Homoeostatic imbalance during cellular ageing: altered responsiveness.

The inability of normal cells to maintain themselves for ever is a reflection of homoeostatic imbalance and a progressive failure of maintenance. Ageing cells respond less to growth stimulants whereas they show increased sensitivity to toxic agents including antibiotics, phorbol esters, radiation and other physical stresses. No major quantitative and qualitative defects in the receptor systems have been detected that could explain the reasons for altered responsiveness during ageing. Random metabolic defects in the processes involved in maintaining homoeostasis may be critical for causing homoeostatic imbalance, cellular ageing and death.     

Oxidants and antioxidants in proliferative senescence.

In terms of the amount of experimental research it has generated the free radical theory of ageing is one of the most popular hypotheses to explain this ubiquitous phenomenon. From the theory two postulates were derived: either cellular defence mechanisms against free radical-dependent oxidants deteriorate during ageing of cells, or essential, unrepairable damages are imparted to the cell by oxidants regardless of the activity of antioxidant defence systems. The many reports dealing with a putative breakdown in antioxidant defence systems failed to positively support this postulate. However, a minor depletion in cellular glutathione by exposure to a model lipophilic peroxide led to a significant decrement in DNA and protein synthesis. In other words, the glutathione redox cycle is intrinsically fallible with respect to defending the cellular DNA replication system against this model lipophilic peroxide. Interestingly, after ageing in culture cells a partial uncoupling of the NADPH-producing and -consuming systems tends to take place. Experiments involving the addition of antioxidants to the culture medium have failed to significantly extend the lifespan of cultured diploid somatic cells. The level of antioxidants appears to be a modulator rather than a primary determinant of cellular ageing in culture. Several lines of evidence suggest that DNA damages accumulate during ageing of the organism, but no oxidant-related DNA damage has been pinpointed in the cultured cell system. Human mutants with defects in antioxidant enzymes have not shown conclusive signs of accelerated ageing. Cells from patients with Werner's syndrome (progeria of the adult), on the other hand, do not suffer from a defect in their antioxidant defence system, nor do they accumulate more than normal amounts of autofluorescent products resulting from lipid peroxidation. The recent finding that Werner's syndrome constitutes a mutator phenotype may prompt the comparison of oxidant- and ageing-related mutation spectra in order to investigate a mutational theory of ageing as a new derivative from the free radical hypothesis.     

Lamin A-dependent misregulation of adult stem cells associated with accelerated ageing

The premature-ageing disease Hutchinson-Gilford Progeria Syndrome (HGPS) is caused by constitutive production of progerin, a mutant form of the nuclear architectural protein lamin A. Progerin is also expressed sporadically in wild-type cells and has been linked to physiological ageing. Cells from HGPS patients exhibit extensive nuclear defects, including abnormal chromatin structure and increased DNA damage. At the organismal level, HGPS affects several tissues, particularly those of mesenchymal origin. How the cellular defects of HGPS cells lead to the organismal defects has been unclear. Here, we provide evidence that progerin interferes with the function of human mesenchymal stem cells (hMSCs). We find that expression of progerin activates major downstream effectors of the Notch signalling pathway. Induction of progerin in hMSCs changes their molecular identity and differentiation potential. Our results support a model in which accelerated ageing in HGPS patients, and possibly also physiological ageing, is the result of adult stem cell dysfunction and progressive deterioration of tissue functions.     

Role of Fanconi DNA repair pathway in neural stem cell homeostasis

Defects in DNA repair pathways have been involved in collapse of early neurogenesis leading to brain development abnormalities and embryonic lethality. However, consequences of DNA repair defects in adult neural stem and progenitor cells and their potential contribution in ageing phenotype are poorly understood. The Fanconi anaemia (FA) pathway, which functions primarily as a DNA damage response system, has been examined in neural stem and progenitor cells during developmental and adult neurogenesis. We have shown that loss of fanca and fancg specifically provokes neural progenitor apoptosis during forebrain development, related to DNA repair defects, which persists in adulthood leading to depletion of the neural stem cell pool with ageing. In addition, neural stem cells from FA mice had a reduced capacity to self-renew in vitro. Here, we expand upon our recent work and give further data examining possible implication of oxidative stress. Therefore, FA phenotype might be interpreted as a premature ageing of stem cells, DNA damages being among the driving forces of ageing.     

