Beyond the Mitochondrial Tune Up: Part I

Part I: Delaying the Mitochondrial Decay of Aging

Introduction

Mitochondrial decay resulting from oxidative damage is known to accumulate with age and is universally recognized as a major contributing factor to the whole range of functional decline and tissue deterioration associated with aging.^{1 2 3 4 5 6 7} Age-associated changes in mitochondria are characterized by increased generation of oxidants during oxidative phosphorylation and a decline in energy production, due in part to impaired enzyme activity and also to a decrease in cardiolipin, a phospholipid concentrated in the inner mitochondrial membrane that is essential for the function of key enzymes in the electron transport chain (see Glossary for more complete definition of italicized terms).⁸

Simplified Structure of a Mitochondrion

During the phase of ATP production in which oxidative phosphorylation takes place within the mitochondria, electrons from NADH or FADH2 are transferred through the electron transport chain (Complexes I through IV), with the end result that ATP is generated and molecular oxygen (O2) is reduced to water. However, in this process, even in youth, ~ 2% of the electrons escape the electron transport chain (ETC) and reduce O2 to the highly reactive oxidant species, superoxide radical (O2−) and hydrogen peroxide (H2O2). This basal leakage of oxidants from the ETC not only appears to be unavoidable, but increases with age, rendering mitochondria the main endogenous center of superoxide radical formation.⁸

A side effect of being the primary source of these toxic oxidants is that mitochondria become their immediate targets. Mitochondria’s proximity to the oxidants they produce (O2− and H2O2), combined with their intricate structure, and the age-associated decline in antioxidant capacity (whose underlying mechanisms are discussed in Parts II and III of this review), render these organelles especially vulnerable to oxidative damage. In addition, mitochondria lack catalase, which catalyzes the decomposition of H2O2 to water and oxygen; the ability to synthesize glutathione (GSH), the body’s premier intracellular antioxidant; the ability to transport glutathione disulfide (GSSG), the oxidized form of GSH, out of the matrix; and chelators for heavy metals – all of which serve as key mechanisms through which cells decrease oxidant production.⁸

Tuning Up Mitochondrial Function with Acetyl L-Carnitine and A-Lipoic Acid

For more than thirty years, the legendary Bruce Ames, PhD, whose more than 450 scientific publications have resulted in his being among the few hundred most-cited scientists in any field—he certainly qualifies for, and has, his own entry on Wikipedia⁹—has been conducting research focused on tuning up mitochondrial metabolism to optimize health and longevity.

In recent studies, Ames and his team at the University of California, Berkeley, have made significant progress, reversing indicators of age-associated mitochondrial dysfunction in old rats by feeding them high doses of two crucial mitochondrial metabolites, acetyl L-carnitine and α-lipoic acid. Specifically, in old compared to young rats, mitochondrial membrane potential, cardiolipin level, respiratory control ratio, and cellular O2 uptake are lower, while levels of oxidants, and mutagenic aldehydes from lipid peroxidation are higher. Yet when old rats are given a diet containing high levels of acetyl-L-carnitine and α-lipoic acid, within just a few weeks, these signs of mitochondrial decay are reversed, restoring their mitochondrial function to a much more youthful level.^{10 11 12 13 14}

While their mechanisms of action differ, acetyl L-carnitine and α-lipoic acid complement one another, in some cases synergistically.

Acetyl L-carnitine has been described as a conditionally essential nutrient for humans. In addition to glycolysis, acetyl-CoA is produced from the oxidation of fatty acids by acyl-CoA synthetase enzymes in the outer mitochondrial membrane, and then transported into the inner mitochondrial matrix by acetyl L-carnitine for β oxidation and ATP production. Acetyl L-carnitine also facilitates removal from the mitochondria of the excess short- and medium-chain fatty acids that accumulate during fat metabolism. Acetyl-L-carnitine is the preferred form of the nutrient in aging or conditions of disease since it is better absorbed and more efficiently crosses the blood–brain barrier compared to L-carnitine.¹⁵

