A growing body of evidence indicates that the human virome, comprising both the gut and circulating viral communities, plays a critical role in shaping host physiology across the lifespan. In the context of aging, this complex viral ecosystem is increasingly recognized as a key modulator of immune function, inflammation, and metabolic balance, with direct implications for healthspan and longevity. While much attention has traditionally focused on bacterial components of the microbiota, recent advances in metagenomics have uncovered age-related shifts in the composition and function of the virome, including expansion of specific bacteriophage families, reactivation of latent viruses, and the persistence of commensal viral pathobionts. These changes are tightly linked to immunosenescence, chronic inflammation, and neurodegeneration, hallmarks of unhealthy aging. Notably, centenarians appear to harbor a unique virome signature marked by increased viral diversity, enhanced lytic activity, and the enrichment of phage-encoded metabolic functions, suggesting a potential protective role in extreme longevity. Despite these insights, significant challenges remain in virome profiling, including technical biases, database limitations, and the vast proportion of taxonomically unassigned sequences known as “viral dark matter”. This review highlights emerging data on the aging virome, underscores its relevance within the Geroscience framework, and discusses current barriers and future directions for translating virome research into clinical aging studies.

The glyoxalase pathway detoxifies reactive dicarbonyls generated during hyperglycemia, but the role of its epigenetic regulation in renal dysfunction and inflammatory dysregulation in older adults remains unclear. We investigated CpG-specific DNA methylation within the glyoxal detoxification pathway, focusing on the GLO1 gene, and examined associations with glycemic status, renal function, and systemic inflammation in hospitalized older adults. We identified a single CpG site within the GLO1 gene (cg26053840) significantly associated with fasting glycemia, suggesting that methylation levels at this locus reflects metabolic stress. Higher methylation at cg26053840 was also associated with impaired renal function, including increased serum creatinine and reduced estimated glomerular filtration rate. Additionally, GLO1 methylation correlated with multiple inflammatory indices, including C-reactive protein, erythrocyte sedimentation rate, neutrophil-to-lymphocyte ratio, and the CRP-to-albumin ratio. Associations with circulating cytokines and immune activation markers such as IL-6, IL-17A, GDF-15, CXCL9, CD163, and soluble RAGE further indicated broader immune-metabolic dysregulation. In silico analyses revealed a significant inverse correlation between cg26053840 methylation and GLO1 mRNA expression in the Broad Institute GDAC Firehose dataset. Genomic annotation further identified putative CEBPD and MYF6 transcription factor binding sites in proximity to the CpG site, suggesting a potential regulatory context. These findings support a model in which glycemic dysregulation increases methylglyoxal production, while reduced renal clearance enhances dicarbonyl stress, potentially driving epigenetic modulation of GLO1. These findings suggest the presence of a metabolic-epigenetic-inflammatory axis, although longitudinal and mechanistic studies are required to determine whether it contributes to organ dysfunction and vulnerability in hospitalized older adults.

Biomarkers of ageing are defined as age-related changes in body function or composition that could serve as a measure of 'biological' age and predict the onset of age-related diseases and/or residual life expectancy. We conducted the MARK-AGE Study, a European population study (3300 subjects aged 35-74) to identify a powerful set of biomarkers of ageing. A total of 362 clinical-chemistry, genetic, cellular or molecular biomarkers were analysed for each subject. Using statistical models as well as machine learning we derived mathematical formulas for females and for males that yield a 'bioage score' of an individual, based on sets of 10 biomarkers for females and 10 for males. Collectively, these biomarkers model chronological age of our study population and, thus yield the 'biological' age of a certain person. 'Age difference' (defined as biological minus chronological age) should then identify biologically older or younger individuals. Using our set of biomarkers, subjects with Down Syndrome and smoking females are biologically older, whereas postmenopausal females taking hormone replacement therapy are biologically younger. Strikingly, our data reveal that age difference of MARK-AGE subjects, but not chronological age, is linearly correlated with levels of HDL, 25-hydroxy-Vitamin D, and CD3+ CD4+/CD45+ ratio in such a way that biologically younger subjects display values that are favourable to good health, whereas other markers such as glucose and HbA1c are correlated with chronological age, but not age difference. This dichotomy of correlations may point to different roles of such markers, that is, drivers of the ageing process versus bystanders of ageing.

