Amino acid restriction, aging, and longevity: an update.

Various so-called dietary restriction paradigms have shown promise for extending health and life. All such paradigms rely on ad libitum (hereafter ad lib) feeding, something virtually never employed in animals whose long-term health we value, either as a control or, except for food restriction itself, for both control and treatment arms of the experiment. Even though the mechanism(s) remain only vaguely understood, compared to ad lib-fed animals a host of dietary manipulations, including calorie restriction, low protein, methionine, branched-chain amino acids, and even low isoleucine have demonstrable health benefits in laboratory species in a standard laboratory environment. The remaining challenge is to determine whether these health benefits remain in more realistic environments and how they interact with other health enhancing treatments such as exercise or emerging geroprotective drugs. Here we review the current state of the field of amino acid restriction on longevity of animal models and evaluate its translational potential.

Comment on "The plateau of human mortality: Demography of longevity pioneers".

Barbi et al (Reports, 29 June 2018, p. 1459) reported that human mortality rate reached a "plateau" after the age of 105, suggesting there may be no limit to human longevity. We show, using their data, that potential lifespans cannot increase much beyond the current 122 years unless future biomedical advances alter the intrinsic rate of human aging.

Superior proteome stability in the longest lived animal.

Bivalve mollusks have several unique traits, including some species with exceptionally long lives, others with very short lives, and the ability to determine the age of any individual from growth rings in the shell. Exceptionally long-lived species are seldom studied yet have the potential to be particularly informative with respect to senescence-resistance mechanisms. To this end, we employed a range of marine bivalve mollusk species, with lifespans ranging from under a decade to over 500 years, in a comparative study to investigate the hypothesis that long life requires superior proteome stability. This experimental system provides a unique opportunity to study closely related organisms with vastly disparate longevities, including the longest lived animal, Arctica islandica.Specifically, we investigated relative ability to protect protein structure and function, both basally and under various stressors in our range of species. We found a consistent relationship between species longevity, resistance to protein unfolding, and maintenance of endogenous enzyme (creatine kinase) activity. Remarkably, our longest-lived species, Arctica islandica (maximum longevity >500 years), had no increase in global proteome unfolding in response to several stressors. Additionally, the global proteome of shorter-lived species exhibited less resistance to temperature-induced protein aggregation than longer-lived species. A reporter assay, in which the same protein's aggregation properties was assessed in lysates from each study species, suggests that some endogenous feature in the cells of long-lived species, perhaps small molecular chaperones, was at least partially responsible for their enhanced proteome stability. This study reinforces the relationship between proteostasis and longevity through assessment of unfolding, function, and aggregation in species ranging in longevity from less than a decade to more than five centuries.

Chronic inhibition of mammalian target of rapamycin by rapamycin modulates cognitive and non-cognitive components of behavior throughout lifespan in mice.

Aging is, by far, the greatest risk factor for most neurodegenerative diseases. In non-diseased conditions, normal aging can also be associated with declines in cognitive function that significantly affect quality of life in the elderly. It was recently shown that inhibition of Mammalian TOR (mTOR) activity in mice by chronic rapamycin treatment extends lifespan, possibly by delaying aging {Harrison, 2009 #4}{Miller, 2011 #168}. To explore the effect of chronic rapamycin treatment on normal brain aging we determined cognitive and non-cognitive components of behavior throughout lifespan in male and female C57BL/6 mice that were fed control- or rapamycin-supplemented chow. Our studies show that rapamycin enhances cognitive function in young adult mice and blocks age-associated cognitive decline in older animals. In addition, mice fed with rapamycin-supplemented chow showed decreased anxiety and depressive-like behavior at all ages tested. Levels of three major monoamines (norepinephrine, dopamine and 5-hydroxytryptamine) and their metabolites (3,4-dihydroxyphenylacetic acid, homovanillic acid, and 5-hydroxyindolacetic acid) were significantly augmented in midbrain of rapamycin-treated mice compared to controls. Our results suggest that chronic, partial inhibition of mTOR by oral rapamycin enhances learning and memory in young adults, maintains memory in old C57BL/6J mice, and has concomitant anxiolytic and antidepressant-like effects, possibly by stimulating major monoamine pathways in brain.

Maximum shell size, growth rate, and maturation age correlate with longevity in bivalve molluscs.

