanisms. Several of them are known to have destructive protein aggregates as well. Many of these diseases are thought to be what is called a NO/ONOO(−) cycle etiology, including several cardiovascular and neurodegenerative diseases, asthma, multiple sclerosis, and epilepsy.54-56 Heart failure is now the best documented NO/ONOO(−) cycle disease.56 The 23rd and most recent disease proposed to be caused by the local impact of the cycle is glaucoma.57 Because the cycle involves oxidative stress including peroxynitrite elevation, inflammatory aspects, and mitochondrial dysfunction, it should not be surprising that apparent NO/ONOO(−) cycle diseases may be prevented and/or treated by raised Nrf2.
The list of health-promoting factors that each act at least in part by raising Nrf2 (Table 2) is also truly stunning. And the fact that the two most healthful diets known, the traditional Mediterranean diet and the traditional Okinawan diet, are both rich in nutrients that act to raise Nrf2 emphasizes the fact that the Nrf2 regulatory system is likely to be important in real human populations. The role of Nrf2 in determining animal lifespans and healthspans also supports this view. It is probable that the Paleolithic diet was much higher in Nrf2-raising nutrients than are our modern diets. By allaying deficiencies in such nutrients, we may be able to lower the prevalence of many of our chronic inflammatory diseases.
It is our belief that raising Nrf2 is likely to be the most important health-promoting approach into the foreseeable future. That is not to say that it is a magic bullet. More is not always better, and other health-promoting nutrients and other agents acting in other ways are likely to act along with Nrf2. Agents that lower NF-kB via Nrf2-independent ways, nutrients that are health promoting in other ways, such as B vitamins and vitamin C, magnesium, and some trace elements are likely to be useful, as are agents such as high doses of the hydroxocobalamin form of B12, which lowers peroxynitrite by lowering its two precursors. Other agents that act to improve mitochondrial function independent of Nrf2 are likely to be useful as well.
It has become almost a fad to denigrate the importance of oxidative stress to medicine. However, the exquisite coordination of the antioxidant enzymes controlled by Nrf2 clearly shows that the importance of oxidative stress may be even greater than many scientists working in the field have long believed. Such a complex and clearly highly coordinated set of mechanisms could not have evolved and been maintained throughout metazoan evolution if it had not been of great importance to our health. The Nrf2 studies also show three other related lessons:
1. Regulatory control of antioxidant proteins may be considerably more important than are direct chain-breaking antioxidant mechanisms.
2. Antioxidant mechanisms should be viewed as functioning together with other cytoprotective mechanisms, including anti-inflammatory mechanisms, improvement of mitochondrial function, autophagy, and detoxification of environmental toxicants.
3. Our modern diets should be viewed as being deficient in Nrf2-raising nutrients, with such deficiencies producing much of the chronic inflammatory disease that afflicts us over much of the world.
Notes added in proof: Since this article was submitted, a much longer paper on the same topic has been published by the authors, a paper that is available full text from the PubMed database.58 In addition, several other important features of Nrf2 have become clear to the authors. Two genes that regulate human health, the ApoE gene, which influences the occurrence of chronic inflammatory disease (increased susceptibility when carrying the epsilon 4 allele), and the BRCA1 (breast cancer gene), both regulate Nrf2, with lowered Nrf2 activity being associated with increased disease. Additional health-promoting factors, including vitamin D and alpha-lipoic acid, also raise Nrf2.
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