Role of mismatch repair in aging

A common feature of aging is the accumulation of genetic damage throughout life. DNA damage can lead to genomic instability. Many diseases associated with premature aging are a result of increased accumulation of DNA damage. In order to minimize these damages, organisms have evolved a complex network of DNA repair mechanisms, including mismatch repair (MMR). In this review, we detail the effects of MMR on genomic instability and its role in aging emphasizing on the association between MMR and the other hallmarks of aging, serving to drive or amplify these mechanisms. These hallmarks include telomere attrition, epigenetic alterations, mitochondrial dysfunction, altered nutrient sensing and cell senescence. The close relationship between MMR and these markers may provide prevention and treatment strategies, to reduce the incidence of age-related diseases and promote the healthy aging of human beings.     

Mitochondrial Dysfunction: Common Final Pathway in Brain Aging and Alzheimer’s Disease—Therapeutic Aspects

As a fully differentiated organ, our brain is very sensitive to cumulative oxidative damage of proteins, lipids, and DNA occurring during normal aging because of its high energy metabolism and the relative low activity of antioxidative defense mechanisms. As a major consequence, perturbations of energy metabolism including mitochondrial dysfunction, alterations of signaling mechanisms and of gene expression culminate in functional deficits. With the increasing average life span of humans, age-related cognitive disorders such as Alzheimer’s disease (AD) are a major health concern in our society. Age-related mitochondrial dysfunction underlies most neurodegenerative diseases, where it is potentiated by disease-specific factors. AD is characterized by two major histopathological hallmarks, initially intracellular and with the progression of the disease extracellular accumulation of oligomeric and fibrillar β-amyloid peptides and intracellular neurofibrillary tangles composed of hyperphosphorylated tau protein. In this review, we focus on findings in AD animal and cell models indicating that these histopathological alterations induce functional deficits of the respiratory chain complexes and therefore consecutively result in mitochondrial dysfunction and oxidative stress. These parameters lead synergistically with the alterations of the brain aging process to typical signs of neurodegeneration in the later state of the disease, including synaptic dysfunction, loss of synapses and neurites, and finally neuronal loss. We suggest that mitochondrial protection and subsequent reduction of oxidative stress are important targets for prevention and long-term treatment of early stages of AD.     

The spatial association of gene expression evolves from synchrony to asynchrony and stochasticity with age

For multicellular organisms, different tissues coordinate to integrate physiological functions, although this systematically and gradually declines in the aging process. Therefore, an association exists between tissue coordination and aging, and investigating the evolution of tissue coordination with age is of interest. In the past decade, both common and heterogeneous aging processes among tissues were extensively investigated. The results on spatial association of gene changes that determine lifespan appear complex and paradoxical. To reconcile observed commonality and heterogeneity of gene changes among tissues and to address evolution feature of tissue coordination with age, we introduced a new analytical strategy to systematically analyze genome-wide spatio-temporal gene expression profiles. We first applied the approach to natural aging process in three species (Rat, Mouse and Drosophila) and then to anti-aging process in Mouse. The results demonstrated that temporal gene expression alteration in different tissues experiences a progressive association evolution from spatial synchrony to asynchrony and stochasticity with age. This implies that tissue coordination gradually declines with age. Male mice showed earlier spatial asynchrony in gene expression than females, suggesting that male animals are more prone to aging than females. The confirmed anti-aging interventions (resveratrol and caloric restriction) enhanced tissue coordination, indicating their underlying anti-aging mechanism on multiple tissue levels. Further, functional analysis suggested asynchronous DNA/protein damage accumulation as well as asynchronous repair, modification and degradation of DNA/protein in tissues possibly contributes to asynchronous and stochastic changes of tissue microenvironment. This increased risk for a variety of age-related diseases such as neurodegeneration and cancer that eventually accelerate organismal aging and death. Our study suggests a novel molecular event occurring in aging process of multicellular species that may represent an intrinsic molecular mechanism of aging.     

