Amino Acid Restriction Triggers Angiogenesis via GCN2/ATF4 Regulation of VEGF and H2S Production.

Angiogenesis, the formation of new blood vessels by endothelial cells (ECs), is an adaptive response to oxygen/nutrient deprivation orchestrated by vascular endothelial growth factor (VEGF) upon ischemia or exercise. Hypoxia is the best-understood trigger of VEGF expression via the transcription factor HIF1α. Nutrient deprivation is inseparable from hypoxia during ischemia, yet its role in angiogenesis is poorly characterized. Here, we identified sulfur amino acid restriction as a proangiogenic trigger, promoting increased VEGF expression, migration and sprouting in ECs in vitro, and increased capillary density in mouse skeletal muscle in vivo via the GCN2/ATF4 amino acid starvation response pathway independent of hypoxia or HIF1α. We also identified a requirement for cystathionine-γ-lyase in VEGF-dependent angiogenesis via increased hydrogen sulfide (H2S) production. H2S mediated its proangiogenic effects in part by inhibiting mitochondrial electron transport and oxidative phosphorylation, resulting in increased glucose uptake and glycolytic ATP production.

Impairment of an Endothelial NAD+-H2S Signaling Network Is a Reversible Cause of Vascular Aging.

A decline in capillary density and blood flow with age is a major cause of mortality and morbidity. Understanding why this occurs is key to future gains in human health. NAD precursors reverse aspects of aging, in part, by activating sirtuin deacylases (SIRT1-SIRT7) that mediate the benefits of exercise and dietary restriction (DR). We show that SIRT1 in endothelial cells is a key mediator of pro-angiogenic signals secreted from myocytes. Treatment of mice with the NAD+ booster nicotinamide mononucleotide (NMN) improves blood flow and increases endurance in elderly mice by promoting SIRT1-dependent increases in capillary density, an effect augmented by exercise or increasing the levels of hydrogen sulfide (H2S), a DR mimetic and regulator of endothelial NAD+ levels. These findings have implications for improving blood flow to organs and tissues, increasing human performance, and reestablishing a virtuous cycle of mobility in the elderly.

Slowing ageing by design: the rise of NAD+ and sirtuin-activating compounds.

The sirtuins (SIRT1-7) are a family of nicotinamide adenine dinucleotide (NAD+)-dependent deacylases with remarkable abilities to prevent diseases and even reverse aspects of ageing. Mice engineered to express additional copies of SIRT1 or SIRT6, or treated with sirtuin-activating compounds (STACs) such as resveratrol and SRT2104 or with NAD+ precursors, have improved organ function, physical endurance, disease resistance and longevity. Trials in non-human primates and in humans have indicated that STACs may be safe and effective in treating inflammatory and metabolic disorders, among others. These advances have demonstrated that it is possible to rationally design molecules that can alleviate multiple diseases and possibly extend lifespan in humans.

Declining NAD(+) induces a pseudohypoxic state disrupting nuclear-mitochondrial communication during aging.

Ever since eukaryotes subsumed the bacterial ancestor of mitochondria, the nuclear and mitochondrial genomes have had to closely coordinate their activities, as each encode different subunits of the oxidative phosphorylation (OXPHOS) system. Mitochondrial dysfunction is a hallmark of aging, but its causes are debated. We show that, during aging, there is a specific loss of mitochondrial, but not nuclear, encoded OXPHOS subunits. We trace the cause to an alternate PGC-1α/ß-independent pathway of nuclear-mitochondrial communication that is induced by a decline in nuclear NAD(+) and the accumulation of HIF-1α under normoxic conditions, with parallels to Warburg reprogramming. Deleting SIRT1 accelerates this process, whereas raising NAD(+) levels in old mice restores mitochondrial function to that of a young mouse in a SIRT1-dependent manner. Thus, a pseudohypoxic state that disrupts PGC-1α/ß-independent nuclear-mitochondrial communication contributes to the decline in mitochondrial function with age, a process that is apparently reversible.

Reprogramming to recover youthful epigenetic information and restore vision.

