Meeting Summary of The NYO3 5th NO-Age/AD Meeting and the 1st Norway-UK Joint Meeting on Aging and Dementia: Recent Progress on the Mechanisms and Interventional Strategies.

Unhealthy aging poses a global challenge with profound healthcare and socioeconomic implications. Slowing down the aging process offers a promising approach to reduce the burden of a number of age-related diseases, such as dementia, and promoting healthy longevity in the old population. In response to the challenge of the aging population and with a view to the future, Norway and the United Kingdom are fostering collaborations, supported by a "Money Follows Cooperation agreement" between the 2 nations. The inaugural Norway-UK joint meeting on aging and dementia gathered leading experts on aging and dementia from the 2 nations to share their latest discoveries in related fields. Since aging is an international challenge, and to foster collaborations, we also invited leading scholars from 11 additional countries to join this event. This report provides a summary of the conference, highlighting recent progress on molecular aging mechanisms, genetic risk factors, DNA damage and repair, mitophagy, autophagy, as well as progress on a series of clinical trials (eg, using NAD+ precursors). The meeting facilitated dialogue among policymakers, administrative leaders, researchers, and clinical experts, aiming to promote international research collaborations and to translate findings into clinical applications and interventions to advance healthy aging.

The central role of DNA damage in immunosenescence.

Ageing is the biggest risk factor for the development of multiple chronic diseases as well as increased infection susceptibility and severity of diseases such as influenza and COVID-19. This increased disease risk is linked to changes in immune function during ageing termed immunosenescence. Age-related loss of immune function, particularly in adaptive responses against pathogens and immunosurveillance against cancer, is accompanied by a paradoxical gain of function of some aspects of immunity such as elevated inflammation and increased incidence of autoimmunity. Of the many factors that contribute to immunosenescence, DNA damage is emerging as a key candidate. In this review, we discuss the evidence supporting the hypothesis that DNA damage may be a central driver of immunosenescence through senescence of both immune cells and cells of non-haematopoietic lineages. We explore why DNA damage accumulates during ageing in a major cell type, T cells, and how this may drive age-related immune dysfunction. We further propose that existing immunosenescence interventions may act, at least in part, by mitigating DNA damage and restoring DNA repair processes (which we term "genoprotection"). As such, we propose additional treatments on the basis of their evidence for genoprotection, and further suggest that this approach may provide a viable therapeutic strategy for improving immunity in older people.

New hallmarks of ageing: a 2022 Copenhagen ageing meeting summary.

Genomic instability, telomere attrition, epigenetic alterations, mitochondrial dysfunction, loss of proteostasis, deregulated nutrient-sensing, cellular senescence, stem cell exhaustion, and altered intercellular communication were the original nine hallmarks of ageing proposed by López-Otín and colleagues in 2013. The proposal of these hallmarks of ageing has been instrumental in guiding and pushing forward research on the biology of ageing. In the nearly past 10 years, our in-depth exploration on ageing research has enabled us to formulate new hallmarks of ageing which are compromised autophagy, microbiome disturbance, altered mechanical properties, splicing dysregulation, and inflammation, among other emerging ones. Amalgamation of the 'old' and 'new' hallmarks of ageing may provide a more comprehensive explanation of ageing and age-related diseases, shedding light on interventional and therapeutic studies to achieve healthy, happy, and productive lives in the elderly.

Crosstalk Between Senescent Bone Cells and the Bone Tissue Microenvironment Influences Bone Fragility During Chronological Age and in Diabetes.

Bone is a complex organ serving roles in skeletal support and movement, and is a source of blood cells including adaptive and innate immune cells. Structural and functional integrity is maintained through a balance between bone synthesis and bone degradation, dependent in part on mechanical loading but also on signaling and influences of the tissue microenvironment. Bone structure and the extracellular bone milieu change with age, predisposing to osteoporosis and increased fracture risk, and this is exacerbated in patients with diabetes. Such changes can include loss of bone mineral density, deterioration in micro-architecture, as well as decreased bone flexibility, through alteration of proteinaceous bone support structures, and accumulation of senescent cells. Senescence is a state of proliferation arrest accompanied by marked morphological and metabolic changes. It is driven by cellular stress and serves an important acute tumor suppressive mechanism when followed by immune-mediated senescent cell clearance. However, aging and pathological conditions including diabetes are associated with accumulation of senescent cells that generate a pro-inflammatory and tissue-destructive secretome (the SASP). The SASP impinges on the tissue microenvironment with detrimental local and systemic consequences; senescent cells are thought to contribute to the multimorbidity associated with advanced chronological age. Here, we assess factors that promote bone fragility, in the context both of chronological aging and accelerated aging in progeroid syndromes and in diabetes, including senescence-dependent alterations in the bone tissue microenvironment, and glycation changes to the tissue microenvironment that stimulate RAGE signaling, a process that is accelerated in diabetic patients. Finally, we discuss therapeutic interventions targeting RAGE signaling and cell senescence that show promise in improving bone health in older people and those living with diabetes.

