Caloric (CR) or dietary (DR) restriction improves health and extends lifespan in multiple species. However, the beneficial effects of DR may diminish if introduced late in life, emphasizing the importance of timing for promoting healthspan and avoiding adverse outcomes. Using a metabolomics approach, we investigated the metabolic responses in plasma, liver, and kidney of mice on acute and chronic DR at various ages. Two hundred and five mice including young (2-month-old; n = 72), middle-aged (6-month-old; n = 76), and old (17-month-old; n = 57) mice for DR were involved. No significant metabolic distinctions were observed during acute DR across different ages. Throughout chronic DR, hepatic glucose, glycogen, and glutathione levels-all of which decreased with age-were elevated in all mice, demonstrating an improvement in energy metabolism and enhanced protection against oxidative stress. We also found age-dependent metabolic responses to DR. Specifically, in young mice, amino acids and lactate contributed to gluconeogenesis in the liver during chronic DR. In contrast, in middle-aged and older mice, only fatty acids played a role in the energy supply within the liver. We noted significant hepatic glycogen accumulation in old mice, along with decreased levels of hepatic betaine and sarcosine in young mice, indicating the negative impact of chronic DR on liver function. The findings suggest that the most substantial benefits of DR occur in the middle stage of life, highlighting the need for tailored dietary intervention strategies to promote health span at different life stages.

Senescent cells are characterized by a stable proliferation arrest and a senescence-associated secretory phenotype or SASP. Although these cells can have some beneficial effects, including protecting from tumor formation, their accumulation is deleterious during aging as it promotes age-related diseases, including cancer initiation and progression. Although the SASP has a critical role, its composition, regulation and dual role in cancer remain largely misunderstood. Here, we show that ANGPTL4 is one of the rare secreted factors induced in many different types of senescent cells. Importantly, ANGPTL4 knockdown during senescence or its constitutive expression, respectively inhibits or induces classical proinflammatory SASP factors, such as IL1A, IL6 and IL8. The latter effect is mediated upstream of IL1A, an early SASP factor, suggesting an upstream role of ANGPTL4 in SASP induction. This ANGPTL4-dependent proinflammatory SASP can promote human neutrophil activation in ex vivo assays, or tumor initiation in a KRAS-dependent lung tumorigenesis model in mice. This upstream activity of ANGPTL4 in regulating the proinflammatory SASP depends on its upregulation following a hypoxia-like response and HIF2A activation, and its proteolytic processing by the FURIN proprotein convertase. Altogether these findings shed light on a two-step activation of ANGPTL4 by HIF2A and FURIN in senescent cells and its upstream role in promoting the proinflammatory SASP, cancer and potentially other senescence-associated diseases.

Major depressive disorder (MDD) is linked to a higher risk of premature aging, but the mechanisms underlying this association remain unclear. Using data from two population cohorts (UK Biobank and Finnish Twin Cohort), we evaluate the relationship between systemic and organ-specific proteomic and epigenetic aging acceleration and MDD. A lifetime history of MDD was associated with accelerated proteomic aging at both systemic and organ-specific levels-including the brain-in both cohorts, with stronger associations than those observed with systemic epigenetic aging. Systemic and brain-specific proteomic aging acceleration were linked to higher risks of incident MDD and a greater risk of Alzheimer's disease, related dementia, and mortality among individuals with MDD in the UK Biobank. Evidence of depressive episode remission attenuated the association between MDD and systemic and brain-specific proteomic aging acceleration. Finally, Mendelian randomization analyses revealed a causal effect of MDD on systemic and brain-specific proteomic aging acceleration. Our results suggest a strong bidirectional association between MDD and biological aging acceleration. Biological aging acceleration, assessed by proteomic systemic and organ-specific clocks, can serve as a novel therapeutic target for treating MDD and for mitigating the long-term risks of adverse health outcomes associated with this condition.