Memory declines in healthy aging, although deficits are heterogeneous. Implicit, procedural, and semantic memory, for example, are relatively preserved, whereas spatial abilities are particularly vulnerable. This vulnerability may reflect changes in the mechanisms underlying spatial navigation. Older adults tend to show impairments in allocentric processing, which encodes object-to-object relationships and environmental features, while relying more on egocentric, route-based strategies. Rodent studies are essential for investigating these changes. However, standard behavioral paradigms developed for young or adult animals may yield misleading or non-translatable results if applied without adaptation to aged mice and rats. This review focuses on four spatial memory tasks: the Morris water maze, the Barnes maze, the object location, and the spontaneous location recognition. These tests primarily rely on allocentric strategies and are sensitive to age-related alterations in the hippocampal-entorhinal cortex, the neural substrate for forming allocentric cognitive maps. The review highlights age-related characteristics, such as reduced physical fitness, sensory decline, and hormonal alterations, that require specific adjustments to experimental protocols, and outlines methodological considerations for adapting procedures, selecting additional assessments, and applying scoring systems in an unbiased manner. An exploratory survey of the literature is also presented to assess how frequently such adaptations are reported. Overall, the synthesis underscores the importance of methodological refinement for improving the validity and translational relevance of aging research.

Studies aimed at preventing age-associated diseases are fundamental in addressing the challenges posed by an aging population. However, biomedical and technological advancements have now reached a stage where it appears increasingly possible to repair the damage caused by severe pathologies and reverse the functional decline that accompanies aging. This perspective highlights the significance of using aging models, specifically non-transgenic geriatric mice (aged over 24 months), to study interventions aimed at reversing or ameliorating age-related pathologies. While most research typically utilizes young, adult, and mid-aged mice to investigate aging mechanisms and develop preventive strategies, geriatric models provide unique insights into the efficacy and safety of treatments in conditions that mimic the complexities of multiple concurrent diseases or syndromes. This manuscript highlights the importance of considering timing responses in aging interventions, illustrated by recent findings such as those involving canagliflozin. These studies reveal that the timing of intervention can significantly influence the outcomes, highlighting aspects often overlooked. Practical challenges and resource demands associated with geriatric mouse studies including concerns related to animal husbandry and aging phenotypes are also discussed. This perspective aims to foster a deeper understanding of the potential benefits and limitations of geriatric mice models in geroscience research and emphasizes the need for continued innovation in this field to meet the critical need to develop effective treatments for age-related diseases.

Although cancer treatment strategies have made considerable progress in recent decades, the challenge of selectively killing tumor cells while minimizing damage to healthy tissue remains. Radiotherapy (RT) continues to be crucial for tumor growth control when combined with surgery, chemotherapy, and immunotherapy. RT effectiveness depends on factors such as cancer type, tumor features, and the choice of external or internal treatment. Given its non-invasive nature and low systemic toxicity, RT is a suitable strategy for elderly patients. However, despite its efficacy, variations in cell sensitivity to radiation influence treatment outcomes, and normal cells surrounding the tumor can also be affected. Radiation-induced DNA damage can trigger cellular senescence, a permanent cell cycle arrest with a pro-inflammatory secretory phenotype, contributing to tissue damage in healthy cells and radio-resistance in tumor cells. RT causes not only DNA breakage but also epigenetic remodeling. Epigenetic pathways are involved in RT response, and the radiosensitivity of tumor cells can be modulated using epigenetic compounds. Emerging treatments based on more precise forms of RT, such as boron neutron capture therapy (BNCT), offer promising results by selectively targeting tumors while potentially preserving healthy tissues. BNCT has shown efficacy in glioblastoma, recurrent head and neck carcinoma, and melanoma. This review i) examines the mechanisms and challenges of conventional RT, focusing on aging and its role in treatment efficacy and tolerability, ii) describes the impact of senescence and epigenetic factors on resistance, iii) explores advances in BNCT, and iv) considers the potential of epigenetic drugs to optimize RT outcomes.

