Interview with
GeriGene Founder Don Kleinsek, Ph.D.

Optimizing Mitochondrial Function
and the Gene Connection
Part I

Recently, at a worldwide meeting of antiaging physicians and scientists in Las Vegas, Don Kleinsek, Ph.D., presented a talk on pharmaceutical targets for age-related disease, based on gene discovery, that included a skillful defense of the cross-link theory of aging, originally created by life-extension pioneer Dr. Johan Bjorksten. Through his association with Bjorksten and his work as president and director of the Bjorksten Research Foundation, Dr. Kleinsek has come into contact with an innovator, whom two-time Nobel laureate Linus Pauling called "one of the most active and effective students of longevity in the world."

Kleinsek's degrees, in chemistry, physiological chemistry, cell biology, hormone action and molecular genetics, have prepared him to go on and investigate new possibilities for molecular, biological and genetically based therapies. Among his advances to date are the discovery and product development of cholesterol-lowering agents and proof of altered gene expression during cell senescence. His firm, GeriGene Medical Corp, is a front-runner in the area of gerontological genetics - how to prolong life through genetic engineering. In an exclusive interview with Life Enhancement's Will Block, Dr. Don Kleinsek talks about what can be done now, while we prepare for the arrival of genetic-based therapies. As Don substantiates, there are powerful reasons to make the most of dietary supplements that the unfolding genetic revolution is helping us to understand more fully and use more intelligently.

When the genetic paradigm starts to take hold (re: genetic supplements), it is important that your mental and physical health be as enhanced and/or intact as possible, otherwise you could miss the boat. As changes occur, it is imperative that one take advantage at the earliest time. Already there are things, to which genetics lends support, that may portend your course of action over the immediate future. This issue was our concern as we began the following interview, which took place by phone in January 1998 between Bogota, Columbia (one of a number of places where Don carries out his research) and Petaluma, California.

WILL: How did you get interested in gerontology and life extension?

DON: Curiosity and confronting the reality of death. When I was 16, both my grandparents died at the age of 70 and I thought they had a lot of life left. But they passed away from our current number one killer in the United States - heart disease and atherosclerosis. It struck me as odd that one is born with no options in terms of life and death. You're born to die. And a lot of good people go through the life cycle and then they die. And everybody just accepts it. It just seemed odd. So basically, I think a philosophy of countering nature's way, the life and death cycle, prompted me into this area.

But the question that really inspired me was, "Can we do something about aging?" There's no sense in working in an area where there's nothing that can be done. Although it has been a source of interest to mankind throughout the ages, when I first became interested in the sixties, the science looked pretty bleak. The cross-linking and free-radical theories [of aging] were strongly postured, the involvement of hormones, calories and nutrition were suspected, but there was nothing about genetics. The data was all very weak and the field fragmented. I thought what needed to be done to properly approach this field was to get a very strong background in a number of disciplines - technical background - so that I would have tools to work with to approach gerontology.

"In five to ten years if not sooner ... we will understand all of the biochemical reactions of aging by first understanding the fundamental level of the gene."

Therefore I earned several degrees. I started out with chemistry. Chemistry is a great premise for anything in cell biology or genetics. From there, I went into biochemistry and cell biology, hormone action, physiology and genetics. I wanted to get a very strong background - and by strong I mean not only doing the homework or studying these disciplines, but the actual lab bench work in those areas, which I've done. It became apparent that by cross-breeding disciplines, new ideas are promoted and often this makes all the difference in understanding how to approach and solve problems.

As an example, early on in my career, there was a pressing problem in the cholesterol field. The purification of an enzyme that was responsible for cholesterol synthesis could not be accomplished. The enzyme was very sensitive to a number of parameters. So I blended enzymology with physical chemistry, not normally done at that time, integrating pertinent thoughts, techniques and information from the two disciplines. I then took it one step further to develop a hybrid technology that allowed me to discover how to isolate the enzyme at the very high purity required. This provided the premise for the development of the statin inhibitors, which lower cholesterol in the serum. This is an outstanding example of how discipline cross-breeding can promote a new technology that didn't exist before.


