Targeting biological processes of aging is a central goal of geroscience; however, limited data exist regarding the feasibility of incorporating biological aging biomarkers into dietary intervention trials. We conducted a pilot feasibility study among 34 adults aged 48-81 years with metabolic syndrome, a condition associated with elevated risk of age-related cardiometabolic disease and advanced biological aging. Participants consumed 1 oz of tree nuts and two tablespoons of extra virgin olive oil daily for 4 weeks. The primary objectives were to evaluate feasibility, adherence, and participant acceptability of epigenetic aging assessments. Exploratory outcomes included DunedinPACE, a measure of the pace of aging, and AgeAccelGrim, a measure of biological age relative to chronological age. At baseline, all participants exhibited a faster pace of biological aging than average as assessed by DunedinPACE, supporting metabolic syndrome as a promising target population for geroscience interventions. Adherence to the dietary intervention exceeded 95%, and most participants reported willingness to participate in a similar longer-term trial. Participants expressed a strong interest in learning their biological age and indicated that evidence of slowed aging would motivate sustained dietary change. No significant changes in epigenetic aging were observed over the 4-week intervention. These findings demonstrate the feasibility and acceptability of incorporating epigenetic aging biomarkers into dietary intervention research and suggest that biological aging measures may serve not only as surrogate outcomes but also as tools to support participant engagement. The results also support metabolic syndrome as a relevant population for dietary geroscience trials and provide practical guidance for designing longer-term studies evaluating whether dietary interventions can slow biological aging and promote healthy longevity. ClinicalTrials.gov Identifier: NCT04361617 (date of registration: 04-23-2020).
Genotype-first screening compares patient genomes to a standard reference. The GRCh38 linear assembly contains reference minor alleles (RMAs), posing a critical structural vulnerability for automated clinical variant classification. To determine whether RMAs systematically generate false-positive annotations, we conducted an observational case series of 20 healthy adults undergoing preventive genome sequencing. Automated bioinformatic processing was performed using the GRCh38 linear reference to identify "high-impact" variant annotations generated at loci where GRCh38 differs from the population consensus major allele. Among all 20 participants (100%), linear alignment to GRCh38 systematically misclassified functional major alleles as false-positive "high-impact" annotations at three distinct loci (SLC37A4, CIMIP2A, and GPR33). These artifacts occurred solely because analysis software mathematically defined the healthy wild-type state as a deviation from the rare RMA. Cross-referencing with gnomAD confirmed these as population-dominant benign alleles. Furthermore, this inherent reference bias introduces a theoretical risk of false-negative classifications if RMAs mathematically mask true pathogenic variants. Automated pipelines using linear references systematically misclassify healthy alleles as high-impact functional annotations. While downstream population-frequency filters manage false-positive artifacts, this retrospective patching creates an unsustainable bottleneck for population-scale screening. Transitioning to graph-based pangenome references represents a highly promising approach to directly resolve these diagnostic vulnerabilities at the alignment level, though computational and standardization challenges must first be addressed to ensure the accuracy of large-scale aging research.
Inflammaging, defined as the persistent, low-grade sterile inflammation accompanying aging, represents a central driver of age-related pathology, including cardiovascular dysfunction, neurodegeneration, metabolic disorders, and frailty. This review discusses the most recent advances in understanding its mechanistic basis, encompassing cellular senescence, the senescence-associated secretory phenotype (SASP), mitochondrial dysfunction, immune cell senescence, innate immune hyperactivation, defective inflammatory resolution, and nutrient-sensing dysregulation. Single-cell and spatial transcriptomics reveal tissue-specific and context-dependent patterns, highlighting the systemic complexity of inflammaging. Preclinical interventions demonstrate that inflammaging is modifiable through senolytics, which selectively eliminate senescent cells, and senomorphics, which suppress SASP without inducing cell death. Metabolic modulators such as metformin and rapamycin attenuate inflammatory signaling, while immune-directed therapies and microbiome-targeted interventions provide synergistic benefits through combinatorial approaches. Early-phase clinical trials in frail older adults show feasibility, safety, and preliminary efficacy, including reductions in circulating inflammatory markers, improved physical function, and enhanced immune responsiveness. Inflammaging trajectories are shaped by lifestyle, environmental exposures, and evolutionary factors, underscoring the need for personalized interventions. Remaining challenges include biomarker development, long-term safety evaluation, and heterogeneity across aging populations. Addressing these through interdisciplinary research supports a precision geroscience paradigm, where multimodal targeting of inflammaging can extend healthspan and reduce chronic disease burden.