Torquetenovirus (TTV) is a prevalent virus whose clinical significance remains unclear, potentially linked to immunosenescence. This study examines TTV species in relation to immune impairment, inflammation, and cellular stress response in aging. A subset of recruited age-stratified individuals (RASIG) from the MARK-AGE study was divided into three groups: Cohort 1 A (healthy young adults), Cohort 1B (older adults with mild immune decline), and Cohort 1 C (older adults with marked immune impairment). Analyses included TTV load, species diversity, lymphocyte subpopulations, inflammatory markers, Poly-(ADP-ribose) polymerase (PARP-1) expression/activity. Alpha- and beta-diversity analyses showed the highest TTV species diversity in Cohort 1 C, with significant cohort-dependent differences and partially cohort-specific clustering patterns. Increased TTV species number correlated with higher TTV load, elevated CMV IgG levels, and greater immune impairment risk. Specific TTV species were associated with CD4/CD8, and reduced T-cell receptor excision circles, suggesting impaired T-cell homeostasis. TTV viremia positively correlated with C-reactive protein (CRP) and α2-macroglobulin. PARP-1 expression and activity increased in individuals with higher TTV diversity, particularly in the presence of TTV9 and TTV20. TTV load and species diversity are associated with immunosenescence, inflammation, and PARP-1 activation suggesting their potential as biomarkers of age-related immune decline. Longitudinal studies are needed to clarify underlying mechanisms.

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.

Genomic instability markers are important hallmarks of aging, as previously evidenced within the European study of biomarkers of human aging, MARK-AGE; however, establishing the specific metabolic determinants of vascular aging is challenging. The objective of the present study was to evaluate the impact of the susceptibility to oxidation of serum LDL particles (LDLox) and the plasma metabolization products of nitric oxide (NOx) on relevant genomic instability markers. The analysis was performed on a MARK-AGE cohort of 1326 subjects (635 men and 691 women, 35–75 years old) randomly recruited from the general population. The Inverse Probability of Treatment Weighting causal inference algorithm was implemented in order to assess the potential causal relationship between the LDLox and NOx octile-based thresholds and three genomic instability markers measured in mononuclear leukocytes: the percentage of telomeres shorter than 3 kb, the initial DNA integrity, and the DNA damage after irradiation with 3.8 Gy. The results showed statistically significant telomere shortening for LDLox, while NOx yielded a significant impact on DNA integrity. Overall, the effect on the genomic instability markers was higher than for the confirmed vascular aging determinants, such as low HDL cholesterol levels, indicating a meaningful impact even for small changes in LDLox and NOx values.

Aging is a natural, multifactorial biological process characterized by progressive cellular and tissue damage in response to various stressors, leading to functional decline that often affects multiple organs, contributing to the development of age-related diseases. Although life expectancy has increased significantly, age-related conditions have become the leading causes of impairment and disability in the elderly, becoming a major global health concern. This highlights the need for innovative, multitarget strategies to modulate the aging process and extend healthspan. In recent years, researchers have explored natural solutions to counteract the hallmarks of aging. Among these, marine-derived molecules represent an up-and-coming niche of bioactive compounds, distinguished by their unique structural diversity and multifunctional properties. Marine products are increasingly studied for their antioxidant, anti-inflammatory and cytoprotective properties, targeting key pathways involved in aging, such as cellular senescence, genomic instability, impaired autophagy, and chronic inflammation. In this review, we aim to (i) explore the field of marine-derived bioactive molecules which demonstrated effects on lifespan extension, (ii) summarize studies showing their capacity to target one or more hallmarks of aging, (iii) highlight those that exhibit therapeutic potential in age-related diseases - including neurodegenerative, cardiovascular, metabolic, cancer, musculoskeletal, and chronic pulmonary disorders. Their multitarget activity makes them attractive candidates for the prevention or treatment of age-related diseases, and several have shown promising results in preclinical studies. However, only a limited number of these compounds have progressed to late-stage clinical trials, highlighting the need for further translational research, which may pave the way for novel anti-aging therapeutic strategies.