Fight Aging
With Mitochondrial Enhancement
By Gail Valentine, D.O.

Don't look now, but our bodies have been invaded by aliens! And if that weren't bad enough, we can't live without them (nor can they live without us). If we don't provide them with the nutrients they need to function, we will become ill and eventually die.

These "aliens" are called mitochondria, and many scientists believe they are independent creatures with their own DNA that are descended from ancient symbiotic bacteria. Somehow, billions of years ago during the earliest stirrings of life on this planet, these primitive organisms insinuated themselves into the first living cells and have remained an essential part of cellular structure ever since. Every living cell contains at least some mitochondria, and some cells, such as those that comprise heart muscle, contain enormous numbers of them.

What is it that mitochondria bring to the living organism that makes them so valuable? Quite simply, energy. These microscopic sausage-shaped structures are the cell's power plants. They take the high-energy biological fuels, carbohydrates and fatty acids, and convert them into cellular energy in the form of ATP via a process known as oxidative phosphorylation.

Given this vital function, it stands to reason that mitochondrial dysfunction can have profound effects on virtually every body system, a fact that science is just beginning to understand. Evidence now indicates, for example, that mitochondrial defects, many of which are age-related, are at least partly responsible for Parkinson's disease, Alzheimer's disease, heart disease, fatigue syndromes, and a variety of genetic syndromes.

It is also becoming apparent that at least some types of mitochondrial dysfunction may be due to a failure to supply them with enough of the nutrients they need to carry out their energy-generating function. The converse may also be true: By supplying adequate amounts of these substances, it may be possible to protect against, or even treat, some serious degenerative diseases. Nutrients identified as being vital to mitochondrial function include:

•Carnitine, which transports long-chain fatty acids into mitochondria so they can be "burned" for fuel and used to make a substance called cardiolipin. Cardiolipin is a special phospholipid that resides in the inner mitochondrial membrane and provides structural support to several important enzymes. Both cardiolipin levels and mitochondrial efficiency decline with age. Studies in animals indicate that supplemental acetyl-L-carnitine can reverse this cardiolipin deficiency and help restore mitochondrial energy production as well.

•Coenzyme Q10, which plays a crucial role in electron transport within mitochondria. CoQ10 deficiencies have been linked to several pathologic states, including cardiomyopathy, a serious disease of the heart muscle that can result in congestive heart failure. Restoration of CoQ10 is often successful in improving this condition.

•Lipoic acid, an essential cofactor in the citric acid cycle, one of the major energy-producing activities of mitochondria. Lipoic acid is also a powerful antioxidant that scavenges both water-soluble and lipid-soluble free radicals.

•DHEA, an essential hormone which declines with age, can help increase transport of carnitine into the mitochondria.

•Medium-chain triglycerides (MCTs), while not essential for mitochondrial function, can be very helpful, because they can be easily transported across mitochondrial membranes and used as a source of fuel for the rapid generation of energy.

•Phosphatidylserine, an essential component of the cell membrane, increases membrane strength, permeability, elasticity, and resistance to stress, all of which are of critical importance in the trafficking of lipids between intracellular organelles, including the mitochondria.

WILL: Didn't this work result in one of the principal mainstream cholesterol-lowering drugs?