Tendon stem/progenitor cell ageing: Modulation and rejuvenation

Tendon ageing is a complicated process caused by multifaceted pathways and ageing plays a critical role in the occurrence and severity of tendon injury. The role of tendon stem/progenitor cells (TSPCs) in tendon maintenance and regeneration has received increasing attention in recent years. The decreased capacity of TSPCs in seniors contributes to impaired tendon functions and raises questions as to what extent these cells either affect, or cause ageing, and whether these age-related cellular alterations are caused by intrinsic factors or the cellular environment. In this review, recent discoveries concerning the biological characteristics of TSPCs and age-related changes in TSPCs, including the effects of cellular epigenetic alterations and the mechanisms involved in the ageing process, are analyzed. During the ageing process, TSPCs ageing might occur as a natural part of the tendon ageing, but could also result from decreased levels of growth factor, hormone deficits and changes in other related factors. Here, we discuss methods that might induce the rejuvenation of TSPC functions that are impaired during ageing, including moderate exercise, cell extracellular matrix condition, growth factors and hormones; these methods aim to rejuvenate the features of youthfulness with the ultimate goal of improving human health during ageing.     

The role of DNA damage in neural stem cells ageing

Neural stem cells (NSCs) are pluripotent stem cells with the potential to differentiate into a variety of nerve cells. NSCs are susceptible to both intracellular and extracellular insults, thus causing DNA damage. Extracellular insults include ultraviolet, ionizing radiation, base analogs, modifiers, alkyl agents and others, while intracellular factors include Reactive oxygen species (ROS) radicals produced by mitochondria, mismatches that occur during DNA replication, deamination of bases, loss of bases, and more. When encountered with DNA damage, cells typically employ three coping strategies: DNA repair, damage tolerance, and apoptosis. NSCs, like many other stem cells, have the ability to divide, differentiate, and repair DNA damage to prevent mutations from being passed down to the next generation. However, when DNA damage accumulates over time, it will lead to a series of alterations in the metabolism of cells, which will cause cellular ageing. The ageing and exhaustion of neural stem cell will have serious effects on the body, such as neurodegenerative diseases. The purpose of this review is to examine the processes by which DNA damage leads to NSCs ageing and the mechanisms of DNA repair in NSCs.     

A review and appraisal of the DNA damage theory of ageing

Given the central role of DNA in life, and how ageing can be seen as the gradual and irreversible breakdown of living systems, the idea that damage to the DNA is the crucial cause of ageing remains a powerful one. DNA damage and mutations of different types clearly accumulate with age in mammalian tissues. Human progeroid syndromes resulting in what appears to be accelerated ageing have been linked to defects in DNA repair or processing, suggesting that elevated levels of DNA damage can accelerate physiological decline and the development of age-related diseases not limited to cancer. Higher DNA damage may trigger cellular signalling pathways, such as apoptosis, that result in a faster depletion of stem cells, which in turn contributes to accelerated ageing. Genetic manipulations of DNA repair pathways in mice further strengthen this view and also indicate that disruption of specific pathways, such as nucleotide excision repair and non-homologous end joining, is more strongly associated with premature ageing phenotypes. Delaying ageing in mice by decreasing levels of DNA damage, however, has not been achieved yet, perhaps due to the complexity inherent to DNA repair and DNA damage response pathways. Another open question is whether DNA repair optimization is involved in the evolution of species longevity, and we suggest that the way cells from different organisms respond to DNA damage may be crucial in species differences in ageing. Taken together, the data suggest a major role of DNA damage in the modulation of longevity, possibly through effects on cell dysfunction and loss, although understanding how to modify DNA damage repair and response systems to delay ageing remains a crucial challenge.     

Hallmarks and detection techniques of cellular senescence and cellular ageing in immune cells

The ageing of the global population brings about unprecedented challenges. Chronic age‐related diseases in an increasing number of people represent an enormous burden for health and social care. The immune system deteriorates during ageing and contributes to many of these age‐associated diseases due to its pivotal role in pathogen clearance, tissue homeostasis and maintenance. Moreover, in order to develop treatments for COVID‐19, we urgently need to acquire more knowledge about the aged immune system, as older adults are disproportionally and more severely affected. Changes with age lead to impaired responses to infections, malignancies and vaccination, and are accompanied by chronic, low‐degree inflammation, which together is termed immunosenescence. However, the molecular and cellular mechanisms that underlie immunosenescence, termed immune cell senescence, are mostly unknown. Cellular senescence, characterised by an irreversible cell cycle arrest, is thought to be the cause of tissue and organismal ageing. Thus, better understanding of cellular senescence in immune populations at single‐cell level may provide us with insight into how immune cell senescence develops over the life time of an individual. In this review, we will briefly introduce the phenotypic characterisation of aged innate and adaptive immune cells, which also contributes to overall immunosenescence, including subsets and function. Next, we will focus on the different hallmarks of cellular senescence and cellular ageing, and the detection techniques most suitable for immune cells. Applying these techniques will deepen our understanding of immune cell senescence and to discover potential druggable pathways, which can be modulated to reverse immune ageing.     