The aging stress response

Aging is the outcome of a balance between damage and repair. The rate of aging and the appearance of age-related pathology are modulated by stress response and repair pathways that gradually decline, including the proteostasis and DNA damage repair networks and mitochondrial respiratory metabolism. Highly conserved insulin/IGF-1, TOR, and sirtuin signaling pathways in turn control these critical cellular responses. The coordinated action of these signaling pathways maintains cellular and organismal homeostasis in the face of external perturbations, such as changes in nutrient availability, temperature, and oxygen level, as well as internal perturbations, such as protein misfolding and DNA damage. Studies in model organisms suggest that changes in signaling can augment these critical stress response systems, increasing life span and reducing age-related pathology. The systems biology of stress response signaling thus provides a new approach to the understanding and potential treatment of age-related diseases.     

Chromatin Regulation of DNA Damage Repair and Genome Integrity in the Central Nervous System

With the continued extension of lifespan, aging and age-related diseases have become a major medical challenge to our society. Aging is accompanied by changes in multiple systems. Among these, the aging process in the central nervous system is critically important but very poorly understood. Neurons, as post-mitotic cells, are devoid of replicative associated aging processes, such as senescence and telomere shortening. However, because of the inability to self-replenish, neurons have to withstand challenge from numerous stressors over their lifetime. Many of these stressors can lead to damage of the neurons' DNA. When the accumulation of DNA damage exceeds a neuron's capacity for repair, or when there are deficiencies in DNA repair machinery, genome instability can manifest. The increased mutation load associated with genome instability can lead to neuronal dysfunction and ultimately to neuron degeneration. In this review, we first briefly introduce the sources and types of DNA damage and the relevant repair pathways in the nervous system (summarized in Fig. 1). We then discuss the chromatin regulation of these processes and summarize our understanding of the contribution of genomic instability to neurodegenerative diseases.     

A Third Link connecting Aging with Double Strand Break Repair

Until recently, the connection between aging and DNA repair has rested on two classes of observation. First, DNA damage and unrepaired double-strand breaks (DSBs) accumulate with age. Second, several defects in DNA repair genes are associated with early onset of age-related diseases and other signs of premature aging. Now, a third link has emerged: The mechanisms by which cells repair DSB damage can change dramatically with age, shifting from simpler end-joining processes in younger organisms to homologous mechanisms in which missing genetic information is restored through use of a template. So far this third link between aging and DNA repair has only been observed in a small number of experimental systems, and cannot yet claim the generality of the other two. Here we review the evidence for this phenomenon and present new data testing models for the underlying causes. If the generality of age-related changes in DSB repair pathway usage can be established, it will provide a new insight into the underlying molecular basis of aging and how evolution has shaped these processes.     

Robustness and aging—A systems-level perspective

The theory of robustness describes a system level property of evolutionary systems, which predicts tradeoffs of great interest for the systems biology of aging, such as accumulation of non-heritable damage, occurrence of fragilities and limitations in performance, optimized allocation of restricted resources and confined redundancies. According to the robustness paradigm cells and organisms evolved into a state of highly optimized tolerance (HOT), which provides robustness to common perturbations, but causes tradeoffs generally characterized as “robust yet fragile”. This raises the question whether the ultimate cause of aging is more than a lack of adaptation, but an inherent fragility of complex evolutionary systems. Since robustness connects to evolutionary designs, consideration of this theory provides a deeper connection between evolutionary aspects of aging, mathematical models and experimental data. In this review several mechanisms influential for aging are re-evaluated in support of robustness tradeoffs. This includes asymmetric cell division improving performance and specialization with limited capacities to prevent and repair age-related damage, as well as feedback control mechanisms optimized to respond to acute stressors, but unable to halt nor revert aging. Improvement in robustness by increasing efficiencies through cellular redundancies in larger organisms alleviates some of the damaging effects of cellular specialization, which can be expressed in allometric relationships. The introduction of the robustness paradigm offers unique insights for aging research and provides novel opportunities for systems biology endeavors.     

