In Focus4 In Focus May 2015 molecular switch that is exquisite - ly sensitive to environmental and molecular triggers. Nrf2 turns out to be a key modu-lator of glutathione

For more information call 800-545-9960 1

NutriCology® Newsletter May 2015

The Genius of Nrf2

Smart About Sugar

Surprising discoveries into an exquisitely sensitive, master cellularregulator shed new light on everything from longevity diets to agingand neurodegeneration ...................................................................................... Page 2

• Nrf2 Explains It All To Us: Evolution’s Master Antioxidant Switch Gives Us New Insight Into Health and Illness

Histamine Intolerance: The Hidden Culprit in Many Health Problems ......................Page 26

A world expert on Nrf2 explains why this pathway is critical formaintaining cellular redox homeostasis, and how it activates profoundly protective genes .................................................................................................Page 11

Clinical nutritionist Elizabeth Jeffery gives us her top tips for preparing broccoli and tomato to maximize their health benefits ..............................................Page 14

The distinguished scientist discusses a wholly unexpected bridge he discovered last year between the NO/ONOO- cycle and Nrf2 .....................................Page 15

New research uncovers unexpected nutrients and molecules that, in combination, safely support healthy glucose levels ....................................Page 17

A report from Todd Born, ND, on the most effective tests and approaches for pre-diabetes and diabetes ................................................................................Page 24

• The Two Faces of the Master Regulator Nrf2, an Interview with Donna Zhang, PhD

• Raw, Cooked, Steamed or Microwaved? an Interview with Elizabeth Jeffery, PhD

• From NO/ONOO- to Nrf2: A Remarkable Bridge, an Interview with Martin Pall, PhD

• Sweet Success: Novel Pathways to Stable Blood Sugar

• Practice Points: A Step by Step Guide to Testing and Treating Impaired Glucose Regulation

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®In Focus

2 In Focus May 2015

Nrf2 Explains It All To Us:Evolution’s Master Antioxidant Switch

Gives Us New Insight into Health and IllnessSixty years ago cardiologist and chemist Denham Harman pub-lished a revolutionary paper that changed the way we viewed health and longevity.1 Harman suggested an idea that is now second nature to us all—he pro-posed aging is due in part to the damage caused by “free radi-cals,” molecules that have a sin-gle unpaired electron in an outer shell, rendering them highly reac-tive and damaging. Free radicals are generated by cellular metabo-lism, as well as by toxicants from the environment.

The energy of life is like a fire, and fire must be controlled or it even-tually it will consume the forest. Harman’s theory got a huge boost in 1969, when a University of Ala-bama graduate student named Joe McCord, and his mentor, Irwin Fridovich, published monumen-tal findings on the potency and necessity of the endogenous an-tioxidant, superoxide dismutase (SOD), which helps quench the important free radical, super-oxide. In fact, later research has shown that catalase, another po-tent antioxidant enzyme, present in all aerobic cells, can turn 6 mil-

lion molecules of hydrogen per-oxide to oxygen and water every single minute.2 Now that’s one potent antioxidant!

Harman was never able to actually extend the maximum lifespan—the age at which the oldest known member of a species has died. He thought about why that was so, and in 1972 published another remarkable paper suggesting that mitochondria, the energy powerhouses inside our cells, produce and are damaged by free radicals when they create energy. It is understood that it is very hard to migrate antioxidants through our own cell membranes and then through the double-membrane of the mitochondria. Harman concluded that mitochondrial integrity and function determine our maximum lifespan.3

He was likely right on all counts. Science now understands that aging, and the degenerative dis-eases associated with aging, are in part due to our slowly declin-ing ability to defend ourselves against free radical damage. These free radicals include su-peroxide, hydrogen peroxide, and peroxynitrite.4 Although

these molecules have the ability to help kill invading microorgan-isms, they also have potential for damaging our cells and their lipid membranes. We now un-derstand that through a healthy diet and exercise, as well as judi-cious supplementation with anti-oxidant nutrients, we may slow that process and help our bodies repair that inevitable damage.5 Antioxidants familiar to us all in-clude Vitamins E and C, glutathi-one, lipoic acid, melatonin, carot-enoids, and natural flavonoids.6,7

Nrf2: Master Antioxidant SwitchThe free radical theory offered us a new lens by which to under-stand aging and degenerative ill-ness. But one huge question has always remained: how does a cell manage to sense oxidative dam-age and then generate or utilize antioxidants to repair itself? To-day, we are beginning to unravel the answer, and it centers on a po-tent master regulator of cellular repair, an intracellular transcrip-tion factor called Nuclear factor erythroid-derived 2-like 2 (Nrf2).8

Evolution generated a master-piece when it crafted Nrf2. Nrf2

The Genius of Nrf2The Genius of Nrf2


is the name for a cellular switch that orchestrates antioxidant, de-toxification, and cellular defenses. When activated, Nrf2 can switch on over 200 genes that help the cell generate its own highly pro-tective molecules.9,10,11 These mol-ecules are crucial in the metab-olism of drugs and toxins. They protect against oxidative stress and inflammation, remove dam-aged proteins, and initiate DNA repair. In fact, the Nrf2 pathway is emerging as an astonishingly rich area of research in immuno-logical, neurological disorders, and in cancer research. Currently, there are over 1,000 peer reviewed studies published a year on this pathway alone.

Nrf2 is widely distrib-uted in all mammals. It is the result of a billion years of evolution, and is highly complex and highly conserved as a system. The diversity of Nrf2’s many target genes shows its vital importance in cell survival and in life itself and is expressed in all tissues, al-though the key detoxification or-gans (the kidney and liver) exhibit the highest levels.12

Nrf2 provides us unexpected in-sight into the dance of life itself—the dance of fire and water—of oxidants and antioxidants. Our Nrf2 system is self-healing. It has evolved to protect us against the slings and arrows of life, and it is consequentially stimulated into action by these and other stressors. Many health-enhanc-ing molecules that we have long

thought of as potent antioxidants, work in part by upregulating Nrf2. Even some of our most potent cancer-fighters work be-cause they activate our own sub-stantive cell-protective defenses. These molecules offer protection that goes far beyond their known roles as antioxidants and singlet oxygen scavengers. They may help explain how health and lon-gevity diets may work (because they are rich in Nrf2 stimulators). By acting as ‘low-dose stressors,’ these constituents of healthy diets may actually prepare us to resist more severe stress. Nrf2 is likely one reason that sulforaphane in broccoli is a potent cancer and inflammation fighter—sulfora-

phane is now known to success-fully activate Nrf2.13,14,15

Nrf2 teaches us that life, and our body, is in a dynamic balance. Some scientists think that Nrf2 is the master regulator of the aging process itself, and that the pro-tective role of Nrf2 against tox-ins and the growth of tumors ac-tively determines longevity and healthspan.4 Raising Nrf2 activity is likely to be of particular impor-tance to the hundreds of millions of people around the globe who are regularly exposed to toxic chemicals that contribute to dis-eases characterized by oxidative stress, inflammation and mito-chondrial dysfunction, diseases

which include many of the chron-ic diseases of 21st century life.21

How Does Nrf2 Work? A Quick GuideNrf2 first emerged from obscurity in 1997, when a biochemist named Masayuki Yamamoto at the University of Tsukuba in Japan showed that it activates an entire class of detoxifying enzymes.16

Nrf2 usually hums along in the cell at low levels and is activated only when a cell is under stress. Until that moment, it is kept qui-etly in check by a molecule known as Keap1.17 When a cell is suitably stressed (including by pro-oxi-dants), Keap1 is stimulated to let

go of its hold on Nrf2, which then moves into the nucleus of the cell and triggers a wide range of genes called antioxidant response

element (ARE)-bearing genes. These ARE-bearing genes churn out powerful natural antioxidants and Phase II liver detoxifying en-zymes, which safely denature and quench electrophiles, reactive ox-ygen and nitrogen species, and help the cell remove damaged proteins, quench inflammation, and repair itself.18

As soon as the trigger passes, Keap1 escorts Nrf2 out of the nu-cleus and it is broken down. Nrf2 then returns to its normal, low-lev-el resting state, and the cell’s mo-lecular switch is set to “off” until the next stressor comes along. The Keap1-Nrf2 bond is an ingenious invention of evolution, creating a

Evolution orchestrated a masterpiece when it crafted Nrf2–a cellular switch that orchestrates antioxidant, detoxifica-tion, and cellular defenses.

The Genius of Nrf2

4 In Focus May 2015

molecular switch that is exquisite-ly sensitive to environmental and molecular triggers.

Nrf2 turns out to be a key modu-lator of glutathione. Reduced glu-tathione (GSH) has often been de-scribed as the most important low molecular weight antioxidant pro-duced in the human body.19 Each of the three genes that encode en-zymes required for the synthesis of GSH are activated by Nrf2, as is the gene for glutathione reduc-tase, the enzyme that converts oxidized glutathione (GSSG) to GSH. But Nrf2 doesn’t stop there. The most potent anti-oxidant enzymes in our body include superox-ide dismutase and cat-alase. Two superoxide dismutase genes (SOD1 and SOD2) are activat-ed by Nrf2, with each SOD lowering oxidative stress by lowering superoxide. Catalase and two glutathione per-oxidase genes are each induced by Nrf2, with each of these enzymes acting to lower hydrogen perox-ide produced from superoxide.20

To the hundreds of millions of people around the world ex-posed daily to substantial levels of toxicants, detoxification may be Nrf2’s most remarkable gift to us. Numerous Nrf2 genes contrib-ute to detoxification of xenobiot-ics (toxic chemical compounds) and heavy metals.21 Upregulation of Nrf2 also has astounding an-ti-inflammatory effects including lowering many inflammatory me-diators such as cytokines, chemo-

kines, and adhesion molecules.22

Nrf2 also helps spur the forma-tion of new mitochondria—the energy and immune system pow-erhouses that stud our cells and regulate cellular respiration and metabolism.23 This is called mi-tochondrial biogenesis, and it is impaired as we age. Nrf2 activa-tion increases the efficient use of fuel (fatty acids and glucose) by mitochondria and increases ener-gy production. It also promotes muscle repair and recovery and reduces muscle injury.

In sum, Nrf2 responds to a wide range of cellular stresses. This includes toxicants of all kinds, some pro-oxidants, exercise, and the many health-promoting phy-tochemicals in fruits and vege-tables. At first glance, this seems counterintuitive. Why would our most healing foods, our richest source of vitamins, minerals and antioxidants, also contain mild toxicants that initiate our cell’s master healing switch?

On further thought, this makes sense. These molecules are mes-sengers to us about our environ-ment. How does any fruit, vege-table, or plant survive and thrive while being attacked by patho-

gens, predators and weather ex-tremes? One way is by churning out a wide range of powerful molecules to protect itself—not just self-repairing molecules but bitters, tannins, toxicants and ox-idants to defend against harsh conditions. Fruits and vegetables are not only power-packed with nutrition; they contain a chem-ical map of their own immuno-logical survival mechanisms, and the ability to trigger our own Nrf2. They are messengers and allies.24,25 A grape vine stressed by too much sun, too many pred-

ators or pathogens, churns out resveratrol and protective tannins. We eat the grapes and our body responds to the phytochemicals by elevating Nrf2, protect-ing us and theoretical-ly increasing our own resistance to disease

and stress. Is it any wonder that a diet generous in nutrient rich, Nrf2-stimulating fruits and vege-tables slashes rates of cancer and all-cause mortality?26

In fact, the traditional Mediterra-nean diet and the traditional Oki-nawan diet are both heavy in Nrf2 stimulators. They are thought to be two of the most healthful hu-man diets known to humans, asso-ciated with high overall lifespans, large numbers of centenarians and low incidences of cancer and cardiovascular disease. The tradi-tional Mediterranean diet relies heavily on olives and olive oil, which contain very high levels of Nrf2 stimulating phenols and

The traditional Mediterranean diet and the traditional Okinawan diet are both heavy in Nrf2 stimulators. They are thought to be two of the most health-ful human diets known to humans, and include some of the longest living and healthiest older adults.


terpenes. The purple sweet pota-to is the staple of the traditional Okinawan diet and is very high in carotenoids and anthocyanins, which are potent Nrf2 activators. Nrf2 likely has a major role in the health promotion in each of these two diets.27,28,29

It’s No Surprise: Nrf2 Research is ExplodingIn recent years, researchers of ev-ery ilk have begun to explore the power of Nrf2. Some are elucidat-ing its basic mechanisms. Some are exploring its ability to oscillate at different speeds, or defining its true molecular weight.30 But perhaps most excit-ing is the research into Nrf2 and many devas-tating human illnesses: Multiple Sclerosis, Par-kinson’s, Huntington’s, cancer, and diabetes. These diseases each appear sep-arate—with distinct causes and symptom pictures—and yet all involve oxidative stress. No won-der a wealth of Nrf2 research is fo-cusing on so many of them, with fascinating new findings suggest-ing that Nrf2 can eventually be tweaked to possibly relieve and slow many terrible illnesses—and maybe even slow aging.

