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Abstract Parkinson’s disease is increasingly prevalent. It progresses from the pre-motor stage (characterised by non-motor symptoms like REM sleep behaviour disorder), to the disabling motor stage. We need objective biomarkers for early/pre-motor disease stages to be able to intervene and slow the underlying neurodegenerative process. Here, we validate a targeted multiplexed mass spectrometry assay for blood samples from recently diagnosed motor Parkinson’s patients (n = 99), pre-motor individuals with isolated REM sleep behaviour disorder (two cohorts: n = 18 and n = 54 longitudinally), and healthy controls (n = 36). Our machine-learning model accurately identifies all Parkinson patients and classifies 79% of the pre-motor individuals up to 7 years before motor onset by analysing the expression of eight proteins—Granulin precursor, Mannan-binding-lectin-serine-peptidase-2, Endoplasmatic-reticulum-chaperone-BiP, Prostaglaindin-H2-D-isomaerase, Interceullular-adhesion-molecule-1, Complement C3, Dickkopf-WNT-signalling pathway-inhibitor-3, and Plasma-protease-C1-inhibitor. Many of these biomarkers correlate with symptom severity. This specific blood panel indicates molecular events in early stages and could help identify at-risk participants for clinical trials aimed at slowing/preventing motor Parkinson’s disease.

Summary: Recent studies have shown that elevated concentrations of unconjugated bilirubin (UCB) may be a protective host factor against the development of noncommunicable diseases (NCDs), whereas low levels of UCB are associated with the opposite effect. The results of this European study, in which 2,489 samples were tested for their UCB concentration using high-performance liquid chromatography (HPLC) and additional data from the MARK-AGE database were used for analysis, provide further evidence that elevated UCB concentrations are linked to a lower risk of developing NCDs and may act as a predictive marker of biological aging as individuals with elevated UCB concentrations showed favorable outcomes in metabolic health and oxidative-stress-related biomarkers. These findings underline the significance of studying individuals with moderate hyperbilirubinemia and investigate UCB routinely, also in the setting of aging, since this condition affects millions of people worldwide but has been underrepresented in clinical research and practice until now.

Abstract Background Yakuts are one of the indigenous populations of the subarctic and arctic territories of Siberia characterized by a continental subarctic climate with severe winters, with the regular January average temperature in the regional capital city of Yakutsk dipping below − 40 °C. The epigenetic mechanisms of adaptation to such ecologies and environments and, in particular, epigenetic age acceleration in the local population have not been studied before. Results This work reports the first epigenetic study of the Yakutian population using whole-blood DNA methylation data, supplemented with the comparison to the residents of Central Russia. Gene set enrichment analysis revealed, among others, geographic region-specific differentially methylated regions associated with adaptation to climatic conditions (water consumption, digestive system regulation), aging processes (actin filament activity, cell fate), and both of them (channel activity, regulation of steroid and corticosteroid hormone secretion). Further, it is demonstrated that the epigenetic age acceleration of the Yakutian representatives is significantly higher than that of Central Russia counterparts. For both geographic regions, we showed that epigenetically males age faster than females, whereas no significant sex differences were found between the regions. Conclusions We performed the first study of the epigenetic data of the Yakutia cohort, paying special attention to region-specific features, aging processes, age acceleration, and sex specificity.

Introduction: DNA methylation clocks presents advantageous characteristics with respect to the ambitious goal of identifying very early markers of disease, based on the concept that accelerated ageing is a reliable predictor in this sense.Methods: Such tools, being epigenomic based, are expected to be conditioned by sex and tissue specificities, and this work is about quantifying this dependency as well as that from the regression model and the size of the training set.Results: Our quantitative results indicate that elastic-net penalization is the best performing strategy, and better so when—unsurprisingly—the data set is bigger; sex does not appear to condition clocks performances and tissue specific clocks appear to perform better than generic blood clocks. Finally, when considering all trained clocks, we identified a subset of genes that, to the best of our knowledge, have not been presented yet and might deserve further investigation: CPT1A, MMP15, SHROOM3, SLIT3, and SYNGR.Conclusion: These factual starting points can be useful for the future medical translation of clocks and in particular in the debate between multi-tissue clocks, generally trained on a large majority of blood samples, and tissue-specific clocks.