Bivalve molluscs are newly discovered models of successful aging, and this invertebrate group includes Arctica islandica, with the longest metazoan life span. Despite an increasing biogerontological focus on bivalves, their life history traits in relation to maximum age are not as comprehensively understood as those in vertebrate model aging organisms. We explore the allometric scaling of longevity and the relationship between development schedules (time to maturity and growth rate) and longevity in the Bivalvia. Using a traditional nonphylogenetic approach and the phylogenetically independent contrasts method, the relationship among these life history parameters is analyzed. It is demonstrated that in bivalves, maximum shell size, development, and growth rates all associate with longevity. Our findings support the observations of life history patterns in mammals and fish. This is the first investigation into the relationship among longevity, size, and development schedules throughout this group, and the results strengthened by the control for phylogenetic independence.

Methusaleh's Zoo: how nature provides us with clues for extending human health span.

As impressive as the accomplishments of modern molecular biologists have been in finding genetic alterations that lengthen life in short-lived model organisms, they pale in comparison to the remarkable diversity of lifespans produced by evolution. Some animal species are now firmly documented to live for more than four centuries and even some mammals, like the bowhead whale, appear to survive 200 years or more. Another group of species may not be as absolutely long-lived, but they are remarkably long-lived for their body size and metabolic rate. These species include a number of bats, some of which live for at least 40 years in the wild, as well as the naked mole-rat, which is the same size, but lives nearly 10 times as long as the laboratory mouse. Together these exceptionally long-lived organisms have important roles to play in our future understanding of the causal mechanisms and modulation of ageing. Bats and naked mole-rats in particular have already contributed in the following ways: (1) they have contributed to the abandonment of the rate-of-living theory and weakened enthusiasm for the oxidative stress hypothesis of ageing, (2) they have helped evaluate how the tumour-suppressing role of cellular senescence is affected by the evolution of diverse body sizes as well as diverse longevities, (3) they have shed light on the relationship between specific types of DNA repair and ageing and (4) they have yielded insight into new processes, specifically the maintenance of the proteome and hypotheses concerning how evolution shapes ageing. The continuing acceleration of progress in genome sequencing and development of more and more cross-species investigatory techniques will facilitate even more contributions of these species in the near future.

Polymorphic microsatellite loci for the common marmoset (Callithrix jacchus) designed using a cost- and time-efficient method.

We describe a cost- and time-efficient method for designing new microsatellite markers in any species with substantial genomic DNA sequence data available. Using this technique, we report 14 new polymorphic dinucleotide microsatellite loci isolated from the common marmoset. The relative yield of new polymorphisms was higher with less labor than described in previous marmoset studies. Of 20 loci initially evaluated, 14 were polymorphic and amplified reliably (70% success rate). The number of alleles ranged from 3 to 9 with heterozygosity varying from 0.48 to 0.83.

Exceptional cellular resistance to oxidative damage in long-lived birds requires active gene expression.

Previous studies indicated that renal tubular epithelial cells from some long-lived avian species exhibit robust and/or unique protective mechanisms against oxidative stress relative to murine cells. Here we extend these studies to investigate the response of primary embryonic fibroblast-like cells to oxidative challenge in long- and short-lived avian species (budgerigar, Melopsittacus undulatus, longevity up to 20 years, vs Japanese quail, Coturnix coturnix japonica, longevity up to 5 years) and short- and long-lived mammalian species (house mouse, Mus musculus, longevity up to 4 years vs humans, Homo sapiens, longevity up to 122 years). Under the conditions of our assay, the oxidative-damage resistance phenotype appears to be associated with exceptional longevity in avian species, but not in mammals. Furthermore, the extreme oxidative damage resistance phenotype observed in a long-lived bird requires active gene transcription and translation, suggesting that specific gene products may have evolved in long-lived birds to facilitate resistance to oxidative stress.

Effect of capture and season on fecal glucocorticoid levels in deer mice (Peromyscus maniculatus) and red-backed voles (Clethrionomys gapperi).

The effect of confinement and season on fecal glucocorticoid (GC) levels in deer mice (Peromyscus maniculatus) and red-backed voles (Clethrionomys gapperi) was determined. Deer mice confined in a Sherman trap more than 4 h had fecal GC levels that were significantly higher than those in individuals that remained in a trap 4 h or less. However, this treatment may not be stressful for red-backed voles as neither plasma nor fecal GC levels were significantly elevated after 12 h of confinement. In addition, a clear temporal pattern in the secretion of fecal GCs was observed between mid June and early November in both species.