Alzheimer's disease (AD) is characterized by progressive memory decline. Converging evidence indicates that hippocampal mRNA translation (protein synthesis) is defective in AD. Here, we show that genetic reduction of the translational repressors, Fragile X messenger ribonucleoprotein (FMRP) or eukaryotic initiation factor 4E (eIF4E)-binding protein 2 (4E-BP2), prevented the attenuation of hippocampal protein synthesis and memory impairment induced by AD-linked amyloid-β oligomers (AβOs) in mice. Moreover, genetic reduction of 4E-BP2 rescued memory deficits in aged APPswe/PS1dE9 (APP/PS1) transgenic mouse model of AD. Our findings demonstrate that strategies targeting repressors of mRNA translation correct hippocampal protein synthesis and memory deficits in AD models. Results suggest that modulating pathways controlling brain mRNA translation may confer memory benefits in AD.

Alzheimer's disease (AD) is characterized by progressive cognitive decline, amyloid β (Aβ) deposition, and synaptic dysfunction. However, the mechanisms underlying neurodegeneration remain poorly understood. In this study, we investigated the therapeutic potential of PBX1, a transcriptional regulator implicated in neurodevelopment and neuroprotection, against AD. PBX1 expression was significantly downregulated in postmortem hippocampal tissues from patients with AD and in the APP/PS1 mouse model. In vitro, PBX1a knockdown reduced neurite complexity and increased apoptosis. PBX1a overexpression reversed these effects and reduced soluble Aβ and Aβ levels. In vivo, hippocampal overexpression of PBX1a restored spatial learning and memory, reduced Aβ burden by 41%, and increased neurite length by 1.5-fold. These behavioral and structural improvements were accompanied by reduced levels of hyperphosphorylated Tau and toxic Aβ oligomers. Mechanistically, PBX1 directly activated the transcription of CRTC2-a coactivator of CREB, thereby increasing CRTC2 expression and its nuclear colocalization with phosphorylated CREB. Restoration of the PBX1-CRTC2-CREB axis enhanced neuronal survival and synaptic integrity. Notably, CRTC2 knockdown blocked PBX1-mediated reductions in Aβ deposition, apoptosis, and hyperphosphorylated Tau expression, confirming the role of the PBX1-CRTC2-CREB axis in conferring neuroprotection. Together, our findings indicate that PBX1 is a key modulator of neuronal resilience in AD and that it functions through transcriptional activation of the CRTC2/CREB pathway. By unraveling a mechanism that links transcriptional regulation to amyloid clearance and cognitive function, this study highlights PBX1 as a promising therapeutic target for AD.

Epigenetic clocks are machine learning models that predict an organism's chronological age (the time elapsed since birth) or biological age (a proxy for physiological integrity) based on methylation levels from multiple genomic sites. To date, all epigenetic clocks rely exclusively on C5-methylcytosine (5 mC), the predominant DNA methylation mark in vertebrates. However, not all species possess detectable 5 mC levels. Here, we used N6-methyladenine (6 mA), a less-characterized DNA modification type, to develop a series of epigenetic clocks in the buff-tailed bumblebee (Bombus terrestris). Using long-read Nanopore sequencing, we generated genome-wide, base-resolution profiles of 6 mA and 5 mC in males of different ages (n = 15), and developed multiple epigenetic clocks based on distinct features of the aging DNA methylome. All clocks showed strong correlations between predicted epigenetic and chronological age. Moreover, they also detected pharmacologically induced lifespan extension, reflected by a reduction in predicted epigenetic age relative to chronological age, indicating that these clocks capture biological aging. These findings demonstrate that 6 mA can be used to build accurate epigenetic clocks and establish 6 mA as a promising biomarker of aging in animals.

Controlled settings may offer limited insight into the complexities of aging in natural and variable ecosystems. Artwork by Zahida Sultanova.

Understanding metabolic changes across the human lifespan is essential for addressing age-related health challenges. However, comprehensive metabolomic and lipidomic analyses, particularly in human plasma, remain underexplored. Herein, we performed untargeted metabolomics and lipidomics profiling of plasma collected from 136 individuals aged 0-84 years. This analysis reveals distinct metabolic signatures across life stages, with newborns displaying unique sphingosine (SPH) profiles, while aging was found to be characterized by elevated amino acid levels and lipid imbalances. Notably, we identified linear and nonlinear metabolic trajectories across the lifespan, highlighting critical transition points reflecting the key stages of metabolic reprogramming. By integrating these metabolic patterns, we developed an "aging clock" based on plasma metabolite profiling, thus providing a powerful tool to predict biological age. These findings offer new insights into the dynamic metabolic landscape of aging, paving the way for targeted interventions to improve healthspan and prevent age-related diseases.