Cellular senescence is a conserved cellular program characterized by a permanent cell cycle arrest triggered by a variety of stressors. Originally described as a tumor-suppressive mechanism, it is now recognized to exert pleiotropic and context-dependent functions, contributing to key physiological processes such as embryogenesis and tissue repair, as well as to processes associated with aging and the development of age-related diseases. Unlike normal cells, senescent cells remain metabolically active despite their non-dividing state. They significantly impact their environment through the Senescence-Associated Secretory Phenotype (SASP), a complex mix of cytokines, growth factors, and proteases. This secretory profile can promote tissue repair and regeneration but, if persistent, contributes to chronic inflammation, fibrosis, and tissue dysfunction. Two major pathways primarily regulate senescence: the p53/p21 and p16^INK4a^/Rb axes. These respond to stress signals like DNA damage, oxidative stress, and oncogenic activation, enforcing stable cell cycle arrest to prevent uncontrolled proliferation. However, as senescent cells accumulate over time, their ongoing SASP activity disrupts tissue homeostasis, driving inflammation and age-related diseases. Recent advances in multi-omics technologies, including metabolomics, proteomics, and lipidomics, have provided deeper insights into the complex molecular changes within senescent cells, revealing new biomarkers and potential therapeutic targets. These approaches offer a comprehensive understanding of cellular senescence, but challenges remain in distinguishing the causal relationships within these data and translating findings into clinical applications. This review integrates recent multi-omics discoveries, highlighting their potential to refine our understanding of senescence and support the development of targeted interventions to extend healthspan and combat age-related pathologies.

BACKGROUND:Grip strength is a key functional marker of musculoskeletal aging, widely used to assess sarcopenia. In preclinical research, multiple measurement methods are often combined to enhance reliability, but standardization remains challenging. To improve measurement robustness, we previously developed a composite strength score (SS5) that integrates five different grip strength tests into a single variable. While SS5 provides a comprehensive evaluation, its implementation is time-consuming, limiting feasibility in large-scale studies. In this study, we also examine two simplified composite scores, SS2 and SS3, as potential streamlined alternatives. Additionally, although normalizing grip strength to body weight is widely used, its appropriateness in geriatric mouse models has never been formally validated. METHODS:Forelimb grip strength was assessed in a cohort of 160-aged C57BL/6J mice using five methods: Weight Lift Tests (Deacon protocol with sponge weights and a modified version with metal wire weights), the Cage Lift Test and the Grip Strength Meter (trapeze bar and grid). Additionally, a cross-sectional group of 173 mice was analysed to assess the correlation between grip strength and muscle size. Each method was evaluated for its correlation with age, ability to detect sex differences, variability and association with muscle size. RESULTS:All methods strongly correlated with age (-0.518 ≤ rs ≤ -0.306). The Grip Strength Meter (trapeze bar) and modified Deacon method were the most effective in detecting sex differences (p < 0.001). While all methods correlated with muscle size (0.153 ≤ rs ≤ 0.332), the modified Deacon method and Grip Strength Meter showed the strongest associations. The mean coefficient of variation (CV%) ranged from 7% to 17%, demonstrating good repeatability. Notably, despite being widely used, normalization of grip strength to body weight was found to introduce bias in geriatric mice, as age-related weight loss distorts strength assessments. Absolute values proved to be a more reliable measure. To improve efficiency while maintaining reliability, we developed two new composite scores (SS2 and SS3) by integrating a subset of methods from SS5. These scores preserved the strong correlation with age observed in SS5 while reducing the number of required tests, enhancing feasibility. CONCLUSIONS:Combining multiple grip strength assessments improves measurement reliability in aging studies. The newly proposed SS2 and SS3 scores provide a streamlined yet robust alternative to SS5, improving standardization and facilitating future comparisons in preclinical sarcopenia research. Our findings also challenge the routine normalization of grip strength to body weight in geriatric mice, emphasizing the importance of using absolute values to avoid bias.

Cytomegalovirus (CMV) infection has been linked to accelerated biological aging, potentially increasing the risk of cardiovascular disease. DNA methylation of the gene Elongation Of Very Long Chain Fatty Acids-Like 2 (ELOVL2) is a molecular biomarker for aging, and its gene product is involved in polyunsaturated fatty acid synthesis, which impacts immune and inflammatory responses. This study, conducted in the MARK-AGE population, aimed to investigate the relationship between CMV infection and ELOVL2 methylation in adults aged 35-75, as well as the influence of CMV IgG levels on lipid metabolism, inflammation, DNA damage, and DNA repair. Our data revealed a higher prevalence of ischemic heart disease, atrial fibrillation, hypertension, and diabetes in CMV-positive individuals. CMV IgG levels were positively associated with ELOVL2 methylation at specific CpG sites and with increased expression of DNA methyltransferase-1 (DNMT1). CMV IgG was linked to lipid imbalances, such as increased BMI, VLDL-cholesterol, triglycerides, and HDL1-cholesterol. Additionally, ELOVL2 methylation was associated with systemic inflammation markers, lipid parameters and altered T-cell subsets. A negative correlation was observed between CMV IgG levels and both baseline DNA integrity and repair capacity. These results suggest that CMV infection might promote cardiovascular disease through ELOVL2 hypermethylation, lipid dysregulation, inflammation, and DNA damage.

