Hitoshi Suda

Age-related changes of mitochondrial structure and function in Caenorhabditis elegans

A number of observations have been made to examine the role that mitochrondrial energetics and superoxide anion production play in the aging of wild-type Caenorhabditis elegans. Ultrastructural analyses reveal the presence of swollen mitochondria, presumably produced by fusion events. Two key mitochondrial functions - the activity of two electron transport chain complexes and oxygen consumption - decreased as animals aged. Carbonylated proteins, one byproduct of oxidative stress, accumulated in mitochondria much more than in the cytoplasm. This is consistent with the notion that mitochondria are the primary source of endogenous reactive oxygen species. However, the level of mitochondrially generated superoxide anion did not change significantly during aging, suggesting that the accumulation of oxidative damage is not due to excessive production of superoxide anion in geriatric animals. In concert, these data support the notion that the mitochondrial function is an important aging determinant in wild-type C. elegans.

Basic principle of the lifespan in the nematode C. elegans.

We present a biophysical model based on the principles of fluctuation and regulation to explain the effect of stochastics on survival. The model is a good fit for the survivorship and mortality rates observed in the nematode Caenorhabditis elegans. A parameter included in the theory, which is called the fluctuation constant, correlates well with a change (or declining rate) of respiration with age, which we term the physiological decline rate. The square of the physiological decline rate is proportional to the reciprocal of the fluctuation constant as revealed in a diffusion equation. In addition, the maximum and mean life spans are proportional to the reciprocal of the decline rate. The framework involved in the fluctuation theory is compatible with the existence of a regulatory system such as that acting in the insulin/insulin-like growth factor-1 (IGF-1) signaling pathway during adulthood, and that sensing, switching, and memorizing the rate of mitochondrial respiration early in life.

Interrelationships between mitochondrial fusion, energy metabolism and oxidative stress during development in Caenorhabditis elegans

Mitochondria are known to be dynamic structures with the energetically and enzymatically mediated processes of fusion and fission responsible for maintaining a constant flux. Mitochondria also play a role of reactive oxygen species production as a byproduct of energy metabolism. In the current study, interrelationships between mitochondrial fusion, energy metabolism and oxidative stress on development were explored using a fzo-1 mutant defective in the fusion process and a mev-1 mutant overproducing superoxide from mitochondrial electron transport complex II of Caenorhabditis elegans. While growth and development of both single mutants was slightly delayed relative to the wild type, the fzo-1;mev-1 double mutant experienced considerable delay. Oxygen sensitivity during larval development, superoxide production and carbonyl protein accumulation of the fzo-1 mutant were similar to wild type. fzo-1 animals had significantly lower metabolism than did N2 and mev-1. These data indicate that mitochondrial fusion can profoundly affect energy metabolism and development.

Decline in oxygen consumption correlates with lifespan in long-lived and short-lived mutants of Caenorhabditis elegans

In humans, the basal energy metabolism is thought to decline linearly with age. On the other hand, in the nematode Caenorhabditis elegans, two research groups reported independently that it declined exponentially. In this study, furthermore, we used various lifespan-mutant strains to determine whether the previous conclusion is more likely to be true. We can indirectly estimate the metabolic energy by conveniently measuring the oxygen consumption rates of C. elegans using an optical apparatus. From the profile of respiratory rates as a function of age, we can quantitatively isolate the physiological decline rate, lambda, that exponentially represents the decay rate of respiratory activity with age. In addition, quantitative analysis indicates that the respiratory activity of worms has a finite value in advanced age. We also show that the maximum and mean lifespans strongly correlate with the reciprocal of the lambda. These findings offer crucial biochemical evidence for a molecular mechanism at work in biological aging. Consequently, we here propose a mechanism based on a chemical reaction and offer a definition of the physiological decline rate and the finiteness of respiratory activity in advanced age.

Timing mechanism and effective activation energy concerned with aging and lifespan in the long-lived and thermosensory mutants of Caenorhabditis elegans.

The lifespans of many poikilothermic animals, including the nematode Caenorhabditis elegans, depend significantly on environmental temperature. Using long-living, thermosensory mutants of C. elegans, we tested whether the temperature dependency of the mean lifespan is compatible with the Arrhenius equation, which typically represents one of the chemical reaction rate theories. The temperature dependency of C. elegans was the Arrhenius type or normal, but daf-2(e1370) mutants were quite different from the others. However, taking into account the effect of the thermal denaturation of DAF-2 with the temperature, we showed that our analyzed results are compatible with previous ones. We investigated the timing mechanism of one parameter (the onset of biodemographic aging (t(0))) in the lifespan equation by applying the RNAi feeding method to daf-2 mutants in order to suppress daf-16 activity at different times during the life cycle. In summary, we further deepened the biological role of two elements, t(0) and z (the inverse of the aging rate), in the lifespan equation and mean lifespan formulated by our diffusion model z(2) = 4Dt(0), where z is composed of t(0) and D (the diffusion constant).