Multiple Sclerosis: Multiple scle-rosis (MS) affects 2.1 million peo-ple around the world, and there is currently no treatment that will cure the disease, though medica-tions can help slow its progres-sion and ease symptoms. A newly approved oral medication for MS, Dimethyl fumarate (DMF), acti-

vates the Nrf2 pathway and low-ers oxidative stress. It significantly reduces relapses and brain lesions. DMF increases the concentration of Nrf2 in the cell, and promotes the activity of ARE-genes down-stream of Nrf2. Extensive and rig-orous clinical trials have assessed the efficacy and safety of DMF.31,32

Parkinson’s Disease: Nrf2 re-search has received grants from The Michael J. Fox Foundation. Numerous studies suggest that activation of the Nrf2–ARE path-way in the brain offers protection to vulnerable neurons, making neuronal Nrf2 up-regulation an

attractive therapeutic strategy for Parkinson’s Disease (PD). Re-search is focusing on novel com-pounds that cross the blood–brain barrier and activate the Nrf2-ARE pathway in the brain. The hope is such compounds could halt the progression of PD.33

Diabetes: As it progresses, diabe-tes impairs antioxidant capacity in the heart, including Nrf2. The American Diabetes Foundation is sponsoring a study on Nrf2 and cardiac health. Researchers at the University of Louisville in Ken-tucky are looking at both Type 1 and Type 2 diabetes. Research has already shown that Nrf2 pro-tects the heart from high levels of glucose. Nrf2 is down regulated

in diabetic patients. Studies in mice show that a novel Nrf2 ac-tivator, known as dh404, protects the animals from heart damage due to diabetes. If Nrf2 can help prevent diabetes-related heart problems, the researchers plan to develop dietary supplements that upregulate cardiovascular levels of Nrf2.34

Epilepsy: In 2013, Epilepsy Re-search UK awarded Matthew Walker and colleagues at Universi-ty College London a grant to study Nrf2 pathways in seizure induced cell death by two pathways.35,36

Huntington’s Disease: The rapidity with which damaged proteins are cleared from neurons may affect cell survival and may explain why some cells die in neuro-degenerative disorders.

To test this idea, researchers acti-vated Nrf2, which allowed neu-rons to live longer.37 The scientists used optical pulse-labeling, a new technology that allowed them to actually monitor individual neu-rons in real time.

In Huntington’s disease and many other neurodegenerative disor-ders, proteins that are misfolded (have abnormal shapes), accumu-late inside and around neurons and are thought to damage and kill nearby brain cells. Normally, cells sense the presence of mal-formed proteins and clear them away before they do any damage. This study was supported by a grant from the National Institute of Neurological Disorders and

As we gain a richer understanding of Nrf2, we can focus on exciting new ways of raising Nrf2, when appropriate, and substantially lowering many health risks of modern life.

6 In Focus May 2015

Stroke (NINDS). “Nrf2 seems like a potentially exciting therapeutic target. It is profoundly neuropro-tective in our Huntington’s mod-el and it accelerates the clearance of mutant huntingtin [a gene po-tentially important for long-term memory storage],” said principal investigator Steven Finkbeiner.

Cardiovascular Disease: Nrf2 also suppresses multiple proin-flammatory genes, including tumor necrosis factor α (TNF-α), and the interleukins (IL-1b, IL-6) as well as prostaglandins, ma-trix-metalloproteinases, and ni-tric oxide synthase.38 Downregu-

lating inflammation benefits the entire cardiovascular system. Nrf2 has been shown to protect arterial endothelial cells from inflammation.39,40 The Nrf2 path-way is crucial in protecting the cardiovascular system against oxidative damage by reactive oxygen species.41

Longevity: “The strong relation-ship between species longevity and cellular resistance to oxida-tive insults has been reported in a wide variety of organisms across the animal kingdom,” write re-searchers at the University of Tex-as at San Antonio. Mice who have

no Nrf2 at all look normal, but are more susceptible than their Nrf2 carrying peers to hyperoxia, li-popolysaccharide (LPS), cigarette smoke, diesel exhaust fumes, ul-traviolet irradiation (UVA and UVB), as well as toxic chemicals.42

Top Nrf2 NutrientsAvailable NowIn spite of the flurry of research on Nrf2, safe pharmaceuticals that increase its activity are a long way off. Yet diet, lifestyle and targeted nutrients can gently and safely support healthy Nrf2 manage-ment now. Many polyphenols in plants stimulate Nrf2, as do to-

Ingredients:

• 200ml/7floz/scant1cupnatural yogurt,soyayogurtorcoconutyogurt

• 1scoopofProBerryAmlaPowder

• 125g/4½oz/heaped¾cup strawberries

• 125g/4½oz/1cupraspberries

• 225g/8oz/heaped1cupfreshor frozencherries,pitted

Directions:

Put all the ingredients into a highspeed blender and process untilsmoothandcreamy.

Pour into an ice creammaker andchurnaccordingtothemanufactur-ers instructions. Alternatively pourintoashallowfreezer-proofcontain-erandfreeze for2–3hoursoruntilfirm.

Removefromthefreezer30minutesbefore serving to allow it to softenslightly. (Store in the freezer for upto3months.)

This delicious, refreshing sorbet is naturally sweet with an array of berries. It was created by Christine Bailey, MSc, PGCE, MBANT, CNHC, for raising Nrf2 in a nourishing and healthy dessert that is rich in probiotics and fiber. The combination of ProBerry Amla powder,

which is packed with Nrf2 stimulating berries, along with regular fresh or frozen berries, makes for a delicious treat. Christine is Director of Advance Nutrition in the UK, television chef and health consultant. Her website is www.christinebailey.co.uk.

Nutritional information per serving Protein 4g, Carbohydrate 12.2g of which sugars 11.5g, Fat 5.3g of which saturates 3.4g, Kcals 110

Serves 4Preparation time: 5 minutes, plus freezing time

SuperBerry Yogurt Sorbet

3

2

1


copherols and tocotrienols, sulfur compounds from the garlic and onion family, carotenoids, fish oil, and molecules such as Ginkgo bi-loba, pomegranate extract, and si-lymarin, among others. Here is a list of some potent, dietary Nrf2 inducers with substantial peer re-view research already available. These nutrients are easily found in fruits, vegetables, and as di-etary supplements.

Sulforaphane: Sulforaphane is a phytochemical found in crucifer-ous vegetables such as broccoli, cabbage, kale, collard greens, cau-liflower, bok choy, and Brussels sprouts. It is especially concen-trated in broccoli sprouts. Sul-foraphane is a hotbed of research: over 1200 peer-reviewed studies have been published on this one phytochemical since 1992.

Sulforaphane is one of the most potent natural inducers of Nrf2, acting first on Keap1. It is able to accumulate in cells and actually swap function with glutathione, effectively handling oxidation.43 Its protective role in cancer is par-ticularly profound. Recent studies on sulforaphane and Nrf2 span the gamut, demonstrating its abil-ity to protect the retina, kidney, lung, liver and skin specifically by activating Nrf2.44

Sulforaphane from broccoli sprouts has been studied as pos-sible liver cancer prevention in China where rates of this cancer are particularly high. Residents of Qidong often suffer from chronic infection with hepatitis B virus, and are exposed to high levels of

aflatoxins in mold-contaminated produce. As a result, heptatocellu-lar cancer is responsible for up to 10% of the adult deaths in areas.45

Clinical trials in the Qidong prov-ince of China found that brocco-li sprouts increase excretion of many toxins, from mold toxins to pollutants. Freeze-dried standard-ized sprout extracts from select cultivars were utilized in multiple studies, and hot water extracts in others.46 A team from Johns Hop-kins University School of Med-icine and Bloomberg School of Public Health, in collaboration with scientists at the Qidong Liv-er Cancer Institute in Shanghai gave a hundred individuals five ounces of broccoli sprouts extract, and another hundred individuals drank a placebo. Tests of subjects’ urine showed carcinogens were being excreted from the body in those who drank the active ex-tract. Statistically significant in-creases of 20-50% in the levels of excretion of chemicals from air-borne pollutants (acrolein, cro-tonaldehyde and benzene) were seen in individuals receiving sul-foraphane from broccoli sprouts.47 The researchers conclude that, “Prevention trials of whole foods or simple extracts offer prospects for reducing an expanding global burden of cancer effectively with minimal cost.”48 In another study, Researchers found a significant association between excretion of total sulforaphane metabolites and excretion of aflatoxin.49

Resveratrol: Resveratrol is a po-tent antioxidant found in more

than 70 species of plants, includ-ing grapes, cranberries, blueber-ries, mulberries, lingonberries, bilberries, jackfruit, peanuts, choc-olate, as well as Asian plant roots. It is a plant-derived polyphenol that is beneficial for the neurolog-ical, hepatic, and cardiovascular systems and seems to slow down aging, mimicking the effects of ca-loric restriction. It is reported to extend longevity in yeast, nema-todes (worms) and flies.50

For at least ten years now, resver-atrol has stimulated wide-ranging research into its potential thera-peutic benefits. In fact, resvera-trol is thought to be the red wine molecule that explains the famous “French paradox” (the French, who consume a lot of red wine, have less coronary heart disease than expected, given their high fat diets).51 And yet until we began to unravel its role in upregulat-ing Nrf2, the actual molecular and cellular mechanisms of this potent antioxidant were elusive. Resver-atrol, it appears, is a strong trigger of Nrf2, enhancing function of the endocrine, cardiovascular, and nervous systems.52

According to a 2010 review article by researchers at NIH and New Jersey Medical School, resvera-trol may offer a safe way to target several pathways involved in the development of cardiovascular diseases.53 Resveratrol has been shown to help attenuate insulin resistance, and may be a useful strategy for protecting against dia-betes.54 It may help prevent breast cancer; in animal studies, supple-

8 In Focus May 2015

mentation with resveratrol alone or in combination with the protec-tive form of estrogen significantly upregulated expression of Nrf2 in mammary tissue. Resveratrol also prevented estrogen-medi-ated inhibition of detoxification genes. Resveratrol treatment in-duced death of malignant cells and inhibited an estrogen-medi-ated increase in DNA damage in mammary tissues. Taken togeth-er, these results suggest that res-veratrol inhibits estrogen-linked breast cancer via Nrf2-mediated protective pathways.55

Resveratrol may help protect against lung cancer associated with cigarette smoking. Cigarette smoke depletes glutathione (GSH)

levels in the lung’s alveolar epi-thelial cells. Resveratrol restored glutathione levels in human al-veolar epithelial cells in part through the Nrf2 pathway. The researchers conclude that “these data may have implications in di-etary modulation of antioxidants in treatment of chronic obstruc-tive pulmonary disease.”56 There are many other powerful studies suggesting that resveratrol’s ben-efits are linked to Nrf2.