Interconnections between Inflammageing and Immunosenescence during Ageing.

Acute inflammation is a physiological response to injury or infection, with a cascade of steps that ultimately lead to the recruitment of immune cells to clear invading pathogens and heal wounds. However, chronic inflammation arising from the continued presence of the initial trigger, or the dysfunction of signalling and/or effector pathways, is harmful to health. While successful ageing in older adults, including centenarians, is associated with low levels of inflammation, elevated inflammation increases the risk of poor health and death. Hence inflammation has been described as one of seven pillars of ageing. Age-associated sterile, chronic, and low-grade inflammation is commonly termed inflammageing-it is not simply a consequence of increasing chronological age, but is also a marker of biological ageing, multimorbidity, and mortality risk. While inflammageing was initially thought to be caused by "continuous antigenic load and stress", reports from the last two decades describe a much more complex phenomenon also involving cellular senescence and the ageing of the immune system. In this review, we explore some of the main sources and consequences of inflammageing in the context of immunosenescence and highlight potential interventions. In particular, we assess the contribution of cellular senescence to age-associated inflammation, identify patterns of pro- and anti-inflammatory markers characteristic of inflammageing, describe alterations in the ageing immune system that lead to elevated inflammation, and finally assess the ways that diet, exercise, and pharmacological interventions can reduce inflammageing and thus, improve later life health.

Intercellular Transfer of Mitochondria between Senescent Cells through Cytoskeleton-Supported Intercellular Bridges Requires mTOR and CDC42 Signalling.

Cellular senescence is a state of irreversible cell proliferation arrest induced by various stressors including telomere attrition, DNA damage, and oncogene induction. While beneficial as an acute response to stress, the accumulation of senescent cells with increasing age is thought to contribute adversely to the development of cancer and a number of other age-related diseases, including neurodegenerative diseases for which there are currently no effective disease-modifying therapies. Non-cell-autonomous effects of senescent cells have been suggested to arise through the SASP, a wide variety of proinflammatory cytokines, chemokines, and exosomes secreted by senescent cells. Here, we report an additional means of cell communication utilised by senescent cells via large numbers of membrane-bound intercellular bridges-or tunnelling nanotubes (TNTs)-containing the cytoskeletal components actin and tubulin, which form direct physical connections between cells. We observe the presence of mitochondria in these TNTs and show organelle transfer through the TNTs to adjacent cells. While transport of individual mitochondria along single TNTs appears by time-lapse studies to be unidirectional, we show by differentially labelled co-culture experiments that organelle transfer through TNTs can occur between different cells of equivalent cell age, but that senescent cells, rather than proliferating cells, appear to be predominant mitochondrial donors. Using small molecule inhibitors, we demonstrate that senescent cell TNTs are dependent on signalling through the mTOR pathway, which we further show is mediated at least in part through the downstream actin-cytoskeleton regulatory factor CDC42. These findings have significant implications for the development of senomodifying therapies, as they highlight the need to account for local direct cell-cell contacts as well as the SASP in order to treat cancer and diseases of ageing in which senescence is a key factor.

Structural basis of the anti-ageing effects of polyphenolics: mitigation of oxidative stress.

Ageing, and particularly the onset of age-related diseases, is associated with tissue dysfunction and macromolecular damage, some of which can be attributed to accumulation of oxidative damage. Polyphenolic natural products such as stilbenoids, flavonoids and chalcones have been shown to be effective at ameliorating several age-related phenotypes, including oxidative stress, inflammation, impaired proteostasis and cellular senescence, both in vitro and in vivo. Here we aim to identify the structural basis underlying the pharmacology of polyphenols towards ROS and related biochemical pathways involved in age-related disease. We compile and describe SAR trends across different polyphenol chemotypes including stilbenoids, flavonoids and chalcones, review their different molecular targets and indications, and identify common structural ground between chemotypes and mechanisms of action. In particular, we focus on the structural requirements for the direct scavenging of reactive oxygen/nitrogen species such as radicals as well as coordination of a broader antioxidant response. We further suggest that it is important to consider multiple (rather than single) biological activities when identifying and developing new medicinal chemistry entities with utility in modulating complex biological properties such as cell ageing.

Correction to: Optimisation of a screening platform for determining IL-6 inflammatory signalling in the senescence-associated secretory phenotype (SASP).

In the original publication of the article, Fig. 2 was published incorrectly. The corrected Figure is given below. The original article has been corrected.

Optimisation of a screening platform for determining IL-6 inflammatory signalling in the senescence-associated secretory phenotype (SASP).