The accumulation of senescent cells contributes to aging and related diseases; therefore, discovering safe senolytic agents-compounds that selectively eliminate senescent cells-is a critical priority. Heat shock protein 90 (HSP90) inhibitors (HSP90i), traditionally investigated for cancer treatment, have shown potential as senolytic agents. However, inhibitors face formulation, toxicity, and cost challenges. To overcome these limitations, we employed a virtual screening approach combining structure-based prefiltering with a ligand-based pharmacophore model to identify novel, potentially safe HSP90 alpha isoform inhibitors exhibiting senolytic properties. This strategy identified 14 candidate molecules evaluated for senolytic activity in primary human fetal pulmonary fibroblasts. Four compounds exhibited significant HSP90i and senolytic activity, including two novel compounds, namely K4 and K5. The latter, 1-benzyl-3-(2-methylphenyl)-3,7-dihydro-1H-purine-2,6-dione, structurally related to the xanthinic family, emerged as a promising, well-tolerated senolytic agent. K5 demonstrated senolytic activity across various cellular senescence models, including human fibroblasts, mesenchymal stem cells, and breast cancer cells. It was also effective in vivo, extending lifespan in Drosophila and reducing senescence markers in geriatric mice. Additionally, the xanthinic nature of K5 implicates a multimodal action, now including the inhibition of HSP90α, that might enhance its efficacy and selectivity towards senescent cells, Senolytic index SI > 1320 for IMR90 cells, and SI > 770 for WI38 cells, underscoring its therapeutic potential. These findings advance senolytic therapy research, opening new avenues for safer interventions to combat age-related inflammaging and diseases, including cancer, and possibly extend a healthy lifespan.

The choroid plexus (ChP) is a complex ventricular structure that forms a semi-permeable barrier between the blood and cerebrospinal fluid (CSF). It is responsible for CSF secretion and clearance, contains macrophages, and is one of the few sites within the central nervous system (CNS) where T cells are present. Additionally, the ChP plays a role in detecting peripheral inflammation, which leads to the modulation of its epithelial cell function. Despite its critical importance in maintaining brain homeostasis, the ChP is often overlooked, particularly concerning the changes it undergoes with aging, such as reduced barrier function, impaired CSF production, and altered immunosurveillance. These age-related alterations may contribute to several harmful effects, including neuroinflammation and oxidative damage, potentially predisposing individuals to neurodegenerative conditions. Although knowledge is still limited, gut dysbiosis and decreased Klotho levels-of which the ChP is one of the main sources-appear to be significant contributors to ChP impairments. This narrative review will examine the impact of age-related gut dysbiosis on the CNS, focusing on the ChP, and explore the effects of reduced Klotho levels in this brain structure. We will also propose the hypothesis that combining the administration of probiotics capable of restoring gut microbiota eubiosis with gene therapy to upregulate Klotho in the ChP could help preserve the structural and functional integrity of the aging brain. Finally, we will provide a technical overview to ensure that vectors encoding Klotho cDNA achieve maximum specificity for the ChP, thereby avoiding off-target effects.

Torque Teno Virus (TTV) is a widespread commensal virus within the human virome, characterized by a high prevalence in human population and an unclear pathogenic role. Over the past three decades, TTV has garnered increasing attention due to its ability to establish lifelong chronic viremia, which intriguingly fluctuates among individuals in relation to immune competence status, with a typical peak after an organ transplantation, followed by a plateau and a slow decrease. The regulatory mechanisms underlying TTV infection remain elusive, and factors influencing its interactions with the immune system have yet to be identified. To explore this complex interplay, we analyzed DNA methylation patterns associated with TTV load in older adult hospitalized patients (mean age: 83.15 ± 7.49) from the PROMOTERA cohort. In this study, we present for the first time the identification of differentially methylated probes (DMPs) correlated to TTV load in our cohort. The statistically significant DMPs were located in genes involved in immune regulation and lipid metabolism. To further characterize these findings, we performed an exploratory enrichment analysis by applying several p-value thresholds, which yielded multiple gene lists derived from the sets of significant probes. Genes associated with this epigenetic signature were found to enrich functional pathways related to immune activation, leukocyte differentiation, and cytokine production, while additional significantly enriched gene sets were involved in cell–cell adhesion and cell migration processes. Since our analysis followed an exploratory approach, these results should be interpreted as hypothesis-generating and warrant further investigation.

