Functional analyses of human DNA repair proteins important for aging and genomic stability using yeast genetics

Model systems have been extremely useful for studying various theories of aging. Studies of yeast have been particularly helpful to explore the molecular mechanisms and pathways that affect aging at the cellular level in the simple eukaryote. Although genetic analysis has been useful to interrogate the aging process, there has been both interest and debate over how functionally conserved the mechanisms of aging are between yeast and higher eukaryotes, especially mammalian cells. One area of interest has been the importance of genomic stability for age-related processes, and the potential conservation of proteins and pathways between yeast and human. Translational genetics have been employed to examine the functional roles of mammalian proteins using yeast as a pliable model system. In the current review recent advancements made in this area are discussed, highlighting work which shows that the cellular functions of human proteins in DNA repair and maintenance of genomic stability can be elucidated by genetic rescue experiments performed in yeast.     

Regulatory molecular mechanisms of the neurochemical processes. History and the present time

Multiple neurochemical mechanisms (neurotransmitters, regulatory peptides, neurotrophic growth factors, and proteins of the signaling transducer systems) maintain the integrity of nerve cell circuits, facilitate the responses to environmental demands and promote the recovery of a function after injury. The recent application of modern approaches of molecular and cellular biology to the problem of "diseased (bad) brain" reveals a remarkable capacity within brain cells for adaptation to aging and resistance to a disease. The death of neurons in different neurological disorders involves apoptotic biochemical cascades leading to mitochondrial alterations, upstream pro-apoptotic effectors, and caspases activation. At the cellular level, neuronal apoptosis in ischemic and neurodegenerative disorders may be triggered by oxidative stress, mitochondrial compromise and disruption of calcium homeostasis. Both genetic and environmental factors, and the aging process itself, contribute to initiation of such neuronal apoptosis. Neuroprotective (antiapoptotic) signal pathways involving neurotrophic factors, neuropeptides, and mediators able to counteract with effects of aging and genetic predisposition in experimental models and clinical events of neuro-destructive disorders.     

The impact of age on oncogenic potential: tumor‐initiating cells and the brain microenvironment

Paradoxically, aging leads to both decreased regenerative capacity in the brain and an increased risk of tumorigenesis, particularly the most common adult‐onset brain tumor, glioma. A shared factor contributing to both phenomena is thought to be age‐related alterations in neural progenitor cells (NPCs), which function normally to produce new neurons and glia, but are also considered likely cells of origin for malignant glioma. Upon oncogenic transformation, cells acquire characteristics known as the hallmarks of cancer, including unlimited replication, altered responses to growth and anti‐growth factors, increased capacity for angiogenesis, potential for invasion, genetic instability, apoptotic evasion, escape from immune surveillance, and an adaptive metabolic phenotype. The precise molecular pathogenesis and temporal acquisition of these malignant characteristics is largely a mystery. Recent studies characterizing NPCs during normal aging, however, have begun to elucidate mechanisms underlying the age‐associated increase in their malignant potential. Aging cells are dependent upon multiple compensatory pathways to maintain cell cycle control, normal niche interactions, genetic stability, programmed cell death, and oxidative metabolism. A few multi‐functional proteins act as ‘critical nodes’ in the coordination of these various cellular activities, although both intracellular signaling and elements within the brain environment are critical to maintaining a balance between senescence and tumorigenesis. Here, we provide an overview of recent progress in our understanding of how mechanisms underlying cellular aging inform on glioma pathogenesis and malignancy.     

Recent Progress in Regulation of Aging by Insulin/IGF-1 Signaling in Caenorhabditis elegans

Caenorhabditis elegans has been used as a major model organism to identify genetic factors that regulate organismal aging and longevity. Insulin/insulin-like growth factor 1 (IGF-1) signaling (IIS) regulates aging in many species, ranging from nematodes to humans. C. elegans is a nonpathogenic genetic nematode model, which has been extensively utilized to identify molecular and cellular components that function in organismal aging and longevity. Here, we review the recent progress in the role of IIS in aging and longevity, which involves direct regulation of protein and RNA homeostasis, stress resistance, metabolism and the activities of the endocrine system. We also discuss recently identified genetic factors that interact with canonical IIS components to regulate aging and health span in C. elegans. We expect this review to provide valuable insights into understanding animal aging, which could eventually help develop anti-aging drugs for humans.     