People with HIV (PWH) on effective antiretroviral therapy continue to experience a disproportionate burden of age-related comorbidities and multimorbidity. This review explores HIV-associated aging as a systems-level dysregulation and loss of resilience and evaluates emerging biomarker strategies for capturing its biological and clinical heterogeneity. Recent evidence indicates that aging in HIV is driven by interacting immune, metabolic, regenerative, structural, and inter-organ pathways that generate measurable signatures across biological scales. Composite biomarker approaches integrating molecular, physiological, and functional measures are increasingly being applied to cardiovascular, neurocognitive, physical, and co-infection-related outcomes. Large cohort studies, risk prediction models, and intervention trials support their clinical relevance, while emerging gerotherapeutic studies suggest that biologic aging markers may serve as responsive treatment endpoints. No single biomarker adequately captures HIV-associated aging. Future progress will depend on multimodal, outcome-anchored composites that integrate biological and functional domains and are coupled with artificial intelligence- and machine learning-based approaches to improve risk stratification, therapeutic monitoring, and evaluation of geroscience-informed interventions in PWH.
Caveolin-1 (CAV1), the principal structural protein of caveolae, is a critical regulator of endothelial homeostasis, vascular aging, and frailty in older adults. By scaffolding multiple signaling pathways and controlling endothelial nitric oxide synthase activity, CAV1 shapes nitric oxide bioavailability, oxidative stress responses, and vascular stiffness. In parallel, extracellular vesicles (EVs) derived from endothelial cells, platelets, and other vascular cells act as key mediators of intercellular communication, propagating senescence, inflammation, and thrombosis across the vascular tree. This narrative review synthesizes current evidence on the CAV1-EV axis as a mechanistic nexus linking endothelial dysfunction, cardiovascular disease, and geriatric syndromes, with a particular focus on frailty. We first summarize the structure, regulation, and post-translational modifications of CAV1, especially tyrosine-14 (Tyr-14) phosphorylation, and their impact on endothelial senescence and dysfunction. We then examine how CAV1 controls EV biogenesis, cargo loading, and uptake, and how CAV1-enriched EVs contribute to pro-inflammatory and prothrombotic phenotypes in aging and frail populations. Finally, we discuss the translational potential of CAV1-related EVs as circulating biomarkers for early vascular aging and as targets for geroscience-guided interventions, including engineered or mesenchymal stem cell-derived EVs with vasculoprotective cargo. By integrating molecular, vascular, and geriatric perspectives, this review proposes the CAV1-EV axis as a promising framework for risk stratification and vascular-targeted therapies aimed at preserving function and independence in older adults.
Geriatricians are aware that the clinical presentation of older people is often with 'atypical' symptoms of commonly encountered conditions. In this Commentary, we explore the emergence of this description as geriatric medicine developed during the era that the disease-diagnosis paradigm dominated medical discourse. We suggest that this recognition of atypicality may have increased the sensibility of non-experts to the more subtle clinical complexity of unwell older adults and added legitimacy to the role of specialist geriatrics, and therefore contributed to improvements in the access and quality of care of older people. Then, noting the growing understanding of the multidimensional nature of illness causation, with age-related frailty now widely appreciated, and insights emerging from geroscience, we offer a critique of its conceptual limitations. Finally, we suggest that holding to the notion of atypicality might be an obstacle to a more assertive reorientation to equitable and age-attuned healthcare, and that the time may be right to offer atypicality a well-deserved retirement.