A reliable assessment of 8-oxo-2-deoxyguanosine levels in nuclear and mitochondrial DNA using the sodium iodide method to isolate DNA.

A major controversy in the area of DNA biochemistry concerns the actual in vivo levels of oxidative damage in DNA. We show here that 8-oxo-2-deoxyguanosine (oxo8dG) generation during DNA isolation is eliminated using the sodium iodide (NaI) isolation method and that the level of oxo8dG in nuclear DNA (nDNA) is almost one-hundredth of the level obtained using the classical phenol method. We found using NaI that the ratio of oxo8dG/10(5 )deoxyguanosine (dG) in nDNA isolated from mouse tissues ranged from 0.032 +/- 0.002 for liver to 0.015 +/- 0.003 for brain. We observed a significant increase (10-fold) in oxo8dG in nDNA isolated from liver tissue after 2 Gy of gamma-irradiation when NaI was used to isolate DNA. The turnover of oxo8dG in nDNA was rapid, e.g. disappearance of oxo8dG in the mouse liver in vivo after gamma-irradiation had a half-life of 11 min. The levels of oxo8dG in mitochondrial DNA isolated from liver, heart and brain were 6-, 16- and 23-fold higher than nDNA from these tissues. Thus, our results showed that the steady-state levels of oxo8dG in mouse tissues range from 180 to 360 lesions in the nuclear genome and from one to two lesions in 100 mitochondrial genomes.

History and prospects: symposium on organisms with slow aging.

We discuss the background concepts which lead to this issue of Experimental Gerontology. On one hand, genetic and molecular studies of short-lived worms, flies, and mice are yielding remarkable discoveries on gene systems that regulate the life span. On the other hand, little is known about the nature of aging in other vertebrates, with life spans extending into the human range or beyond the record 122y human life span, which may have aging processes that are so slow as to be 'negligible'. We point out that organisms with these vastly different life spans have essentially identical cells within an evolutionary group and that the cellular tool kit that existed by 600 million years ago allowed the evolution of life spans ranging up to one million-fold difference in length. The possibility of negligible senescence has not been widely discussed, and may be in conflict with mathematical deductions from population genetics theory. We propose minimal criteria for the lack of senescence: (1) no observable increase in age-specific mortality rate or decrease in reproduction rate after sexual maturity; and (2) no observable age-related decline in physiological capacity or disease resistance. We also introduce some of the species discussed in subsequent chapters which are unfamiliar models to most biomedical researchers.

An experimental paradigm for the study of slowly aging organisms.

An experimental paradigm for the study of mechanisms of resistance to aging in long-lived organisms has been developed. The paradigm assumes, in concert with accumulating empirical data, that resistance to the aging processes at the organismal level will be reflected in resistance to various stressors at the cellular level. The advantage of this paradigm is that it requires neither the long-term monitoring of individuals nor the use of exceptionally old individuals. The research approach consists of: (1) verifying that primary cell cultures from the long-lived organism exhibit better resistance to key stressors than cells from related, short-lived organisms; (2) assessing differences in gene-expression before and after stress exposure in cultured cells from the long- and short-lived species in order to identify key genes involved in the stress-resistance response; (3) transfecting putative key genes from long-lived species into cells or cell lines of defined stress-resistance and hope to observe that the stress-resistance phenotype has thereby been transferred with the gene(s); (4) generating transgenic model animals containing the gene(s) of interest and look for extended life/health span.

Comparative biology of aging in birds: an update.

The long life spans and slow aging rates of birds relative to mammals are paradoxical in view of birds' high metabolic rates, body temperatures and blood glucose levels, all of which are predicted to be liabilities by current biochemical theories of aging. Available avian life-table data show that most birds undergo rapid to slow "gradual" senescence. Some seabird species exhibit extremely slow age-related declines in both survival and reproductive output, and even increase reproductive success as they get older. Slow avian senescence is thought to be coupled evolutionarily with delayed maturity and low annual fecundity. Recent research in our lab and others supports the hypothesis that birds have special adaptations for preventing age-related tissue damage caused by reactive oxygen species (ROS) and advanced glycosylation endproducts, or AGEs, as well as an unusual capacity for neurogeneration in brain. Much of this work is in its early stages, however, and reliable biomarkers for comparing avian and mammalian aging need more thorough development.

Does caloric restriction in the laboratory simply prevent overfeeding and return house mice to their natural level of food intake?