Cell culture aging

Summary Cellular research in aging has been stimulated by the observation that human diploid cells have a limited number of cell divisions in culture. This loss of cellular proliferation (in vitro senescence) has been extensively studied by biochemical, clonal, and genetic analysis. Studies of human skin fibroblast cultures have revealed thatin vitro senescense is related toin vivo human cellular aging. Recently differentiated cells have been proposed for aging studies. These cells may provide additional information on aging since alterations ofin vitro cellular functions may be related to thein vivo behavior of specific differented cell types.     

Integrating cellular senescence with the concept of damage accumulation in aging: Relevance for clearance of senescent cells

Understanding the aging process and ways to manipulate it is of major importance for biology and medicine. Among the many aging theories advanced over the years, the concept most consistent with experimental evidence posits the buildup of numerous forms of molecular damage as a foundation of the aging process. Here, we discuss that this concept integrates well with recent findings on cellular senescence, offering a novel view on the role of senescence in aging and age‐related disease. Cellular senescence has a well‐established role in cellular aging, but its impact on the rate of organismal aging is less defined. One of the most prominent features of cellular senescence is its association with macromolecular damage. The relationship between cell senescence and damage concerns both damage as a molecular signal of senescence induction and accelerated accumulation of damage in senescent cells. We describe the origin, regulatory mechanisms, and relevance of various damage forms in senescent cells. This view on senescent cells as carriers and inducers of damage puts new light on senescence, considering it as a significant contributor to the rise in organismal damage. Applying these ideas, we critically examine current evidence for a role of cellular senescence in aging and age‐related diseases. We also discuss the differential impact of longevity interventions on senescence burden and other types of age‐related damage. Finally, we propose a model on the role of aging‐related damage accumulation and the rate of aging observed upon senescent cell clearance.     

Donor age-dependent acceleration of cellular aging by repeated ultraviolet A irradiation of human dermal fibroblasts derived from a single donor

The relationship between cellular aging and aging of entire organisms has been studied extensively. The findings are confusing, however, and no clear relationships have been demonstrated. The conflicting data may be due to individual differences among the donors of the studied cells. It is crucial to identify the changes in cellular properties that are the result of the aging process. Here, we used human dermal fibroblast cell lines established from a single donor at different ages to assess the influence of ultraviolet A (UVA) on cellular aging. These cell lines have the same genetic background and were obtained from a restricted body region. The results indicated that cellular aging was accelerated by UVA irradiation in a donor age-dependent manner. The ratio of lifespan shortening increased with donor age. Increased donor age not only decreased cell division, but also increased the growth arrest response to UVA irradiation. The characteristics of the cultured cells reflected the age-related changes in dermal fibroblasts.     

Dietary antiaging phytochemicals and mechanisms associated with prolonged survival

Aging is well-known an inevitable process that is influenced by genetic, lifestyle and environmental factors. However, the exact mechanisms underlying the aging process are not well understood. Increasing evidence shows that aging is highly associated with chronic increase in reactive oxygen species (ROS), accumulation of a low-grade proinflammatory phenotype and reduction in age-related autophagy, suggesting that these factors may play important roles in promoting aging. Indeed, reduction of ROS and low-grade inflammation and promotion of autophagy by calorie restriction or other dietary manipulation can extend lifespan in a wide spectrum of model organisms. Interestingly, recent studies show that some food-derived small molecules, also called phytochemicals, can extend lifespan in various animal species. In this paper, we review several recently identified potential antiaging phytochemicals that have been studied in cells, animals and humans and further highlight the cellular and molecular mechanisms underlying the antiaging actions by these molecules.     