Preovulatory follicle aging is the period between formation and ovulation of a mature follicle. Previous studies had shown that mammalian preovulatory follicle aging is associated with chromosomal abnormalities and developmental defects such as decreased implantation, increased malformation and mortality and lower embryonic weight. Our understanding of the molecular events governing this process has been hampered by the difficulty in accessing them in vivo under natural conditions. We hypothesize that the quality of the mature oocyte is regulated by crosstalk between the oocyte and the somatic microenvironment during extended storage prior to ovulation. By combining temporal profiling and tissue-specific functional analyzes in Drosophila, we characterize a spatiotemporal crosstalk between the oocyte and the granulosa cells that governs preovulatory follicle aging in vivo. Preovulatory follicle aging is characterized by two distinct phases-early oocyte protective and late degenerative phases. The degenerative phase involves a positive feedback loop between oocyte mitochondrial dysfunction mediated by a mitochondrial-localized microprotein PIGBOS, and granulosa cell functional decline through a circular RNA circdlg1. Activation of the feedback loop is suppressed by germline Sestrin during the early phase. Our findings highlight that natural preovulatory follicle aging in vivo is governed by a mechanism that represses an oocyte-degenerative positive feedback loop between oocyte and granulosa cells.

The amyloid precursor protein (APP) plays a pivotal role in the pathogenesis of Alzheimer's disease (AD). While the production of Amyloid beta (Aβ) has traditionally been considered the primary cause of AD, the role of the APP intracellular domain (AICD) remains largely elusive. In this study, we established a novel model in the adult fly wing by expressing human APP, recapitulating AD-associated axon degeneration. Using this model, we discovered that ectopic APP expression in Drosophila wing margin neurons led to age-dependent axon degeneration. APP's effect depended on AICD production, and AICD overexpression alone was sufficient to induce axon degeneration in adult wings. Further investigations indicated that APP- or AICD-induced axon degeneration could be alleviated by blocking autophagy, but not apoptosis. Additionally, we identified a FoxO/Snail-Atg1 axis as an essential mediator of APP/AICD-induced autophagy-dependent axon degeneration. Finally, we demonstrated that administration of chloroquine, an autophagy inhibitor, effectively ameliorates APP- or AICD-induced axon degeneration. Our findings provide crucial insights into how APP induces autophagy-dependent axon degeneration through AICD production, laying a foundation for future investigations into AD pathogenesis.

During brain aging, terminally differentiated neuroglia exhibit metabolic dysfunction and increased oxidative damage, compromising their function. These cellular and molecular alterations impair their ability to maintain myelin sheath integrity, contributing to age-related white matter degradation. Calorie restriction (CR) is a well-established intervention that can slow biological aging and may reduce age-related metabolic alterations, thereby preserving the molecular function of aging glia. Here we present a single nucleus resolution, transcriptomics dataset evaluating the molecular profile of oligodendrocytes and microglia in the brain of aging rhesus monkeys following lifelong, 30% calorie restriction. Oligodendrocytes from CR subjects exhibited increased expression of myelin-related genes and showed enrichment in glycolytic and fatty acid biosynthetic pathways. In CR subjects, a subpopulation of oligodendrocytes upregulated cell adhesion gene, NLGN1 and were in closer proximity to axons. Microglia from CR subjects upregulated amino acid and peptide metabolism pathways and showed a reduced myelin debris signature. Our findings reveal cell-type specific transcriptional reprogramming in response to long term CR and highlight potential protective mechanisms against myelin pathology in the aging primate brain.

Studies in laboratory organisms typically minimize all environmental and genetic variation other than the intervention of interest. In aging studies, these highly controlled conditions have yielded profound insights into aging. But even within isogenic cohorts of lab animals in controlled environments, we observe substantial variation in lifespan. Here we exploited the climbing behavior of Drosophila to study variation in mortality among isogenic populations in a controlled environment. We show that fractionating large cohorts of relatively young isogenic flies by climbing behavior predicts future mortality risk and stress sensitivity. Using metabolomics to dissect this variation, we found metabolites whose abundances differ among the fractions. We also took advantage of the large number of individuals in each fraction, and the ease with which they can be collected, to explore the covariance structure of metabolites in flies that are genetically identical, but divisible into short-lived and long-lived fractions. In doing so, we identified metabolites and metabolic pathways as candidate biomarkers of intrinsic mortality risk.