Silymarin: Silymarin, from the plant milk thistle, is a unique blend of three flavonoids, silybin, silydianin, and silychrisin. Sily-marin is well known as a potent antioxidant that has profound protective effects on the liver. It

has been demonstrated to help generate new liver cells damaged by alcohol or drugs, stimulate bile flow and reduce liver/gall-bladder stagnation, increase the survival rate of patients suffering from cirrhosis, and protect the liver against all kinds of poisons and pharmaceuticals and protect the kidneys in diabetic nephrop-athy.57,58,59,60 In healthy individu-als, silymarin raises glutathione in the liver by more than a third.61 Silymarin also increases the level of the important antioxidant en-zyme superoxide dismutase in cell cultures.62

Silymarin also appears to protect against cancer, including prostate cancer.63 It also shows potentially protective effects in an astound-ing range of other cancers: colon, ovarian, skin, lung, breast and cervical cancers.64,65,66 But sily-marin appears to offer protection far beyond the liver and potential-ly cancer. New research suggests that it may benefit the kidneys, protect the heart, guard against insulin resistance, and even help prevent Alzheimer’s disease.67

How can one phytochemical from one single plant pack all this pro-tective power? One way is via Nrf2 activation. Silymarin in-creases the movement of Nrf2 into the nucleus.68

Oleuropein: The olive tree is highly resistant to disease, in large part due to a bitter compound in both raw olives and olive leaf called oleuropein. It has been shown to lower blood pressure, increase blood flow in the arteries,

Endogenous & ExogenousStress Signals

Extension of Longevity &Health Span

CytoprotectiveEnzymes

Detoxification

Cytoplasm

Nucleus

ARE

Maf

The General Scheme for the Inductionof Nrf2-Signaling Pathway

Figure 1: Nrf2 (Nuclear factor (erythroid-derived 2), Keap1 (Kelch-like ECH-associated protein 1), ARE (antioxidant response elements), Maf (proteins that help regulate expres-sion and induction of detoxifying enzyme genes)


and inhibit the growth of patho-gens through bactericidal, anti-viral and antifungal activity. It is antioxidant, antiatherogenic, an-ti-cancer, anti-inflammatory and has antimicrobial properties.69,70,71 It is neuroprotective as well.72

It is not surprising that oleuro-pein appears to upregulate Nrf2. In new research, both oleacin (a polyphenol in olive oil) and oleuropein were restorative via increasing heme oxygenase-1 (HO-1) by upregulating Nrf2.73 Synthetic analogues of oleuropein are impressively potent, and also work by upregulating Nrf2.74

Ginkgo biloba, Pomegranate, and Green Tea PolyphenolMany other nutraceuticals stim-ulate Nrf2 activity. Here are just a few:

Ginkgo biloba is one of the lon-gest living tree species in the world. Trees of this species can live a thousand years. Ginkgo bi-loba is well known as a powerful antioxidant with a special affinity for vascular tissue.75 Folklore uses include stroke, memory prob-lems, erectile dysfunction, vision problems, and asthma. It may also protect against radiation damage. New research shows its protective ability may be me-diated by Nrf2, which under the influence of Ginkgo helps stimu-late Phase II detoxifying enzymes in the liver. A recent study found that Nrf2 content in the nucleus of the cell was raised by Ginkgo biloba extract. Ginkgo appears to act on Keap1, inhibiting its reten-

tion related activity so that more Nrf2 is released.76

Another study suggests that Gink-go biloba’s ability to activate Nrf2 is responsible for its vascular pro-tective effects, which triggers the increased activity of a gene called Heme-oxygenase-1.77 Heme ox-ygenase is an enzyme that de-grades heme. Heme is the deep red, oxygen-carrying, nonprotein, iron component of hemoglobin. Heme oxygenase is induced by oxidative stress, and in animal studies it has been shown that in-creasing this expression seems to be protective.78

Pomegranate is an ancient fruit, well known for its antioxidant features. Pomegranate possesses remarkable antioxidant proper-ties capable of protecting normal cells from oxidative stress and cell death. The fruit contains at least three powerful anthocyani-dins (delphinidin, cyanidin, and pelargonidin), which quench free radicals.79

New research points pomegran-ate in the direction of Nrf2. A 2011 animal study, for instance, shows that pomegranate extract reduced the incidence, number, size and volume of liver nodules, which are a precursor to liver cancer. The study also found that pomegran-ate extract increased Nrf2 expres-sion. The potent activity of pome-granate is in part achieved by “upregulation of several house-keeping genes under the control of Nrf2 without toxicity,” the re-searchers conclude.80 This may

be particularly relevant to liver diseases in general, since many Nrf2 target genes are expressed in the liver. Nrf2 plays complex and multicellular roles in hepatic in-flammation, fibrosis, hepatic can-cer, and regeneration.81

Green tea contains a potent poly-phenol, Epigallocatechin-3-gal-late (EGCG), which has proven neuroprotective effects in animal studies. EGCG protects mouse neurons in Parkinson’s disease, and in mouse models of ALS.82 Other mice studies found that EGCG inhibits amyloid plaque formation and reduces cognitive impairment.83 EGCG increases distribution of Nrf2 in the nu-cleus of the cell, and modestly upregulates numerous Nrf2 tar-get genes.84 EGCG may also help prevent cancer; tea polyphenols have been extensively studied in cell culture and animal models where they inhibited tumor on-set and progression.85 Green tea consumption is linked to heart health, and appears in part to be due to Nrf2, activating heme ox-ygenase-1 (HO-1). At the molecu-lar level, EGCG treatment signifi-cantly activates Nrf2 activity.86

New Insights YieldNew ApproachesEvery health conscious individual recognizes that organic produce is generally healthier than conven-tional produce. Organic produce offers higher levels of vitamins and minerals, and lower levels of toxic metals and pesticides, according to the most comprehensive scien-

10 In Focus May 2015

tific analysis to date.87 The team, led by Newcastle University sci-entist Carlo Leifert, found that organic food was 19-69% richer in antioxidants. Switching to organ-ic fruit and vegetables could give the same benefits as adding one or two portions of the recommended “five a day” known to dramatical-ly lower all-cause mortality.

But why are organic foods richer in antioxidants? The likelihood is that without the influence of heavy pesticides, the fruits and vegetables are forced to mount their own defenses. And we reap the benefits, through our high-ly responsive, self-healing Nrf2 system. Our master antioxidant switch, Nrf2, is naturally and intermittently activated by con-suming healthy foods and phy-tochemicals, and by exercise, some of which are best described as being pro-oxidant in nature. This new insight helps us refine Harman’s revolutionary under-standing of what we have uni-versally called “anti-oxidants.” Some of these anti-oxidants ac-tually assist us because they are Nrf2-stimulating pro-oxidants. Consuming Nrf2 activators in foods and spices avoids inappro-priate overstimulation of Nrf2, so that you have appropriate and proportional physiological re-sponse as needed.88

The Nrf2 master antioxidant switch embedded deep in our mo-lecular chemistry acts as an envi-ronmental response agent unique-ly adapted to assist mammals to survive in the face of numerous

challenges and stressors. Its role as a gene amplifier and cellular detoxifier has been extensively explored, and further comprehen-sion is added every few weeks. But one simple observation we can distill from all of the remarkable new science is that antioxidant mechanisms are among the most important in the evolution of life on earth. Nrf2 produces complex and well-coordinated detoxifica-tion, immune, anti-inflammatory, mitogenesis and other healthful effects. As we gain a richer under-standing of Nrf2, we can focus on exciting new ways of raising Nrf2 and substantially lowering many health risks of modern life, with our constant exposures to xeno-biotic toxicants, pollutants, toxic metals and other adverse sources of oxidative stress.

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In Focus: You’ve assembled an extraordinary body of research on the Nrf2 pathway, going all the way back to 2001. You were way ahead of your time. How did you get interested in this pathway in the first place?

DZ: I was trained as toxicologist at New York University, and was always interested in how a cell re-sponds to toxicants. At the time we knew cells had two detoxify-ing pathways involving phase I and phase II genes. They work in

concert to protect cells from envi-ronmental insults. But we knew very little about how the expres-sion of phase II genes was regulat-ed, and as I was doing a literature search on that, I came across a study about Nrf2 regulating phase

The Two Faces of Nrf2, The Cell’s Master Regulator

An Interview with Donna Zhang, PhD

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II detoxification. That’s how I got interested. Studying Nrf2 allows me to combine my interest in ex-ploring mechanistic regulation of cellular signaling pathways with my training in toxicology.

In Focus: In the years since that time, interest in Nrf2 has exploded. There are over a thousand stud-ies published annually. Can you explain, in the simplest of terms, why this pathway is so important, and how it gets activat-ed or quieted down?

DZ: In one sense, the Nrf2 pathway is our very life. It is critical for maintaining cellu-lar redox homeostasis, and it activates a whole range of genes that protect the cell by signaling our body to produce antioxidants and detoxifying en-zymes. Nrf2 can protect cells and tissues from a variety of toxicants and carcinogens by increasing the expression of innumerable genes. As a result, several Nrf2 activa-tors are currently being tested as chemopreventive and protective compounds in clinical trials. This is a very promising strategy for cancer prevention.

Nrf2 is activated by many di-etary ingredients, and you can eat certain foods or take dietary supplements to help enhance the pathway. One example is sulfora-phane, a molecule with anti-can-cer activity, found in broccoli and broccoli sprouts. This molecule has been demonstrated to protect against liver cancer in over fifteen years of clinical trials in China. Scientists have shown that broc-coli sprouts made into a tea are

effective in helping lower the risk of liver cancer. The sprouts do this by activating Nrf2. That’s just one of many examples of dietary in-fluence. We are now working on a potent and nontoxic molecule de-rived from carrots, which upreg-ulates Nrf2 and is showing great promise in preliminary research.

In Focus: How do these dietary compounds upregulate Nrf2 activity?

DZ: Nrf2 is normally regulat-ed by a molecule called Keap1.

Keap1 keeps Nrf2 at low levels by targeting Nrf2 degradation in your body most of the time. You only need Nrf2 when your body encounters stress in the form of toxicants, carcinogens, even exer-cise. At that point Keap1 loosens its hold on Nrf2 and in a sense sets it free from destruction. When you eat something like broccoli sprouts, they inactivate Keap1, al-lowing Nrf2 to increase temporar-ily. The upregulation of Nrf2 is, by

nature, intermittent, and tightly regulated.

In Focus: In some of your papers, you’ve highlighted the double edged sword of Nrf2. It can protect the cell from

all kinds of insults—but that the pathway can also be hijacked by can cer cells for their benefit.

DZ: Yes. Just as Nrf2 protects nor mal cells, Nrf2 may also pro-tect cancer cells from chemo-therapeutic agents and facilitate cancer progression. Nrf2 is ab-errantly accumulated in many types of cancer, and its expression is associated with a poor progno-sis in patients. This is what I call the ‘dark side’ of Nrf2. Nrf2 and

Biography: Donna D. Zhang, PhD, is a molecular toxicologist and oncologist at the University of Arizona. Her specialty is the Nrf2/Keap1 signaling pathway, and she has published over 86 peer reviewed papers, many on this pathway. In 1999 she received a DuPont Young Investigator Award, in 2006 she was the recipient of an NIH Outstanding New Environmental Health Scientist Award, and in 2012 she received the Society of Toxicology Achievement Award. She receives over 1,000 citations a year of her work with a total citation of over 6,000. She is currently the Principal Investigator for a study characterizing the anti-cancer properties of brusatol,

an inhibitor of the Nrf2 pathway being tested as a possible adjunctive therapy in chemotherapy. She is also the principal investigator for a study on the protective role of Nrf2 in arsenic-induced toxicity and carcinogenicity. She is a member of the National Institutes of Health Review Committee on Chemo/Dietary Prevention, is a reviewer for NIH, and is a member of the Distinguished Editorial Panel for Botanical Dietary Supplement Research Centers (BDSRC) and the Distinguished Editorial Panel for Centers for Advancing Natural Products Innovation and Technology Centers (CANPIT).

The Nrf2 pathway is our very life. It is critical for maintaining cellular redox homeostasis, and it activates a whole range of genes that protect the cell.


its downstream genes are overex-pressed in many can cer cell lines and human cancer tis sues, giving cancer cells an advantage for sur-vival and growth. Furthermore, Nrf2 is upregulated in resistant cancer cells and is thought to be responsible for both intrinsic and acquired resis tance to chemother-apy. Therefore, it may actually be helpful to inhibit the Nrf2 path-way during chemotherapy.

In Focus: How do cancer cells manage to take advantage of Nrf2, and how does it help them?

DZ: Many cancer cells carry a mutation in Nrf2 or Keap1, which diminishes the ability of Keap1 to keep Nrf2 destroyed. Thus, Nrf2 is constitutively overexpressed at maximal levels. With all that extra Nrf2 to protect them, these cells are more resistant to chemo- and radio- therapy. When we down-regulate Nrf2, the cells are more susceptible to anti-cancer drugs.

In Focus: Does that mean that an individual who has cancer should not be eating healthy, Nrf2 promoting foods and di-etary compounds?

DZ: I get this question quite of-ten after I give a talk. My answer is, no. Diet is a gentle and inter-mittent stimulant of Nrf2, and your diet wouldn’t really impact the cancer cell, because the cancer cell has evolved a somatic mu-tation so that Keap1 is no longer effectively binding to Nrf2. Nrf2 is continually upregulated in the cancer cell. So in those cancer cells, Nrf2 is always there, always high-ly expressed. Eating Nrf2 stimu-lating foods in your diet would

not further upregulate Nrf2 levels in cancer cells at all.

In Focus: That’s good to hear. So tell us about your work on in-hibiting Nrf2 in the cancer cell.

DZ: Our laboratory has devel-oped the first Nrf2 inhibitor in the world, called Brusatol. We have been working on it since 2005. In our studies, it appears to be re-ally great for sensitizing cancer cells to chemotherapy, and may eventually prove an effective ad-juvant to enhance chemotherapy. We aren’t at the stage of a clini-cal trial yet, but we have shown it works beautifully in mice lung cancer models.

In Focus: What is your final message?