Cellular senescence has been shown to be sufficient for the development of multiple age-related pathologies. Senescent cells adopt a secretory phenotype (the SASP) which comprises a large number of pro-inflammatory cytokines, chemokines and proteases. The SASP itself is thought to be causative in many pathologies of age-related diseases, and there is growing interest in developing seno-modifying agents that can suppress the SASP. However, in order to identify new agents, it is necessary to conduct moderate to high throughput screening with robust assays for the required outcome. Here, we describe optimisation and validation of a cell-based biosensor HEK cell line for measurement of IL-6 concentrations within the range secreted into conditioned medium by primary senescent fibroblasts, adapted for a 384 well plate format suitable for library screening applications. We further show that the assay can measure changes in IL-6 secretion dependent on cell population age, and that the assay is responsive to mTOR inhibition in the senescent cells, which reduces the SASP, including IL-6. Hence, we propose that this optimised biosensor, which we term HEK-SASP, may prove of value in studies requiring robust, renewable and relatively inexpensive assays for measuring SASP factors.

Generation of a novel model of primary human cell senescence through Tenovin-6 mediated inhibition of sirtuins.

Cell senescence, a state of cell cycle arrest and altered metabolism with enhanced pro-inflammatory secretion, underlies at least some aspects of organismal ageing. The sirtuin family of deacetylases has been implicated in preventing premature ageing; sirtuin overexpression or resveratrol-mediated activation of sirtuins increase longevity. Here we show that sirtuin inhibition by short-term, low-dose treatment with the experimental anti-cancer agent Tenovin-6 (TnV6) induces cellular senescence in primary human fibroblasts. Treated cells cease proliferation and arrest in G1 of the cell cycle, with elevated p21 levels, DNA damage foci, high mitochondrial and lysosomal load and increased senescence-associated ß galactosidase activity, together with actin stress fibres and secretion of IL-6 (indicative of SASP upregulation). Consistent with a histone deacetylation role of SIRT1, we find nuclear enlargement, possibly resulting from chromatin decompaction on sirtuin inhibition. These findings highlight TnV6 as a drug that may be useful in clinical settings where acute induction of cell senescence would be beneficial, but also provide the caveat that even supposedly non-genotoxic anticancer drugs can have unexpected and efficacy-limiting impacts on non-transformed cells.

mTORC Inhibitors as Broad-Spectrum Therapeutics for Age-Related Diseases.

Chronological age represents the greatest risk factor for many life-threatening diseases, including neurodegeneration, cancer, and cardiovascular disease; ageing also increases susceptibility to infectious disease. Current efforts to tackle individual diseases may have little impact on the overall healthspan of older individuals, who would still be vulnerable to other age-related pathologies. However, recent progress in ageing research has highlighted the accumulation of senescent cells with chronological age as a probable underlying cause of pathological ageing. Cellular senescence is an essentially irreversible proliferation arrest mechanism that has important roles in development, wound healing, and preventing cancer, but it may limit tissue function and cause widespread inflammation with age. The serine/threonine kinase mTOR (mechanistic target of rapamycin) is a regulatory nexus that is heavily implicated in both ageing and senescence. Excitingly, a growing body of research has highlighted rapamycin and other mTOR inhibitors as promising treatments for a broad spectrum of age-related pathologies, including neurodegeneration, cancer, immunosenescence, osteoporosis, rheumatoid arthritis, age-related blindness, diabetic nephropathy, muscular dystrophy, and cardiovascular disease. In this review, we assess the use of mTOR inhibitors to treat age-related pathologies, discuss possible molecular mechanisms of action where evidence is available, and consider strategies to minimize undesirable side effects. We also emphasize the urgent need for reliable, non-invasive biomarkers of senescence and biological ageing to better monitor the efficacy of any healthy ageing therapy.

Small molecule modulation of splicing factor expression is associated with rescue from cellular senescence.

BACKGROUND: Altered expression of mRNA splicing factors occurs with ageing in vivo and is thought to be an ageing mechanism. The accumulation of senescent cells also occurs in vivo with advancing age and causes much degenerative age-related pathology. However, the relationship between these two processes is opaque. Accordingly we developed a novel panel of small molecules based on resveratrol, previously suggested to alter mRNA splicing, to determine whether altered splicing factor expression had potential to influence features of replicative senescence. RESULTS: Treatment with resveralogues was associated with altered splicing factor expression and rescue of multiple features of senescence. This rescue was independent of cell cycle traverse and also independent of SIRT1, SASP modulation or senolysis. Under growth permissive conditions, cells demonstrating restored splicing factor expression also demonstrated increased telomere length, re-entered cell cycle and resumed proliferation. These phenomena were also influenced by ERK antagonists and agonists. CONCLUSIONS: This is the first demonstration that moderation of splicing factor levels is associated with reversal of cellular senescence in human primary fibroblasts. Small molecule modulators of such targets may therefore represent promising novel anti-degenerative therapies.

