Superoxide dismutase and aging

Aging can be viewed as the process of continuous deterioration characteristic of every object in nature, whether organic or inorganic. This process is described by the 2^nd law of thermodynamics where every organism is an open thermodynamic system, with an outflow of heat and inflow of free energy from chemical or thermal sources. This constant power flow, as well as the continuous reproduction of non-obsolescent genetic information and structures during cell division, ensures that the complex structure of a living organism can exist and be maintained.

Age-induced changes in body cells are associated with a declining potential and increasing free-radical reactions, which mainly originate from the oxygen restoration performed by the mitochondria, microsomes and NADN, which are oxidizing systems in phagocytes or other specialized cells. The volume of active oxygen flow is related to the basal metabolism intensity; therefore, damage accumulation in cells depends on the rate difference between the processes that generate active oxygen and those that neutralize it. According to the 2^nd law of thermodynamics, a process of this sort cannot be 100% effective; hence, generation, leakage and diffusion of active oxygen is unavoidable. Comparing the specific activity of superoxide dismutase, an enzyme catalyst that divides a superoxide into oxygen and hydrogen peroxide, in human granulocytes, platelets, red cells and lymphocytes with their propensity to generate superoxide radicals has revealed a correlation between the above-mentioned factors and cell lifespan, which may vary from 12 hours to several years.

The theory of free-radical-induced aging originated from observations in radiology and its applications for medicine. Today it is becoming a primary line of investigation, which promises to reveal the fundamental biochemical principles underlying aging. After being out of favor for many years, a hypothesis proposed by D. Harman received attention when in 1969 McCord and I. Fridovich discovered superoxide dismutase. That discovery not only provided conclusive evidence for one-electron oxygen reduction in live intermediate cells; it also stimulated research into the role of the complex multilevel system of anti-oxidant protection. While its application is constantly extending, the theory of free-radical-induced aging organically includes many of earlier hypotheses and theories of aging. Current theories on aging are based on the concepts of modern thermodynamics which describe the development and evolution of complex living and non-living systems.

The present understanding of the aging mechanism in continuously dividing cells (e.g. bacteria) is still controversial, despite the fact that the question was raised over a hundred years ago. According to the fundamental principles postulated by A.Weismann, unicellular organisms do not age. In evolutionary terms, the conflicting characteristics of cell specialization and cell immortality as a result of endless division has resulted in a categorical division of unicellular organism into body cells, which differentiate, age and die, and immortal, elementary gametal cells. The giant complex of multicellular organisms, containing trillions of body cells, is, in essence, aimed at nothing more than to ensure the immortality of gametes. Research by L. Hayflick has provided convincing evidence for the A. Weismann theory when he showed that normal body cells have limited mitotic potential and finite lifespan. Obviously, infinitely dividing cells also age, although it is unnoticeable as they have relatively simple structures and are continuously reproducing their genetic information. On the other hand, the aging of multicellular organisms develops from a general decline in potential energy, as well as from the aging and death of its completely differentiated cells.

The replication theory of aging has been validated by the observed correlation between the lifespan of various species and the lifespan of their cells. Presently, a large amount of information has been collected about mechanisms that body cells use to limit the number of their divisions. Furthermore, the hypothesis that the shortening of telomere DNA impacts the longevity of body cells has been confirmed; and remarkably, free radical processes affect the speed of aging even at the molecular level of DNA. Hyperoxia reduces the lifespan of continuously dividing cells dramatically and stimulates the accumulation of lipofuscin, a pigment characteristic of aging. Exposure of fibroblasts to a solution with a concentration of 40% oxygen results in damage and loss of telomere DNA, while, a low partial pressure of oxygen has been correlated with the increased lifespan of fibroblasts in vitro. An anti-oxidant, when added to incubating medium, is efficient in increasing the lifespan of the fibroblast culture, depending to the dosage. The role of active oxygen in regulating cell division, differentiation, aging and carcinogenesis has been under discussion for a long time. Moreover, recent published reports explain how carcinogenic genes trigger the generation of oxidants in cells.

The essence of cell aging may be demonstrated with the example of high-specialized cells found in peripheral blood systems, which are either unable to divide or divide relatively seldom. Human blood cell life spans vary widely, from several hours to several years. There is the evidence that the level of specific activity of superoxide dismutase in different blood corpuscles also varies considerably. Superoxide dismutase concentrations have been reported to be lowest in granulocytes; at the same time, these cells aggressively generate exogenous superoxide radicals used in bactericidal activity. That is what the majority of oxygen consumed by granulocytes is used for; thus, the lifespan of these cells are, not surprisingly, short.

Increased phagocytosis accompanied by a surge in respiratory metabolism and rocketing superoxide radical generation accelerates the death of phagocytes, while exogenous superoxide dismutase increases the survival rate for phagocytes to that of resting cells.