The notion of notable anatomical, biochemical, and behavioral distinctions within male and female brains has been a contentious topic of interest within the scientific community over several decades. Advancements in neuroimaging and molecular biological techniques have increasingly elucidated common mechanisms characterizing brain aging while also revealing disparities between sexes in these processes. Variations in cognitive functions; susceptibility to and progression of neurodegenerative conditions, notably Alzheimer’s and Parkinson’s diseases; and notable disparities in life expectancy between sexes, underscore the significance of evaluating aging within the framework of gender differences. This comprehensive review surveys contemporary literature on the restructuring of brain structures and fundamental processes unfolding in the aging brain at cellular and molecular levels, with a focus on gender distinctions. Additionally, the review delves into age-related cognitive alterations, exploring factors influencing the acceleration or deceleration of aging, with particular attention to estrogen’s hormonal support of the central nervous system.

Abstract Aging is a gradual, continuous series of natural changes in biological, physiological, immunological, environmental, psychological, behavioral, and social processes. Aging entails changes in the immune system characterized by a decrease in thymic output of naïve lymphocytes, an accumulated chronic antigenic stress notably caused by chronic infections such as cytomegalovirus (CMV), and immune cell senescence with acquisition of an inflammatory senescence-associated secretory phenotype (SASP). For this reason, and due to the SASP originating from other tissues, aging is commonly accompanied by low-grade chronic inflammation, termed “inflammaging”. After decades of accumulating evidence regarding age-related processes and chronic inflammation, the domain now appears mature enough to allow an integrative reinterpretation of old data. Here, we provide an overview of the topics discussed in a recent workshop “Aging and Chronic Inflammation” to which many of the major players in the field contributed. We highlight advances in systematic measurement and interpretation of biological markers of aging, as well as their implications for human health and longevity and the interventions that can be envisaged to maintain or improve immune function in older people.

Abstract Isolated REM Sleep Behavior Disorder (iRBD) is the strongest prodromal marker for α-synucleinopathies. Overt α-synucleinopathies and aging share several mechanisms, but this relationship has been poorly investigated in prodromal phases. Using DNA methylation-based epigenetic clocks, we measured biological aging in videopolysomnography confirmed iRBD patients, videopolysomnography-negative and population-based controls. We found that iRBDs tended to be epigenetically older than controls, suggesting that accelerated aging characterizes prodromal neurodegeneration.

Subcutaneous adipocytes are crucial for mammary gland epithelial development during pregnancy. Our and others’ previous data have suggested that adipo-epithelial transdifferentiation could play a key role in the mammary gland alveolar development. In this study, we tested whether adipo-epithelial transdifferentiation occurs in vitro. Data show that, under appropriate co-culture conditions with mammary epithelial organoids (MEOs), mature adipocytes lose their phenotype and acquire an epithelial one. Interestingly, even in the absence of MEOs, extracellular matrix and diffusible growth factors are able to promote adipo-epithelial transdifferentiation. Gene and protein expression studies indicate that transdifferentiating adipocytes exhibit some characteristics of milk-secreting alveolar glands, including significantly higher expression of milk proteins such as whey acidic protein and β-casein. Similar data were also obtained in cultured human multipotent adipose-derived stem cell adipocytes. A miRNA sequencing experiment on the supernatant highlighted mir200c, which has a well-established role in the mesenchymal–epithelial transition, as a potential player in this phenomenon. Collectively, our data show that adipo-epithelial transdifferentiation can be reproduced in in vitro models where this phenomenon can be investigated at the molecular level.

