Identifying the relation between food groups and biological ageing: a data-driven approach.

BACKGROUND: Heterogeneity in ageing rates drives the need for research into lifestyle secrets of successful agers. Biological age, predicted by epigenetic clocks, has been shown to be a more reliable measure of ageing than chronological age. Dietary habits are known to affect the ageing process. However, much remains to be learnt about specific dietary habits that may directly affect the biological process of ageing. OBJECTIVE: To identify food groups that are directly related to biological ageing, using Copula Graphical Models. METHODS: We performed a preregistered analysis of 3,990 postmenopausal women from the Women's Health Initiative, based in North America. Biological age acceleration was calculated by the epigenetic clock PhenoAge using whole-blood DNA methylation. Copula Graphical Modelling, a powerful data-driven exploratory tool, was used to examine relations between food groups and biological ageing whilst adjusting for an extensive amount of confounders. Two food group-age acceleration networks were established: one based on the MyPyramid food grouping system and another based on item-level food group data. RESULTS: Intake of eggs, organ meat, sausages, cheese, legumes, starchy vegetables, added sugar and lunch meat was associated with biological age acceleration, whereas intake of peaches/nectarines/plums, poultry, nuts, discretionary oil and solid fat was associated with decelerated ageing. CONCLUSION: We identified several associations between specific food groups and biological ageing. These findings pave the way for subsequent studies to ascertain causality and magnitude of these relationships, thereby improving the understanding of biological mechanisms underlying the interplay between food groups and biological ageing.

A torpor-like state (TLS) in mice slows blood epigenetic aging and prolongs healthspan.

Torpor and hibernation are extreme physiological adaptations of homeotherms associated with pro-longevity effects. Yet the underlying mechanisms of how torpor affects aging, and whether hypothermic and hypometabolic states can be induced to slow aging and increase health span, remain unknown. We demonstrate that the activity of a spatially defined neuronal population in the avMLPA, which has previously been identified as a torpor-regulating brain region, is sufficient to induce a torpor like state (TLS) in mice. Prolonged induction of TLS slows epigenetic aging across multiple tissues and improves health span. We isolate the effects of decreased metabolic rate, long-term caloric restriction, and decreased core body temperature (Tb) on blood epigenetic aging and find that the pro-longevity effect of torpor-like states is mediated by decreased Tb. Taken together, our findings provide novel mechanistic insight into the pro-longevity effects of torpor and hibernation and support the growing body of evidence that Tb is an important mediator of aging processes.

MammalMethylClock R package: software for DNA methylation-based epigenetic clocks in mammals.

MOTIVATION: Epigenetic clocks are prediction methods based on DNA methylation levels in a given species or set of species. Defined as multivariate regression models, these DNA methylation-based biomarkers of age or mortality risk are useful in species conservation efforts and in preclinical studies. RESULTS: We present an R package called MammalMethylClock for the construction, assessment, and application of epigenetic clocks in different mammalian species. The R package includes the utility for implementing pre-existing mammalian clocks from the Mammalian Methylation Consortium. AVAILABILITY AND IMPLEMENTATION: The source code and documentation manual for MammalMethylClock, and clock coefficient .csv files that are included within this software package, can be found on Zenodo at https://doi.org/10.5281/zenodo.10971037.

Meta-analysis of epigenetic aging in schizophrenia reveals multifaceted relationships with age, sex, illness duration, and polygenic risk.