DZ: Eat your broccoli or oth-er tasty foods and spices that we have discovered are Nrf2 induc-ers, from cinnamon to carrots!

Selected Recent BibliographyRiahi R, Sun J, Wang S, Long M, Zhang DD, and Wong PK. 2015. Notch1-Dll4 signaling and mechanical force regulate leader cell formation during collective cell migration. Nature Communication 2015 Mar 13; 6:6556. PMID: 25766473

Canet MJ, Merrell MD, Harder BG, Maher JM, Wu T, Lickteig AJ, Jackson JP, Zhang DD, Yamamoto M, Cherrington NJ. 2014. Iden-tification of a Functional Antioxidant Response Element within the Eighth Intron of the Human ABCC3 Gene. Drug Metab Dispos [Epub ahead of print]. PMID: 25349122

Tao S, Wang S, Moghaddam SJ, Ooi A, Chapman E, Wong PK, Zhang DD. 2014. Oncogenic KRAS confers chemoresistance by upregulat-ing NRF2. Cancer Res [Epub ahead of print]. PMID: 25339352

Nam KH, Jamilpour N, Mfoumou E, Wang FY, Zhang DD, Wong PK. 2014 Probing mechanoregulation of neuronal differentiation by plasma lithography patterned elastomeric substrates. Sci Rep. 4:6965-6974. PMID: 25376886

Bao L, Jaramillo MC, Zhang Z, Zheng Y, Yao M, Zhang DD, Yi X. 2014. Induction of autophagy contributes to cisplatin resistance in human ovarian cancer cells. Mol Med Rep [Epub ahead of print]. PMID: 25322694

Kang MJ, Wu T, Wijeratne EM, Lau EC, Mason DJ, Mesa C, Tillotson J, Zhang DD, Gunatilaka AA, La Clair JJ, Chapman E. 2014. Func-tional chromatography reveals three natural products that target the same protein with distinct mechanisms of action. Chembiochem [Epub ahead of print]. PMID: 25125376

Wu T, Harder BG, Wong PK, Lang JE, Zhang DD. 2014. Oxidative stress, mammospheres and Nrf2-new implication for breast cancer therapy? Molecular Carcinogenesis [Epub ahead of print]. PMID: 25154499

Wu T, Zhao F, Gao B, Tan C, Yagishita N, Nakajima T, Wond PK, Chapman E, Fang D, and Zhang DD. 2014. Hrd1 suppresses Nrf2-mediated cellular protection during liver cirrhosis. Genes Dev 28(7):708-722. PMID: 24636985

Riahi R, Wang S, Long M, Li N, Chiou PY, Zhang DD, Wong PK. 2014. Mapping Photothermally induced gene expression in living cells and tissues by nanorod-locked nucleic acid complexes. ACS Nano 8(4):3597-3605. PMID: 24645754

Bao LJ, Jaramillo MC, Zhang ZB, Zheng YX, Yao M, Zhang DD, Yi XF. 2014. Nrf2 induces cisplatin resistance through activation of au-tophagy in ovarian carcinoma. Int J Clin Exp Pathol. 7(4):1502-1513. PMID: 24817946

Shen T, Chen XM, Harder B, Long M, Wang XN, Lou HX, Wondrak, GT, Ren DM, Zhang DD. 2014. Plant extracts of the family Laura-ceae: a potential resouce for chemopreventive agents that activate the nuclear factor-erythroid 2-related factor 2/antioxidant response element pathway. Planta Med 80(5):426-434. PMID: 24585092

Tao S, Justiniano R, Zhang DD, Wondrak GT. 2013. The Nrf2-induc-ers tanshinone I and dihydrotanshinone protect human skin cells and reconstructed human skin against solar simulated UV. Redox biology 1:532-541. PMID: 24273736

Jiang T, Tian F, Zheng H, Whitman, SA, Lin Y, Zhang X, Zhang N, Zhang DD. 2013. Nrf2 suppresses lupus nephritis through inhibi-tion of oxidative injury and the NF-κB-mediated inflammatory re-sponse. Kidney Int 82(2):333-343. PMID: 24025640

Riahi R, Long M, Yang Y, Dean Z, Zhang DD, Slepian MJ, Wong PK. 2014. Single cell gene expression analysis in injury-induced collective cell migration. Integrative biology 6(2):192-202. PMID: 24336811

Wu T, Wang XJ, Tian W, Jaramillo MC, Lau A, Zhang DD. 2014. Poly(ADP-ribose) polymerase-1 modulates Nrf2-dependent tran-scription. Free Radic Biol Med 67:69-80. PMID: 24140708

Qiao S, Tao S, Rojo de la Vega M, Park SL, Vonderfecht AA, Jacobs SL, Zhang DD, Wondrak GT. 2013. The antimalarial amodiaquine causes autophagic-lysosomal and proliferative blockade sensitizing human melanoma cells to starvation- and chemotherapy-induced cell death. Autophagy 9:2087-2102. PMID: 24113242

Jaramillo MC, Zhang DD. 2013. The emerging role of the Nrf2-Keap1 signaling pathway in cancer. Genes Dev 27(20):2179-2191. PMID: 24142871

Whitman SA, Long M, Wondrak GT, Zheng H, Zhang DD. 2013. Nrf2 modulates contractile and metabolic properties of skeletal muscle in streptozotocin-induced diabetic atrophy. Exp Cell Res 319(17):2673-2683. PMID: 23896025

Qiu Q, Zheng Z, Chang L, Zhao YS, Tan C, Dandekar A, Zhang Z, Lin Z, Gui M, Li X, Zhang T, Kong Q, Li H, Chen S, Chen A, Kaufman RJ, Yang WL, Lin HK, Zhang DD, Perlman H, Thorp E, Zhang K, Fang D. 2013. Toll-like re ceptor-mediated IRE1alpha ac-tivation as a therapeutic target for inflammatory arthritis. EMBO J 32(18):2477-2490. PMID: 23942232

Chen W, Wang H, Tao S, Zheng Y, Wu W, Lian F, Jaramillo M, Fang D, Zhang DD. 2013. Tumor protein translationally controlled 1 is a p53 target gene that promotes cell survival. Cell Cycle 12(14):2321-2328. PMID: 23797580

Villeneuve NF, Tian W, Wu T, Sun Z, Lau A, Chapman E, Fang D, Zhang DD. 2013. USP15 Negatively Regulates Nrf2 through Deu-biquitination of Keap1. Mol Cell 51(1):68-79. PMID: 23727018

Ji L, Li H, Gao P, Shang G, Zhang DD, Zhang N, Jiang T. 2013. Nrf2 pathway regulates multidrug-resistance-associated protein 1 in small cell lung cancer. PLoS One 8:e63404. PMID: 23667609

Riahi R, Dean Z, Wu TH, Teitell MA, Chiou PY, Zhang DD, Wong PK. 2013. Detection of mRNA in living cells by double-strand-ed locked nucleic acid probes. Analyst 138(17):4777-4785. PMID: 23772441

Lau A, Zheng Y, Tao S, Wang H, Whitman SA, White E, Zhang DD. 2013. Arsenic Inhibits Autophagic Flux, Activating the Nrf2-Keap1 Pathway in a p62-Dependent Manner. Mol Cell Biol 33:2436-2446. PMID: 23589329

Lu Y, Gao J, Zhang DD, Gau V, Liao JC, Wong PK. 2013. Single cell antimicrobial susceptibility testing by confined microchannels and electrokinetic loading. Anal Chem 85:3971-3976. PMID: 23445209

Wu W, Qiu Q, Wang H, Whitman SA, Fang D, Lian F, Zhang DD. 2013. Nrf2 is crucial to graft survival in a rodent model of heart transplantation. Oxid Med Cell Longev 2013:919313. PMID: 23533698

Tao S, Zheng Y, Lau A, Jaramillo MC, Chau BT, Lantz RC, Wong PK, Wondrak GT, Zhang DD. 2013. Tanshinone I Activates the Nrf2-De-pendent Antioxidant Response and Protects Against As(III)-In-duced Lung Inflammation In Vitro and In Vivo. Antioxid Redox Signal 19(14):1647-1661. PMID: 23394605 (This paper was highlight-ed as the top story in Pulmonary Cell News 2.06. February 14, 2013).

Zhang DD. 2013. Bardoxolone Brings Nrf2-Based Therapies to Light. Antioxid Redox Signal 19(5):517-518. PMID: 23227819

Lau A, Whitman SA, Jaramillo MC, Zhang DD. 2013. Arsenic-medi-ated activation of the Nrf2-Keap1 antioxidant pathway. J Biochem Mol Toxicol 27:99-105. PMID: 23188707

Lau A, Tian W, Whitman SA, Zhang DD. 2013. The predicted mo-lecular weight of Nrf2: it is what it is not. Antioxid Redox Signal 18:91-93. PMID: 22703241

Zheng Y, Tao S, Lian F, Chau BT, Chen J, Sun G, Fang D, Lantz RC, Zhang DD. 2012. Sulforaphane prevents pulmonary damage in re-sponse to inhaled arsenic by activating the Nrf2-defense response. Toxicol Appl Pharmacol 265:292-299. PMID: 22975029

Wang F, Tian F, Whitman SA, Zhang DD, Nishinaka T, Zhang N, Jiang T. 2012. Regulation of transforming growth factor beta1-de-pendent aldose reductase expression by the Nrf2 signal pathway in human mesangial cells. Eur J Cell Biol 91:774-781. PMID: 22951256

Bray K, Mathew R, Lau A, Kamphorst JJ, Fan J, Chen J, Chen HY, Ghavami A, Stein M, DiPaola RS, Zhang DD, Rabinowitz JD, White E. 2012. Autophagy suppresses RIP kinase-dependent necrosis enabling survival to mTOR inhibition. PLoS One 7:e41831. PMID: 22848625

Chen W, Jiang T, Wang H, Tao S, Lau A, Fang D, Zhang DD. 2012. Does Nrf2 contribute to p53-mediated control of cell survival and death? Antioxid Redox Signal 17:1670-1675. PMID: 22559194

Riahi R, Yang Y, Zhang DD, Wong PK. 2012. Advances in wound-healing assays for probing collective cell migration. J Lab Autom 17:59-65. PMID: 22357609

Ma J, Cai H, Wu T, Sobhian B, Huo Y, Alcivar A, Mehta M, Cheung KL, Ganesan S, Kong AN, Zhang DD, Xia B. 2012. PALB2 interacts with KEAP1 to promote NRF2 nuclear accumulation and function. Mol Cell Biol 32:1506-1517. PMID:22331464

Lin Z, Yang H, Kong Q, Li J, Lee SM, Gao B, Dong H, Wei J, Song J, Zhang DD, Fang D. 2012. USP22 antagonizes p53 transcriptional activation by deubiquitinating Sirt1 to suppress cell apoptosis and is required for mouse embryonic development. Mol Cell 46:484-494. PMID: 22542455

Hu Q, Zhang DD, Wang L, Lou H, Ren D. 2012. Eriodictyol-7-O-glu-coside, a novel Nrf2 activator, confers protection against cispla-tin-induced toxicity. Food Chem Toxicol 50:1927-1932. PMID: 22465804

Zheng H, Whitman SA, Wu W, Wondrak GT, Wong PK, Fang D, Zhang DD. 2011. Therapeutic potential of Nrf2 activators in strep-tozotocin-induced diabetic nephropathy. Diabetes 60:3055-3066. PMID:22025779 (This paper was featured by Judy B. de Haan in the same issue 2683-4. PMID: 22025774).


Broccoli powder was slightly better than tomato powder, and broccoli and tomato powder together were the most effec-tive at inhibiting cancer growth—a very impressive 52 percent.

Biography: Elizabeth Jeffery, PhD, is a nutritionist at the University of Illinois in Urbana, IL. Her laboratory’s focus is biochemical and nutritional pharmacology, with an emphasis on cancer prevention. Epidemiological studies show that a diet rich in cruciferous vegetables, such as broccoli and Brussels sprouts, can lower the incidence of several cancers, including liver, prostate and colorectal. Jeffery’s experiments identify how components

in crucifers alter the synthesis of detoxification enzymes, resulting in more rapid clearance of deleterious compounds from the body before they can initiate a toxic or carcinogenic response. A major goal is to develop cruciferous vegetables with increased concentrations of chemopreventive agents and she works closely with an interdisciplinary group to meet this goal.

Raw, Cooked, Steamed or Microwaved?Top Tips on Preparing Broccoli and Tomato from Clinical Nutritionist Elizabeth Jeffery

In Focus: What got you inter-ested in studying broccoli?