Abstract Background There are several mechanisms via which increased protein intake might maintain or improve bone mineral density (BMD), but current evidence for an association or effect is inconclusive. The objectives of this study were to investigate the association between dietary protein intake (total, plant and animal) with BMD (spine and total body) and the effects of protein supplementation on BMD. Methods Individual data from four trials that included either (pre‐)frail, undernourished or healthy older adults (aged ≥65 years) were combined. Dietary intake was assessed with food records (2, 3 or 7 days) and BMD with dual‐energy X‐ray absorptiometry (DXA). Associations and effects were assessed by adjusted linear mixed models. Results A total of 1570 participants [57% women, median (inter‐quartile range): age 71 (68–75) years] for which at least total protein intake and total body BMD were known were included in cross‐sectional analyses. In fully adjusted models, total protein intake was associated with higher total body and spine BMD [beta (95% confidence interval): 0.0011 (0.0006–0.0015) and 0.0015 (0.0007–0.0023) g/cm2, respectively]. Animal protein intake was associated with higher total body and spine BMD as well [0.0011 (0.0007–0.0016) and 0.0017 (0.0010–0.0024) g/cm2, respectively]. Plant protein intake was associated with a lower total body and spine BMD [−0.0010 (−0.0020 to −0.0001) and −0.0019 (−0.0034 to −0.0004) g/cm2, respectively]. Associations were similar between sexes. Participants with a high ratio of animal to plant protein intake had higher BMD. In participants with an adequate calcium intake and sufficient serum 25(OH)D concentrations, the association between total protein intake with total body and spine BMD became stronger. Likewise, the association between animal protein intake with total body BMD was stronger. In the longitudinal analyses, 340 participants [58% women, median (inter‐quartile range): age 75 (70–81) years] were included. Interventions of 12 or 24 weeks with protein supplementation or protein supplementation combined with resistance exercise did not lead to significant improvements in BMD. Conclusions An association between total and animal protein intake with higher BMD was found. In contrast, plant protein intake was associated with lower BMD. Research is warranted to further investigate the added value of dietary protein alongside calcium and vitamin D for BMD improvement, especially in osteopenic or osteoporotic individuals. Moreover, more research on the impact of a plant‐based diet on bone health is needed.

Abstract Sex differences in human survival have been extensively investigated in many studies that have in part uncovered the biological determinants that promote a longer life in females with respect to males. Moreover, researches performed in the past years have prompted increased awareness about the biological effects of environmental factors that can modulate the magnitude of the sex gap in survival. Besides the genetic background, epigenetic modifications like DNA methylation, that can modulate cell function, have been particularly studied in this framework. In this review, we aim to summarize the role of the genetic and epigenetic mechanisms in promoting female advantage from the early in life (“INNATE” features), and in influencing the magnitude of the gap in sex differences in survival and ageing (“VARIABLE” features). After briefly discussing the biological bases of sex determination in humans, we will provide much evidence showing that (i) “innate” mechanisms common to all males and to all females (both genetic and epigenetic) play a major role in sex differences in lifespan; (ii) “variable” genetic and epigenetic patterns, that vary according to context, populations and exposures to different environments, can affect the magnitude of the gap in sex differences in survival. Then we will describe recent findings in the use of epigenetic clocks to uncover sex differences in biological age and thus potentially in mortality. In conclusion, we will discuss how environmental factors cannot be kept apart from the biological factors providing evidence from the field of human ecology.

Abstract Transcriptomics in Parkinson’s disease (PD) offers new insights into the molecular mechanism of PD pathogenesis. Several pathways, such as inflammation and protein degradation, have been identified by differential gene expression analysis. Our aim was to identify gene expression differences underlying the disease etiology and the discovery of pre-symptomatic risk biomarkers for PD from a multicenter study in the context of the PROPAG-AGEING project. We performed RNA sequencing from 47 patients with de novo PD, 10 centenarians, and 65 healthy controls. Using identified differentially expressed genes, functional annotations were assigned using gene ontology to unveil significant enriched biological processes. The expression of 16 selected genes was validated using OpenArray® assays and samples from independent cohorts of 201 patients with advanced PD, 340 healthy siblings of PD patients, and 177 healthy controls. Differential gene expression analysis identified higher FCGBP expression in patients with de novo PD compared with healthy controls and compared with centenarians. Furthermore, FCGBP showed no differences in terms of population origin or aging process. The increased FCGBP expression was validated in patients with advanced PD and their siblings. Thus, we provided evidence for an upregulation of FCGBP mRNA levels not only in patients with PD but also in individuals at putative higher risk of PD, suggesting that it could be important in gut–brain PD interaction, mediating the connection between microbiota and intestinal inflammatory processes, as well as neuroinflammation and neurodegeneration.