BACKGROUND: The study of biological age acceleration may help identify at-risk individuals and reduce the rising global burden of age-related diseases. Using DNA methylation (DNAm) clocks, we investigated biological aging in schizophrenia (SCZ), a mental illness that is associated with an increased prevalence of age-related disabilities and morbidities. In a whole blood DNAm sample of 1090 SCZ cases and 1206 controls across four European cohorts, we performed a meta-analysis of differential aging using three DNAm clocks (i.e., Hannum, Horvath, and Levine). To dissect how DNAm aging contributes to SCZ, we integrated information on duration of illness and SCZ polygenic risk, as well as stratified our analyses by chronological age and biological sex. RESULTS: We found that blood-based DNAm aging is significantly altered in SCZ independent from duration of the illness since onset. We observed sex-specific and nonlinear age effects that differed between clocks and point to possible distinct age windows of altered aging in SCZ. Most notably, intrinsic cellular age (Horvath clock) is decelerated in SCZ cases in young adulthood, while phenotypic age (Levine clock) is accelerated in later adulthood compared to controls. Accelerated phenotypic aging was most pronounced in women with SCZ carrying a high polygenic burden with an age acceleration of + 3.82 years (CI 2.02-5.61, P = 1.1E-03). Phenotypic aging and SCZ polygenic risk contributed additively to the illness and together explained up to 14.38% of the variance in disease status. CONCLUSIONS: Our study contributes to the growing body of evidence of altered DNAm aging in SCZ and points to intrinsic age deceleration in younger adulthood and phenotypic age acceleration in later adulthood in SCZ. Since increased phenotypic age is associated with increased risk of all-cause mortality, our findings indicate that specific and identifiable patient groups are at increased mortality risk as measured by the Levine clock. Our study did not find that DNAm aging could be explained by the duration of illness of patients, but we did observe age- and sex-specific effects that warrant further investigation. Finally, our results show that combining genetic and epigenetic predictors can improve predictions of disease outcomes and may help with disease management in schizophrenia.

Atherosclerotic Plaque Epigenetic Age Acceleration Predicts a Poor Prognosis and Is Associated With Endothelial-to-Mesenchymal Transition in Humans.

BACKGROUND: Epigenetic age estimators (clocks) are predictive of human mortality risk. However, it is not yet known whether the epigenetic age of atherosclerotic plaques is predictive for the risk of cardiovascular events. METHODS: Whole-genome DNA methylation of human carotid atherosclerotic plaques (n=485) and of blood (n=93) from the Athero-Express endarterectomy cohort was used to calculate epigenetic age acceleration (EAA). EAA was linked to clinical characteristics, plaque histology, and future cardiovascular events (n=136). We studied whole-genome DNA methylation and bulk and single-cell transcriptomics to uncover molecular mechanisms of plaque EAA. We experimentally confirmed our in silico findings using in vitro experiments in primary human coronary endothelial cells. RESULTS: Male and female patients with severe atherosclerosis had a median chronological age of 69 years. The median epigenetic age was 65 years in females (median EAA, -2.2 [interquartile range, -4.3 to 2.2] years) and 68 years in males (median EAA, -0.3 [interquartile range, -2.9 to 3.8] years). Patients with diabetes and a high body mass index had higher plaque EAA. Increased EAA of plaque predicted future events in a 3-year follow-up in a Cox regression model (univariate hazard ratio, 1.7; P=0.0034) and adjusted multivariate model (hazard ratio, 1.56; P=0.02). Plaque EAA predicted outcome independent of blood EAA (hazard ratio, 1.3; P=0.018) and of plaque hemorrhage (hazard ratio, 1.7; P=0.02). Single-cell RNA sequencing in plaque samples from 46 patients in the same cohort revealed smooth muscle and endothelial cells as important cell types in plaque EAA. Endothelial-to-mesenchymal transition was associated with EAA, which was experimentally confirmed by TGFß-triggered endothelial-to-mesenchymal transition inducing rapid epigenetic aging in coronary endothelial cells. CONCLUSIONS: Plaque EAA is a strong and independent marker of poor outcome in patients with severe atherosclerosis. Plaque EAA was linked to mesenchymal endothelial and smooth muscle cells. Endothelial-to-mesenchymal transition was associated with EAA, which was experimentally validated. Epigenetic aging mechanisms may provide new targets for treatments that reduce atherosclerosis complications.

Young Plasma Rejuvenates Blood DNA Methylation Profile, Extends Mean Lifespan, and Improves Physical Appearance in Old Rats.