Jeffery: I used to be a toxicol-ogist and I got very interested in foods that stimulate detoxifica-tion enzymes in the liver. I actu-ally sent a grant proposal to the USDA to study Brussels sprouts, but the grant was turned down, as they said too few people eat Brus-sels sprouts. So I changed the veg-etable to broccoli, and they funded the study. And ever since I’ve been fascinated by broccoli. It’s such an easy vegeta-ble to eat. It’s becoming more and more popular. It’s one of the most effective can-cer-preventing vegetables there is, both because of its levels of Indole 3 carbinol and sulforaphane. It also accumulates selenium.

In Focus: What have your studies re-vealed about eating broccoli raw, ver-sus steamed, microwaved, or cooked?

Jeffery: We’ve done numerous studies, and ultimately we’ve

found that light steaming for 3-4 minutes is ideal. That releases enough of an enzyme that releas-es sulforaphane, without destroy-ing it. Light steaming is more ef-fective than eating raw broccoli, though of course I don’t want to tell anyone not to eat their broc-coli raw. It’s still good for them raw. But steaming makes it even

better. Many people boil their broccoli, but we found in a study earlier this year that the enzyme that releases the sulforaphane is destroyed within one minute of either microwaving or boiling. Steaming is far gentler.

In Focus: So boiled brocco-li doesn’t do the trick? Not even

when you digest it?

Jeffery: There is definitely a discussion going on in the scien-tific literature about whether the microbes that live in your lower gut could release sulforaphane for you. We did two small clini-cal studies to test that, with four male students in each study. None of them were able to form more

than about 10% of the total possible amount of sulforaphane by eating boiled broccoli. However, if we gave them another food in the same family with the same enzyme that

releases the sulforaphane, that helped. For instance, just eating a side dish or salad with wasabi, or radish, will help release sulfora-phane from your broccoli.

In Focus: What about frozen broccoli?

Jeffery: We have been worried about frozen broccoli, since so many people seem to be buying and cooking it. So we are study-


Introduction: Martin L. Pall, PhD, is Professor Emeritus of Biochemistry and Basic Medical Sciences at Wash-ington State University and widely known for his novel theory, dubbed NO/ONOO- cycle (pronounced no-oh-no!). In the cycle, which is based on 34 well-documented mechanisms, elevated levels of nitric oxide (NO) and its highly damaging metabolite, peroxynitrite (ONOO-), are at the crux of a runaway cycle of free-radical damage in which inflammatory molecules are chronically elevated and we have mitochondrial dysfunction as well. Peroxynitrite initiates a complex biochemical vicious cycle, known as the NO/ONOO- cycle, which is responsible for many chronic inflammatory conditions. Pall developed a highly specific and effective antioxidant protocol, called The Pall Protocol, to help reverse the NO/ONOO- cycle and restore health and immune balance. Here, he discusses the remarkable and wholly unexpected bridge he discovered last year between the NO/ONOO- cycle and Nrf2, along with his own dietary choices. His review article on Nrf2 appeared in Acta Physiologica Sinica in February of 2015.1

From NO/ONOO- Cycle to Nrf2:A Remarkable Bridge

An Interview with Martin Pall, PhD, about NO/ONOO- and Nrf2

ing it, and we find that people get virtually no indole-3-carbinol or sulforaphane from the varieties of frozen broccoli we have studied so far. A student of mine named Ed Dosz is writing up this research at the moment. We think the reason is that all across the food industry, vegetables are blanched before being frozen. That kills degra-dative enzymes, and in the case of broccoli, turns it an attractive bright green color. Unfortunately though, the blanching process is far harsher than necessary, and just destroys the enzymes needed to release sulforaphane and in-dole-3-carbinol. We’re working on coming up with some improve-ments in this process for the food industry, but for the meantime, eat a radish in your salad at the same meal.

In Focus: You also did a study on tomatoes and broccoli, and found they work better together.

Jeffery: Yes, I have a colleague named John Erdman who has been studying tomatoes and ly-

copene and prostate cancer for a long time. So we’ve always had a sort of friendly sparring going on. We’d pass in the corridor and I’d say, “Broccoli is better” and he’d say, “No, tomato is better.” So we decided to do a study together. We took animals that were de-veloping prostate cancer and fed them a diet that included either 1) a lycopene supplement 2) tomato powder with the same amount of lycopene as the supplement 3) a lycopene supplement that was ten times stronger 4) broccoli pow-der and 5) broccoli and tomato powder together. We got about 35% protection with the tomato powder, almost no effect with the weak lycopene supplement and a modest effect with the strong lycopene supplement. Broccoli powder was slightly better than tomato powder, and broccoli and tomato powder together were the most effective at inhibiting can-cer growth—a very impressive 52 percent.

In Focus: So the next time you

go to your kitchen, lightly steam your cruciferous veggies, and add a dash of wasabi to flavor, and a salad garnished with radish-es and tomatoes. Or, dribble to-mato sauce on your food. You’re allowed to boil tomatoes, aren’t you? They don’t need light steam-ing like broccoli and other cruci-ferous veggies?

Jeffery: Yes, it’s fine to boil, fry, bake or cook your tomatoes how-ever you like. But remember, ly-copene is lipid soluble. So add a bit of oil to your tomato sauce, or your salad.

Selected BibliographyCramer J and Jeffery EH Sulforaphane Absorption and Excretion Following Ingestion of a Semi Purified Broccoli Powder Rich in Glucoraphanin and Broccoli Sprouts in Healthy Men Nutr Cancer 2011.523495 (2011). PMID: 21240766

Lai R-H., Miller MM, Jeffery EH Glucoraphanin hydrolysis by mi-crobiota in the rat cecum results in sulforaphane absorption. Food and Function 1: 161-166 (2010). PMID: 21776467

Jeffery E.H. and Keck A.S., Translating knowledge generated by epidemiological and in vitro studies into dietary cancer prevention. Mol. Nutr. Fd. Res. 52:S7-S17 (2008). PMID: 18327874

Canene-Adams, K., B.L. Lindshield, S. Wang, E.H. Jeffery, S.K. Clin-ton, J.W. Erdman, Jr. (2007) Combinations of tomato and broccoli enhance antitumor activity in Dunning R3327-H prostate adenocar-cinomas. Cancer Res. 67(2):836-843. PMID: 17213256

Eberhardt, M.V. and E.H. Jeffery. (2006) When dietary antioxidants perturb the thiol redox. J. Sci. Food Agric. 86(13):1996-1998.

Gills, J.J., E.H. Jeffery, N.V. Matusheski, R.C. Moon, D.D. Lantvit, and J.M. Pezzuto. (2006) Sulforaphane prevents mouse skin tumorigen-esis during the stage of promotion. Cancer Lett. 236(1):72-79. PMID: 15993536

Matusheski, N.V., J.A. Juvik, and E.H. Jeffery. (2004) Heating de-creases epithiospecifier protein activity and increases sulforaphane in broccoli. Phytochemistry 65(9):1273-1281. PMID: 15184012

Nho, C.W. and E.H. Jeffery. (2004) Crambene, a bioactive nitrile derived from glucosinolate hydrolysis, acts via the antioxidant re-sponse element to upregulate quinone reductase alone or synergisti-cally with indole-3-carbinol. Toxicol. Appl. Pharmacol. 198(1):40-48. PMID: 15207647


In Focus: You’ve published 31 papers and a 16 chapter book on the NO/ONOO- cycle and related topics. What led you in a different direction, to Nrf2?

MP: The two are actually deep-ly connected, something I learned when I published a paper suggest-ing the NO/ONOO- cycle plays a central role in heart failure. That was 2013.2 But there were a few aspects that still puzzled me, and one was how peroxynitrite inhib-ited the cyclooxygenase (COX) pathway. The COX enzymes trig-ger the production of inflammato-ry molecules such as prostaglan-dins, and though they are critical for many functions they can also be involved in chronic inflamma-tory states. And as I was hunting for an answer, I stumbled onto Nrf2. I discovered that the cyclo-oxygenase pathway actually rais-es the levels of Nrf2 in the cell. So I thought, let me look into Nrf2. I

had no idea what was in store for me. I was com pletely blown away by what I found.

In Focus: Can you summarize what you discovered about Nrf2?

MP: Nrf2 does far more than raise endogenous antioxidants. It has highly significant anti-in-flammatory effects. It improves mitochondrial function. It’s an incredibly important central node of regulation for the body, one where all these health promoting factors come together and pro-duce wide-ranging responses to help protect us from essentially all the chronic inflammatory states. I said to myself, okay, if that’s the overall pattern I’d better look at the details to see how they play out. So I had a flurry of activity where I looked at about a hun-dred papers, and read about fifty of them in depth.

To me, the most exciting aspect of Nrf2 is its connection with the

NO/ONOO- cycle that I’d been working on for so many years. Nrf2 is how evolution helps us avoid NO/ONOO- inflammatory conditions. I’ve published now on 23 different diseases that are linked in some way to NO/ONOO-. But a lot of those publications are nec-essarily somewhat superficial. My best, most detailed paper is prob-ably the one on heart failure. I’m aware that critics of my theory often feel I’m over-reaching, and that NO/ONOO- cannot possibly be involved in all those diseases. It sounds too wide ranging. That’s a reasonable response until you look more closely at the data. You discover that Nrf2 is this incredi-bly complex, highly evolved sys-tem that very effectively quiets the NO/ONOO- cycle. You only get that kind of well-coordinated and highly evolved set of respons-es when nature has decided some-thing is very important.

In Focus: What is the best


Smart About SugarSmart About Sugar

way to use our insights into Nrf2 therapeutically?

MP: I always say Nrf2 is best used for preventing disease. We have less evidence that it can re-verse disease, especially cancer, although we may find that it’s also very useful for treatment of many diseases. I think that diet, exercise and supplements are all useful. (Caveat, in some conditions, such as chronic fatigue syndrome, ex-ercise may not be helpful.) So let me just point out something that nobody else has yet pointed out, to my knowledge. Three of the classes of nutrients that raise Nrf2 actually work through oxidation. Oxidative stress activates Nrf2, and some classes of nutrients ac-tually produce low levels of oxi-dative stress. Those are the active phenolics, the fish oils and Ome-ga 3’s, and the carotenoids. I think it’s important to use those groups specifically in treatment.

In Focus: If the carotenoids are

so important, why do you think beta-carotene hasn’t panned out in supplements?

MP: I believe one reason is that people usually take synthetic beta-carotene. About half of the natural beta-carotene molecules contain a cis double bond, but the synthetic ones do not, so they don’t oxidize as easily. The syn-thetic molecules may not raise Nrf2 at all. That’s my hypothesis, at any rate.

In Focus: What are your favor-ite Nrf2 foods?

MP: I tend to eat things that I know have components that raise Nrf2. I eat a fair amount of carot-enoids including tomatoes and tomato products, a fair amount of onions and garlic, a lot of herbs, and lately I’ve been eating many black olives. I also found a source of the purple sweet po-tatoes that are part of the famed Okinawan longevity diet. Pur-ple sweet potatoes are high in

phenolics (the anthocyanins that give them their deep purple col-or) and in carotenoids. So they’re very healthy. I also eat black rice with lots of phenolics, and Chi-nese broccoli, which has a slight tinge of bitterness but not if you cook it properly.

In Focus: Sounds delicious. So what is your takeaway on Nrf2?

MP: You have to pay close at-tention to what evolution is tell-ing you. Nrf2 is a very complex, incredibly well coordinated re-sponse to protect us from oxida-tive/nitrosative stress, inflamma-tion, mitochondrial dysfunction and toxic chemicals in the body. So evolution is telling us that survival, health and longevity depend on our ability to handle these challenges.

Selected Bibliography1. Pall ML, Levine S. Nrf2, a master regulator of detoxification and also antioxidant, anti-inflammatory and other cytoprotective mechanisms, is raised by health promoting factors. Sheng Li Xue Bao. 2015 Feb 25;67(1):1-18. PMID: 25672622

2. Pall ML.The NO/ONOO-cycle as the central cause of heart fail-ure. Int J Mol Sci. 2013 Nov 13;14(11):22274-330. PMID: 24232452

Sweet SuccessNovel Pathways to Stable Blood Sugar

What’s the first thing that comes to mind when you think about blood sugar? Usually, it’s a magic number.

For physicians, the Holy Grail is a number that lies between 70 and 99

mg/dL (milligrams of glucose for

every deciliter of blood) when fast-

ing. Or it’s that magic “post-pran-

dial” number that hovers below

140 mg/dL two hours after eating.