BackgroundThe aging process affects all systems of the human body, and the observed increase in inflammatory components affecting the immune system in old age can lead to the development of age-associated diseases and systemic inflammation.ResultsWe propose a small clock model SImAge based on a limited number of immunological biomarkers. To regress the chronological age from cytokine data, we first use a baseline Elastic Net model, gradient-boosted decision trees models, and several deep neural network architectures. For the full dataset of 46 immunological parameters, DANet, SAINT, FT-Transformer and TabNet models showed the best results for the test dataset. Dimensionality reduction of these models with SHAP values revealed the 10 most age-associated immunological parameters, taken to construct the SImAge small immunological clock. The best result of the SImAge model shown by the FT-Transformer deep neural network model has mean absolute error of 6.94 years and Pearson ρ = 0.939 on the independent test dataset. Explainable artificial intelligence methods allow for explaining the model solution for each individual participant.ConclusionsWe developed an approach to construct a model of immunological age based on just 10 immunological parameters, coined SImAge, for which the FT-Transformer deep neural network model had proved to be the best choice. The model shows competitive results compared to the published studies on immunological profiles, and takes a smaller number of features as an input. Neural network architectures outperformed gradient-boosted decision trees, and can be recommended in the further analysis of immunological profiles.

Summary: Background: People with Down syndrome (DS) show clinical signs of accelerated ageing. Causative mechanisms remain unknown and hypotheses range from the (essentially untreatable) amplified-chromosomal-instability explanation, to potential actions of individual supernumerary chromosome-21 genes. The latter explanation could open a route to therapeutic amelioration if the specific over-acting genes could be identified and their action toned-down. Methods: Biological age was estimated through patterns of sugar molecules attached to plasma immunoglobulin-G (IgG-glycans, an established “biological-ageing-clock”) in n = 246 individuals with DS from three European populations, clinically characterised for the presence of co-morbidities, and compared to n = 256 age-, sex- and demography-matched healthy controls. Isogenic human induced pluripotent stem cell (hiPSCs) models of full and partial trisomy-21 with CRISPR-Cas9 gene editing and two kinase inhibitors were studied prior and after differentiation to cerebral organoids. Findings: Biological age in adults with DS is (on average) 18.4–19.1 years older than in chronological-age-matched controls independent of co-morbidities, and this shift remains constant throughout lifespan. Changes are detectable from early childhood, and do not require a supernumerary chromosome, but are seen in segmental duplication of only 31 genes, along with increased DNA damage and decreased levels of LaminB1 in nucleated blood cells. We demonstrate that these cell-autonomous phenotypes can be gene-dose-modelled and pharmacologically corrected in hiPSCs and derived cerebral organoids. Using isogenic hiPSC models we show that chromosome-21 gene DYRK1A overdose is sufficient and necessary to cause excess unrepaired DNA damage. Interpretation: Explanation of hitherto observed accelerated ageing in DS as a developmental progeroid syndrome driven by DYRK1A overdose provides a target for early pharmacological preventative intervention strategies. Funding: Main funding came from the “Research Cooperability” Program of the Croatian Science Foundation funded by the European Union from the European Social Fund under the Operational Programme Efficient Human Resources 2014–2020, Project PZS-2019-02-4277, and the Wellcome Trust Grants 098330/Z/12/Z and 217199/Z/19/Z (UK). All other funding is described in details in the “Acknowledgements”.

Abstract In Italy, at least in the cohorts born up to the beginning of the twentieth century, women’s mortality in post-reproductive ages was influenced by fertility, with large progenies (and, to a lesser extent, childlessness) leading to markedly lower survival chances. This relationship proved strong enough to affect the female-to-male ratio in old age as fertility declined. In this paper, we show that various measures of extra female survival at high ages are closely connected to the fertility transition in Italy, and to its peculiar historical and geographical evolution.

Abstract Parkinson’s disease (PD) is the neurological disorder showing the greatest rise in prevalence from 1990 to 2016. Despite clinical definition criteria and a tremendous effort to develop objective biomarkers, precise diagnosis of PD is still unavailable at early stage. In recent years, an increasing number of studies have used omic methods to unveil the molecular basis of PD, providing a detailed characterization of potentially pathological alterations in various biological specimens. Metabolomics could provide useful insights to deepen our knowledge of PD aetiopathogenesis, to identify signatures that distinguish groups of patients and uncover responsive biomarkers of PD that may be significant in early detection and in tracking the disease progression and drug treatment efficacy. The present work is the first large metabolomic study based on nuclear magnetic resonance (NMR) with an independent validation cohort aiming at the serum characterization of de novo drug-naive PD patients. Here, NMR is applied to sera from large training and independent validation cohorts of German subjects. Multivariate and univariate approaches are used to infer metabolic differences that characterize the metabolite and the lipoprotein profiles of newly diagnosed de novo drug-naive PD patients also in relation to the biological sex of the subjects in the study, evidencing a more pronounced fingerprint of the pathology in male patients. The presence of a validation cohort allowed us to confirm altered levels of acetone and cholesterol in male PD patients. By comparing the metabolites and lipoproteins levels among de novo drug-naive PD patients, age- and sex-matched healthy controls, and a group of advanced PD patients, we detected several descriptors of stronger oxidative stress.