As global populations age and lifespan increases, health systems too often conceptualize "healthy aging" and "end-of-life care" as distinct phases. Yet emerging evidence from geroscience, palliative care, and health systems science suggests that aging and dying lie on a continuum of adaptation and care. This Perspective proposes a framework that treats system-supported, goal-aligned end-of-life care as a public health outcome of integrated life-course care and argues for system redesign to introduce earlier palliative approaches, align care models across the life-course, and reduce health disparities. This article outlines key mechanisms, evidence for integration, a conceptual model, and implications.
A central challenge in computational geroscience is to distinguish molecular quantities that predict mortality from those that causally drive it. Epigenetic clocks and aging biomarkers are increasingly used as if they were that mechanism, yet this is rarely tested directly. This distinction also bears on competing theories of aging: damage/reliability (A), hyperfunction/mTOR-IIS (B-1), and information loss (B-2). Although individual aging proteins have been tested piecemeal, no study has asked, in one framework, what fraction of mortality is measurable, whether it is causal, and whether it is reversible. Using only public, de-identified data, we evaluate this three ways. First, a Markov generator-matrix model of hallmark-load dynamics with death as an absorbing state, fitted by Bayesian inference through a joint biomarker-and-mortality likelihood to NHANES with linked mortality (n=23,844) and replicated in the Health and Retirement Study (HRS), decomposes Gompertz acceleration into visible (measured-biomarker-driven) and latent components. Second, a positive-control-calibrated, two-platform cis-pQTL Mendelian-randomization and colocalization design (UKB-PPP, deCODE) against parental-lifespan GWAS tests whether the latent's measurable components are causal. Third, a clock battery (Horvath, chronological; DamAge, causality-enriched damage) tests reversibility in cellular reprogramming. Within the model, ~92% of Gompertz acceleration is assigned to a latent component not captured by measured blood-biomarker axes (NHANES 92.5%, HRS 91.6%); the latent is partly encoded in DNA-methylation signatures but not transcription. The known causal proteins are detected (LPA p=9×10-12; IL6R p=2.8×10-5), yet the latent's components, across inflammatory, renal and growth-signalling (IGFBP3, IGF-1) axes, are null and do not colocalize on either platform. Reprogramming reverses the chronological clock (-11 to -22 yr) but not the causality-enriched damage clock. The model-inferred mortality-driving component is largely latent to accessible biomarkers; its measurable molecular proxies show no supported causal effect where the design detects known causes; and the causality-enriched damage-clock signal is resistant to partial reprogramming.
Vitamin D is increasingly recognized as a multisystem hormone whose actions converge on physiological domains central to frailty. Low circulating 25-hydroxyvitamin D [25(OH)D] concentrations are associated with sarcopenia, bone loss, immune and cognitive dysfunction, and excess mortality, while observational cohort studies link vitamin D deficiency to frailty onset and progression. Mechanistic evidence provides a biologically coherent framework for these associations. Vitamin D receptor (VDR) signaling regulates myogenic differentiation, neuromuscular junction integrity, osteoblast-osteoclast coupling, and osteocyte-mediated mechanotransduction. It also modulates innate and adaptive immune responses, restrains inflammasome activation, and preserves mitochondrial function and metabolic flexibility. Disruption of VDR signaling promotes muscle atrophy, skeletal fragility, immunosenescence, and impaired bioenergetic capacity, processes that mirror key biological features of frailty. Despite this strong mechanistic plausibility, randomized controlled trials have largely yielded neutral results in community-dwelling adults with adequate vitamin D status. Clinical benefits appear confined to individuals with deficiency or heightened vulnerability, whereas intermittent high-dose bolus regimens have been associated with increased risks of falls and fractures. These discordant findings reflect persistent methodological limitations, including heterogeneous frailty definitions, insufficient follow-up duration, inter-assay variability in 25(OH)D measurement, and uncertainty regarding thresholds relevant to non-skeletal outcomes. In this narrative review, we integrate clinical, epidemiological, and mechanistic evidence to position vitamin D as a biomarker and potential modulator of vulnerability within a broader geroscience framework.