To epidemiologists–who study

health in the population at large–

blood sugar is a very different

number: 29 million. That’s how

many Americans are current-


ly diagnosed with diabetes. But, like a ghostly afterimage, eight million more diabetics lurk in the shadows: undiagnosed, walking around oblivious to their potential-ly deadly blood sugar problems. And globally, 371 million individ-uals suffer from diabetes, while 4.8 million die of diabetes every year.1

Then there are the 86 million Amer-icans with pre-diabetes. Pre-diabe-tes manifests as insulin resistance, hyperinsulinemia and hypergly-cemia, and can lead to vascular dysfunction, an abnormal lipid profile, hypertension, and vascu-lar inflammation, all of which pro-mote the development of cardiovascular dis-ease.2,3 It is a pro-inflam-matory, pro-thrombotic state. Obese people with metabolic syndrome have a 10-fold increased risk for diabetes and a two-fold increased risk for car-diovascular disease (CVD) relative to normal weight people without the metabolic syndrome.5 Normal weight people with metabolic syn-drome have a four-fold increased risk for diabetes and a three-fold increased risk for CVD.3 Metabol-ic syndrome puts an individual at high risk of full-fledged diabetes, and at increased risk for ocular, renal, neurologic, cardiovascular and metabolic disease—as well as early death.4,5,6

Finally, there’s a number measured only in money. In 2012, when we last looked, the economic burden of diabetes and pre-diabetes ex-ceeded $322 million.7,8

These numbers are powerful and sobering. But the number on a

blood test or census doesn’t take into account the complex orches-tration of sugar storage through-out your body. Diabetes impacts the blood vessels, heart, liver, pancreas, kidneys, muscles, brain and nerves. It takes place in broad strokes (when insulin surges out from the pancreas after we eat a meal, for instance) and in tiny mo-lecular reactions at an individual cell’s insulin receptor.

What follows is an up-to-date look at the newest research into aston-ishing and novel mechanisms of nutrients, each of which works on different pathways to support sta-

ble blood sugar. With scientifically based nutritional intervention, we can actually shift our biology, and end up with more than a number: we can return to good health.

Berberine: A Marvel ofa MoleculeGoldenseal is a perennial and popular herb, with potent antimi-crobial activity familiar to ancient Ayurveda and Chinese medicine, and quantified in numerous stud-ies.9 It is also native to America and was in use by Native Amer-icans when settlers arrived here. It is often used to help treat bac-terial, protozoal and fungal infec-tions. The root and rhizomes con-tain highly concentrated unique alkaloids such as hydrastine, ber-berine, and canadine.

Berberine, particularly in its hy-

drochloride (HCl) form, is emerg-ing as a novel molecule to help treat diabetes and related car-diovascular dysfunction. New research has uncovered its sur-prising ability to impact blood sugar by working on the insulin receptor itself. It also has a po-tent impact on the cardiovascular system, lipids and the liver, sug-gesting that berberine is a potent oral hypoglycemic agent that also has beneficial effects for disorders tightly linked to diabetes.10

Astonishingly, a recent study found that berberine actually in-creases insulin sensitivity through

activation of the insulin receptor itself (InsR).11 It is working at the molec-ular level of the cell as

a truly unique phytochemical. By enhancing InsR sensitivity, ber-berine increases the cell’s glucose consumption. It also up-regulates the expression of the InsR gene in the liver and muscle cells. Rather impressively, in studies of numer-ous cell lines, berberine increased InsR in all of them.15 In animal models, rats with type 2 diabetes were treated with berberine and showed lowered fasting blood glucose and fasting serum insulin, and increased insulin sensitivity. In a human study by the same re-searchers, ninety-seven patients with Type 2 diabetes were given one gram of berberine daily for two months and compared to patients taking either the drugs metformin or rosiglitazone. Berberine therapy (1 g/d for 2 months) lowered fasting blood glucose by nearly 26%, compara-

Berberine lowers fasting blood glucose by nearly 26% with no adverse effects.


ble to that of the drugs, and with no adverse effects. Serum insulin levels dropped by nearly 30% in the berberine group, which indi-cates increased insulin sensitivi-ty. The researchers looked at InsR protein expression on the surface of blood cells by using sophisti-cated flow cytometry, and found an astonishing 3.6-fold increase. That increase was directly cor-related with the reduction in blood sugar.15

Prescription drugs for diabetics can have many adverse effects, depending on the class of drug implemented, including liver ef-fects. Berberine lowers elevated liver enzymes. In fact, it turns out to be an appropriate inter-vention for hyperglyce-mic patients with liver problems. Thirty five chronic hepatitis pa-tients with Type 2 Diabetes were given berberine for two months. Berberine significantly lowered fasting blood glucose in the pa-tients, and at the same time, their elevated liver enzymes declined significantly.15

Berberine has a strong impact on carbohydrate and lipid metab-olism, and a potent effect on the stability of blood glucose. In fact, berberine increases insulin recep-tor expression in cultured human liver cells and skeletal muscle.12 Studies of diabetic rats found their insulin levels and insulin sensitivity normalized when they were treated with berberine. The pancreatic islets (insulin produc-ing cells) were atrophied in dia-betic rats, but less pathological

change was observed in rats treat-ed with berberine. The study con-cluded that “berberine has a pro-tective effect for diabetes through increasing insulin expression, b-cell regeneration, antioxidant enzyme activity and decreasing lipid peroxidation.”13

An impressive human study followed two groups of diabet-ics. The first contained 36 newly diagnosed Type 2 diabetics for three months. They received ei-ther berberine or metformin (0.5 g 3 times a day). Their glycated hemoglobin (a test that estimates the levels of blood glucose over

3 months), fasting blood sugar, postprandial blood sugar (after a meal) and their triglycerides were measured. All parameters im-proved significantly on berberine, and the drop in blood sugar was equivalent to that of metformin. In the second group, 48 individ-uals with poorly controlled Type 2 diabetes were given the same program and experienced im-provement in all measures. In both groups, about a third of the patients experienced transitional gastrointestinal side effects.14

Berberine doesn’t just lower blood sugar. It improves lipid me-tabolism, which often goes astray in metabolic syndrome and dia-betes. Here, too, it is uniquely ef-fective as a phytochemical, acting on the LDL receptor in the liver, as well as other pathways.15 In a

study of 91 individuals with ele-vated cholesterol (52 males and 39 females) 1,000 mg of berber-ine taken daily for three months lowered cholesterol by 18%, tri-glycerides by 28% and LDL by 20%. HDL (the good cholesterol) remained unchanged. Liver func-tion was also improved. A con-trol group, in contrast, showed no change.16,17

Finally, berberine lowers blood pressure in animal studies, and hypertension is another issue in metabolic syndrome and diabe-tes. Berberine acts on both the

endothelium and the vascular smooth muscle through multiple cellu-lar mechanisms.18,19 It also helps patients with acute coronary syn-

drome (ACS)—which is caused by plaque that ruptures, forms clots, and occludes vessels. ACS is usually treated with implantation of a stent to expand the occluded vessel, but stents can cause vas-cular injury and an inflammatory response, so prevention is a key strategy. Numerous studies have shown that diabetics with ACS have a worse outcome. This has been attributed, above all, to the fact that diabetes is a pro-inflam-matory and pro-thrombotic state. In a recent study, 130 ACS patients receiving a stent were followed. 69 patients received standard thera-py, while 61 were given standard therapy plus 300 mg of berberine three times a day for a month. In the berberine group, several im-portant markers of inflammation declined significantly, helping

Berberine taken daily for three months lowered cholesterol by 18%, triglycerides by 28% and LDL by 20%.


protect against the vascular injury caused by stent placement. Ber-berine clearly appears to be clini-cally useful for both diabetes and heart disease.20,21

Truly, berberine, like its mother plant, is golden.

Common Ash: The Liver’sPerfect HelperThe liver choreographs the storage and manufacture (called gluco-neogenesis) of sugar in the body. When we eat, the liver stores sug-ar as glycogen. Between meals and at night, the liver turns that stored glycogen into glucose (called gly-colysis or glycogenolysis). It’s no surprise that a burdened liver be-comes less efficient at managing glucose regulation, par-ticularly in those who are overweight or suf-fering from the increas-ingly common fatty liv-er diseases.

The seeds and fruits of the com-mon ash tree (Fracinus excelsior) seem to have been crafted by na-ture to aid the liver in handling sugar. Long used by Mediterra-nean populations for their potent anti-hyperglycemia and anti-obe-sity effects, science now backs up folklore and reveals many, potent mechanisms of action. F. excelsior contains potent molecules such as coumarins, which thin the blood and increase blood flow in veins; as well as secoiridoids, a class of plant chemicals that can be neu-roprotective, hepatoprotective, cardioprotective, hypoglycemic, hypolipidemic, and antispasmod-ic.22 They also contain phenyletha-

noids, which can be hepatoprotec-tive and antioxidant.23, 24

Extracts of F. excelsior seed display potent hypoglycemic and anti-hy-perglycemic activity in laboratory animals without affecting blood insulin concentrations. The seeds also appear to inhibit the kidneys’ reabsorption of glucose.25 In a 2011 study, laboratory mice were protected from weight gain, high blood sugar and high insulin by an F. excelsior seed extract. Mice were fed a high-fat diet over 16 weeks plus F. excelsior extract, and compared with low-fat and high-fat diet controls. Weight gain, fat deposition, fasting blood glucose, and fasting blood insulin were measured. Mice on the extract

gained 32% less weight, and their fasting blood glucose levels were lowered by 76%. Plasma insu-lin levels dropped by 53%. Liver weight gain was reduced by 63% and fatty liver curbed by 66.67%. The researchers concluded that the extract protects against obesi-ty-related fatty liver.26

The seeds also activate receptors called PPAR-alpha (Peroxisome proliferator-activated receptors). PPAR-alpha is mainly present in the liver, where it is a master reg-ulator of fatty acid oxidation.27 In a variety of mouse models, substances acting on PPAR-al-pha receptors lower plasma tri-

glycerides, reduce fat and de-creases hepatic and muscle fat, improving insulin sensitivity and reducing glucose in blood.28

In humans, F. excelsior reduces the “post-prandial” jump in blood sugar (a rise that occurs after eat-ing). In a 2009 study, a standard-ized extract of F. excelsior seeds was given to volunteers in a dou-ble-blind, randomized, crossover design. Sixteen healthy volun-teers were given either a gram of wheat bran (placebo) or a gram of F. excelsior seed extract after consuming 50 grams of glucose. Blood glucose levels (“postpran-dial plasma glucose”) were mea-sured at 0, 15, 30, 45, 60, 90 and 120 minutes after consuming sugar or

placebo. Insulin levels were measured at 0, 30, 60, 90 and 120 minutes. The extract significantly lowered glucose at both 45 and 120 minutes.29

Finally, in an unpub-lished study, 22 overweight indi-viduals aged 50-80 were studied for a total of 7 weeks in a random-ized, crossover, and double-blind nutritional intervention, with a one week wash-out period be-tween two separate intervention periods of 3 weeks each. Experi-mental groups were administered 1 gram daily of either placebo capsules or capsules containing an extract from Fraxinus excelsior L. seeds. The seeds significantly lowered the glucose “curve” (the speed and amount that the blood sugar rises). The researchers con-clude: “The administration of an extract from Fraxinus excelsior L.

Extracts of F. excelsior seed display potent hypoglycemic and anti-hyperglycemic activity without affecting blood insulin concentrations.


seeds in combination with a low calorie diet induced favorable effects in glucose homeostasis in overweight/obese subjects.”30

Golden Oldies Offer upNew Scientific ProofSilymarin, from the herb milk thistle, is well known for its liver protective properties. It has been extensively studied for inflam-mation and as an adjunctive in cancer treatment, yet much less is known about its therapeutic effect on diabetes. It turns out that sily-marin helps regulate glucose and protect against related liver and cardiac dysfunction.