DON: That's right. This discovery is the basis for 35 patents on statins, all of which credit this work. This work also led to the control of cholesterol synthesis by inhibition of the enzyme that is the rate-limiting step. It is the basis for the drugs marketed by Merck, Bristol Myers and Rhone-Poulenc, and is now a six billion dollar a year market in the United States. The statins have been shown to lower the chance of heart attacks by 50%. For people in high-risk categories this is significant. I've personally known patients, with cholesterol counts at 325, who were put on lovastatin, simvastatin or pravastatin for a couple of months, to go down to 150. So it helped remove them from the risk group for heart disease, at least when cholesterol was the culprit, as it is in a large percent of cases.
The strategy to use now, rooted in the genetics of aging, is identical to what was accomplished earlier at the lab bench - the cholesterol discovery that has resulted in a considerable health-care solution for one age-related affliction, cardiovascular disease. That strategy is to understand the key steps in body metabolism and chemical reactions in the body. We have thousands of reactions going on that are ultimately governed by our genetics. We have a hundred thousand genes in every cell. Of those hundred thousand, each cell probably has anywhere from fifteen to twenty thousand turned on. The rest are mute because only those that are differentially active determine tissue specificity; what makes a particular brain cell, such as a glial cell, different from a skin cell, is that many of the genes are shut off or on in a differential fashion that determines the particular structural and functional relationships in those tissues. There are, roughly, about twenty thousand genes turned on in any cell at any one time. Right now, science understands about three percent of what those genes do. So there's a tremendous amount of new information that needs to be gained, and which will be gained in the next five to ten years, if not sooner.

WILL: Does the strategy of the genetics of aging involve cross-breeding between different disciplines?

DON: Some of the technology our company uses is based on cross-breeding of disciplines. This approach constitutes a powerful hybrid technology, to apply to some of these genetic problems. By understanding the genetic blueprint of the cell, we'll be able to understand all the biochemical reactions taking place. The genes are the blueprint. The genes code for the protein building blocks of the cell, which in turn control the biochemical reactions by utilizing substrates. Incidentally, nutraceuticals may help enhance insufficient levels of substrates, which can help optimize the biochemical reactions.

So there are several levels here. At level one, we lay the foundation for understanding what the proteins do and what substrates are utilized by the proteins, including vitamins. To understanding aging, and the role that genes play in aging, we need to determine which biochemical steps are important. Thus, we need to map out all the biochemical reactions involved. From there we can examine the steps in each biochemical pathway and determine which are important for either mitigating, eliminating, or preventing an age-related disease, as well as actually stopping and reversing the aging process. Our gene technology will allow us to proceed in this manner.

The question that really inspired me was, "Can we do something about aging?"

Using such technology enables us to identify genes and start determining the answers to level two - which proteins are made by these genes. And that enables us to go to level three, which is to find out what biochemical reactions these proteins act on or influence. Then we can properly approach level four, which is: what substrates are involved in these reactions? This includes vitamins and minerals. When one chemical is converted to another by an enzyme using a vitamin as a co-factor, the complete picture of what takes place in that particular step has four components: the substrate, the product, the enzyme catalyzing the reaction and the vitamin required for enzyme activity.
The framework that I developed 20 years ago is still the same. In genetics, you look for the master gene and the master protein that the master gene is coded to produce, and then focus on the key step gauged by the gene and protein at all four levels to determine every effect on physiology and biochemistry, in our case, as a function of age.

WILL: If I'm correct, you may be able to supply all the needed raw materials - ingestible items such as vitamins, amino acids, hormones and such - but there may still be some rate-limiting biochemical step and overcoming that can't be done merely by means of raw material provision.

DON: Correct. By understanding biochemistry, you can figure out the raw materials, but not the processes as easily.

WILL: What would some of the considerations be, in terms of identifying what the optimum amounts of dietary supplements are? Can this be better understood in reverse, by understanding the gene mechanisms involved?

DON: That's a good question. The gene activities that operate during our life span change. Those of an adolescent are different from those of a mature person. Those gene activities that differ command a cascade of reactions, including hormonal changes, different manufactured proteins and ultimately different chemical reactions requiring different substrates.

If you increase DHEA you increase carnitine transport into the mitochondria.

WILL: How specifically would this apply to mitochondrial health, which appears to be of special concern, given that many aspects of mitochondrial functioning are being identified and understood in terms of their mechanisms?