The role of cellular senescence in ageing of the placenta.

Aberrant placental ageing is implicated in a high percentage of birth complications, stillbirths and neonatal deaths. Understanding how this complex organ is established and maintained for the 9-10 months of pregnancy and then how and why it undergoes the physiological changes that result in labour at term is therefore of enormous clinical importance. In this review, we assess the evidence that placental ageing results from cellular senescence, a state of terminal proliferation arrest accompanied by characteristic morphological and metabolic changes including a shift to a pro-inflammatory phenotype. We discuss how senescence both contributes to placental formation during cytotrophoblast fusion, and to the changes necessary for labour onset, such as cervical remodelling and increased sterile inflammatory signalling. Based on evidence from human clinical studies and experimental interventions in mice, we assess possible biochemical pathways that may drive senescence, and speculate on how aberrant senescence in the placenta may contribute to pre-eclampsia, pre-term birth and stillbirth.

Animal and human models to understand ageing.

Human ageing is the gradual decline in organ and tissue function with increasing chronological time, leading eventually to loss of function and death. To study the processes involved over research-relevant timescales requires the use of accessible model systems that share significant similarities with humans. In this review, we assess the usefulness of various models, including unicellular yeasts, invertebrate worms and flies, mice and primates including humans, and highlight the benefits and possible drawbacks of each model system in its ability to illuminate human ageing mechanisms. We describe the strong evolutionary conservation of molecular pathways that govern cell responses to extracellular and intracellular signals and which are strongly implicated in ageing. Such pathways centre around insulin-like growth factor signalling and integration of stress and nutritional signals through mTOR kinase. The process of cellular senescence is evaluated as a possible underlying cause for many of the frailties and diseases of human ageing. Also considered is ageing arising from systemic changes that cannot be modelled in lower organisms and instead require studies either in small mammals or in primates. We also touch briefly on novel therapeutic options arising from a better understanding of the biology of ageing.

Reversal of phenotypes of cellular senescence by pan-mTOR inhibition.

Cellular senescence, a state of essentially irreversible proliferation arrest, serves as a potent tumour suppressor mechanism. However, accumulation of senescent cells with chronological age is likely to contribute to loss of tissue and organ function and organismal aging. A crucial biochemical modulator of aging is mTOR; here, we have addressed the question of whether acute mTORC inhibition in near-senescent cells can modify phenotypes of senescence. We show that acute short term treatment of human skin fibroblasts with low dose ATP mimetic pan-mTORC inhibitor AZD8055 leads to reversal of many phenotypes that develop as cells near replicative senescence, including reduction in cell size and granularity, loss of SA-ß-gal staining and reacquisition of fibroblastic spindle morphology. AZD8055 treatment also induced rearrangement of the actin cytoskeleton, providing a possible mechanism of action for the observed rejuvenation. Importantly, short-term drug exposure had no detrimental effects on cell proliferation control across the life-course of the fibroblasts. Our findings suggest that combined inhibition of both mTORC1 and mTORC2 may provide a promising strategy to reverse the development of senescence-associated features in near-senescent cells.

Suppression of the senescence-associated secretory phenotype (SASP) in human fibroblasts using small molecule inhibitors of p38 MAP kinase and MK2.

Senescent cells show an altered secretome profile termed the senescence-associated secretory phenotype (SASP). There is an increasing body of evidence that suggests that the accumulation of SASP-positive senescent cells in humans is partially causal in the observed shift to a low-level pro-inflammatory state in aged individuals. This in turn suggests the SASP as a possible therapeutic target to ameliorate inflammatory conditions in the elderly, and thus a better understanding of the signalling pathways underlying the SASP are required. Prior studies using the early generation p38 MAPK inhibitor SB203580 indicated that p38 signalling was required for the SASP. In this study, we extend these observations using two next-generation p38 inhibitors (UR-13756 and BIRB 796) that have markedly improved selectivity and specificity compared to SB203580, to strengthen the evidence that the SASP is p38-dependent in human fibroblasts. BIRB 796 has an efficacy and toxicity profile that has allowed it to reach Phase III clinical trials, suggesting its possible use to suppress the SASP in vivo. We also demonstrate for the first time a requirement for signalling through the p38 downstream MK2 kinase in the regulation of the SASP using two MK2 inhibitors. Finally, we demonstrate that a commercially-available multiplex cytokine assay technology can be used to detect SASP components in the conditioned medium of cultured fibroblasts from both young and elderly donors. This assay is a high-throughput, multiplex microtitre-based assay system that is highly sensitive, with very low sample requirements, allowing it to be used for low-volume human biological fluids. Our initial studies using existing multiplex plates form the basis for a "SASP signature" assay that could be used as a high-throughput system in a clinical study setting. Our findings therefore provide important steps towards the study of, and intervention in, the SASP in human ageing and age-related disease.