Some researchers have speculated that the senescence-retarding effect of caloric restriction on laboratory rodents is an artifact of overfeeding under captive conditions. The argument posits that mice in nature are chronically calorically restricted; therefore, the typical laboratory protocol of restricting animals to 60% of their ad lib food intake more realistically replicates life in the field: the conditions under which the animals' physiology has been designed by natural selection to thrive. The hypothesis concludes that instead of comparing control animals with restricted animals, we are in fact comparing overfed animals with adequately fed ones, and, not surprisingly, the overfed ones die younger. In this Perspective, the author discusses the merits and drawbacks of this hypothesis in light of energy consumption data for various types of mice.

Genetic variability in responses to caloric restriction in animals and in regulation of metabolism and obesity in humans.

Panel 5 focused on genetic factors that might mediate or moderate the effects of caloric restriction (CR) on longevity. Panel members stated that currently there is limited information directly addressing these issues. Therefore, they focused attention on what studies could be done. In addition, the panel believed that certain conceptual issues merited clarification and focused attention on this issue. Human studies and studies of nonhuman model organisms were discussed. The panel found at least three reasons why it would be valuable to find genes that influence the (putative) longevity-promoting effect of CR in humans. Such knowledge would offer: (a) the ability to predict individual responses to CR; (b) increased understanding of physiological mechanisms; and (c) the potential to develop mechanism-based interventions to promote longevity or healthy aging. In addition, the panel emphasized several macro-level recommendations regarding research strategies to avoid, research strategies to emphasize, and resources needing development.

Why do we age?

The evolutionary theory of ageing explains why ageing occurs, giving valuable insight into the mechanisms underlying the complex cellular and molecular changes that contribute to senescence. Such understanding also helps to clarify how the genome shapes the ageing process, thereby aiding the study of the genetic factors that influence longevity and age-associated diseases.

Fecal glucocorticoids: a noninvasive method of measuring adrenal activity in wild and captive rodents.

To determine the utility of fecal corticosteroid concentration as a measure of chronic stress under laboratory and field conditions, we biochemically and physiologically validated a radioimmunoassay for corticosteroids in three rodent species, house mice (Mus musculus), deer mice (Peromyscus maniculatus), and red-back voles (Clethrionomys gapperi). The biochemical validations demonstrated that the assay accurately and precisely measured corticosteroid concentration in the feces. The physiological validation indicated that the assay was sensitive enough to detect the stress associated with (a) brief handling and bleeding of animals, (b) chronic caloric restriction, (c) exposure to a novel environment, and (d) exposure to a novel cold environment. Our results suggest that fecal measurements reflect stress levels experienced by these animals approximately 6-12 h before defecation. Therefore, given a judicious trapping and trap-monitoring protocol, this assay has considerable utility for measuring the stress levels at which animals actually exist in the field.

Cultured renal epithelial cells from birds and mice: enhanced resistance of avian cells to oxidative stress and DNA damage.

Current mechanistic theories of aging would predict that many species of birds, given their unusually high metabolic rates, body temperatures, and blood sugar levels, should age more rapidly than mammals of comparable size. On the contrary, many avian species display unusually long life spans. This finding suggests that cells and tissues from some avian species may enjoy unusually robust and/or unique protective mechanisms against fundamental aging processes, including a relatively high resistance to oxidative stress. We therefore compared the sensitivities of presumptively homologous epithelial somatic cells derived from bird and mouse kidneys to various forms of oxidative stress. When compared to murine cells, we found enhanced resistance of avian cells from three species (budgerigars, starlings, canaries) to 95% oxygen, hydrogen peroxide, paraquat, and gamma-radiation. Differential resistance to 95% oxygen was demonstrated with both replicating and quiescent cultures. Hydrogen peroxide was shown to induce DNA single-strand breaks. There were fewer breaks in avian cells than in mouse cells when similarly challenged.

Comparative aging and life histories in mammals.

A comparative assessment of aging and longevity in mammals has four uses in aging research. These are: (1) hypothesis formulation and evaluation, (2) investigating the generality of putative aging mechanisms, (3) isolating key physiological factors influencing aging rate, and (4) allowing the most appropriate choice of animal models for particular research questions. The first use requires detailed information on a wide variety of species, and I will examine general patterns of aging in a sample of over 600 species of mammals. The second use requires the selection of several models as distantly related to one another as feasible. The third use is best served by evaluating species or populations as closely related to one another as possible, assuming that they differ substantially in aging rate. The fourth use requires a logic of animal model selection, as well detailed information about a wide range of species. Specific examples of each use will be given.