DON: There's no doubt that mitochondria are important. It's a part of cellular function. Without mitochondria, of course, we have no energy generated and the cell ceases to function. Part of our program at GeriGene looks at mitochondria, both the nuclear and mitochondrial gene components, which are intimately involved with each other. Mitochondria cannot exist without nuclear genes and without a nucleus and the nucleus cannot exist without the mitochondria. In fact, in electron transport, which involves the machinery in the mitochondria that generates energy in the form of ATP, there are at least 69 known polypeptides involved, the majority of which come from the nucleus, not from the mitochondria. A strong synergistic interplay exists between the nucleus and the mitochondria.

WILL: Could you explain a little bit why you talk about them as if they're separate? Are different genes operating with regard to both the cell and the mitochondria?

DON: About a third of the genes in, say, electron transport are coded by the mitochondrial DNA, which is separate from the nucleus of the cell. And these are thought to have evolved from bacteria. During our evolution a symbiotic relationship was established between bacteria and eukaryotic cells, such that bacteria were incorporated into the eukaryotic cells. So about a third of the genes in the electron transport chain are still ancient genes being expressed, and they are expressed by the mitochondria, independent of the nucleus of the cell.

WILL: This is very odd. It's as if the bacteria were captured in some sense and then put to work by the cells. Or the cells actually found the bacteria to be beneficial and, curiously enough, never adopted that technology for themselves, at least as far as I understand.

DON: It is not known which theories prevail. But I think the first one seems somewhat likely; bacteria were captured, and it just turned out to be good for both parties, rather than the bacteria looking for a better way to spruce up the machinery. But nonetheless, over time, something had to happen to promote the symbiotic relationship. The bacteria has all its own operative genes, but we know the mitochondria is very reliant on the nuclear gene products being transported into the mitochondria. Somewhere along the line of evolution, the mitochondria represented a modified bacterial system that lost part of their original activity and/or were displaced by nuclear genes.

A person taking carnitine is producing more ATP, and that can be measured by means of a simple skin biopsy.

WILL: A lot of nutrients have been identified that optimize the inner production of energy within the cell by the mitochondria. Other nutrients enhance performance of the mitochondrial membrane, including improved ability to transport energy across the membrane and the ability to enhance the actual process within mitochondria. How do we discover optimalities and whether we're getting enough of these nutrients? Are there tests? What knowledge has been garnered or derived from understanding the mechanisms of the genes themselves?

DON: Excellent examples of nutrients affecting mitochondria are DHEA, which has a high profile at this point, and carnitine. These two interact with each other. If you increase DHEA you increase carnitine transport into the mitochondria. Within limits, transport can also be increased by elevating carnitine levels alone. The DHEA-carnitine relationship is an example of the control interplay previously mentioned. You have a hormone that changes a number of gene activities in the cell, such as the amount of carnitine acetyltransferase, thereby increasing the transport of carnitine into mitochondria.

WILL: Carnitine operates as an escort system for fuel into the cell?

DON: That's correct. Carnitine greatly stimulates the transfer of preferential fatty acids into the mitochondria by the action of carnitine transferases, carnitine and fatty acyl CoA esters, to form fatty acyl carnitine. The energy produced is subsequently derived from fatty acid oxidation.
How you can optimize mitochondrial function can be determined with specific tests at the bench level in the physiology lab. For example, a person taking carnitine is producing more ATP and that can be measured by taking a simple biopsy of skin, for instance. One can obtain cells to examine by swabbing the inside of the mouth - a very non-invasive type of test - and from this sample determine the ATP production at designated points after taking carnitine at various dosages. After determining mitochondrial activity in the lab, whole-body physiological parameters can then be assessed.
There are a number of physiologic tests that can be used to measure the effects of nutrients such as those which look at electrochemical potentials. Other broader test parameters affecting cardiac function, blood pressure, memory, etc. can reflect the level of a given nutrient. There are probably 200 physiologic tests you can do to evaluate what a certain nutrient does at a certain dosage level. Of course, the tests will be compounded by interactions occurring with other nutrients, so such evaluations can become very complex.


There are probably 200 physiologic tests you can do to evaluate what a certain nutrient does at a certain dosage level.