Ageing is a degenerative process that leads to tissue dysfunction and death. A proposed cause of ageing is the accumulation of epigenetic noise that disrupts gene expression patterns, leading to decreases in tissue function and regenerative capacity1-3. Changes to DNA methylation patterns over time form the basis of ageing clocks4, but whether older individuals retain the information needed to restore these patterns-and, if so, whether this could improve tissue function-is not known. Over time, the central nervous system (CNS) loses function and regenerative capacity5-7. Using the eye as a model CNS tissue, here we show that ectopic expression of Oct4 (also known as Pou5f1), Sox2 and Klf4 genes (OSK) in mouse retinal ganglion cells restores youthful DNA methylation patterns and transcriptomes, promotes axon regeneration after injury, and reverses vision loss in a mouse model of glaucoma and in aged mice. The beneficial effects of OSK-induced reprogramming in axon regeneration and vision require the DNA demethylases TET1 and TET2. These data indicate that mammalian tissues retain a record of youthful epigenetic information-encoded in part by DNA methylation-that can be accessed to improve tissue function and promote regeneration in vivo.

Dynamic Acetylation of Phosphoenolpyruvate Carboxykinase Toggles Enzyme Activity between Gluconeogenic and Anaplerotic Reactions.

Cytosolic phosphoenolpyruvate carboxykinase (PCK1) is considered a gluconeogenic enzyme; however, its metabolic functions and regulatory mechanisms beyond gluconeogenesis are poorly understood. Here, we describe that dynamic acetylation of PCK1 interconverts the enzyme between gluconeogenic and anaplerotic activities. Under high glucose, p300-dependent hyperacetylation of PCK1 did not lead to protein degradation but instead increased the ability of PCK1 to perform the anaplerotic reaction, converting phosphoenolpyruvate to oxaloacetate. Lys91 acetylation destabilizes the active site of PCK1 and favors the reverse reaction. At low energy input, we demonstrate that SIRT1 deacetylates PCK1 and fully restores the gluconeogenic ability of PCK1. Additionally, we found that GSK3ß-mediated phosphorylation of PCK1 decreases acetylation and increases ubiquitination. Biochemical evidence suggests that serine phosphorylation adjacent to Lys91 stimulates SIRT1-dependent deacetylation of PCK1. This work reveals an unexpected capacity of hyperacetylated PCK1 to promote anaplerotic activity, and the intersection of post-translational control of PCK1 involving acetylation, phosphorylation, and ubiquitination.

NAD+ in COVID-19 and viral infections.

NAD+, as an emerging regulator of immune responses during viral infections, may be a promising therapeutic target for coronavirus disease 2019 (COVID-19). In this Opinion, we suggest that interventions that boost NAD+ levels might promote antiviral defense and suppress uncontrolled inflammation. We discuss the association between low NAD+ concentrations and risk factors for poor COVID-19 outcomes, including aging and common comorbidities. Mechanistically, we outline how viral infections can further deplete NAD+ and its roles in antiviral defense and inflammation. We also describe how coronaviruses can subvert NAD+-mediated actions via genes that remove NAD+ modifications and activate the NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome. Finally, we explore ongoing approaches to boost NAD+ concentrations in the clinic to putatively increase antiviral responses while curtailing hyperinflammation.

Characteristics of older unpaid carers in England: a study of social patterning from the English Longitudinal Study of Ageing.

BACKGROUND: A growing number of older people provide unpaid care, but contemporary research evidence on this group is limited. AIM: This study aims to describe the characteristics of older people who provide unpaid care and how these vary by socioeconomic position. METHODS: Using recent information from the English Longitudinal Study of Ageing (ELSA wave 9, 2019), we analysed cross-sectional data on 1,282 unpaid carers aged ≥50. Data on sociodemographics, health, social wellbeing, care intensity and caregiver-recipient relationships were extracted. Total net non-pension wealth quintiles were used as a relative measure of socioeconomic position. Differences between the poorest and richest wealth quintiles were examined through logistic regression. FINDINGS: Most older carers in ELSA were female and looking after another older person. Poor mental and physical health and social isolation were common, and socially patterned. Compared with carers in the middle wealth group, the poorest group were more likely to be living with the person they cared for (odds ratio (OR) 1.56 [95% confidence interval (CI) 1.03-2.36]) and more likely to experience loneliness (OR 2.29 [95% CI 1.42-3.69]), dependency (i.e. the need for help with activities of daily living) (OR 1.62 [95% CI 1.05-2.51]), chronic pain (OR 1.81 [95% CI 1.23-2.67]), a higher number of diseases (OR 1.75 [95% CI 1.15-2.65]) and fair/poor self-rated health (OR 2.59 [95% CI 1.79-3.76]). The poorest carers were also less likely to have a high quality of life (OR 0.51 [95% CI 0.33-0.80]) or be in work (OR 0.33 [95% CI 0.19-0.59]). CONCLUSION: Our findings suggest that financially disadvantaged unpaid carers (and their households) may have the greatest needs for intervention and support. Focussing resources on this group has potential to address social inequalities.