Adult Sox2+ stem cell exhaustion in mice results in cellular senescence and premature aging

Aging is characterized by a gradual functional decline of tissues with age. Adult stem and progenitor cells are responsible for tissue maintenance, repair, and regeneration, but during aging, this population of cells is decreased or its activity is reduced, compromising tissue integrity and causing pathologies that increase vulnerability, and ultimately lead to death. The causes of stem cell exhaustion during aging are not clear, and whether a reduction in stem cell function is a cause or a consequence of aging remains unresolved. Here, we took advantage of a mouse model of induced adult Sox2+ stem cell depletion to address whether accelerated stem cell depletion can promote premature aging. After a short period of partial repetitive depletion of this adult stem cell population in mice, we observed increased kyphosis and hair graying, and reduced fat mass, all of them signs of premature aging. It is interesting that cellular senescence was identified in kidney after this partial repetitive Sox2+ cell depletion. To confirm these observations, we performed a prolonged protocol of partial repetitive depletion of Sox2+ cells, forcing regeneration from the remaining Sox2+ cells, thereby causing their exhaustion. Senescence specific staining and the analysis of the expression of genetic markers clearly corroborated that adult stem cell exhaustion can lead to cellular senescence induction and premature aging.     

Identification of evolutionarily conserved genetic regulators of cellular aging

To identify new genetic regulators of cellular aging and senescence, we performed genome-wide comparative RNA profiling with selected human cellular model systems, reflecting replicative senescence, stress-induced premature senescence, and distinct other forms of cellular aging. Gene expression profiles were measured, analyzed, and entered into a newly generated database referred to as the GiSAO database. Bioinformatic analysis revealed a set of new candidate genes, conserved across the majority of the cellular aging models, which were so far not associated with cellular aging, and highlighted several new pathways that potentially play a role in cellular aging. Several candidate genes obtained through this analysis have been confirmed by functional experiments, thereby validating the experimental approach. The effect of genetic deletion on chronological lifespan in yeast was assessed for 93 genes where (i) functional homologues were found in the yeast genome and (ii) the deletion strain was viable. We identified several genes whose deletion led to significant changes of chronological lifespan in yeast, featuring both lifespan shortening and lifespan extension. In conclusion, an unbiased screen across species uncovered several so far unrecognized molecular pathways for cellular aging that are conserved in evolution.     

Nonculturable bacteria: programmed survival forms or cells at death's door?

Upon starvation and growth arrest, Escherichia coli cells gradually lose their ability to reproduce. These apparently sterile/nonculturable cells initially remain intact and metabolically active and the underlying molecular mechanism behind this sterility is something of an enigma in bacteriology. Three different models have been proposed to explain this phenomenon. The first theory suggests that starving cells become nonculturable due to cellular deterioration, are moribund, and show some of the same signs of senescence as aging organisms. The two other theories suggest that genetically programmed pathways, rather than stochastic deterioration, trigger nonculturability. One "program" theory suggests that nonculturability is the culmination of an adaptive pathway generating dormant survival forms, similar to spore formation in differentiating bacteria. The other "program" theory states that starved cells lose viability due to activation of genetic modules mediating programmed cell death. The different models will be reviewed and evaluated in light of recent data on the physiology and molecular biology of growth-arrested E. coli cells.     

Recent developments in the intracellular degradation of oxidized proteins

The accumulation of oxidized proteins in cells and tissues is a feature of a number of age-related diseases and may also occur as a result of the aging process itself. In this article we review recent advances in our understanding of the cellular degradation of oxidized proteins directing our attention primarily to information which directly bears on the behavior of intact eukaryotic cells. We summarize new work on the key intracellular degradative machineries, proteasomes and lysosomes and examine evidence implicating an increase in protein hydrophobicity as the primary signal to the proteasome to initiate degradation. The data identifying the proteasome as the main route of degradation of oxidized proteins is examined, as well as recent data investigating changes in proteasome function after exposure of cells to oxidants and the altered catabolism of oxidized proteins in aging cells. Evidence for the cooperation between the lysosomal and proteasomal systems in the degradation of oxidized proteins is discussed. We conclude that the cellular catabolism of oxidized proteins may be a more complex process than it first appeared and suggest key issues that need to be resolved to improve our understanding of this important process.     

Negative growth effectors and cellular senescence.

Current studies suggest a genetic program governs the lifespan of each organism. Using cellular senescence as a model system, components of this program for aging have been sought. Human diploid fibroblasts, upon reaching senescence, express active inhibitors of DNA synthesis. It is believed that such inhibitors could be members of a new family of negative growth effectors involved in the pathway to senescence. Factors capable of inhibiting DNA synthesis in a similar manner have also been identified from human quiescent fibroblasts and liver cells as well as from quiescent rodent liver cells. The relationship of these inhibitors to previously identified negative growth effectors and aging are discussed.     