Abstract Advanced age represents one of the major risk factors for Parkinson’s Disease. Recent biomedical studies posit a role for microRNAs, also known to be remodelled during ageing. However, the relationship between microRNA remodelling and ageing in Parkinson’s Disease, has not been fully elucidated. Therefore, the aim of the present study is to unravel the relevance of microRNAs as biomarkers of Parkinson’s Disease within the ageing framework. We employed Next Generation Sequencing to profile serum microRNAs from samples informative for Parkinson’s Disease (recently diagnosed, drug-naïve) and healthy ageing (centenarians) plus healthy controls, age-matched with Parkinson’s Disease patients. Potential microRNA candidates markers, emerging from the combination of differential expression and network analyses, were further validated in an independent cohort including both drug-naïve and advanced Parkinson’s Disease patients, and healthy siblings of Parkinson’s Disease patients at higher genetic risk for developing the disease. While we did not find evidences of microRNAs co-regulated in Parkinson’s Disease and ageing, we report that hsa-miR-144-3p is consistently down-regulated in early Parkinson’s Disease patients. Moreover, interestingly, functional analysis revealed that hsa-miR-144-3p is involved in the regulation of coagulation, a process known to be altered in Parkinson’s Disease. Our results consistently show the down-regulation of hsa-mir144-3p in early Parkinson’s Disease, robustly confirmed across a variety of analytical and experimental analyses. These promising results ask for further research to unveil the functional details of the involvement of hsa-mir144-3p in Parkinson’s Disease.

IntroductionGrowth Differentiation Factor 15 (GDF15) is a mitochondrial-stress-responsive molecule whose expression strongly increases with aging and age-related diseases. However, its role in neurodegenerative diseases, including Alzheimer’s disease (AD), is still debated.MethodsWe have characterized the expression of GDF15 in brain samples from AD patients and non-demented subjects (controls) of different ages.ResultsAlthough no difference in CSF levels of GDF15 was found between AD patients and controls, GDF15 was expressed in different brain areas and seems to be predominantly localized in neurons. The ratio between its mature and precursor form was higher in the frontal cortex of AD patients compared to age-matched controls (p

Abstract A prodromal phase of Parkinson’s disease (PD) may precede motor manifestations by decades. PD patients’ siblings are at higher risk for PD, but the prevalence and distribution of prodromal symptoms are unknown. The study objectives were (1) to assess motor and non-motor features estimating prodromal PD probability in PD siblings recruited within the European PROPAG-AGEING project; (2) to compare motor and non-motor symptoms to the well-established DeNoPa cohort. 340 PD siblings from three sites (Bologna, Seville, Kassel/Goettingen) underwent clinical and neurological evaluations of PD markers. The German part of the cohort was compared with German de novo PD patients (dnPDs) and healthy controls (CTRs) from DeNoPa. Fifteen (4.4%) siblings presented with subtle signs of motor impairment, with MDS-UPDRS-III scores not clinically different from CTRs. Symptoms of orthostatic hypotension were present in 47 siblings (13.8%), no different to CTRs (p = 0.072). No differences were found for olfaction and overall cognition; German-siblings performed worse than CTRs in visuospatial-executive and language tasks. 3/147 siblings had video-polysomnography-confirmed REM sleep behavior disorder (RBD), none was positive on the RBD Screening Questionnaire. 173/300 siblings had