Statin optimization in older adults is increasingly relevant within the geroscience framework of cardiovascular ageing. This randomized controlled trial evaluated whether pharmacist-led pharmaceutical care improves lipid outcomes and statin adherence in community-dwelling older adults receiving primary care. A total of 99 patients aged 65 years or older on long-term statin therapy were randomized to usual care or structured pharmacist-led care delivered every 3 months. The primary outcome was change in low-density lipoprotein cholesterol at 6 months. Secondary outcomes included changes in other lipid parameters and dispensing-based adherence assessed by the proportion of days covered. The present analysis represents a pre-specified interim evaluation and was conducted on a per-protocol population with complete 6-month laboratory data. No statistically significant between-group difference in low-density lipoprotein cholesterol reduction was observed (adjusted difference -0.129 mmol/L, p = 0.385). Statin adherence was high and stable in both groups throughout follow-up, with 84-87% of patients meeting adherence thresholds and approximately 75-77% achieving perfect dispensing, consistent with an apparent ceiling effect. Exploratory sex-stratified analyses identified potential differences in lipid responses, but these were not statistically robust. In these highly adherent elderly populations, pharmacist-led pharmaceutical care was not associated with additional short-term improvement in LDL-C or dispensing-based adherence. The findings highlight the importance of baseline adherence when evaluating adherence-focused interventions in older adults.
Epigenetic aging is a new dimensional advancement in geroscience that continually addresses factors beyond genetic determinants of longevity. It occurs through three conventional pathways, one of which is non-coding RNA production. miR-124 is a newly introduced miRNA that was found in aged skin and several tumorigenic cells and is associated with aging. Additionally, SIRT1, a protein from the NAD+-dependent family, is found to have a protective mechanism in the pathways of aging. On the other hand, vitamin D works as an epigenetic factor and often targets pathways related to the aging process. In this study, the aim is to identify the expression of miR-124 in circulating blood to address the epigenetic-related biomarker and link the facets with anti-aging SIRT1 and vitamin D cognate receptor (VDR) to understand vitamin D deficiency interception. A vitamin D deficiency middle-aged rat model was developed by assigning male Albino Wistar Rats. Paricalcitol was administered IP (intraperitoneal) to the experimental group (32 ng/day) to induce deficiency, along with a calcium-rich rescue diet. The study was conducted for 21 days following blood collection and storage. ELISA and RTqPCR were performed to estimate serum 25(OH)D and gene expression analysis, respectively. 25(OH)D comparison between the group p-value of 0.002. The log2 fold changes of SIRT1, VDR, and miR-124 were compared between the groups using boxplot visualisation. A linear regression model was developed to understand the explorative nature of aging index (R2 = 0.561; Adjusted R2 = 0.517; p = 0.005; n = 12). A putative epigenetic relation was found between miR-124 levels and their association with SIRT1. The investigated biomarkers of aging are associated with serum concentration of 25(OH) vitamin D in a middle-aged vitamin D-deficient rat model, which supports the need for mechanistic investigation in future studies.
Aging is commonly framed as a progressive accumulation of cellular and tissue-level damage. However, the aged organism does not decline as a collection of isolated organs. Aging is communicated systemically through blood-borne signals that connect senescent cells, immune remodeling, vascular dysfunction, metabolic stress, dysbiosis, and chronic inflammation. I propose the concept of the circulating senosome to describe the composite network of age-associated circulating mediators, including senescence-associated secretory phenotype proteins, extracellular vesicles, inflammatory cytokines, lipids, metabolites, complement and coagulation mediators, autoantibodies, cell-free nucleic acids, and microbiome-derived products. This framework positions blood as both a biomarker compartment and a therapeutic interface in aging biology. The circulating senosome does not replace established hallmarks of aging; rather, it provides a systemic layer through which multiple hallmarks interact. Defining, measuring, and therapeutically remodeling this circulating network may create new opportunities for translational geroscience.