The structural changes in chromatin at different levels of its organization during aging]

The data about changes in the molecular organization of the cell nucleus during aging are reviewed. Changes in DNA primary and secondary structure include deletions of some sequences, changes in base methylation pattern and the increasing number of DNA breaks. Conditions underlying poor reproducibility of the results obtained in corresponding experiments are discussed. Changes observed in the nucleosomal and supranucleosomal chromatin structure reflect either its increasing compactization or the loss the nucleosomal structure during aging. The data about the increased DNA superhelicity and topoisomerase activity in aging organisms contradict the accepted views about age-related decrease in chromatin activity. It is suggested that the gene activity in aging organisms is specifically altered rather than generally decreased. The data are presented about the ways of preventing or reverting the age-related changes in chromatin structure.     

The oxidative DNA lesions 8,5'-cyclopurines accumulate with aging in a tissue-specific manner

Accumulation of DNA damage is implicated in aging. This is supported by the fact that inherited defects in DNA repair can cause accelerated aging of tissues. However, clear-cut evidence for DNA damage accumulation in old age is lacking. Numerous studies report measurement of DNA damage in nuclear and mitochondrial DNA from tissues of young and old organisms, with variable outcomes. Variability results from genetic differences between specimens or the instability of some DNA lesions. To control these variables and test the hypothesis that elderly organisms have more oxidative DNA damage than young organisms, we measured 8,5'-cyclopurine-2'-deoxynucleosides (cPu), which are relatively stable, in tissues of young and old wild-type and congenic progeroid mice. We found that cPu accumulate spontaneously in the nuclear DNA of wild-type mice with age and to a greater extent in DNA repair-deficient progeroid mice, with a similar tissue-specific pattern (liver > kidney > brain). These data, generated under conditions where genetic and environmental variables are controlled, provide strong evidence that DNA repair mechanisms are inadequate to clear endogenous lesions over the lifespan of mammals. The similar, although exaggerated, results obtained from progeroid, DNA repair-deficient mice and old normal mice support the conclusion that DNA damage accumulates with, and likely contributes to, aging.     

The protein poly(ADP-ribosyl)ation system: Its role in genome stability and lifespan determination

The processes that lead to violation of genome integrity are known to increase with age. This phenomenon is caused both by increased production of reactive oxygen species and a decline in the efficiency of antioxidant defense system as well as systems maintaining genome stability. Accumulation of different unrepairable genome damage with age may be the cause of many age-related diseases and the development of phenotypic and physiological signs of aging. It is also clear that there is a close connection between the mechanisms of the maintenance of genome stability, on one hand, and the processes of spontaneous tumor formation and lifespan, on the other. In this regard, the system of protein poly(ADP-ribosyl)ation activated in response to a variety of DNA damage seems to be of particular interest. Data accumulated to date suggest it to be a kind of focal point of cellular processes, guiding the path of cell survival or death depending on the degree of DNA damage. This review summarizes and analyzes data on the involvement of poly(ADP-ribosyl)ation in various mechanisms of DNA repair, its interaction with progeria proteins, and the possible role in the development of spontaneous tumors and lifespan determination. Special attention is given to the relationship between various polymorphisms of the human poly(ADP-ribose) polymerase-1 gene and longevity.     

Germ line recombination may be primarily a manifestation of DNA repair processes.

Several kinds of DNA repair and reactivation processes have been found to occur by recombination in E. coli and its phages. Recombinational repair appears to be a major mechanism for overcoming lesions induced by various agents including ultraviolet light, X-rays, [³²P]-decay, nitrous acid and psoralen-plus-light. When the lesions caused by the above agents are not repaired by recombinational repair or other accurate repair modes they generally lead to lethality. However a minority of these unrepaired lesions lead to mutations through replication errors or error-prone repair. In higher organisms accumulation of unrepaired lethal lesions or mutations in somatic cells may cause aging. Perpetuation of a species depends on preventing accumulation of these types of defects in the germ line. It is proposed that the recombinational acts occurring during meiosis may largely reflect the recombinational removal or repair of DNA lesions to conserve the germ line. It is further suggested that the repair function of recombination may be as significant as the better studied function of recombination, the generation of diversity. The ubiquitous occurrence of recombination in the biological world implies that it arose very early in evolution. The possibility that recombination initially arose as a repair process is discussed.     