There is converging evidence that young blood conveys cells, vesicles, and molecules able to revitalize function and restore organ integrity in old individuals. We assessed the effects of young plasma on the lifespan, epigenetic age, and healthspan of old female rats. Beginning at 25.6 months of age, a group of 9 rats (group T) was intraperitoneally injected with plasma from young rats until their natural death. A group of 8 control rats of the same age received no treatment (group C). Blood samples were collected every other week. Survival curves showed that from age 26 to 30 months, none of the group T animals died, whereas the survival curve of group C rats began to decline at age 26 months. Blood DNAm age versus chronological age showed that DNAm age in young animals increased faster than chronological age, then slowed down, entering a plateau after 27 months. The DNAm age of the treated rats fell below the DNAm age of controls and, in numerical terms, remained consistently lower until natural death. When rats were grouped according to the similarities in their differential blood DNA methylation profile, samples from the treated and control rats clustered in separate groups. Analysis of promoter differential methylation in genes involved in systemic regulatory activities revealed specific GO term enrichment related to the insulin-like factors pathways as well as to cytokines and chemokines associated with immune and homeostatic functions. We conclude that young plasma therapy may constitute a natural, noninvasive intervention for epigenetic rejuvenation and health enhancement.

Intervention with metabolites emulating endogenous cell transitions accelerates muscle regeneration in young and aged mice.

Tissue regeneration following an injury requires dynamic cell-state transitions that allow for establishing the cell identities required for the restoration of tissue homeostasis and function. Here, we present a biochemical intervention that induces an intermediate cell state mirroring a transition identified during normal differentiation of myoblasts and other multipotent and pluripotent cells to mature cells. When applied in somatic differentiated cells, the intervention, composed of one-carbon metabolites, reduces some dedifferentiation markers without losing the lineage identity, thus inducing limited reprogramming into a more flexible cell state. Moreover, the intervention enabled accelerated repair after muscle injury in young and aged mice. Overall, our study uncovers a conserved biochemical transitional phase that enhances cellular plasticity in vivo and hints at potential and scalable biochemical interventions of use in regenerative medicine and rejuvenation interventions that may be more tractable than genetic ones.

Epigenetic aging of human blood cells is influenced by the age of the host body.

Allogenic hematopoietic stem cell transplantation is a therapeutic procedure performed over a wide range of donor and recipient age combinations, representing natural experiments of how the age of the recipient affects aging in transplanted donor cells in vivo. We measured DNA methylation and epigenetic aging in donors and recipients and found that biological epigenetic clocks are accelerated in cells transplanted into an older body and decelerated in a younger body. This is the first evidence that the age of the circulating environment influences human epigenetic aging in vivo.

Liver-derived plasminogen mediates muscle stem cell expansion during caloric restriction through the plasminogen receptor Plg-RKT.

An intriguing effect of short-term caloric restriction (CR) is the expansion of certain stem cell populations, including muscle stem cells (satellite cells), which facilitate an accelerated regenerative program after injury. Here, we utilized the MetRSL274G (MetRS) transgenic mouse to identify liver-secreted plasminogen as a candidate for regulating satellite cell expansion during short-term CR. Knockdown of circulating plasminogen prevents satellite cell expansion during short-term CR. Furthermore, loss of the plasminogen receptor KT (Plg-RKT) is also sufficient to prevent CR-related satellite cell expansion, consistent with direct signaling of plasminogen through the plasminogen receptor Plg-RKT/ERK kinase to promote proliferation of satellite cells. Importantly, we are able to replicate many of these findings in human participants from the CALERIE trial. Our results demonstrate that CR enhances liver protein secretion of plasminogen, which signals directly to the muscle satellite cell through Plg-RKT to promote proliferation and subsequent muscle resilience during CR.

Alterations of the gut microbiome are associated with epigenetic age acceleration and physical fitness.

Epigenetic clocks can measure aging and predict the incidence of diseases and mortality. Higher levels of physical fitness are associated with a slower aging process and a healthier lifespan. Microbiome alterations occur in various diseases and during the aging process, yet their relation to epigenetic clocks is not explored. To fill this gap, we collected metagenomic (from stool), epigenetic (from blood), and exercise-related data from physically active individuals and, by applying epigenetic clocks, we examined the relationship between gut flora, blood-based epigenetic age acceleration, and physical fitness. We revealed that an increased entropy in the gut microbiome of physically active middle-aged/old individuals is associated with accelerated epigenetic aging, decreased fitness, or impaired health status. We also observed that a slower epigenetic aging and higher fitness level can be linked to altered abundance of some bacterial species often linked to anti-inflammatory effects. Overall our data suggest that alterations in the microbiome can be associated with epigenetic age acceleration and physical fitness.