Although phagocytes have the highest superoxide dismutase levels, they generate exogenous superoxide radicals, which affect their lifespan substantially.

Red blood cells are also characterized by high level of superoxide dismutase; despite their low ability to generate superoxide radicals, red blood cells contain a high quantity of hemoglobin which continuously interacts with oxygen to generate superoxide radicals. They have a longer lifespan than platelets, but not much longer. Lymphocytes do not generate exogenous superoxide radicals; their lifespan is relatively long, apparently due to the high level of superoxide dismutase.

Using superoxide dismutase as an indicator has enabled researchers to identify that the source of superoxide radical generation comes mainly from mitochondria and sub-cellular generating centers. This provided the evidence for D. Harman’s hypothesis that the mitochondria are the cells’ “molecular clock”. In the mitochondria, 90% of oxygen consumed by aerobic cells is metabolized and about 75% of the overall flow of superoxide radicals is generated.

Mitochondria in species with short life spans (e.g. the common house fly) generate 6 to 300 times more oxidants than those in mammals. A comparative study done on myocardium tissues in eight species of mammals with considerably diverse life spans, has shown that relatively short-lived species generate more superoxide radicals in sub-mitochondrial corpuscles than the longer-lived mammals. In mice, rats, rabbits, pigs and cows the ratio of superoxide radical generation to lifespan varies by about six-fold. Aging increases the generation of superoxide radicals and hydrogen peroxide in the mitochondria of insects as well as in the brain, heart and liver of mammals.

American gerontologist R.G. Cutler and his team have been able to calculate a complete correlation between the oxygen consumed per unit mass of an organism and its lifespan in thirty species of mammals including humans. The activity of superoxide dismutase in the liver, brain and heart relative to the basal metabolic rate increases with increasing maximal lifespan and has a strictly linear character with a correlation factor close to 1. Species which have a higher total energy expenditure and longer lifespan than is commensurate with their weight (humans being a key example, as well as the lemur) are characterized by higher superoxide dismutase levels. On the other hand, the lower total energy expenditure in mice and gorillas is associated with their low superoxide dismutase levels.

Free-radical theory can efficiently explain a number of facts known by gerontologists, however there are still many observations that require different approaches. The phenomenon of caloric restriction causing increased lifespan that was discovered about 60 years ago is still not well understood. Intuitively, it has been proposed that it may be associated with a declining metabolic rate. This supposition has been proved by a comparative study that observed the speed of generation of superoxide radicals and hydrogen peroxide by the mitochondria of the brain, heart and liver in two groups of rats. One group of rats was on a calorie restricted diet, while the other was on a normal diet. It has been proposed, however, that this phenomenon is not merely caused by reduced oxygen consumption, but declined oxidation stress and activated anti-oxidant protection play a role, as well. This form of research is currently in the initial stages and cannot be regarded as complete.

A similar correlation has been reported between the speed of aging and body temperature. It is most apparent in cold-blooded organisms, which change their metabolic rate in accordance with their body temperature. When earthworms adapt to a temperature rise from 15 to 30°C, its tissues undergo a 28% increase in the level of superoxide dismutase and its oxygen consumption rises by 135%. Since this level of increased oxygen consumption cannot be accounted for by the rise in superoxide dismutase activity, it has been proposed to be the reason for the considerably higher lifespan of the earthworm at 15°C.

There is the evidence that women, who, on average, live ten years longer than men, have a metabolic rate that is 6% lower than men. Locations with reported unusually long human life spans are situated in highlands where oxygen content is lower than in flat country. This manifests the reason for similarities between the changes characteristic of aging and exposure to ionizing radiation. Despite the fact that water radiolysis and one-electron oxygen reduction differ in quantitative and spatial distribution of the active oxygen produced, they are similar in the damaging action.

Non-protein anti-oxidants may increase lifespan in short-living strains of mice by 30 to 33%. Experiments on transgenic animals are very promising. In a very early study of superoxide dismutase, the DNA of copper-bearing superoxide dismutase was implanted in drosophila by genetic engineering methods. This genetic modification resulted in its increased activity and modest, but significant, lifespan increase. Drosophila with complementary copies of genes of superoxide dismutase and catalase lived longer than control animals by 20 to 37%. Additionally, transgenic drosophila demonstrated signs of improved age-related characteristics; decreased accumulation of carbonyl proteins; decreased oxygen-sensitive enzyme inactivation as well as decreased accumulation of oxygen damaged DNA products; and reduced generation rate of oxidants in mitochondria. Recent studies have shown lifespan increases of 40% in drosophila with abundant human superoxide dismutase gene expressions in motoneurons.