Reduced levels of ADP-ribosylatable elongation factor-2 in aged and SV40-transformed human cell cultures

The elongation step is involved in the regulation of protein synthesis during the cell cycle, environmental stress, ageing and transformation. Using a diphtheria toxin-mediated assay for measuring the levels of ADP-ribosylatable elongation factor EF-2, we have observed an irreversible decrease of up to 64% in the amount of ADP-ribosylatable EF-2 in normal diploid human fibroblasts MRC-5 undergoing ageing in vitro. However, a similar decrease in low serum-associated G0/G1-arrested cells is reversible both in MRC-5 cells and in their SV40-transformed counterparts. Reduced levels of ADP-ribosylatable EF-2 could account for the slowing-down of protein synthesis during cell cycle arrest and during cellular ageing in culture.     

Defending genome integrity during DNA replication: a proposed role for RecQ family helicases

The RecQ family of DNA helicases have been shown to be important for the maintenance of genomic integrity in all organisms analysed to date. In human cells, representatives of this family include the proteins defective in the cancer predisposition disorder Bloom's syndrome and the premature ageing condition, Werner's syndrome. Several pieces of evidence suggest that RecQ family helicases form associations with one or more of the cellular topoisomerases, and together these heteromeric complexes manipulate DNA structure to effect efficient DNA replication, genetic recombination, or both. Here, we propose that RecQ helicases are required for ensuring that structural abnormalities arising during replication, such as at sites where replication forks encounter DNA lesions, are corrected with high fidelity. In mutants defective in these proteins, not only is replication abnormal, but cells display aberrant responses to DNA-damaging agents or inhibitors of DNA synthesis. We suggest that RecQ helicases may be important for the integration of cellular responses to these insults, such as by linking cell cycle checkpoint responses to recombinational repair. BioEssays 21:286–294, 1999. © 1999 John Wiley & Sons, Inc.     

Comparison of Mitochondrial Mutation Spectra in Ageing Human Colonic Epithelium and Disease: Absence of Evidence for Purifying Selection in Somatic Mitochondrial DNA Point Mutations

Human ageing has been predicted to be caused by the accumulation of molecular damage in cells and tissues. Somatic mitochondrial DNA (mtDNA) mutations have been documented in a number of ageing tissues and have been shown to be associated with cellular mitochondrial dysfunction. It is unknown whether there are selective constraints, which have been shown to occur in the germline, on the occurrence and expansion of these mtDNA mutations within individual somatic cells. Here we compared the pattern and spectrum of mutations observed in ageing human colon to those observed in the general population (germline variants) and those associated with primary mtDNA disease. The pathogenicity of the protein encoding mutations was predicted using a computational programme, MutPred, and the scores obtained for the three groups compared. We show that the mutations associated with ageing are randomly distributed throughout the genome, are more frequently non-synonymous or frameshift mutations than the general population, and are significantly more pathogenic than population variants. Mutations associated with primary mtDNA disease were significantly more pathogenic than ageing or population mutations. These data provide little evidence for any selective constraints on the occurrence and expansion of mtDNA mutations in somatic cells of the human colon during human ageing in contrast to germline mutations seen in the general population.     

Mouse models of mitochondrial DNA defects and their relevance for human disease

Qualitative and quantitative changes in mitochondrial DNA (mtDNA) have been shown to be common causes of inherited neurodegenerative and muscular diseases, and have also been implicated in ageing. These diseases can be caused by primary mtDNA mutations, or by defects in nuclear‐encoded mtDNA maintenance proteins that cause secondary mtDNA mutagenesis or instability. Furthermore, it has been proposed that mtDNA copy number affects cellular tolerance to environmental stress. However, the mechanisms that regulate mtDNA copy number and the tissue‐specific consequences of mtDNA mutations are largely unknown. As post‐mitotic tissues differ greatly from proliferating cultured cells in their need for mtDNA maintenance, and as most mitochondrial diseases affect post‐mitotic cell types, the mouse is an important model in which to study mtDNA defects. Here, we review recently developed mouse models, and their contribution to our knowledge of mtDNA maintenance and its role in disease.     

Antizyme suppression leads to an increment of the cellular redox potential and an induction of HIF-1alpha: its involvement in resistance to gamma-radiation

The mammalian antizyme (AZ) promotes ubiqutin-independent degradation of ornithine decarboxylase, a key enzyme in polyamine biosynthesis. This study shows that AZ suppression in human lung carcinoma A549 cells caused growth defects and death, but made the cells resistant to DNA damaging agents such as gamma-radiation and cisplatin. In these cells, the cellular redox potential (glutathione/glutathione disulfide [GSH/GSSG] ratio) was increased and thus intracellular reactive oxygen species were severely diminished, which might cause growth defects and cell death. The increase of cellular redox potential was mainly caused by dramatic increase of the cytoplasmic nicotinamide adenine dinucleotide phosphate (NADP)(+)-dependent isocitrate dehydrogenase, which generates the reducing equivalents NADPH. In the AZ-suppressed cells, the hypoxia inducible factor 1alpha (HIF-1alpha) was also increased. As in other cases which showed an increment of HIF-1alpha and the cellular redox potential, the AZ-suppressed cells showed resistance to gamma-radiation and anticancer drugs. Therefore, these facts might be considered as important for the use of radio- and chemotherapy on tumor cells which show an unbalance in their polyamine levels.     