Validation of biomarkers of aging.

The search for biomarkers that quantify biological aging (particularly 'omic'-based biomarkers) has intensified in recent years. Such biomarkers could predict aging-related outcomes and could serve as surrogate endpoints for the evaluation of interventions promoting healthy aging and longevity. However, no consensus exists on how biomarkers of aging should be validated before their translation to the clinic. Here, we review current efforts to evaluate the predictive validity of omic biomarkers of aging in population studies, discuss challenges in comparability and generalizability and provide recommendations to facilitate future validation of biomarkers of aging. Finally, we discuss how systematic validation can accelerate clinical translation of biomarkers of aging and their use in gerotherapeutic clinical trials.

Causality-enriched epigenetic age uncouples damage and adaptation.

Machine learning models based on DNA methylation data can predict biological age but often lack causal insights. By harnessing large-scale genetic data through epigenome-wide Mendelian randomization, we identified CpG sites potentially causal for aging-related traits. Neither the existing epigenetic clocks nor age-related differential DNA methylation are enriched in these sites. These CpGs include sites that contribute to aging and protect against it, yet their combined contribution negatively affects age-related traits. We established a new framework to introduce causal information into epigenetic clocks, resulting in DamAge and AdaptAge-clocks that track detrimental and adaptive methylation changes, respectively. DamAge correlates with adverse outcomes, including mortality, while AdaptAge is associated with beneficial adaptations. These causality-enriched clocks exhibit sensitivity to short-term interventions. Our findings provide a detailed landscape of CpG sites with putative causal links to lifespan and healthspan, facilitating the development of aging biomarkers, assessing interventions, and studying reversibility of age-associated changes.

Diet Quality and Epigenetic Aging in the Women's Health Initiative.

BACKGROUND: Higher diet quality scores are associated with a lower risk for many chronic diseases and all-cause mortality; however, it is unclear if diet quality is associated with aging biology. OBJECTIVE: This study aimed to examine the association between diet quality and a measure of biological aging known as epigenetic aging. DESIGN: A cross-sectional data analysis was used to examine the association between three diet quality scores based on self-reported food frequency questionnaire data and five measures of epigenetic aging based on DNA methylation (DNAm) data from peripheral blood. PARTICIPANTS/SETTING: This study included 4,500 postmenopausal women recruited from multiple sites across the United States (1993-98), aged 50 to 79 years, with food frequency questionnaire and DNAm data available from the Women's Health Initiative baseline visit. MAIN OUTCOME MEASURES: Five established epigenetic aging measures were generated from HumanMethylation450 Beadchip DNAm data, including AgeAccelHannum, AgeAccelHorvath, AgeAccelPheno, AgeAccelGrim, and DunedinPACE. STATISTICAL ANALYSES PERFORMED: Linear mixed models were used to test for associations between three diet quality scores (Healthy Eating Index, Dietary Approaches to Stop Hypertension, and alternate Mediterranean diet scores) and epigenetic aging measures, adjusted for age, race and ethnicity, education, tobacco smoking, physical activity, Women's Health Initiative substudy from which DNAm data were obtained, and DNAm-based estimates of leukocyte proportions. RESULTS: Healthy Eating Index, Dietary Approaches to Stop Hypertension, and alternate Mediterranean diet scores were all inversely associated with AgeAccelPheno, AgeAccelGrim, and DunedinPACE (P < 0.05), with the largest effects with DunedinPACE. A one standard deviation increment in diet quality scores was associated with a decrement (ß ± SE) in DunedinPACE z score of -0.097 ± 0.014 (P = 9.70 x 10-13) for Healthy Eating Index, -0.107 ± 0.014 (P = 1.53 x 10-14) for Dietary Approaches to Stop Hypertension, and -0.068 ± 0.013 (P = 2.31 x 10-07) for the alternate Mediterranean diet. CONCLUSIONS: In postmenopausal women, diet quality scores were inversely associated with DNAm-based measures of biological aging, particularly DunedinPACE.