WILL: When you add in certain things like lipoic acid, phosphatidylserine, and CoQ10 - there are others as well - it gets very complicated.

DON: Very complicated. It's typical in the literature to see a study done with one component, one nutrient, assessing one parameter. It is complicated when you try to determine a number of parameters with one nutrient. And it gets horrendously complicated when you start bringing in two or three nutrients.

But evaluating the test conditions is surmountable when performed in a systematic fashion at the lab bench and using the appropriate physiologic assays. Determinations can be made as to proper dosages and how compounds interact with each other or by themselves and what they actually are doing at a cell level. Ultimately, what we want to understand is the molecular level. It always comes down to the molecular level because those are the building blocks of the cell, just like atoms are building blocks for molecules. This type of very detailed information is invaluable.

WILL: But this is no more complicated than looking at genes and discovering the relationships of genes to body functions; in fact, isn't it significantly less complicated?

DON: I think it can get very complicated, but we can discover the answers. There's nothing stopping us. The tools in the lab are there. A lot of compounds are known. And their antecedents are very clear and simple. As we understand the genes, we'll have more determinations to make, more molecular levels to look at. We'll also begin to understand new compounds that might be necessary for proper cellular function. All these explorations really work in concert with the genetic work.
Again, I'll go back to our goal. We want to understand the blueprint of the cell by knowing the genetics. That's level one, the foundation level. Level two, then, is determining what the genes code for - proteins, enzymes and hormones. Level three is establishing the biochemical pathways in which chemicals (substrates) are transformed and conducted by the proteins and hormones. Level four can be regarded as developing the fine-tuning mechanism that will allow us to alter rates of chemical conversions in the biochemical pathways by adding appropriate nutraceuticals to assist in attaining a desired physiologic endpoint, whether it is energy production, free-radical quenching, or metabolic shifts.



Interview with
GeriGene Founder Don Kleinsek, Ph.D.

Optimizing
Mitochondrial Function
and the Gene Connection
Part II

In Optimizing Mitochondrial Function and the Gene Connection: Part I - March 1998, Dr. Don Kleinsek began talking about what can be done now while we prepare for the arrival of genetic-based supplement therapies. In this second part of a three issue interview, Dr. Kleinsek states clearly the positive case for dietary supplements. He makes explicit what we can do to fend off the slings and arrows of the aging process while awaiting the inevitable gene revolution.  

When the genetic paradigm starts to take hold it is important that your mental and physical health be as enhanced and/or intact as possible otherwise you could miss the boat. As changes occur, it is imperative that one take advantages at the earliest time. Already there are things that genetics lend support to that may portend your course of action over the immediate future. So we continue with our concerns as we begin this interview which took place by phone in January 1998 between Bogota, Columbia (one of a number of places where Dr. Kleinsek carries out his research) and Petaluma, California.  

Please note that Part I is the pre-read to the following interview and is available for your reading. As some of you may already know, this interview is not an "easy read," but please know that the material is complex, yet very rewarding if you stay with it. And it is very likely to become more important as the future unfolds. 

WILL: For those of us interested in improving mitochondrial function, what do genes tell us, say with regard to the citric acid cycle? 

DON: The citric acid cycle (also known as the Krebs cycle), discovered in the 1930s, is a central part of the body's chemistry which converts food components like carbohydrates, fats, and proteins into a high energy compound adenosine triphosphate (ATP), the universal energy storage molecule in the body which provides energy to carry out biological functions. 

The citric acid cycle functions in the mitochondria within every cell where it undergoes feedback regulation similar to that observed in other parts of the cell. Feedback is the influence of the output or result of a system on the input or stimulus. Understanding this process first helped define classical enzyme kinetics, how enzymes work and how they influence boichemical processes in the body. An example of feedback regulation is the relationship between the rate at which the citric acid cycle operates and the cell's requirements for ATP production: feedback corresponds between the rate and the needs, so ATP energy is produced via the Krebs cycle at a rate needed by the cell. The mechanisms presiding over this process involve interactions of metabolic products with enzymes that drive (catalyse) the cycle. 