Combining Magnetic Sorting of Mother Cells and Fluctuation Tests to Analyze Genome Instability During Mitotic Cell Aging in Saccharomyces cerevisiae

Saccharomyces cerevisiae has been an excellent model system for examining mechanisms and consequences of genome instability. Information gained from this yeast model is relevant to many organisms, including humans, since DNA repair and DNA damage response factors are well conserved across diverse species. However, S. cerevisiae has not yet been used to fully address whether the rate of accumulating mutations changes with increasing replicative (mitotic) age due to technical constraints. For instance, measurements of yeast replicative lifespan through micromanipulation involve very small populations of cells, which prohibit detection of rare mutations. Genetic methods to enrich for mother cells in populations by inducing death of daughter cells have been developed, but population sizes are still limited by the frequency with which random mutations that compromise the selection systems occur. The current protocol takes advantage of magnetic sorting of surface-labeled yeast mother cells to obtain large enough populations of aging mother cells to quantify rare mutations through phenotypic selections. Mutation rates, measured through fluctuation tests, and mutation frequencies are first established for young cells and used to predict the frequency of mutations in mother cells of various replicative ages. Mutation frequencies are then determined for sorted mother cells, and the age of the mother cells is determined using flow cytometry by staining with a fluorescent reagent that detects bud scars formed on their cell surfaces during cell division. Comparison of predicted mutation frequencies based on the number of cell divisions to the frequencies experimentally observed for mother cells of a given replicative age can then identify whether there are age-related changes in the rate of accumulating mutations. Variations of this basic protocol provide the means to investigate the influence of alterations in specific gene functions or specific environmental conditions on mutation accumulation to address mechanisms underlying genome instability during replicative aging.     

Accumulation of nuclear DNA damage or neuron loss: molecular basis for a new approach to understanding selective neuronal vulnerability in neurodegenerative diseases

According to a long-standing hypothesis, aging is mainly caused by accumulation of nuclear (n) DNA damage in differentiated cells such as neurons due to insufficient nDNA repair during lifetime. In line with this hypothesis it was until recently widely accepted that neuron loss is a general consequence of normal aging, explaining some degree of decline in brain function during aging. However, with the advent of more accurate procedures for counting neurons, it is currently widely accepted that there is widespread preservation of neuron numbers in the aging brain, and the changes that do occur are relatively specific to certain brain regions and types of neurons. Whether accumulation of nDNA damage and decline in nDNA repair is a general phenomenon in the aging brain or also shows cell-type specificity is, however, not known. It has not been possible to address this issue with the biochemical and molecular-biological methods available to study nDNA damage and nDNA repair. Rather, it was the introduction of autoradiographic methods to study quantitatively the relative amounts of nDNA damage (measured as nDNA single-strand breaks) and nDNA repair (measured as unscheduled DNA synthesis) on tissue sections that made it possible to address this question in a cell-type-specific manner under physiological conditions. The results of these studies revealed a formerly unknown inverse relationship between age-related accumulation of nDNA damage and age-related impairment in nDNA repair on the one hand, and the age-related, selective, loss of neurons on the other hand. This inverse relation may not only reflect a fundamental process of aging in the central nervous system but also provide the molecular basis for a new approach to understand the selective neuronal vulnerability in neurodegenerative diseases, particularly Alzheimer's disease.     

Induction of oxidative stress by low doses of lead in human hepatic cell line WRL-68

Even though the molecular mechanisms by which lead induces toxicity and cancer have been intensely studied for many years, its carcinogenic mechanisms are not well understood yet. Several possible mechanisms have been examined to gain understanding on the carcinogenic properties of lead, which include mitogenesis, alteration of gene expression, and oxidative damage, among others. The aim of the present study was to explore the induction of oxidative damage at low lead concentrations using human embryonic hepatic cells WRL-68. Our results showed induction of reactive oxygen species, changes in the superoxide dismutase and catalase activity, as well as an induction of lipidperoxidation and DNA damage. However, after 5 weeks of exposure, these alterations returned to their basal levels. These results taking together indicate that at low concentrations, lead is able to establish an oxidative stress scenario; however under optimal antioxidant defense the oxidative scenario could be abolished through an adaptative process.     