DNA methylation clocks for clawed frogs reveal evolutionary conservation of epigenetic aging.

To address how conserved DNA methylation-based epigenetic aging is in diverse branches of the tree of life, we generated DNA methylation data from African clawed frogs (Xenopus laevis) and Western clawed frogs (Xenopus tropicalis) and built multiple epigenetic clocks. Dual species clocks were developed that apply to both humans and frogs (human-clawed frog clocks), supporting that epigenetic aging processes are evolutionary conserved outside mammals. Highly conserved positively age-related CpGs are located in neural-developmental genes such as uncx, tfap2d as well as nr4a2 implicated in age-associated disease. We conclude that signatures of epigenetic aging are evolutionary conserved between frogs and mammals and that the associated genes relate to neural processes, altogether opening opportunities to employ Xenopus as a model organism to study aging.

Higher testosterone and testosterone/estradiol ratio in men are associated with decreased Pheno-/GrimAge and DNA-methylation based PAI1.

Sex hormones are hypothesized to drive sex-specific health disparities. Here, we study the association between sex steroid hormones and DNA methylation-based (DNAm) biomarkers of age and mortality risk including Pheno Age Acceleration (AA), Grim AA, and DNAm-based estimators of Plasminogen Activator Inhibitor 1 (PAI1), and leptin concentrations. We pooled data from three population-based cohorts, the Framingham Heart Study Offspring Cohort, the Baltimore Longitudinal Study of Aging, and the InCHIANTI Study, including 1,062 postmenopausal women without hormone therapy and 1,612 men of European descent. Sex-stratified analyses using a linear mixed regression were performed, with a Benjamini-Hochberg (BH) adjustment for multiple testing. Sex Hormone Binding Globulin (SHBG) was associated with a decrease in DNAm PAI1 among men (per 1 standard deviation (SD): -478 pg/mL; 95%CI: -614 to -343; P:1e-11; BH-P: 1e-10), and women (-434 pg/mL; 95%CI: -589 to -279; P:1e-7; BH-P:2e-6). The testosterone/estradiol (TE) ratio was associated with a decrease in Pheno AA (-0.41 years; 95%CI: -0.70 to -0.12; P:0.01; BH-P: 0.04), and DNAm PAI1 (-351 pg/mL; 95%CI: -486 to -217; P:4e-7; BH-P:3e-6) among men. In men, testosterone was associated with a decrease in DNAm PAI1 (-481 pg/mL; 95%CI: -613 to -349; P:2e-12; BH-P:6e-11). SHBG was associated with lower DNAm PAI1 among men and women. Higher testosterone and testosterone/estradiol ratio were associated with lower DNAm PAI and a younger epigenetic age in men. A decrease in DNAm PAI1 is associated with lower mortality and morbidity risk indicating a potential protective effect of testosterone on lifespan and conceivably cardiovascular health via DNAm PAI1.

Associations between cardiorespiratory fitness and lifestyle-related factors with DNA methylation-based ageing clocks in older men: WASEDA'S Health Study.

DNA methylation-based age estimators (DNAm ageing clocks) are currently one of the most promising biomarkers for predicting biological age. However, the relationships between cardiorespiratory fitness (CRF), measured directly by expiratory gas analysis, and DNAm ageing clocks are largely unknown. We investigated the relationships between CRF and the age-adjusted value from the residuals of the regression of DNAm ageing clock to chronological age (DNAmAgeAcceleration: DNAmAgeAccel) and attempted to determine the relative contribution of CRF to DNAmAgeAccel in the presence of other lifestyle factors. DNA samples from 144 Japanese men aged 65-72 years were used to appraise first- (i.e., DNAmHorvath and DNAmHannum) and second- (i.e., DNAmPhenoAge, DNAmGrimAge, and DNAmFitAge) generation DNAm ageing clocks. Various surveys and measurements were conducted, including physical fitness, body composition, blood biochemical parameters, nutrient intake, smoking, alcohol consumption, disease status, sleep status, and chronotype. Both oxygen uptake at ventilatory threshold (VO2 /kg at VT) and peak oxygen uptake (VO2 /kg at Peak) showed a significant negative correlation with GrimAgeAccel, even after adjustments for chronological age and smoking and drinking status. Notably, VO2 /kg at VT and VO2 /kg at Peak above the reference value were also associated with delayed GrimAgeAccel. Multiple regression analysis showed that calf circumference, serum triglyceride, carbohydrate intake, and smoking status, rather than CRF, contributed more to GrimAgeAccel and FitAgeAccel. In conclusion, although the contribution of CRF to GrimAgeAccel and FitAgeAccel is relatively low compared to lifestyle-related factors such as smoking, the results suggest that the maintenance of CRF is associated with delayed biological ageing in older men.