Abstract Background The assessment of muscle mass is a key determinant of the diagnosis of sarcopenia. We introduce for the first time an ultrasound imaging method for diagnosing sarcopenia based on changes in muscle geometric proportions. Methods Vastus lateralis muscle fascicle length (Lf) and thickness (Tm) were measured at 35% distal femur length by ultrasonography in a population of 279 individuals classified as moderately active elderly (MAE), sedentary elderly (SE) (n = 109), mobility impaired elderly (MIE) (n = 43), and in adult young controls (YC) (n = 60). The ratio of Lf/Tm was calculated to obtain an ultrasound index of the loss of muscle mass associated with sarcopenia (USI). In a subsample of elderly male individuals (n = 76) in which corresponding DXA measurements were available (MAE, n = 52 and SE, n = 24), DXA‐derived skeletal muscle index (SMI, appendicular limb mass/height2) was compared with corresponding USI values. Results For both young and older participants, USI values were found to be independent of sex, height and body mass. USI values were 3.70 ± 0.52 for YC, 4.50 ± 0.72 for the MAE, 5.05 ± 1.11 for the SE and 6.31 ± 1.38 for the MIE, all significantly different between each other (P 5.82 were classified as severely sarcopenic (prevalence 9.6%). The DXA‐derived SMI was found to be significantly correlated with USI (r = 0.61, P

An inadequate selenium (Se) status can accelerate the aging process, increasing the vulnerability to age-related diseases. The study aimed to investigate plasma Se and Se species in a large population, including 2200 older adults from the general population (RASIG), 514 nonagenarian offspring (GO), and 293 GO Spouses (SGO). Plasma Se levels in women exhibit an inverted U-shaped pattern, increasing with age until the post-menopausal period and then declining. Conversely, men exhibit a linear decline in plasma Se levels with age. Subjects from Finland had the highest plasma Se values, while those from Poland had the lowest ones. Plasma Se was influenced by fish and vitamin consumption, but there were no significant differences between RASIG, GO, and SGO. Plasma Se was positively associated with albumin, HDL, total cholesterol, fibrinogen, and triglycerides and negatively associated with homocysteine. Fractionation analysis showed that Se distribution among plasma selenoproteins is affected by age, glucometabolic and inflammatory factors, and being GO or SGO. These findings show that sex-specific, nutritional, and inflammatory factors play a crucial role in the regulation of Se plasma levels throughout the aging process and that the shared environment of GO and SGO plays a role in their distinctive Se fractionation.

The aim of the present study was to provide a comprehensive characterization of whole genome DNA methylation patterns in replicative and ionizing irradiation- or doxorubicin-induced premature senescence, exhaustively exploring epigenetic modifications in three different human cell types: in somatic diploid skin fibroblasts and in bone marrow- and adipose-derived mesenchymal stem cells. With CpG-wise differential analysis, three epigenetic signatures were identified: (a) cell type- and treatment-specific signature; (b) cell type-specific senescence-related signature; and (c) cell type-transversal replicative senescence-related signature. Cluster analysis revealed that only replicative senescent cells created a distinct group reflecting notable alterations in the DNA methylation patterns accompanying this cellular state. Replicative senescence-associated epigenetic changes seemed to be of such an extent that they surpassed interpersonal dissimilarities. Enrichment in pathways linked to the nervous system and involved in the neurological functions was shown after pathway analysis of genes involved in the cell type-transversal replicative senescence-related signature. Although DNA methylation clock analysis provided no statistically significant evidence on epigenetic age acceleration related to senescence, a persistent trend of increased biological age in replicative senescent cultures of all three cell types was observed. Overall, this work indicates the heterogeneity of senescent cells depending on the tissue of origin and the type of senescence inducer that could be putatively translated to a distinct impact on tissue homeostasis.