Loss of cognitive function is a significant challenge in aging, and developing models to understand and target cognitive decline is crucial for the development of Geroscience-based interventions. Aged mice offer a valuable model as they share features of cognitive decline with humans. Despite numerous studies, knowledge of longitudinal age-related cognitive changes and cognitive frailty in naturally aging mice is limited, particularly in cohorts exceeding 30 months of age, where cognitive decline is more pronounced. Moreover, the impaired physical function of aged mice is known to affect latency-based strategies to measure cognitive performances. Here, we show a comprehensive longitudinal assessment using the Barnes Maze test in a large cohort of 424 aged (≥ 21 months) C57BL/6J mice. We introduced a new metric, the Cognitive Frailty Index (CoFI), which summarizes different age-associated Barnes Maze parameters into a unique function. CoFI strongly associates with advancing age and mortality, offering a reliable ability to discriminate long- and short-lived mice. We also established a CoFI cut-off and a physically adjusted CoFI, both of which can distinguish between physical and cognitive frailty. This is further supported by the enhanced predictive power when physical and cognitive frailty are combined to assess short-term mortality. Moreover, the computation method for CoFI is adaptable to various cognitive assessment tests, leveraging procedures akin to those used for calculating other frailty indices. In conclusion, through robust longitudinal tracking, CoFI has the potential to become an important ally in assessing the effectiveness of Geroscience-based interventions to counteract age-related cognitive impairment.

Aging is accompanied by chronic low-grade inflammation ("inflammaging"), which contributes to increased morbidity and mortality in older adults. This study evaluated the prognostic value of circulating inflammatory biomarkers, i.e. interleukin-6 (IL-6), interleukin-10 (IL-10), and CXCL9, and their integration with frailty for long-term risk stratification. We analyzed 1,009 patients (median age, 84 years) hospitalized in acute care wards of three Italian geriatric hospitals as part of the ReportAGE cohort. Frailty was assessed using a deficit accumulation–based Frailty Index (FI), and serum cytokines were measured by immunoassay. Cytokine-specific risk categories were combined into a composite I3 score (range, 3–9). Cox proportional hazards models adjusted for age, sex, comorbidity burden, polypharmacy, and laboratory variables were used to assess associations with 10-year mortality. In a subset of 237 patients, DNA methylation–based estimates of cytokine levels were also analyzed. Higher I3 scores were independently associated with increased mortality (hazard ratio [HR] 2.42, 95

Torque teno virus (TTV) is a ubiquitous virus whose viremia increases in conditions of immune dysfunction and aging, suggesting its potential role as a biomarker of immunosenescence. This study investigated the association between TTV viremia and all-cause mortality risk over seven years in a hospitalized older cohort, and its relationship with inflammatory markers including osteopontin (OPN) and growth differentiation factor 15 (GDF15). Data from 956 patients were analyzed, with high TTV load defined as ≥5 log DNA copies/mL. High TTV viremia was significantly associated with increased mortality risk at 1, 3, and 7 years independently of age, sex, comorbidities, and inflammatory markers. In stratified analyses, this association was significant at one year in both males and females, but persisted at three and seven years only in males. The strongest association was observed in participants aged 80-89 years, remaining significant across all follow-up periods. When patients were stratified by a composite immune score reflecting degrees of immunosenescence, high TTV viremia predicted increased mortality among those with intermediate or severe immune dysfunction, persisting up to seven years in the most immunosenescent subgroup. Patients with elevated TTV loads exhibited increased erythrocyte sedimentation rate (ESR), decreased serum albumin and hemoglobin, and significantly higher plasma levels of OPN and GDF15, whereas IL-10 tended to decrease. No significant differences were observed for neutrophil-to-lymphocyte ratio, IL-6, CD163, CCL22, or CXCL9 between high and low TTV viremia groups. These findings indicate that high TTV viremia independently predicts mortality risk and reflects a pro-inflammatory and immunosenescent state.