Fidelity of protein synthesis does not decline during aging of cultured human fibroblasts

To ascertain whether the fidelity of protein synthesis declines during cellular aging in vitro, we have developed a cell-free protein synthesizing system from cultured human fibroblasts which actively incorporates phenylalanine into acid-insoluble material upon addition of poly (U). The accuracy of poly(U)-directed protein synthesis was determined by comparing the ratio of leucine to phenylalanine incorporation in extracts of early- and late-passage fibroblasts derived from normal persons and from subjects with two genetic disorders of premature aging, progeria, and Werner syndrome. The results show no decline in translational fidelity at late passage or in prematurely aging cells, and thus fail to support the error catastrophe theory of cellular aging.     

Effect of stem cell turnover rates on protection against cancer and aging

Tissue stem cells are responsible for replenishing and maintaining a population of cells which make up a functioning organ. They divide by asymmetric cell division where one daughter remains a stem cell while the other daughter becomes a transit cell, which divides a defined number of times and differentiates. A fully differentiated cell has a finite life-span. A tissue can be maintained by various strategies. Stem cells can divide often and differentiated cells die often (fast turnover). Alternatively, stem cells can divide infrequently, and the differentiated cells are long lived (slow turnover). Genetic alterations and mutations can interfere with tissue homoeostasis. Mutations can induce senescence and apoptosis, and this can result in a reduction of the number of functioning tissue cells which could correlate with tissue aging. Alternatively, mutations can result in the carcinogenic transformation of cells and the formation of a tumour. Using mathematical models, I find that the cellular turnover rate affects the ability of genetic alterations to induce aging and the development of cancer. If mutations occur as a result of errors during cell division, the model suggests that a low cellular turnover rate protects both against aging and the development of cancer. On the other hand, if mutations occur independent from cell division (e.g. if DNA is hit by damaging agents), I find that a high cellular turnover rate protects against aging, while it promotes the development of cancer. Implications for optimal tissue design are discussed.     

Cloning adult animals - what is the genetic age of the clones?

The rapid progress in cloning research along with its many ramifications will soon have a significant beneficial impact on basic research, agriculture, and biomedicine. However, for the nuclear transfer technology to reach its fullest potential, it is important to understand whether the cloning procedure can reverse cellular aging and generate clones with normal genetic and physiological age, similar to those produced from natural reproduction. Telomere shortening is believed to correlate with cellular aging both in vitro and in vivo. Telomere lengths in cells of cloned individuals thus may reflect their genetic age. However, controversies have developed over whether the eroded telomere in somatic cells used for nuclear transfer can be restored during the cloning process.     

Cellular senescence: A reflection of normal growth control,differentiation, or aging?

Normal cells, with few exceptions, cannot proliferate indefinitely. Cell populations--in vivo and in culture--generally undergo only a limited number of doublings before proliferation invariably and irreversibly ceases. This process has been termed the finite lifespan phenotype or cellular senescence. There is long-standing, albeit indirect, evidence that cellular senescence plays an important role in complex biological processes as diverse as normal growth control, differentiation, development, aging, and tumorigenesis. In recent years, it has been possible to develop a molecular framework for understanding some of the fundamental features of cellular senescence. This framework derives primarily from the physiology, genetics, and molecular biology of cells undergoing senescence in culture. Our understanding of senescence, and the mechanisms that control it, is still in its infancy. Nonetheless, recent data raise some intriguing possibilities regarding potential molecular bases for the links between senescence in culture and normal and abnormal growth control, differentiation, and aging.     

Cellular and molecular aspects of immune system aging

Summary We begin with a brief discussion of the importance and advantages of immune studies to the problem of aging. This is followed by a short over-view of immune system aging at the systemic level. The major portion of the article is a review of observations, both at the cellular and molecular level, of changes in aging immune cells, with sections on intercellular communication, membrane phenomena, cyclic nucleotides, and molecular genetic changes. Publication number 043 from Wadsworth GRECC.