Collateral circulation that determines infarct progression in large vessel occlusion (LVO) is implicitly regarded as stationary. We investigated collateral circulation changes during interfacility transfer for endovascular thrombectomy (EVT) and its association with functional outcome in anterior circulation LVO stroke. Seventy consecutive patients with middle cerebral artery occlusion transferred for EVT from January 2017 to December 2018 within a regional stroke network underwent repeated CTA at the comprehensive center allowing longitudinal assessment of collateral status, quantified by the Tan score, and AI-derived collateral percentage. Demographics, NIHSS, intravenous thrombolysis, blood pressure, stroke-to-reperfusion time, recanalization status, and 90-day mRS were recorded. Ordinal logistic regression was used to identify predictors of outcome. Collateral status was dynamic rather than stationary, with changes observed in 51/70 (73%) patients (median = -3.69% (IQR =  - 22.28-4.83)). Collateral change emerged as a significant independent predictor of 90-day functional outcome (OR = 0.98, p = 0.008), with patients showing improvement in collaterals having better outcomes. Additionally, older age (OR = 1.04, p = 0.030), higher baseline NIHSS (OR = 1.20, p = 0.001), and longer stroke-to-reperfusion time (OR = 1.01, p = 0.014) were each significantly associated with worse 90-day functional outcome. Patients with improved/stable collaterals had significantly better 90-day functional outcomes compared to decliners (median mRS 2[IQR 0-4] vs. 4 [1.25-6], p = 0.008). Collateral status is not stationary during a stroke event, and its improvement independently predicts good functional outcome, highlighting the importance of therapeutic efforts to improve collateral circulation.
As a major contributor to cardiac diseases, thrombosis, a pathological process in which the formation of a blood clot restricts blood flow, is a leading cause of mortality worldwide. A central event in thrombosis involves thrombin-mediated cleavage of soluble fibrinogen into fibrin monomers, which polymerize into an insoluble fibrin mesh that forms the thrombus. While the risk of thrombosis increases markedly with age, the underlying cellular mechanisms linking aging and thrombus formation remain incompletely understood. Here, we identify a potential mechanism whereby cellular senescence in cardiac cells exacerbates thrombus formation via increased thrombin generation and impairs fibrinolysis, together promoting a pro-thrombotic state. We examined the pro-thrombotic phenotype of doxorubicin-induced senescence of human cardiac fibroblasts and cardiac endothelial cells. In both cell types, senescence accelerated and amplified fibrinogen polymerization. Treatment with a thrombin inhibitor reduced this polymerization and the cleavage of a thrombin-specific substrate by senescent cells, implicating thrombin as a key mediator of their pro-thrombotic phenotype. Complementary to their procoagulant effects, senescent cells also impaired thrombus resolution by suppressing plasminogen activation and downstream fibrinolysis. Gene expression analysis revealed senescence-associated upregulation of procoagulant and antifibrinolytic mediators, consistent with the observed functional effects. Finally, we show that the senescence-mediated polymerization of fibrinogen is dose dependent and attenuated by the senostatic agent, rapamycin. Collectively, these findings establish a direct mechanistic link between cellular senescence and thrombosis in cardiac cells, highlighting senescence as a potential therapeutic target in age-related thrombotic disease.