In a four-month, randomized, double-blind clinical trial it proved highly beneficial in im-proving the blood sugar profile of Type 2 diabet-ics. The study looked at 51 diabetics, divided into two groups. The first group of 25 diabet-ics received 200 milligrams of si-lymarin 3 times a day, along with conventional therapy. The second group of 26 diabetics received a placebo. Each month the research-ers tested for glycosylated hemo-globin (HbA1c), fasting blood glucose, insulin, total cholester-ol, LDL and HDL, triglycerides, and liver enzymes such as SGOT (AST) and SGPT (ALT). Those on silymarin had a significant de-crease in all but the “good” cho-lesterol (HDL).31

Diabetic nephropathy is a leading cause of end-stage kidney disease. It is usually managed by trying to control blood sugar, blood pres-

sure and proteinuria with med-ications. Silymarin reduces uri-nary excretion of albumin, tumor necrosis factor alpha (TNF)-α, and a toxic substance called malondi-aldehyde in patients with diabetic nephropathy, according to a recent review article. Malondiadelhyde is a particularly useful indicator, since it is a naturally occurring byproduct of lipid peroxidation, and is both mutagenic and car-cinogenic. It is a commonly used, accurate indicator of lipid peroxi-dation. This suggests the kidneys are functioning better, since, for example, an increased loss of al-bumin in the urine is caused by renal dysfunction.32 Another year-long study of two-well matched groups of diabetics (30 patients

each) with alcoholic cirrhosis showed remarkable results from silymarin. All the patients were being treated with insulin, but 30 patients received only standard therapy, while another 309 re-ceived standard therapy plus 600 mg silymarin per day. After only four months of treatment, the sily-marin group demonstrated a sig-nificant decrease in fasting blood glucose levels, mean daily blood glucose levels, daily glucosuria and HbA1c levels. The silymarin group also had decreased fasting insulin levels and needed a lower insulin dosage, while the untreat-ed group showed a significant increase in fasting insulin levels

and a stabilized insulin need. Finally, the treated group had a significant decrease in malondi-aldehyde levels compared to the treated group.33

Finally, in an ingenious study, berberine and silymarin were combined. Sixty-nine patients with Type 2 diabetes were treated with the standard approach: diet, hypoglycemic drugs, and when necessary, lipid-lowering medi-cations. All patients also received a supplement: either a standard-ized extract of berberine (1000 mg a day), or a combination of berberine and silymarin (1000 mg a day and 210 mg a day re-spectively). Both treatments im-proved fasting glucose, total cho-lesterol, LDL, triglycerides and

liver enzyme levels. However, the combina-tion formula reduced glycosylated hemoglo-bin (HbA1c) values to a greater extent. The

researchers conclude that the combination is more effective for Type 2 diabetic patients.34

Like silymarin, resveratrol is widely known as a potent anti-oxidant, and particularly known for its cardioprotective effects. Its impact on diabetes is now emerg-ing. A 2013 study found that res-veratrol supplementation exert-ed strong anti-diabetic effects in patients with type 2 diabetes. Sixty-six individuals with Type 2 diabetes were randomly assigned to a group that received either a gram of resveratrol daily, or a gram of placebo. Body weight, blood pressure, fasting blood

A combination of silymarin and ber-berine improves fasting glucose, total cholesterol, LDL, triglycerides and liver enzyme levels.


glucose (FBG), hemoglobin A1c (HbA1c), insulin, insulin resis-tance, triglycerides, total choles-terol, LDL, HDL, and markers of liver and kidney damage were measured at the beginning of the study and after 45 days. Res-veratrol treatment significantly decreased fasting blood glucose, HbA1c, insulin, and insulin re-sistance. HDL was significantly increased. Liver and kidney func-tion markers were unchanged.35

Resveratrol has also proved re-markably helpful in obese patients with Type 2 diabetes. In one 2011 study, 11 obese males received a daily dose of 150 mg resveratrol or placebo for 30 days in a double-blind crossover design. Significant im-provements were seen in insulin sensitivity, blood pressure, metabolic rate, and fatty liver.36

Resveratrol is thought to mimic the benefi-cial effects of caloric restriction on glucose regulation and health by activating sirtuins, a group of powerful enzymes that are key to maintaining cellular homeostasis. Sirtuins are increasingly being recognized for their role in dia-betes, especially in protecting the heart. Caloric restriction leads to overexpression of sirtuins—and improved health and longevi-ty. Increasing the cell’s store of NAD+, a molecule that is now at the center of a hotbed of anti-ag-ing research, also stimulates sir-tuin activity, and this may be the mechanism of action of resver-atrol. Sirtuins influence glucose

metabolism in the liver, pancreas, muscle and fat. There is evidence they can improve insulin sensi-tivity and mitochondrial func-tion.37 In a 2014 study, resveratrol was shown to activate one of the many sirtuins and improve cardi-ac function and fibrosis in mice, as well as in heart cells in vitro.38

Vitamins, Minerals, and DietAre PowerfulKey vitamins and minerals bring up the rear guard in natural constituents that help maintain healthy blood sugar. Chromium, biotin (a B vitamin) and magne-sium are all known to improve glucose regulation. In a recent

study on 43 individuals with poorly controlled blood sugar in spite of medication, a combination of biotin and chromium picolinate led to significant improvement in the blood sugar curve, as well as reductions in triglycerides and LDL.39 In another study, 447 adults with poorly controlled Type 2 di-abetes were given either a com-bination of chromium picolinate (600 micrograms) with biotin (2 mg), or a placebo. Both groups were given oral anti-diabetic medications as well, and followed for 90 days. Glycosylated hemo-globin (HbA1c), fasting glucose, and lipids were measured. HbA1c

in the chromium picolinate/bio-tin group decreased by over 50% and fasting glucose levels were re-duced in the entire chromium pi-colinate/biotin group versus pla-cebo.40 In addition, niacin-bound chromium (known as chromium polynicotinate), has a positive im-pact on lipids and blood sugar. In one randomized, double-blind, placebo controlled study of 40 individuals with high cholesterol (total cholesterol 210-300 mg/dL), a combination of grape seed pro-anthocyanidin extract (GSE) and chromium polynicotinate (CP) was given for two months. It was significantly more helpful than placebo at lowering LDL levels.41

Finally, magnesium is very well tolerated and effective in diabetes, and plays an import-ant role in insulin sen-sitivity and homeosta-sis. When 50 patients with Type 2 diabetes

were analyzed for fasting and post-prandial glucose, HbA1c and magnesium levels, serum magne-sium levels were found to decline with rise in HbA1c levels and with duration of the disease.42 In other research, Type 2 diabetic patients show lower serum magnesium levels than non-diabetic patients.43 Not surprisingly, magnesium sup-plementation has beneficial effects on blood glucose, lipid profile, and blood pressure in patients with Type 2 diabetes. In a 2014 double-blind, placebo controlled study, 54 diabetic patients were followed for 3 months. They re-ceived either 300 mg of magne-

Resveratrol treatment significantly decreases fasting blood glucose, hemoglobin A1c, insulin, and insulin resistance. Resveratrol is thought to mimic the health benefits of caloric restriction, working on the same pathways.


sium sulfate daily, or a placebo. Daily administration of mag-nesium significantly improved fasting blood glucose, 2-hour post prandial glucose, lipid profile, blood pressure and he-patic enzymes.44

No discussion of diabetes is com-plete without a nod to the over-whelming evidence that diet modification, weight loss, and ex-ercise can stabilize and often re-verse the condition. A meta-anal-ysis of ten studies of physical activity and Type 2 diabetes re-ported a lower risk of developing diabetes with regular moderate physical activity, including brisk walking, compared with being sedentary.45 Losing weight can substantially improve blood sug-ar control in Type 2 diabetes and prevent progression from pre-di-abetes to Type 2 diabetes. This is shown in meta-analyses of tri-als that look at exercise, diet and standard drug therapy.46,47,48

A profound change in diet can ac-tually reverse Type 2 diabetes in as little as 8 weeks, according to a 2011 study.49 Eleven people with Type 2 diabetes were followed for 1, 4 and 8 weeks on a restrictive, 600 calorie a day diet. Liver glu-cose output, liver and peripheral insulin sensitivity, and pancreatic beta cell function were measured. An age-, sex- and weight-matched group of eight non-diabetic par-ticipants studied. Blood glucose normalized in one week, and by the end of the study, all other measures had normalized as well. Though this highly restricted diet would be virtually impossible for

most folks to follow long term, it does demonstrate the power of dietary change.

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30. Zulet MA, Navas-Carretero S, Lara Y, Sánchez D, Abete I, Roller, M, Issaly N, Flanagan J, Martínez J. A Fraxinus Excelsior L. Seed Extract, Glucevi, Has Favorable Effects on Glucose Ho-meostasis in Elderly Overweight Subjects. Dpt of Nutrition, Food Science and Physiology, University of Navarra Pamplona, Spain.

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33. Velussi M, Cernigoi AM, De Monte A, Dapas F, Caffau C, Zilli M. Long-term (12 months) treatment with an anti-oxidant drug (silymarin) is effective on hyperinsulinemia, exogenous insulin need and malondialdehyde levels in cirrhotic diabetic patients. J Hepatol. 1997 Apr;26(4):871-9. PMID: 9126802

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35. Movahed A, Nabipour I, Lieben Louis X, Thandapilly SJ, Yu L, Kalantarhormozi M, Rekabpour SJ, Netticadan T. Antihypergly-cemic effects of short term resveratrol supplementation in type 2 diabetic patients. Evid Based Complement Alternat Med. 2013; 013:851267. PMID: 24073011

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39. Singer GM, Geohas J. The effect of chromium picolinate and biotin supplementation on glycemic control in poorly controlled patients with type 2 diabetes mellitus: a placebo-controlled, double-blinded, randomized trial. Diabetes Technol Ther. 2006 Dec;8(6):636-43. PMID: 17109595

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41 Preuss HG, Wallerstedt D, Talpur N, Tutuncuoglu SO, Echard B, Myers A, Bui M, Bagchi D Effects of niacin-bound chromium and grape seed proanthocyanidin extract on the lipid profile of hypercholesterolemic subjects: a pilot study. J Med. 2000;31(5-6):227-46. PMID: 11508317

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Practice PointsA Step by Step Guide to Testing and

Treating Impaired Glucose RegulationBy Dr. Todd A. Born, ND

Diabetes is a silent killer. Undi-agnosed pre-diabetics and diabet-ics have no idea they have blood sugar problems. They may have had uncontrolled glucose (blood sugar) for years before they go to the doctor with obvious symp-toms (e.g. increased thirst, feet pain, wounds that don’t heal well, vision problems, etc.) In the early stages of diabetes, metformin is typically the drug of choice. It is easy to take, fairly well tolerated, and works well. As insulin resis-tance progresses, so do the medi-cations that are initiated. Once the beta cells (insulin producing) of the pancreas become irreparably damaged, exogenous insulin is required-most often for life. If the glucose and insulin resistance is not controlled, end-organ damage occurs, such as nephropathy, neu-ropathies and retinopathies.

A more holistic and integrative ap-

proach to diabetes is far more ef-fective at preventing and reversing the condition. Here are examples of some step-by-step guidelines.

• Use glycated hemoglobin as a screening test, along with fast-ing glucose and insulin, in any adult at high risk for diabetes (particularly those with metabolic syndrome). Glycated hemoglobin is sugar bonded to your hemo-globin, and is measured by a lab test called HbA1C. The results in-dicate the average level of blood sugar (glucose) over the previ-ous 3 months. Though this test is commonly used by physicians to monitor active diabetes, it is also useful as a screening test for adults in pre-diabetes or at risk of diabetes. There is an increased risk of developing diabetes when the HbA1C levels are 5.7% to 6.4%. Normal is < 5.7%.1

• Use the c-peptide test when in-

dicated. C-peptide is a surrogate marker that tells us how well a di-abetic on insulin is doing—wheth-er their beta cells are still function-ing well enough to use the insulin. If insulin is elevated and C peptide is normal, we know the pancreas is still working (when not taking exogenous insulin). If c-peptide is low, we can infer some pancreat-ic insufficiency,2 which is a much more perilous situation.

• Educate your patients, so that they understand that diabetes and pre-diabetes are a systemic phenomenon of impaired glucose tolerance (in part due to insulin resistance) which includes the liv-er3 and muscle4, not just the pan-creas and their beta cells. Both the liver and muscle store and release glycogen. Though the standard American diet (SAD) is a diet high in simple carbohydrates that are nutrient depleted and calorie

Biography: Dr. Todd A. Born, ND is a naturopathic doctor, and co-owner and medical director of Born Naturopathic Associates, Inc., in Alameda, California. He is also Product Manager at Allergy Research Group, LLC, and a Thought Leader for the UK-based “Clinical Education,” a free peer-to-peer forum for clinicians sponsored by Nutri-Link, a subsidiary of Allergy Research Group. Dr. Born graduated from Bastyr University in Seattle, and completed his residency at the Bastyr Center for

Natural Health and its thirteen teaching clinics, with rotations at Seattle-area hospitals. Dr. Born’s clinical focus utilizes integrative medicine to treat chronic disease and refractory cases, supported by the basic medical sciences, including physical medicine (osseous manipulation, craniosacral therapy, hydrotherapy and physiotherapy), botanical medicine, homeopathy, biotherapeutic drainage, Ayurveda, counseling, pharmacology, and diet and nutrient therapies.


dense-a primary trigger of diabe-tes, it is due to more than diet. It’s due to our modern lifestyle: sed-entary, with the constant stress of overwork, long commutes, and financial burdens that impact on numerous genes encoded for de-fense responses, that in turn are no longer mitigated by appropri-ate physical and mental activities.