Aging is a key driver of cardiac dysfunction, promoting structural remodeling, metabolic alterations, and loss of cellular resilience. In aged hearts, extracellular matrix remodeling and collagen accumulation reduce ventricular compliance, impairing both diastolic function and stress adaptability. Cardiomyocytes exhibit diminished regenerative capacity and dysregulated stress responses, with mitochondrial dysfunction emerging as a central contributor to energy imbalance, oxidative stress, and fibrosis. Traditional single-omics approaches are insufficient to capture the complexity of these interconnected changes. To address this, we employed an integrative multi-omics strategy-combining spatial transcriptomics, proteomics, and metabo-lipidomics with electron microscopy-to investigate cardiac aging in mice at three life stages: adult (12 months), middle-aged (24 months), and elderly (30 months). Electron microscopy revealed enlarged, structurally compromised mitochondria. Spatial transcriptomics showed reduced expression of cardioprotective genes (MANF, CISH, and BNP) and increased expression of profibrotic markers like CTGF. Proteomics revealed widespread mitochondrial dysregulation and impaired ATP production. Metabolic and lipidomic profiling identified reduced antioxidant metabolites and accumulation of lipotoxic species, such as ceramides and diacylglycerols. This multiscale analysis highlights key molecular and metabolic alterations driving cardiac aging, identifying potential therapeutic targets to mitigate age-related functional decline. Overall, our findings highlight the value of integrated, system-level approaches for uncovering the complex mechanisms that drive organ aging. Although our study was conducted in mice, validation in human models will be crucial to establish the translational relevance of these results and to guide future research with potential impact across diverse biomedical fields.

COX7RP is a critical factor that assembles mitochondrial respiratory chain complexes into supercomplexes, which is considered to modulate energy production efficiency. Whether COX7RP contributes to metabolic homeostasis and lifespan remains elusive. We here observed that COX7RP-transgenic (COX7RP-Tg) mice exhibit a phenotype characterized by a significant extension of lifespan. In addition, metabolic alterations were observed in COX7RP-Tg mice, including lower blood glucose levels at 120 min during the glucose tolerance test (GTT) without a significant difference in the area under the curve (AUC), as well as reduced serum triglyceride (TG) and total cholesterol (TC) levels. Moreover, COX7RP-Tg mice exhibited elevated ATP and nicotinamide adenine dinucleotide levels, reduced ROS production, and decreased senescence-associated β-galactosidase levels. Single-nucleus RNA-sequencing (snRNA-seq) revealed that senescence-associated secretory phenotype genes were downregulated in old COX7RP-Tg white adipose tissue (WAT) compared with old WT WAT, particularly in adipocytes. This study provides a clue to the role of mitochondrial respiratory supercomplex assembly factor COX7RP in resistance to aging and longevity extension.

C-reactive protein (CRP) and growth differentiation factor 15 (GDF15) are important markers of inflammation associated with brain health. Compared to plasma, DNA methylation (DNAm) measures of CRP and GDF15 may provide stable epigenetic measures of chronic exposure to inflammation and could therefore be robustly predictive of inflammation-related brain aging and neurodegeneration. We leveraged a subsample of Baltimore Longitudinal Study of Aging (BLSA) participants with DNAm/plasma data and longitudinal neuroimaging/cognition data (n = 430-1100). We used a proteome-wide analysis to characterize the biology of DNAm CRP and GDF15, and latent growth curve models to explore the associations with longitudinal trajectories of 19 brain region volumes and five cognitive domains. Finally, we related DNAm/plasma CRP and GDF15 to dementia risk in two external cohorts. DNAm CRP and GDF15 showed a proteomic signature consistent with systemic immune activation. We identified several brain regions with significant associations between elevated DNAm CRP And GDF15 and (a) lower brain volume level (at age 75) and (b) greater rate of atrophy. Compared to plasma CRP, DNAm CRP was more strongly associated with brain volume, cognitive trajectories, and dementia risk. DNAm and plasma GDF15 were similarly associated with several total lobar, total lobar white matter, and AD-relevant region trajectories and dementia risk, but DNAm measures outperformed plasma measures in relation to cognitive trajectories. Epigenetic signatures of CRP and GDF15 reflect immune and inflammation-related pathway activation. These signatures, especially DNAm CRP, were associated with accelerated brain atrophy, cognitive decline, as well as long-term dementia risk.