Feeding old rats an acetyl-L-carnitine-supplemented diet restores tissue levels of free and acyl carnitines to those found in plasma and brain tissues of younger animals and, when combined with α-lipoic acid, significantly increases carnitine acetyltransferase (CAT) activity. More than 70% of CAT, an enzyme present in all mammalian tissues, is located in the mitochondrial matrix, where it catalyses the reversible conversion of acetyl-CoA and carnitine to acetyl carnitine and CoA. Thus CAT regenerates CoA, which allows peroxisomal β-oxidation to proceed, and facilitates transport of acetyl moieties to the mitochondria for oxidation. (β-oxidation of fatty acids generates acetyl-CoA, the entry molecule for the Krebs or citric acid cycle.)¹³

Alpha lipoic acid is a cofactor for pyruvate dehydrogenase and α-ketoglutarate dehydrogenase, two key enzymes involved in the Krebs cycle. Not only does α-lipoic acid function as a mitochondrial coenzyme, but it also is reduced in the mitochondria to a powerful antioxidant, dihydrolipoic acid, which recycles other antioxidants, including vitamins C and E.¹³

In addition, α-lipoic acid is a potent inducer of the ~200 Phase II enzymes with antioxidant activity, including those required for the synthesis of GSH, the body’s most important intracellular antioxidant.

Thus supplementation with α-lipoic acid also increases intracellular GSH levels, which is critical for neuronal function. As a result, in aged rodents, α-lipoic acid supplementation restores long-term potentiation, a synaptic analogue of learning and memory, and partially restores ambulatory activity and memory lost during aging.^{13 16 17 18}

As noted above, when old rats receive high levels of both acetyl L-carnitine and α-lipoic acid, within several weeks, the combination greatly improves mitochondrial function, resulting in restoration of ambulatory activity and cognition (as assayed with the Skinner box and Morris water maze), heart, and immune function to levels seen in young rats.

The KM Concept: Key to Mitochondrial Decay?

Ames’ rationale for the mechanism behind the rejuvenating effects of acetyl-L-carnitine and α-lipoic acid is that, with age, increased oxidative damage to key proteins, causes deformation in their structure, resulting in an increased Michaelis constant/(KM) or decreased binding affinity for a co-enzyme (i.e., the nutrient co-factor for the enzyme), and thus a decrease in enzyme function.¹⁹

Ames’ research has demonstrated that this age-associated effect on enzyme-binding affinity can be mimicked by exposing enzymes to malondialdehyde, a lipid-peroxidation product, levels of which also increase with age. Increasing availability of the substrate acetyl L-carnitine along with α-lipoic acid, a mitochondrial antioxidant, restores the velocity of the reaction (KM) for the enzyme acetyl-L-carnitine transferase and thus improves mitochondrial function.⁸

Common Nutrient Deficiencies Accelerate Mitochondrial Decay

Ames also targets iron and vitamin B6 as two common nutrient deficiencies that accelerate mitochondrial decay.²⁰ Heme biosynthesis occurs predominantly in the mitochondria. Iron insufficiency disrupts heme synthesis resulting in loss of Complex IV and, as a result, significantly increased release of oxidants. Iron deficiency is fairly common among menstruating women in the U.S., 25% of whom ingest ≤50% of the DRI. Vitamin B6 deficiency, also not uncommon (10% of Americans ingest ≤50% of the DRI), can also cause heme deficiency since heme biosynthesis requires B6. A further consideration is that menstruating women may be using oral contraceptives, which have repeatedly been found to be associated with significantly lowered levels of vitamins B6 and B12.²¹ Ames notes that insufficiencies of iron or B6 are likely to result in accelerated aging and neural decay.