Age-related changes in immunity: implications for vaccination in the elderly

Average life expectancy is continuously rising in all developed countries, leading to an ever-increasing elderly population. Of the many functions of the body affected by the complex process of ageing, the immune system in particular undergoes various changes, collectively termed immunosenescence. As a result, elderly people are more susceptible to infections and are frequently less protected by vaccines. This review summarises the effect of ageing on immunity, emphasising the age-associated changes within T and B cells at a molecular and cellular level. Furthermore, it discusses strategies, such as the addition of immunostimulatory adjuvants and the use of potent antigen-delivery systems, that may counteract age-related defects in immune responses to vaccination. A proper understanding of how immunological memory is affected by ageing, and the introduction of strategies to ameliorate vaccine efficacy in the elderly, might reduce the incidence and the severity of infectious disease within this fragile age group and have a strong impact on the quality of life of elderly individuals.     

Autophagy in neuronal cell loss: a road to death

The regulation of ageing has been extensively studied in divergent animal model systems including worms, flies and mice. However, little is known about the cellular pathways that mediate the death of these organisms. Analysing major cellular changes in the ageing nematode Caenorhabditis elegans has revealed a gradual, progressive deterioration of different tissues except for the nervous system, which remarkably preserves its integrity even in advanced old age. In addition, genetic data have shown that, in C. elegans and in the fruit fly Drosophila melanogaster, lifespan is controlled by signals derived from neurons and acting throughout adulthood. Organismal death thus seems to be a consequence of the decline of specific neurons. Accumulating evidence demonstrates that late onset of neuronal cell loss generally occurs via autophagy, a process in which eukaryotic cells self-digest parts of their contents during development or to survive starvation. Here we suggest that overactivation of autophagy in the cells of the nervous system is the eventual cause of "physiological" death.     

The effects of reactive oxygen and nitrogen species during yeast replicative ageing

Free radicals are considered the most important cause of cellular ageing. We have investigated ageing process in the yeast Saccharomyces cerevisiae. We have compared the wild type strain with the mutant cells with constitutively active Ras oncogen, which generates increased amounts of free radicals. Increased generation of oxygen-derived free radicals resulted in the Ras mutant cells accumulation of lipofuscin-like pigments during ageing. Ageing wild type cells did not accumulate lipofuscin-like pigments. This is quite unique feature among known biological models. It may be caused by increased concentration of alpha tocopherol (the most prominent lipophilic antioxidant) in the wild type cells. In contrast, the Ras mutant cells contained decreased levels of alpha tocopherol even in the young cells. This observation indicates that the increased free radical generation can overwhelm the endogenous antioxidant system. We have documented the involvement of nitrogen-derived free radicals in the yeast metabolism. Protein nitrotyrosine, a marker of the reactive nitrogen species, has significantly increased in the senescent Ras mutant cells. The wild type cells contained basic level of nitrotyrosine corresponding to its concentration found in non-activated mammalian macrophages.     

Fibroblast subpopulations: a developmental approach of skin physiology and ageing

Skin is an organ whose function is far beyond a physical barrier between the inside and the outside of the body. Skin as the whole organism is subjected to ageing which concerns skin mostly in its dermal and deepest component which is also its matricial component. The dermis is a tissue rich in matricial elements and poor in cellular content and it is generally admitted that modifications occurring in the matrix are those which mostly contribute to skin ageing, by altering its biomechanical properties. Therefore it is common to address questions related to skin ageing by considering alterations in matrix molecules like collagen. Actually the dermis is a complex tissue both matricial and cellular and is divided between a superficial dermis close to epidermis and a deep dermis much thicker and histologically different. Several years ago we have undertaken investigations related to fibroblasts which are the cells responsible for the formation and maintenance of the dermis, aiming at isolation, culture and characterization of the fibroblasts from the superficial dermis also called papillary dermis and fibroblasts from the deep dermis also called reticular dermis. We were able to show that these fibroblasts in classical culture on plastic exhibit very different morphologies associated with different secretion properties and we have confirmed and expanded such observations revealing different phenotypes by incorporating these cells in reconstructed skin which allows the reproduction of a three-dimensional architecture recalling skin in vivo especially after grafting onto the nude mouse. We also raise the question of how these two dermal regions appear during the formation of the dermis and the question of their fate during ageing. Progress in solving these questions would certainly appear to be very useful for a better understanding of skin physiology and ageing and would hopefully provide new strategies in anti-ageing research.