Aging constitutes the largest risk factor for melanoma progression. While a contribution of factors secreted from senescent skin fibroblasts to the progression of melanoma has been proposed, the nature of such factors and subsequent underlying mechanisms remains elusive. Here we show that the chemokine GCP-2 is excessively released by senescent fibroblasts in vitro and the skin of old melanoma patients. GCP-2 regulates, via phosphorylation of the transcription factor CREB at serine 133, defense-, cell cycle control-, and glycolysis-enhancing genes in melanoma cell lines. GCP-2 promotes oncogenic properties in vitro and in vivo in murine melanoma models. Inhibition of CREB phosphorylation in melanoma cells represses glycolytic target genes and induces a switch from glycolysis to oxidative phosphorylation that translates into a significant decline in tumor size in vivo in murine melanoma models. This study identifies a senescent fibroblast to chemokine to CREB to metabolic axis that drives melanoma progression. Targeting this axis may hold promise for novel therapeutic approaches in difficult-to-treat melanoma in older adults.

Aging is associated with a progressive decline in physiological resilience, often linked to impaired stress responses and metabolic dysfunction. In Caenorhabditis elegans (C. elegans), caloric restriction (CR) and pharmacological interventions are widely used to dissect conserved longevity pathways. Here, we identify the N-methyl-D-aspartate receptor (NMDAR) antagonist memantine as a novel modulator of lifespan and stress tolerance in C. elegans. Memantine, but not ketamine, extends median lifespan and reproductive lifespan, suggesting that the observed effects are not shared with ketamine at the tested concentration. Transcriptomic analysis revealed significant overlap between memantine-treated animals and CR models, particularly eat-2 mutants, implicating shared metabolic and longevity-associated pathways. Functionally, memantine was found to reduce mitochondrial and oxidative stress, while enhancing β-oxidation of fatty acids, and modifying behavioral responses to food cues, delaying food-seeking behavior and increasing locomotion under starvation, without affecting lipid storage. In summary, these findings suggest that memantine promotes stress resilience and healthy aging via metabolic changes that overlap with CR-associated pathways, highlighting its potential as a longevity-modulating intervention.

Tendon degeneration is common, and its risk increases with age both in humans and horses. Tendon regeneration and healing is limited due to inherent low cell density and vascularisation, and current treatments are insufficient as indicated by scar tissue formation and a high re-injury rate. The tendon vasculature plays a crucial role in tendon homeostasis, regeneration and healing, making it a potential therapeutic target. However, the effect of ageing on the tendon microvasculature is poorly understood. Here, we provide the first comprehensive characterisation of the tendon microvasculature. We employed high-resolution 3D imaging techniques, using micro-computed tomography (μCT) and confocal microscopy, to investigate age-related alterations in the vasculature within the equine superficial digital flexor tendon (SDFT), a functional equivalent of the human Achilles tendon. μCT analysis revealed a well-developed vascular network within the interfascicular matrix (IFM) and demonstrated significant age-associated reductions in vascular volume (70%), vessel diameter (30%) and density (74%). 3D immunolabelling showed significant reductions in MYH11- (96%) and desmin-positive (78%) volumes; however, there was a pronounced age-associated increase in von Willebrand factor (VWF)-positive volume (220%), which was accompanied by a significantly higher (249%) pericyte density. Taken together, these results indicate a loss of larger blood vessels in the IFM but an increase in small vessel formation, suggesting that neo-angiogenesis is induced in aged tendon alongside a loss of vascular homeostasis. These insights enhance our understanding of tendon ageing and may contribute to developing new therapeutic approaches for improving tendon health and repair in older individuals.