Oxidative DNA damage — The effects of certain genotoxic and operationally non-genotoxic carcinogens

A wide variety of oxidative DNA lesions are commonly present in untreated human and animal DNA. One of these lesions, 8-hydroxydeoxyguanosine, has been shown to lead to base mispairing (mutation) on DNA replication. Other lesions remain to be investigated in this respect. Oxidative DNA lesions on cell replication may, in appropriate circumstances, lead to proto-oncogene activation. Oxidative DNA damage, on fixation, may also lead to cytotoxicity followed by regenerative proliferation. The probable or possible importance of oxidative DNA damage is reviewed for various classes of carcinogens and natural processes, including metal ions, high-energy radiation, miscellaneous chemicals, tumor-promoting agents, polyhydroxyphenols/quinones, lipid metabolism, peroxisome proliferators and thyroid function. It is concluded that although the evidence needs considerable strengthening in many of these examples, the available information indicates the potential importance of oxidative DNA damage in the induction of tumors by these agents. It is also possible that non-cancerous degenerative diseases associated with aging are the result of the accumulation of lesions resulting from unrepaired oxidative DNA damage.     

Mutation frequency and specificity with age in liver, bladder and brain of lacI transgenic mice

Mutation frequency and specificity were determined as a function of age in nuclear DNA from liver, bladder, and brain of Big Blue lacI transgenic mice aged 1.5-25 months. Mutations accumulated with age in liver and accumulated more rapidly in bladder. In the brain a small initial increase in mutation frequency was observed in young animals; however, no further increase was observed in adult mice. To investigate the origin of mutations, the mutational spectra for each tissue and age were determined. DNA sequence analysis of mutant lacI transgenes revealed no significant changes in mutational specificity in any tissue at any age. The spectra of mutations found in aging animals were identical to those in younger animals, suggesting that they originated from a common set of DNA lesions manifested during DNA replication. The data also indicated that there were no significant age-related mutational changes due to oxidative damage, or errors resulting from either changes in the fidelity of DNA polymerase or the efficiency of DNA repair. Hence, no evidence was found to support hypotheses that predict that oxidative damage or accumulation of errors in nuclear DNA contributes significantly to the aging process, at least in these three somatic tissues.     

Age-related changes of bradykinin B1 and B2 receptors in rat heart

Aging is a major risk factor for the development of vascular diseases, such as hypertension and atherosclerosis, that leads to end organ damage and especially heart failure. Bradykinin has been demonstrated to have a cardioprotective role by affecting metabolic processes and tissue perfusion under conditions of myocardial ischemia. Its actions are exerted via the bradykinin B1- and B2-type receptors (B1Rs and B2Rs), but the functional status of these receptors during the aging process is poorly understood. This study aims to investigate whether changes in B1R and B2R gene and protein expression in rat heart are associated with the age-related alterations of cardiac structure and function. Using real-time PCR, we found that B1R mRNA expression increased 2.9-fold in hearts of older rats (24 mo of age) compared with younger rats (3 mo of age), whereas B2R gene expression remained unchanged. Western blot analysis showed that expression of B2R at the protein level is approximately twofold higher in young rats compared with old rats, whereas the B1R protein is approximately twofold higher in old rats compared with young rats. The present results provide clear functional and molecular evidence that indicate age-related changes of bradykinin B1Rs and B2Rs in heart. Because the cardioprotective actions of bradykinin are physiologically mediated via the B2Rs, whereas the B1Rs become induced by tissue damage, these results suggest that age-related decreases in B2R protein levels may leave the heart vulnerable to ischemic damage, and increases in B1R expression and activity may represent a compensatory reaction in aging hearts.