Galactic cosmic radiation exposure causes multifaceted neurocognitive impairments.

Technological advancements have facilitated the implementation of realistic, terrestrial-based complex 33-beam galactic cosmic radiation simulations (GCR Sim) to now probe central nervous system functionality. This work expands considerably on prior, simplified GCR simulations, yielding new insights into responses of male and female mice exposed to 40-50 cGy acute or chronic radiations relevant to deep space travel. Results of the object in updated location task suggested that exposure to acute or chronic GCR Sim induced persistent impairments in hippocampus-dependent memory formation and reconsolidation in female mice that did not manifest robustly in irradiated male mice. Interestingly, irradiated male mice, but not females, were impaired in novel object recognition and chronically irradiated males exhibited increased aggressive behavior on the tube dominance test. Electrophysiology studies used to evaluate synaptic plasticity in the hippocampal CA1 region revealed significant reductions in long-term potentiation after each irradiation paradigm in both sexes. Interestingly, network-level disruptions did not translate to altered intrinsic electrophysiological properties of CA1 pyramidal cells, whereas acute exposures caused modest drops in excitatory synaptic signaling in males. Ultrastructural analyses of CA1 synapses found smaller postsynaptic densities in larger spines of chronically exposed mice compared to controls and acutely exposed mice. Myelination was also affected by GCR Sim with acutely exposed mice exhibiting an increase in the percent of myelinated axons; however, the myelin sheathes on small calibur (< 0.3 mm) and larger (> 0.5 mm) axons were thinner when compared to controls. Present findings might have been predicted based on previous studies using single and mixed beam exposures and provide further evidence that space-relevant radiation exposures disrupt critical cognitive processes and underlying neuronal network-level plasticity, albeit not to the extent that might have been previously predicted.

The Role of Epigenetics in Accelerated Aging: A Reconsideration of Later-Life Visual Loss After Early Optic Neuropathy.

BACKGROUND: In 2005, we reported 3 patients with bilateral optic nerve damage early in life. These patients had stable vision for decades but then experienced significant bilateral vision loss with no obvious cause. Our hypothesis, novel at that time, was that the late decline of vision was due to age-related attrition of retinal ganglion cells superimposed on a reduced neuronal population due to the earlier injury. EVIDENCE ACQUISITION: The field of epigenetics provides a new paradigm with which to consider the normal aging process and the impact of neuronal injury, which has been shown to accelerate aging. Late-in-life decline in function after early neuronal injury occurs in multiple sclerosis due to dysregulated inflammation and postpolio syndrome. Recent studies by our group in mice have also demonstrated the possibility of partial reversal of cellular aging and the potential to mitigate anatomical damage after injury and even improve visual function. RESULTS: The results in mice and nonhuman primates published elsewhere have shown enhanced neuronal survival and visual function after partial epigenetic reprogramming. CONCLUSIONS: Injury promotes epigenetic aging , and this finding can be observed in several clinically relevant scenarios. An understanding of the epigenetic mechanisms at play opens the opportunity to restore function in the nervous system and elsewhere with cellular rejuvenation therapies. Our earlier cases exemplify how reconsideration of previously established concepts can motivate inquiry of new paradigms.

Sirtuin Evolution at the Dawn of Animal Life.

Sirtuins are a family of proteins that protect against cellular injury and aging; understanding their evolution should reveal fundamental mechanisms governing longevity. "Early-branching" animals such as sea sponges and jellyfish have been understudied in previous analyses of sirtuin diversity. These organisms not only hold important positions at the base of the evolutionary tree, but also have unique aging dynamics that defy convention, such as quasi-immortality and high regenerative capacity. In this study, we survey the evolution of sirtuin proteins in animals, with a focus on the oldest living lineages. We describe previously unrecognized expansions of "Class IV" and "Class I" sirtuins around the origin of animals, raising the number of sirtuin families in the last common ancestor to at least nine. Most of these undescribed sirtuins have been lost in vertebrates and other bilaterian animals. Our work also clarifies the evolution of PNC1 and NAMPT enzymes that carry out the rate-limiting step in sirtuin-related NAD+ biosynthesis. The genes for PNC1 and NAMPT enzymes were both present in the first animals, with the genes being lost a minimum of 11 and 13 times, respectively, over the course of animal evolution. We propose that species with these ancestral gene repertoires are ideal model organisms for studying the genetic regulation of animal longevity and will provide clues to increasing longevity in humans.