We examined whether a genome-wide polygenic lifespan score (PLS) is associated with all-cause mortality and how this association compares with associations between long-term lifestyle factors and mortality. The PLS was computed for the older Finnish Twin Cohort (mean age 57.4 years; 45.2% men; N = 5575). Cox regression was used to estimate the effects of the PLS on all-cause mortality risk, before and after adding sex, physical activity, BMI, alcohol consumption, smoking behavior, and education level. Concordance indices (C-indices) were used to assess each predictor's contribution to the model's discriminatory performance. Over a mean follow-up of 17.5 ± 8.3 years, 1405 deaths (25.2%) occurred. A one standard deviation increase in the PLS was statistically significantly associated with a lower all-cause mortality risk (hazard ratio [HR] = 0.838, 95% confidence interval [CI] = 0.792-0.887). This association remained relatively unchanged after adding all covariates (HR = 0.863, 95% CI = 0.816-0.912). Smoking 20 or more cigarettes per day showed the strongest association with increased mortality risk (HR = 3.341, 95% CI = 2.751-4.056), while female sex was associated with the greatest risk reduction (HR = 0.678, 95% CI = 0.597-0.770). Smoking behavior had the largest impact on model performance (ΔC-index = 0.027); other covariates contributed less (ΔC-index < 0.006). In conclusion, genetic predisposition to a longer lifespan was associated with a modest reduction in all-cause mortality risk, independent of lifestyle and other factors. Smoking behavior and female sex were stronger predictors of mortality than the PLS.
Understanding the links between metabolism, ageing, and age-related phenotypes may clarify the role of ageing in disease onset and improve risk prediction. We conducted a cross-cohort assessment of biological age using broad-spectrum LC-MS metabolomics of 3,686 plasma samples in 2,295 participants, aged 20-89, from the UK Airwave study (N = 960) and the Irish Longitudinal Study of Ageing (N = 1,335). The nucleoside N2,N2-dimethylguanosine, C-glycosyltryptophan, bile acid glucuronides, and the antioxidant zeta-carotene were associated with chronological age, frailty, and mortality. The noradrenergic metabolite 3-methoxy-4-hydroxyphenylglycol sulphate and the oligosaccharide sialyllactose were strongly associated with both age and mortality. We developed a metabolomic clock that was highly predictive of chronological age (r = 0.92) in test samples. Metabolomic age acceleration was strongly correlated between study visits (r > 0.6). Each standard deviation increase in metabolomic age acceleration (~ 5 years) was associated with 43% higher mortality risk, 27% higher risk of mild cognitive impairment, and 10% increased risk of a higher frailty score in fully adjusted models. The metabolites identified here may link ageing and age-related vulnerability and should be further investigated in mechanistic studies. The metabolomic clock has potential for translational applications, including as a prognostic and response marker of generalised age-related disease risk.
Unsupervised serious video games on the SmartMe&You home telemonitoring platform may enable the assessment of cognitive fluctuations in older individuals, with or without cognitive deficits. Here, we tested their validity in cognitively unimpaired older adults (Healthy) and patients with mild cognitive impairment and mild to moderate dementia due to Parkinson's disease (PDCD). Such validity was assessed using Mini-Mental State Examination (MMSE) scores and resting-state eyes-closed electroencephalography (rsEEG) activity. Clinical, demographic, and rsEEG datasets were collected from 34 healthy and 68 PDCD participants. The SmartMe&You video games included 7 unsupervised cognitive tasks implemented on a commercial tablet. The rsEEG source activities in the individual delta, theta, and alpha frequency bands were estimated using the eLORETA freeware. As novel findings, we showed that game performance: (1) discriminated between healthy participants and PDCD patients, with overall accuracy exceeding 90%; (2) was positively associated with MMSE scores across all participants (p < 0.00001); and (3) was negatively associated with global rsEEG source activity in the delta and theta frequency bands (p < 0.00001). These findings suggest that serious videogames of the SmartMe&You home telemonitoring platform were feasible and sensitive to cognitive status across the spectrum from normal aging to Parkinson's disease-related dementia. This approach provides a scalable, low-cost, and ecologically valid tool for remote cognitive monitoring, with future potential applications in both early detection and longitudinal tracking of cognitive decline.