For instance, one of the simpler feedback mechanisms observed is the creation (synthesis) of citrate from oxaloacetate and acetyl CoA, the entry step in the citric acid cycle. This reaction is catalyzed by the enzyme citrate synthase and is inhibited by the interaction of ATP with the enzyme. ATP governs the concentration of substrates thus limiting the reactability for acetyl CoA. When ATP levels increase, less citrate synthase is saturated with acetyl CoA, resulting in less citrate formed. Feedback from this blocks the formation of more ATP production. 

Investigations at the gene level with respect to disease and mitochondrial metabolism are also producing insights into the causality of disease.

WILL: How does this relate to gene discoveries? 

DON: If you start with the biochemistry you can then check to see if the genetics are true. At the levels of the citric acid cycle where the biochemical pathways and the enzyme kinetics are understood, we know which enzymes control the interactions. Underlying this control is the production of the enzymes themselves and that is governed by the genes that code for these enzymes. So the feedback mechanisms observed at the enzyme level will also coordinate (in the long term) the activities of the genes that code for these enzymes in the pathway. Accordingly more or less of the enzyme gets produced. This is an example of the degree and hierarchies of control involved in normal cell metabolism. 

Investigations at the gene level with respect to disease and mitochondrial metabolism are also producing insights into the causality of disease. Mutations (point deletions) in the mitochondrial genes are potential markers for not only myopathies (diseases affecting muscle) but a number of degenerative disease states, including some age-related diseases such as Alzheimer's and Parkinson's. Recent work indicates in Alzheimer's that there is a defect the genes that cripple the last stage of the mitochondria's electron transport chain. 

That thermostat that we have in our cells and in our bodies is there for a reason, and that's to prevent too little or too much of a biochemical process.

Only a small portion the mitochondrial mutation correlates with aging, less than 5%. Studies on cells have indicated that levels of damage greater than 85% of total mitochondrial DNA must occur before a biochemical or clinical abnormality is observed. This means that mitochondrial DNA is extremely recessive, meaning that mitochondrial genes tend to yield to the cell's DNA. 

Some investigators believe that changes in mitochondrial function could be the primary mechanism of aging. Although the mitochondria are the major intracellular source of reactive oxygen species (ROS) and free radicals which can inflict DNA damage and alter enzyme activities in the mitochondria, at this date and time there is no concrete proof of the mitochondrial theory of aging. Any proof forthcoming would need to show interference with production of ATP. The proof should show how interference is directly correlated with the disease state, the mutation type, and where it exists in the level of the mitochondrial DNA. Correlations with what we know about mitochondria are intriguing and represent a springboard for further investigation, but causality has been elusive.
 
WILL: There are studies showing the benefit of acetyl L-carnitine for Alzheimer's disease. 

DON: Correct. And in Parkinson's there's been suggestions that NADH (nicotinamide adenine dinucleotide), a coenzyme found in highest concentrations in the brain and which helps as an electron carrier. The amino acid L-carnitine helps increase fatty acid oxidation and together with NADH would be expected to increase ATP production in Parkinson's diseased cells thus assisting in optimizing cellular function. 

Some investigators believe that changes in mitochondrial function could be the primary mechanism of aging.

But the results are equivocal, suggesting a number of possibilities: additional mitochondrial function boosters are required, higher concentrations of nutrients are needed, or perhaps mitochondria are not as involved in these disease states as is commonly believed.

Biochemical reactions are dictated in part by factors (constants) which control their ability to return to equilibrium and the speeds (maximum velocity) of reactions. These parameters can often be changed simply by concentration gradients, such as citrate. Such a simple compound entering the mitochondria could result in the production of more energy. Two compounds present in the same biochemical pathway, say citric acid and malic acid, or in the same reaction, say NADH and CoQ10, could accelerate ATP production even higher.