Novel feature selection methods for construction of accurate epigenetic clocks.

Epigenetic clocks allow us to accurately predict the age and future health of individuals based on the methylation status of specific CpG sites in the genome and are a powerful tool to measure the effectiveness of longevity interventions. There is a growing need for methods to efficiently construct epigenetic clocks. The most common approach is to create clocks using elastic net regression modelling of all measured CpG sites, without first identifying specific features or CpGs of interest. The addition of feature selection approaches provides the opportunity to optimise the identification of predictive CpG sites. Here, we apply novel feature selection methods and combinatorial approaches including newly adapted neural networks, genetic algorithms, and 'chained' combinations. Human whole blood methylation data of ~470,000 CpGs was used to develop clocks that predict age with R2 correlation scores of greater than 0.73, the most predictive of which uses 35 CpG sites for a R2 correlation score of 0.87. The five most frequent sites across all clocks were modelled to build a clock with a R2 correlation score of 0.83. These two clocks are validated on two external datasets where they maintain excellent predictive accuracy. When compared with three published epigenetic clocks (Hannum, Horvath, Weidner) also applied to these validation datasets, our clocks outperformed all three models. We identified gene regulatory regions associated with selected CpGs as possible targets for future aging studies. Thus, our feature selection algorithms build accurate, generalizable clocks with a low number of CpG sites, providing important tools for the field.

Is polypharmacy associated with difficulty taking medicines in people aged ≥85 living at home? Findings from the Newcastle 85+ Study.

It is unclear whether polypharmacy is associated with difficulty taking medications amongst people aged ≥85 living at home. This is despite the projected decline in availability of family carers, who may support independent living. Using Newcastle 85+ Study data and mixed-effects modelling, we investigated the association between polypharmacy and difficulty taking medications amongst 85-year-olds living at home, over a 10-year time period. Polypharmacy was not associated with difficulty taking medications as either a continuous (OR = 0.99 [0.91-1.08]) or categorical variable (5-9 medications, OR = 0.69 [0.34-1.41]; ≥10 medications, OR = 0.85 [0.34-2.07]). The significant predictors included disability, visual impairment and cognitive impairment. Our results suggest that people aged ≥85 living at home with disability, visual impairment and/or cognitive impairment will have difficulty taking their medications, regardless of how many they are prescribed. Therefore, healthcare professionals should routinely ask about, assess and address problems that these patient groups may have with taking their medicines, independent of the number of drugs taken.

Xenohormesis: sensing the chemical cues of other species.

Many plant molecules interact with and modulate key regulators of mammalian physiology in ways that are beneficial to health, but why? We propose that heterotrophs (animals and fungi) are able to sense chemical cues synthesized by plants and other autotrophs in response to stress. These cues provide advance warning about deteriorating environmental conditions, allowing the heterotrophs to prepare for adversity while conditions are still favorable.

SIRT1 redistribution on chromatin promotes genomic stability but alters gene expression during aging.

Genomic instability and alterations in gene expression are hallmarks of eukaryotic aging. The yeast histone deacetylase Sir2 silences transcription and stabilizes repetitive DNA, but during aging or in response to a DNA break, the Sir complex relocalizes to sites of genomic instability, resulting in the desilencing of genes that cause sterility, a characteristic of yeast aging. Using embryonic stem cells, we show that mammalian Sir2, SIRT1, represses repetitive DNA and a functionally diverse set of genes across the mouse genome. In response to DNA damage, SIRT1 dissociates from these loci and relocalizes to DNA breaks to promote repair, resulting in transcriptional changes that parallel those in the aging mouse brain. Increased SIRT1 expression promotes survival in a mouse model of genomic instability and suppresses age-dependent transcriptional changes. Thus, DNA damage-induced redistribution of SIRT1 and other chromatin-modifying proteins may be a conserved mechanism of aging in eukaryotes.

Household wealth, neighbourhood deprivation and frailty amongst middle-aged and older adults in England: a longitudinal analysis over 15 years (2002-2017).