Reversal of biological age in multiple rat organs by young porcine plasma fraction.

Young blood plasma is known to confer beneficial effects on various organs in mice and rats. However, it was not known whether plasma from young adult pigs rejuvenates old rat tissues at the epigenetic level; whether it alters the epigenetic clock, which is a highly accurate molecular biomarker of aging. To address this question, we developed and validated six different epigenetic clocks for rat tissues that are based on DNA methylation values derived from n = 613 tissue samples. As indicated by their respective names, the rat pan-tissue clock can be applied to DNA methylation profiles from all rat tissues, while the rat brain, liver, and blood clocks apply to the corresponding tissue types. We also developed two epigenetic clocks that apply to both human and rat tissues by adding n = 1366 human tissue samples to the training data. We employed these six rat clocks to investigate the rejuvenation effects of a porcine plasma fraction treatment in different rat tissues. The treatment more than halved the epigenetic ages of blood, heart, and liver tissue. A less pronounced, but statistically significant, rejuvenation effect could be observed in the hypothalamus. The treatment was accompanied by progressive improvement in the function of these organs as ascertained through numerous biochemical/physiological biomarkers, behavioral responses encompassing cognitive functions. An immunoglobulin G (IgG) N-glycosylation pattern shift from pro- to anti-inflammatory also indicated reversal of glycan aging. Overall, this study demonstrates that a young porcine plasma-derived treatment markedly reverses aging in rats according to epigenetic clocks, IgG glycans, and other biomarkers of aging.

DNA repair-deficient premature aging models display accelerated epigenetic age.

Several premature aging mouse models have been developed to study aging and identify interventions that can delay age-related diseases. Yet, it is still unclear whether these models truly recapitulate natural aging. Here, we analyzed DNA methylation in multiple tissues of four previously reported mouse models of premature aging (Ercc1, LAKI, Polg, and Xpg). We estimated DNA methylation (DNAm) age of these samples using the Horvath clock. The most pronounced increase in DNAm age could be observed in Ercc1 mice, a strain which exhibits a deficit in DNA nucleotide excision repair. Similarly, we detected an increase in epigenetic age in fibroblasts isolated from patients with progeroid syndromes associated with mutations in DNA excision repair genes. These findings highlight that mouse models with deficiencies in DNA repair, unlike other premature aging models, display accelerated epigenetic age, suggesting a strong connection between DNA damage and epigenetic dysregulation during aging.

Cross-species and tissue imputation of species-level DNA methylation samples across mammalian species.

DNA methylation data offers valuable insights into various aspects of mammalian biology. The recent introduction and large-scale application of the mammalian methylation array has significantly expanded the availability of such data across conserved sites in many mammalian species. In our study, we consider 13,245 samples profiled on this array encompassing 348 species and 59 tissues from 746 species-tissue combinations. While having some coverage of many different species and tissue types, this data captures only 3.6% of potential species-tissue combinations. To address this gap, we developed CMImpute (Cross-species Methylation Imputation), a method based on a Conditional Variational Autoencoder, to impute DNA methylation for non-profiled species-tissue combinations. In cross-validation, we demonstrate that CMImpute achieves a strong correlation with actual observed values, surpassing several baseline methods. Using CMImpute we imputed methylation data for 19,786 new species-tissue combinations. We believe that both CMImpute and our imputed data resource will be useful for DNA methylation analyses across a wide range of mammalian species.