• Understand the role of stress. When we are chronically stressed, our body assumes there is danger afoot. It releases cortisol, which tells the liver to make extra sugar (gluconeogenesis).5 The cortisol also tells our body to make the neurotransmitters norepineph-rine and epinephrdine, which go instantly to the muscles to help them run from danger if neces-sary. The muscles begin to break down their stored sugar, glycogen (glycolysis),6 to fuel the flight. But in reality, we are simply sitting at our desks, highly stressed, and sedentary. All that excess cortisol, adrenaline, and sugar is flooding our body for no legitimate reason at all. In fact, we may reach for the

nearest comfort food—the cookie or potato chip at that point, high-ly refined, quick to act on key re-ceptors and well known for com-pounding the problem.7

• Craft a lifestyle program. Once our patients, and clinicians, un-derstand the many causes and contributions to diabetes, we need to help them with a program of cognitive reframing, stress reduction, mindfulness medita-tion, regular exercise (whether walking, jogging, swimming, or sports, but must also include weight bearing exercise), and time off for play and rest—as well as dietary remodeling.

• Choose combination formu-las of natural medicines. Rec-ognize that although nutritional interventions and supplements can provide powerful support to pre-diabetics and diabetics, many patients can suffer from supple-ment burnout if they have to take too many pills, too many times a day. Try to look for combination formulas that support many dif-

ferent systems in the body and work on different mechanisms, leading to a healthy blood sugar level without an onerous sup-plement schedule. These include liver support, nutrients and mol-ecules that sensitize insulin re-ceptors, molecules that help pro-tect the heart, and much more. Try to lean toward supplements that work on many mechanisms at once (such as resveratrol and milk thistle).

• Keep abreast of new literature on supplements. See accompany-ing article for the newest research on novel uses of safe supplements that impact blood sugar.

References:1. Wisse, B. A1C Test. MedlinePlus. Aug 2014. http://www.nlm.nih.gov/medlineplus/ency/article/003640.htm

2. Unknown. C-peptide. Lab Tests Online. June 2014. http://labtestsonline.org/understanding/analytes/c-peptide

3. UCSF. The Liver & Blood Sugar. http://dtc.ucsf.edu/types-of-diabetes/type1/understanding-type-1-diabetes/how-the-body-processes-sugar/the-liver-blood-sugar/

4. DeFronzo RA, Tripathy D. Skeletal muscle insulin resistance is the primary defect in type 2 diabetes. Diabetes Care. 2009 Nov;32 Suppl 2:S157-6 PMID: 19875544

5. Khani S & Tayek JA. Cortisol increases gluconeogenesis in hu-mans: its role in the metabolic syndrome. Clin Sci (Lond). 2001 Dec;101(6):739-47. PMID: 11724664

6. Raz I, Katz A, Spencer MK. Epinephrine inhibits insulin-medi-ated glycogenesis but enhances glycolysis in human skeletal mus-cle. Am J Physiol. 1991 Mar;260(3 Pt 1):E430-5. PMID: 1900669

7. http://www.hsph.harvard.edu/nutritionsource/carbohy-drates/carbohydrates-and-blood-sugar/

The insulin receptor (InsR) is essential for the action of that all important hormone—insulin—throughout our body. Like all cell surface receptors, it helps ferry messages from the outside of the cell to the inside. Binding of insulin to InsR in the liver, muscles, or fat activates multiple cellular pathways that cause glycogen synthesis and glucose uptake increase, as well as reducing the amount of glucose the liver and muscles put out. Via the InsR, the blood glucose can be quickly lowered, keeping the entire body stable and healthy. Type 2 diabetes is a hyperglycemic state characterized by insulin resistance in many tissues, particularly the liver, muscles, adipocytes, and pancreatic beta cells. Eventually the insulin resistance leads to pancreatic β-cell burnout, where the cells actually start to die. Individuals with insulin resistance have either decreased levels or absence of InsR expression, or the receptor fails to activate an important substance called tyrosine kinase, which triggers a series of important cellular responses. Clearly, the InsR is the supremely important eye of the tornado we call diabetes. Upregulating and optimizing the InsR and restoring its homeostatic capacity is a key goal when treating and reversing diabetes.

A Quick Guide to the Essential Insulin Receptor


Histamine is a natural molecule that, when in balance, is extremely important for maintaining health. It functions as a neurotransmitter, and helps regulate stomach acid, muscle contraction, brain function, and the permeability of blood ves-sels. It assists our immune system in defending itself against patho-gens. Histamine is stored within special immune system cells called mast cells, along with a smaller amount that is stored in basophils. When the im-mune system is activat-ed in response to foreign material entering the body, histamine is the first inflammatory mol-ecule that is released, as a kind of call-to-arms. Our body metabolizes endoge-nous histamine via a naturally-oc-curring enzyme called Histamine N-methyl transferase (HMT). HMT appears to work on the his-tamine receptors in our body.

Histamine is also present in many foods and some food additives (such as tartrazine, sulfite, or ben-zoate). Cheese, alcoholic bever-ages, vinegar, fermented vegeta-bles, soy sauce, processed meats, strawberry and raspberry, toma-toes, apricots, cherry, plums, and pumpkin are all common sources of dietary histamine. Other foods, such as egg white, seem to release histamine. Most healthy people can easily detoxify food sources of the molecule via an enzyme

called Diamine oxidase (DAO). DAO breaks down excess dietary or gut histamine, and excretes the breakdown products through the kidneys into the urine.1

However, if an individual is nat-urally low in DAO, eats too many histamine-rich foods at once, or suffers from an imbalanced gut microbiome that tends to pro-duce excess histamine, he or she may experience diarrhea, head-

ache, stuffy nose, asthma, hives, low blood pressure, tissue swell-ing and throat tightening, racing heart, anxiety, fatigue, and other unpleasant symptoms. In addi-tion, chronically elevated levels of histamine will intensify allergic responses. In histamine-sensitive patients with reduced DAO activ-ity, symptoms can take up to eight hours after ingestion of a food to appear, leading to chronically cy-cling symptoms that appear mys-terious and disabling.2,3,4,5

Histamine intolerance is not a classical allergy, where the pres-ence of an antigen, such as flower pollen, results in subsequent an-tibody (usually IgE) release plus the release of our own natural

histamine, along with symptoms. Yet the rise of traditional allergies impacts histamine intolerance, by increasing the total load of hista-mine. The prevalence and intensi-ty of pollen is on the rise in many areas, because of global warming and pollution. According to a 2010 report by the National Wildlife Federation, “Unchecked global warming will worsen respirato-ry allergies for approximately 25

million Americans. Rag-weed—–the primary al-lergen trigger of fall hay fever—–grows faster, produces more pollen per plant, and has high-er allergenic content under increased carbon dioxide levels. Longer

growing seasons under a warmer climate allow for bigger ragweed plants that produce more pollen later into the fall. Springtime al-lergies to tree pollens also could get worse. Warmer temperatures could allow significant expansion of the habitat suitable for oaks and hickories, which are two highly allergenic tree species. Changing climate conditions may even affect the amount of fungal allergens in the air.”6 And as pollution increas-es, our allergies worsen, as well. Humans tend to produce more al-lergic responses in the presence of pollution, such as diesel fumes.7,8 No wonder that, these days, hista-mine excess and histamine intoler-ance are genuine concerns.

A low histamine diet, DAO and targeted probiotic supplementation may be very useful tools in the growing problem of histamine intolerance in a high-pollen, high-pollutant world.

Histamine Intolerance: The Hidden Culprit in Many Health Problems


Vertigo

Hypotonia,Hypertension

Anaphylaxis

Arrhythmia

Pruritus

Flush

Urticaria

Congestion of the noserhinorrhea, sneezing Bronchoconstriction, dyspnea Dysmenorrhea Bloating

Stomach ache,cramps

Headache Nausea, vomiting

Mast cell secretion

Diarrhea

Circadian rhythm, arousalContribution to the

regulation ofbody temperature,

food intake,locomotion,

learning, memoryCentralnervous system

Neurotransmitterrelease

Regulation ofhematopoiesis

cGMP

cAMP

Gastric acidsecretion

Smooth musclecontraction

Mucussecretion

EndothelialPermeability

Stimulation ofnociceptionnerve fibers

Tachycardia,Arrhythmias

Estrogen

VasodilationCardiovascularsystem

Respiratorytract

Bone marrow

Leukocytes

Histamine

Skin

Uterus

Gastrointestinal Tract

Histamine

Diagram adapted from: Histamine and histamine intolerance, by Laura Maintz and Natalija Novak, American Journal of Clinical Nutrition, May 2007.

One simple answer may be a long-term, low-histamine diet.9 This can also be supplemented with DAO, available as an oral enzyme. DAO can be seen as a first aid treatment for people who are aware of their difficulty in successfully breaking down and eliminating endogenous and ex-ogenous histamine, after suitable confirmation by blood and urine tests. However, many people pres-ent to their clinicians with a series of mild to moderate immune re-sponses to environment and food selections that defy easy analysis and diagnosis. A trial with the enzyme DAO may prove to be a quick and economic way to deter-mine if symptoms improve – fur-ther investigations may also then

be ordered, as well as the start of a low histamine lifestyle.

DAO can even be taken by nor-mal individuals before a very high-histamine meal (pizza, rich in tomatoes and cheese, along with red wine, for instance). In addition, since gut microbes can produce histamine, oral probi-otics may help lower histamine intolerance.10 The probiotic Lac-tobacillus rhamnosus, for instance, downregulates the Histamine 4 receptor.11 Thus, select probiot-ic bacteria might diminish his-tamine-mediated, mast cell al-lergy-related activation. A low histamine diet, DAO and proper probiotic supplementation may be very useful tools in the grow-

ing problem of histamine intoler-ance in a high-pollen, high-pol-lutant world.

References:1 Maintz L, Novak N. Histamine and histamine intolerance. Am Jnl Clin Nutr, 2007. PMID: 17490952

2 Jarisch R, Wantke F. Wine and headache. Int Arch Allergy Im-munol 1996;110:7–12. PMID:8645981

3 Steinbrecher I, Jarisch R. Histamin und Kopfschmerz. (Hista-mine and headache. ) Allergologie 2005;28:84–91 (in German).

4. Lessof MH, Gant V, Hinuma K, Murphy GM, Dowling RH. Re-current urticaria and reduced diamine oxidase activity. Clin Exp Allergy 1990;20:373– 6. PMID 2115815

5 The Pivotal Role of Histamine in the Symptoms of Food Intoler-ance.Janice M. Joneja, Ph.D., Powerpoint Lecture

6 http://www.nwf.org/pdf/Reports/NWF_AllergiesFinal.pdf

7 Brandt EB, Kovacic MB, Lee GB, Gibson AM, Acciani TH, Le Cras TD, Ryan PH, Budelsky AL, Khurana Hershey GK. Diesel exhaust particle induction of IL-17A contributes to severe asth-ma. J Allergy Clin Immunol. 2013 Nov;132(5):1194-1204.e2. doi: 10.1016/j.jaci.2013.06.048. PMID: 24060272

8 Plé C, Chang Y, Wallaert B, Tsicopoulos A. Environmental pol-lution and allergy: immunological mechanisms]. Rev Pneumol Clin. 2013 Feb;69(1):18-25. PMID: 23333049

9 Wantke F, Gotz M, Jarisch R. Histamine-free diet: treatment of choice for histamine-induced food intolerance and supporting treatment for chronic headaches. Clin Exp Allergy 1993;23:982–5. PMID: 10779289

10 Joneja, J. Histamine Intolerance, Diamine Oxidase Activity, and Probiotics. June 2004. http://www.allergynutrition.com/journal-type/histamine-sensitivity/

11 Oksaharju A, Kankainen M, Kekkonen RA, Lindstedt KA, Ko-vanen PT, Korpela R, Miettinen M. Probiotic Lactobacillus rham-nosus downregulates FCER1 and HRH4 expression in human mast cells. World J Gastroenterol. 2011 Feb 14;17(6):750-9. PMID: 21390145


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IN FOCUS publishes emerging nutritional science and scientific theories that should not be construed to be conclusive scientific proof of any specific cause, effect, or relationship. The publication is for the educational use of healthcare practitioners and physicians. The articles in the publication are the independent scientific views and theories of the authors. IN FOCUS takes no position on the views and theories expressed but offers them for candid inquiry and debate. The articles are not intended for use in support of the sale of any commercial product and should not be construed as indicative of the use or efficacy of any commercial product. Emerging science and scientific theories do not constitute scientific proof of any specific cause, effect, or relationship. Copyright © 2015 NutriCology®. Special permission is required to reproduce by any manner, in whole or in part, the materials herein contained.

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