Additionally, compounds such as NADH can act as a cofactor in tandem with enzymes, thereby increasing the reaction equilibrium quotient even further to the right (this expression is just a chemist's indication of increasing the speed of the reactions). It should be possible to do a lot of fine tuning with substrates like malic acid, citric acid, the cofactor NADH, acetyl L-carnitine, among others, that can further optimize mitochondrial function and stave off some of the age-related decline that occur in cells.

It should be possible to do a lot of fine tuning with substrates like malic acid, citric acid, the cofactor NADH, acetyl L-carnitine, among others, that can further optimize mitochondrial function and stave off some of the age-related decline that occur in cells.

WILL: In what ways can one learn more about how not to overdo it, how to take the right amount, and what the costs or liabilities are for dealing with excess amounts of dietary supplements? Do you have any particular concern about that?

DON: Yes. That's an excellent question. A little too much or a little too little does not get the job done and can be deleterious either way. The body, as you mentioned before, works on feedback mechanisms between tissues and within the tissues between cells, and within a cell between all the macromolecules, the proteins for instance and the genes.

That thermostat that we have in our cells and in our bodies is there for a reason, and that's to prevent too little or too much of a biochemical process. Just as in your house, you don't want it too warm or too cold. Yet there are problems on either side. Not enough, and the cell quiesces or undergoes apoptosis (cell death). On the other hand, mitochondria that are at full-throttle for long-periods of time, trying to produce as much energy as possible, would release tremendous amounts of ROS and free radicals under circumstances in which the cell might not be able to disarm these high-energy intermediates before sustained oxidative damage is done to the proteins, DNA and moiety (special function) lipids present in the cell. Under another mechanism of high oxidative stress, brought on by over-exercising and probably high caloric intake, some marathon runners are burnt out in their early 40s resulting in an overall lower amount of energy.

Feedback mechanisms under high oxidative stress would be working overtime to produce at the normal gene level, increased amounts of enzymatic quenchers (antioxidants produced in the body such as superoxide dismutase, catalase, peroxidase, etc). Trying to find out what the optimum levels of nutrient intake to the cell is amenable to experimental testing. For starters, nutrients can be increased and introduced into cells by microinjection or by increasing levels in the cell culture media.

Recent experiments in which superoxide dismutase and catalase are overexpressed at the gene level indicate an extension of lifespan in the fruit fly drosophila (by about 30%) and the flatworm C. elegans.

WILL: Endogenous (produced in the body) antioxidants appear to be extremely valuable with regard to longevity. Aren't there some gene companies who are actually working on increasing the internal production of them?

DON: Yes. Endogenous antioxidants would include a number of enzymes that are directly involved in antioxidation reactions as just mentioned: superoxide dismutase, catalase, glutathione peroxidase and the enzymes that produce antioxidants in the cell, such as glutathione reductase which produces the reduced thiol form of glutathione. The importance of these endogenous species (molecules) in aging is yet to be fully determined. Recent experiments in which superoxide dismutase and catalase are overexpressed at the gene level indicate an extension of lifespan in the fruit fly drosophila (by about 30%) and the flatworm C. elegans. 

However, to express these enzymes in humans must be accomplished by gene therapy, in which the enzyme is manufactured by control at the gene level. Supplementation of these enzymes by oral or intravenous routes are unlikely to be effective, primarily due to degradation by proteases enzymes in the GI tract. Alternately, some of these enzymes can be activated by co-factors that are not degraded by oral ingestion. Examples are zinc and copper for general cell interior superoxide dismutase activity, manganese for mitochondrial superoxide dismutase activity and selenium for glutathione peroxidase activity. All metal are pro-oxidative and therefore supplementation must be carefully thought out and justified with research.

Gene therapy will soon be a routine anti-aging therapy. Understanding the genes will more precisely define which nutrients to take and which hormones to supplement our bodies with as a function of aging.