Conclusion

While it has commonly been thought that Americans’ intake of essential micronutrients is adequate, evidence indicates that damage occurs at levels higher than those that cause acute deficiency disease. In addition, as many as one-third of all single nucleotide polymorphisms (SNPs) in a gene result in the corresponding enzyme having an increased KM (decreased binding affinity) for its coenzyme, and therefore a lower rate of reaction. Given that some of these SNPs (to be discussed in Part II of this review: The Methylation Transsulfuration Connection to Mitochondrial Function) are found in ~30-40% of the population,²⁹ it is obvious that higher than DRI levels are necessary for optimal function in significant numbers of people.

Dr. Ames’ KM concept applies both to aging-associated mitochondrial dysfunction, as discussed above in regards to acetyl-L-carnitine and lipoic acid, and to inborn (genetic) weaknesses in metabolism. In both cases, the functional capacity of defective enzymes can be ameliorated by administration of high doses of the corresponding cofactors, which raises coenzyme levels and at least partially restores enzymatic activity and mitochondrial function.

However, a protocol focused on restoration of KM, while certainly helpful in delaying mitochondrial decay, does not address a more fundamental issue – why does human physiology shift from a homeostasic state that repairs and balances itself to one that allows decay to accumulate? A new theory concerning what researcher Wulf Dröge has called “the first cause of death” may provide insight into a vicious cycle responsible for the shift from a state of youthful homeostatic repair to a homeostatic state that promotes mitochondrial decay. This will be the topic of Part III of this review: Reversing the Age-related Metabolic Shift towards Mitochondrial Decay.

Citric Acid (Krebs) Cycle

Glossary

α-ketoglutarate dehydrogenase (aka xoglutarate dehydrogenase)—enzyme involved in the citric acid or Krebs cycle in which it catalyzes the reaction: α-ketoglutarate + NAD+ + CoA → Succinyl CoA + CO2 + NADH

β oxidation—the process by which fatty acids, in the form of Acyl-CoA molecules, are broken down in mitochondria and/or in peroxisomes to generate Acetyl-CoA, the entry molecule for the Krebs cycle.

betaine—in chemistry, any neutral compound with both a positively- and a negatively-charged functional group, so that it carries a total net charge of 0, while carrying formal positive and negative charges, also called a zwitterion. In biological systems, betaines, which are polar and water-soluble, permit water to remain in cells, thus serving as intracellular protectors against osmotic stress. Betaines also serve as methyl donors; trimethylglycine is a betaine.

cardiolipin—an important component accounting for ~20% of the total lipid content of the inner mitochondrial membrane, cardiolipin (so named since it was first identified in beef heart in the 1940s) is essential for the optimal function of numerous enzymes involved in mitochondrial energy production including cytochrome bc1 (Complex III) and cytochrome c oxidase (Complex IV) in the electron transport chain. Decreased cardiolipin synthesis is thought to be associated with mitochondrial dysfunction in Parkinson’s disease, non-alcoholic fatty liver disease, heart failure and diabetes.^{30 31 32 33} Increased mitochondrial ROS production promotes oxidation and depletion of cardiolipin, as well as inhibition of cytochrome c oxidase activity. Peroxidation of cardiolipin has been suggested to impair the barrier function of the inner membrane and facilitate the detachment of cytochrome c from the electron transport chain.²⁴

catalase—a common enzyme found in virtually all organisms exposed to oxygen, catalase catalyzes the decomposition of hydrogen peroxide to water and oxygen. Catalase has one of the highest turnover numbers of all enzymes; one molecule of catalase can convert millions of molecules of hydrogen peroxide to water and oxygen per second.³⁴³

Glutathione—GSH is a tripeptide composed of cysteine, glycine, and glutamate that is synthesized de novo in all cells and serves as the major intracellular antioxidant and redox buffer.

heme—the major functional form of iron, synthesized in the mitochondria.³⁶

pyruvate dehydrogenase—enzyme involved in transforming pyruvate from glycolysis into acetyl-CoA, which is then used in the citric acid or Krebs cycle. Thus, pyruvate dehydrogenase serves as a link from glycolysis to the citric acid or Krebs cycle, which is followed by oxidative phosphorylation in the electron transport chain of the mitochondria.

References