BACKGROUND: frailty is a condition of reduced function and health due to ageing processes and is associated with a higher risk of falls, hospitalisation, disability and mortality. OBJECTIVE: to determine the relationship between household wealth and neighbourhood deprivation with frailty status, independently of demographic factors, educational attainment and health behaviours. DESIGN: population-based cohort study. SETTING: communities in England. SUBJECTS: in total 17,438 adults aged 50+ from the English Longitudinal Study of Ageing. METHODS: multilevel mixed-effects ordered logistic regression was used in this study. Frailty was measured using a frailty index. We defined small geographic areas (neighbourhoods) using English Lower layer Super Output Areas. Neighbourhood deprivation was measured by the English Index of Multiple Deprivation, grouped into quintiles. Health behaviours included in this study are smoking and frequency of alcohol consumption. RESULTS: the proportion of respondents who were prefrail and frail were 33.8% [95% confidence interval (CI) 33.0-34.6%] and 11.7 (11.1-12.2)%, respectively. Participants in the lowest wealth quintile and living in the most deprived neighbourhood quintile had 1.3 (95% CI = 1.2-1.3) and 2.2 (95% CI = 2.1-2.4) times higher odds of being prefrail and frail, respectively, than the wealthiest participants living in the least deprived neighbourhoods Living in more deprived neighbourhood and poorer wealth was associated with an increased risk of becoming frail. Those inequalities did not change over time. CONCLUSIONS: in this population-based sample, living in a deprived area or having low wealth was associated with frailty in middle-aged and older adults. This relationship was independent of the effects of individual demographic characteristics and health behaviours.

Liver-specific overexpression of SIRT3 enhances oxidative metabolism, but does not impact metabolic defects induced by high fat feeding in mice.

The mitochondrial enzyme SIRT3 is an NAD+-dependent deacetylase important in cell metabolism, and a decline in its protein expression or activity has been linked with insulin resistance in obesity, ageing and type 2 diabetes. While studies in SIRT3 knockout mice have dramatically improved our understanding of the function of SIRT3, the impact of increasing SIRT3 levels remains under-examined. In this study we investigated the effects of liver-specific SIRT3 overexpression in mice on mitochondrial function and metabolic profile in both isolated hepatocytes and in vivo. Primary hepatocytes overexpressing SIRT3 displayed increased oxygen consumption and a reduction in triglyceride accumulation. In mice with hepatic SIRT3 overexpression, increased fasting ß-hydroxybutyrate levels were observed, coupled with an increase in oxygen consumption in isolated mitochondria and increased substrate utilization in liver homogenates. However, metabolic profiling of mice exposed to either chow or high-fat diet revealed no effect of hepatic SIRT3 overexpression on glucose tolerance, body composition or tissue triglyceride accumulation. These findings suggest limited whole-body benefit of increasing hepatic SIRT3 during the development of diet-induced insulin resistance.

Measuring frailty and its association with key outcomes in the ambulance setting: a cross sectional observational study.

BACKGROUND: Little is known about frailty in the ambulance setting, or its association with outcomes relevant to ambulance services. We sought to measure frailty in people aged ≥ 50 attended by an ambulance, and describe the relationship between frailty, odds of conveyance to hospital, and duration at scene. METHODS: An observational study between 01/01/2021-30/06/2021 in North East Ambulance Service, England. Participants were aged ≥ 50 attended by an ambulance, excluding patients requiring immediate treatment for a life-threatening condition or with Glasgow Coma Scale < 15. Paramedics (n = 112) measured patient frailty using the Clinical Frailty Scale (CFS). Additional information was extracted from ambulance care records. Weighted regression models examined associations between frailty, hospital conveyance, and duration at scene. RESULTS: Three thousand and fifty-six callouts were observed (mean patient age: 78.1 years, 57.2% female). Frailty prevalence (CFS ≥ 5) was 58.7%. Median duration at scene was 47.0 min (interquartile range 34.0-67.0 min). Ambulances spent a median of 8.2 (95%CI:5.4-11.0) minutes longer with frail patients than non-frail patients. Frail patients were less likely to be conveyed to hospital than non-frail patients (OR:0.75, 95%CI:0.60-0.94). CONCLUSION: Frailty is common among people aged ≥ 50 attended by an ambulance and an important influence on workload. Ambulance services need a good understanding of frailty to meet patient needs. As populations age, community support should be prioritised to deliver appropriate frailty care and reduce demands on ambulance services.