Multiple gene changes due to free radical activity, may contribute to the aging process and age-related disease. The approach to the understanding of the multiple effects can be answered by identifying the underlying genetic make-up and changes with aging. For instance, free radicals increase the mutation rate of DNA which in a random fashion, changes gene activities. This then changes the cell metabolism from a normal state to a deleterious or lethal state. 
WILL: Some free radical theorists claim that their theory explains the maximum life span benefits of caloric deprivation.

DON: That could be. With caloric restriction, we know if we decrease calories by 20%, animals live 20% to 30% longer. With lower caloric intake there is lower metabolic activity and less free radical production with lifespan.

WILL: But at 50% caloric restriction you start reversing the benefits. 

At present I take a spectrum of dietary supplements at megadose levels in the promise that it will add 10-15 years onto the normal lifespan, which may not be so much as a longevity effect than it is postponing or altering a particular disease course.

DON: At 50% an undefined physiological threshold exists. But under 50% caloric deprivation the results with increased lifespan are clear - extension reverses and life shortens. How this is accomplished is not known. Is it something chemical? Less free radicals or less cross-linking generated? Lower glucose levels results in less glycation (glucose damage) of proteins and toxicity effects on cells. 

WILL: So, cross-linking may or may not involve free radicals. In other words, it creates unstable hybrid molecules that are, of themselves, thought of as ... 

DON: Dysfunctional. Crosslinking refers to the formation of irreversible, interlocking connections between two or more macromolecules (protein, DNA, lipid) which results in insoluble and dysfunctional aggregate molecules. This includes impedance of intra- and inter-cellular transport of molecules, the loss of elasticity in the extracellular matrix resulting in tissue brittleness, the inactivation of vital molecules resulting in reduced cell replacement, DNA synthesis, protein turnover and altered gene expression. The nonenzymatic glycation is the result of covalent bonding due to interactions of sugar groups with protein amino groups resulting in what are known as Amadoric products and AGEs (Advanced Glycation End-Products). AGEs have been postulated to not only impair the function of the protein they are attached to, but also to elicit toxic gene damaging responses, induce amyloid deposits (thought to be involved in Alzheimer's disease), induce vascular alterations promoting atherosclerosis, among others. 

WILL: Do these dysfunctional processes benchmark the aging process? 

DON: Indeed. A number of proteins are increasingly cross-linked with age. Crystalline lens protein in the eye, the serum protein albumin and collagen in the skin ... 

WILL: Hemoglobin. 

DON: That's a typical marker in the serum. There are several others and more will be correlated as some of the companies and research groups investigate this particular problem further. 

As for now, it's important to take supplements in the manner that integrates current studies - indications and contraindications - until we understand the genes better.

WILL: What dietary supplements do you take?

DON: Gene therapy will soon be a routine anti-aging therapy. Understanding the genes will more precisely define which nutrients to take and which hormones to supplement our bodies with as a function of aging. At present I ask myself, "What studies indicate any type of health improvement or cell optimization?" And there are studies with vitamins that indicate benefits. Too many to enumerate in this article. Vitamin E is a recent example that is understood by mainstream medicine. These studies showed that 800-1200 IU taken daily decreased risk for heart disease significantly. Antioxidation of LDL (the "bad" cholesterol) to prevent plaque formation is likely to be one of vitamin E's mechanism of action. Others include vasomotor function, inhibition of monocyte adhesion, inhibition of platelet activation and preserving endothelium-derived nitric oxide activity. Vitamin E regulates at multiple levels.

In time there will be more clinical-type studies on dosage levels and types of supplements for preventive and therapeutic measures. In general, there is not much data saying vitamins are bad for you, even megadose type of levels. At present, I take a spectrum of dietary supplements at megadose levels in the promise that it will add 10-15 years onto the normal lifespan, which may not be so much as a longevity effect than it is postponing or altering a particular disease course. Heredity is primary and in my family, there's pretty good health and "graceful" aging. Basically, it is the genes being expressed. That will provide the most help to my body in the future. As for now, it's important to take supplements in the manner that integrates current studies - indications and contraindications - until we understand the genes better.


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