A further test of the equation of lifespan by C. elegans: effective activation energy for aging and lifespan.

We previously proposed a rate theory of chemical reaction as well as a lifespan equation derived by a stochastic fluctuation theory. Both were applied to biodemographic data by C. elegans to quantitatively explain that respiratory activity declines exponentially with age and that it has a physiological decline rate and a finite value (threshold) in advanced age. In this work, using the poikilothermic nature of Caenorhabditis elegans, we demonstrate the further validity of the rate theory of chemical reaction as well as the lifespan equation by changing two methods. First, to test the appropriateness of the lifespan equation from another aspect, lifespan assays were conducted by varying the time interval of observation employing the egl-1 mutant. The results indicate that, as the time interval is reduced, mortality rates gradually approach the force of mortality expected from the fitting equation of the survival curve. Second, based on the dependence of lifespan on the temperature of the culture, the physiological decline rate, and the onset of biodemographic aging, we show that the effective activation energy or energy barrier for aging and lifespan may be closely related to the standard free-energy change of ATP or ADP for a wild type and some lifespan-related mutants of C. elegans.

Analyzing observed or hidden heterogeneity on survival and mortality in an isogenic C. elegans cohort

It is generally difficult to understand the rates of human mortality from biological and biophysical standpoints because there are no cohorts or genetic homogeneity; in addition, information is limited regarding the various causes of death, such as the types of accidents and diseases. Despite such complexity, Gompertz’s rule is useful in humans. Thus, to characterize the rates of mortality from a demographic viewpoint, it would be interesting to research a single disease in one of the simplest organisms, the nematode C. elegans, which dies naturally under identically controlled circumstances without predators. Here, we report an example of the fact that heterogeneity on survival and mortality is observed through a single disease in a cohort of 100% genetically identical (isogenic) nematodes. Under the observed heterogeneity, we show that the diffusion theory, as a biophysical model, can precisely analyze the heterogeneity and conveniently estimate the degree of penetrance of a lifespan gene from the biodemographic data. In addition, we indicate that heterogeneity models are effective for the present heterogeneous data.

Biophysical and biological meanings of healthspan from C. elegans cohort.

Lifespan among individuals ranges widely in organisms from yeast to mammals, even in an isogenic cohort born in a nearly uniform environment. Needless to say, genetic and environmental factors are essential for aging and lifespan, but in addition, a third factor or the existence of a stochastic element must be reflected in aging and lifespan. An essential point is that lifespan or aging is an unpredictable phenomenon. The present study focuses on elucidating the biophysical and biological meanings of healthspan that latently indwells a stochastic nature. To perform this purpose, the nematode Caenorhabditis elegans served as a model animal. C. elegans fed a healthy food had an extended healthspan as compared to those fed a conventional diet. Then, utilizing this phenomenon, we clarified a mechanism of healthspan extension by measuring the single-worm ATP and estimating the ATP noise (or the variability of the ATP content) among individual worms and by quantitatively analyzing biodemographic data with the lifespan equation that was derived from a fluctuation theory.

Impaired p53/CEP‐1 is associated with lifespan extension through an age‐related imbalance in the energy metabolism of C. elegans

In the nematode Caenorhabditis elegans, the mammalian tumor suppressor p53 ortholog CEP‐1 mediates the stress response, activates germ line apoptosis and regulates meiotic chromosome segregation. A reduction in its expression, which frequently occurs in mammalian cancer cells, extends lifespan and induces an adaptive response in C. elegans. However, these effects do not involve an increase in oxidative stress resistance. Here, we showed that intermittent exposure to hyperoxia, which induces oxidative stress resistance and lowers the production of ROS derived from mitochondrial respiration in C. elegans, slightly improved the lifespan extension of cep‐1 mutant. Interestingly, ATP levels were increased without an increase in oxygen consumption in cep‐1 mutant during aging. In the wild‐type, lactate levels and consequentially the lactate/pyruvate ratio decreased during aging in adults. Furthermore, the expression levels of mitochondrial respiration‐related sco‐1, which is a target of p53/CEP‐1, as well as those of gluconeogenesis regulation and mammalian sirtuin ortholog genes, were also increased in the aged and adaptive conditioned wild‐type animals. In contrast, the lactate/pyruvate ratio increased in cells of the cep‐1 mutant and was amplified by intermittent hyperoxia. These results suggest that impaired p53/CEP‐1 leads to an imbalance in the age‐related energy metabolic alteration between mitochondrial oxidative phosphorylation and aerobic glycolysis and plays an important role in the extension of both intact and adaptive lifespans.