There is a growing contradiction between the rising demand for mechanical ventilation among the elderly and their heightened sensitivity to ventilator-induced lung injury (VILI). This discrepancy compels us to explore therapeutic targets for VILI in elderly patients. Our research revealed that aging increases the sensitivity of pulmonary endothelial cells to low-magnitude mechanical stretch. By analyzing transcriptome sequencing data from lung tissues of humans and mice at different ages, as well as published transcriptome sequencing data from senescent endothelial cells, we identified tissue kallikrein-related peptidase 8 (KLK8) as an age-dependent upregulated gene in lung tissues. Using KLK8 knockout mice, intra-pulmonary KLK8-overexpressing mice, and mouse lung vascular endothelial cells (MLVECs) with KLK8 overexpression or knockdown, we demonstrated that age-dependent KLK8 upregulation contributes to pulmonary endothelial senescence and increased susceptibility of aged mice to VILI. Mechanistically, KLK8 promotes pulmonary endothelial senescence by inactivating the fibronectin/focal adhesion kinase (FAK) pathway. Through transcriptional profiling, we identified the poly(ADP-ribose) polymerase 1/2 (PARP1/2) inhibitor olaparib as a potential agent that rescues KLK8-induced pulmonary endothelial cell senescence and alleviates VILI in aged mice. Our findings underscore the critical role of KLK8 in pulmonary endothelial senescence and provide preclinical evidence for PARP1/2 inhibitors as a therapeutic target for VILI in elderly individuals.

Despite the long interspersed nuclear element-1 (LINE1, L1) retrotransposons having been implicated in Alzheimer's disease (AD), a fundamental understanding of the AD-specific lifespan-long trajectory of L1 has been limited. Here, we characterize the content and expression of L1 covering four brain regions (hippocampus, prefrontal cortex, cerebellum, and the rest of brain tissue) of APP/PS1 mice, a murine model of AD, and their wild-type C57BL/6 littermates from 3 to 24 months of age. We report that both L1 content (indicated by DNA copy number) and expression (indicated by protein levels of L1-encoded ORF1 and ORF2) across brain regions had nonlinear, U-shaped associations with age in wild-type and APP/PS1 mice. Compared to age-matched wild-types, APP/PS1 mice constantly have significantly decreased L1 content but increased L1 expression, suggesting L1 differences between wild-type and APP/PS1 mice establish early and remain stable throughout the life course. Strikingly, L1 content and expression in wild-type and APP/PS1 mice are sexually different, depending on age and brain region. The appearance of L1 alteration precedes the onset of β-amyloidosis by 3 months in APP/PS1 mice, and β-amyloidosis is positively correlated with L1 content and expression in males but anti-correlated with L1 content in females of both wild-type and APP/PS1 mice. Overall, this study (i) reveals an unanticipated U-shaped trajectory of L1 content and expression in both normal and pathological aging of mouse brains and (ii) discerns specific changes in L1 content and expression tied to AD neuropathology in a sex-different manner.

Human umbilical cord blood (HUCB) exhibits distinct characteristics compared to adult blood, offering significant potential for medical applications, particularly in antiaging therapies. However, the metabolic profile of HUCB relative to adult blood remains poorly understood. Moreover, the specific metabolites within HUCB that confer antiaging properties have yet to be identified. Here, we conducted an untargeted metabolomic analysis comparing cord plasma and adult plasma. Our results reveal a unique metabolic landscape in cord plasma, characterized by significant differences in 662 out of 1092 total compounds and 43 out of 59 total human metabolic pathways. Notably, 211 abundant cord metabolites decline with age, involving key aging-related processes, including inflammation, oxidative stress, energy and nutrition metabolism, proteostasis and DNA damage responses, implicating their potential role in counteracting aging. Importantly, a proof-of-concept experiment demonstrates that a formula containing five of these metabolites (carnosine, taurocholic acid, inosine, L-Histidine and N-acetylneuraminic acid) significantly extends both lifespan and healthspan in C. elegans. Collectively, our findings provide novel insights into the distinctive characteristics of the human cord plasma metabolome and identify promising metabolites with therapeutic potential for antiaging and other cord blood-based medical applications.