Age-related disease burden accumulates heterogeneously from later midlife to older age, but the biology underlying these divergent trajectories is poorly understood. We analysed 7199 adults aged 40 years and over in the Tsuruoka Metabolomics Cohort Study, Japan, with baseline fasting plasma metabolomics (94 metabolites measured by capillary electrophoresis-mass spectrometry) and linked health insurance claims. Monthly cumulative Charlson Comorbidity Index scores were constructed from aligned cohort entry to 60 months to capture accumulation of newly documented Charlson conditions after follow-up start. K-means clustering identified six trajectories of claims-recorded disease burden, and ordinal logistic regression related metabolites to ordered trajectory severity with adjustment for demographic and lifestyle factors. Six trajectories ranged from minimal accumulation to rapid progression. Nineteen metabolites were associated with greater trajectory severity after false discovery rate correction. Glutamate showed the strongest positive association (odds ratio, 1.18 per standard deviation; 95% confidence interval, 1.12-1.24), whereas cysteine-glutathione disulfide showed the strongest inverse association (odds ratio, 0.89; 95% confidence interval, 0.86-0.93). Eighteen of these metabolites were also associated with time to first newly documented Charlson disease. Disease-specific analyses linked glutamate to diabetes with complications, mild liver disease, and cerebrovascular disease. Exploratory cluster-specific analyses identified hippurate as a distinctive marker of a late-acceleration trajectory. These findings implicate amino acid metabolism, redox balance, and microbiome-host interactions as candidate biological pathways underlying heterogeneous patterns of age-related disease accumulation, and warrant replication in independent cohorts. These signals may inform biomarker development for accelerated disease-burden accumulation.
Aging is accompanied by progressive epigenetic alterations, including changes in DNA methylation affecting both gene-associated regions and repetitive elements such as LINE-1 retrotransposons. Reduced epigenetic repression of transposable elements has been implicated in age-related genomic instability. Circulating cell-free DNA (cfDNA) provides a minimally invasive substrate for monitoring systemic molecular changes; however, age-related LINE-1 methylation dynamics in cfDNA remain poorly characterized in non-human species. Dogs represent a valuable translational model for aging research due to their shared environment with humans and shorter lifespan. We investigated age-associated LINE-1 CpG methylation in plasma-derived cfDNA and in prefrontal cortex genomic DNA from dogs. Sequence analysis of 264 potentially active canine LINE-1 elements identified a highly conserved central region spanning ORF1 and ORF2, with the targeted CpG site located within a locally low-variability segment, supporting multi-copy assessment. Methylation was quantified using methylation-sensitive restriction endonuclease digestion followed by real-time quantitative PCR (MSRED-PCR), with bisulfite conversion-based PCR (BSC-PCR) used for technical validation. In clinically healthy dogs, LINE-1 methylation in blood cfDNA showed a strong negative association with chronological age, best described by a nonlinear (logarithmic) model. In prefrontal cortex samples, older dogs exhibited significantly lower LINE-1 methylation compared to younger individuals. Across tissues, aging was consistently associated with LINE-1 hypomethylation. These findings indicate that age-related LINE-1 hypomethylation is detectable in both brain tissue and circulating cfDNA. While based on cross-sectional cohorts, the results suggest that LINE-1 methylation in cfDNA may represent a candidate minimally invasive marker of chronological aging in dogs.
Brain MRI shows promise for predicting cognitive functioning, but its utility depends on its capacity to capture stable between-person differences (e.g., patient stratification), longitudinal within-person changes (e.g., prognosis, treatment monitoring), or both. Using longitudinal data from 450 adults (aged 21-90; up to three waves, five years apart) in the Dallas Lifespan Brain Study, we benchmarked five modalities, task fMRI, functional connectivity (FC), structural MRI (sMRI), diffusion-weighted imaging (DWI), and arterial spin labeling (ASL), across 37 phenotypes and their combination. Stacking all MRI modalities into one marker predicted cognitive functioning with the highest accuracy (R2 = .51), followed by DWI and FC. Variance decomposition showed MRI markers explained substantial between-person variance (up to 60.3%) but modest within-person changes (up to 17.2%) in cognitive functioning. Commonality analysis revealed most markers, except ASL, overlapped with age-related variance in cognitive functioning. These findings clarify the strengths and limitations of MRI markers for stratifying and monitoring cognitive aging.