Abstract Background Myelofibrosis (MF) is a clonal disorder of hemopoietic stem/progenitor cells (HSPCs) with high prevalence in elderly patients and mutations in three driver genes (JAK2, MPL, or CALR). Around 10–15% of patients are triple-negative (TN) for the three driver mutations and display significantly worse survival. Circulating extracellular vesicles (EVs) play a role in intercellular signaling and are increased in inflammation and cancer. To identify a biomolecular signature of TN patients, we comparatively evaluated the circulating HSPCs and their functional interplay with the microenvironment focusing on EV analysis. Methods Peripheral blood was collected from MF patients (n = 29; JAK2 V617F mutation, n = 23; TN, n = 6) and healthy donors (HD, n = 10). Immunomagnetically isolated CD34+ cells were characterized by gene expression profiling analysis (GEP), survival, migration, and clonogenic ability. EVs were purified from platelet-poor plasma by ultracentrifugation, quantified using the Nanosight technology and phenotypically characterized by flow cytometry together with microRNA expression. Migration and survival of CD34+ cells from patients were also analyzed after in vitro treatments with selected inflammatory factors, i.e. (Interleukin (IL)-1β, Tumor Necrosis Factor (TNF)-α, IL6) or after co-culture with EVs from MF patients/HD. Results The absolute numbers of circulating CD34+ cells were massively increased in TN patients. We found that TN CD34+ cells show in vitro defective functions and are unresponsive to the inflammatory microenvironment. Of note, the plasma levels of crucial inflammatory cytokines are mostly within the normal range in TN patients. Compared to JAK2V617F-mutated patients, the GEP of TN CD34+ cells revealed distinct signatures in key pathways such as survival, cell adhesion, and inflammation. Importantly, we observed the presence of mitochondrial components within plasma EVs and a distinct phenotype in TN-derived EVs compared to the JAK2V617F-mutated MF patients and HD counterparts. Notably, TN EVs promoted the survival of TN CD34+ cells. Along with a specific microRNA signature, the circulating EVs from TN patients are enriched with miR-361-5p. Conclusions Distinct EV-driven signals from the microenvironment are capable to promote the TN malignant hemopoiesis and their further investigation paves the way toward novel therapeutic approaches for rare MF.

Understanding how to “Age Longer and Age Well” is a priority for people personally, for populations and for government policy. Approximately ten percent of nonagenarians reach 90 years and beyond in good condition and seem to have a combination of both age-span and health-span. However, the factors which contribute to human longevity remain challenging. Culture is a shared system of learning ideas, feelings, and survival strategies. It has a strong influence on each person’s psychological development, behavior, values and beliefs. Nonagenarians have rich life experiences that can teach us much about aging well; they are rich reservoirs of genetic, lifestyle and psychological information which can help understanding about how to live longer and better. Sibling or trio nonagenarians are important sources of family beliefs and behaviors upon which individual personalities may have been built. Their personal family histories and narratives are powerful tools that help to determine familial traits, beliefs and social behaviors which may help establish factors important in the siblings’ longevity. Using purposefully selected subjects, recruited to the Genetics of Healthy Ageing (GeHA) project in four European countries, this research used the simple life story and qualitative research methods to analyze contrasting and distinctive questions about the interface between the psychological and social worlds as presented in the nonagenarian siblings’ insights about their longevity. Their stories aimed to give better understanding about which psychological aspects of their common life journey and the degree of emotional support in their sibling relationships may have supported their paths to longevity. The most universal finding in each of the four European countries was that nonagenarians demonstrated high positivity, resilience and coping skills and were supported in social networks. Around this theme, nonagenarians reported “being happy,” “always cheerful,” “never melancholy” and having a contentment with a “rich life” and family relationships which fits with accumulating evidence that life satisfaction comes from a perceived self-efficacy and optimism. Most sibling relationships in this study, when analyzed according to the Gold classification, fit the “congenial” or “loyal” relationship type – demonstrating a healthy respect for the others’ opinion without overt dependence, which may help individual coping and survival mechanisms.

Control of ribosome biogenesis is a critical aspect of the regulation of cell metabolism. As ribosomal genes (rDNA) are organized in repeated clusters on chromosomes 13, 14, 15, 21, and 22, trisomy of chromosome 21 confers an excess of rDNA copies to persons with Down syndrome (DS). Previous studies showed an alteration of ribosome biogenesis in children with DS, but the epigenetic regulation of rDNA genes has not been investigated in adults with DS so far. In this study, we used a targeted deep-sequencing approach to measure DNA methylation (DNAm) of rDNA units in whole blood from 69 adults with DS and 95 euploid controls. We further evaluated the expression of the precursor of ribosomal RNAs (RNA45S) in peripheral blood mononuclear cells (PBMCs) from the same subjects. We found that the rDNA promoter tends to be hypermethylated in DS concerning the control group. The analysis of epihaplotypes (the combination of methylated and unmethylated CpG sites along the same DNA molecule) showed a significantly lower intra-individual diversity in the DS group, which at the same time was characterized by a higher interindividual variability. Finally, we showed that RNA45S expression is lower in adults with DS. Collectively, our results suggest a rearrangement of the epigenetic profile of rDNA in DS, possibly to compensate for the extranumerary rDNA copies. Future studies should assess whether the regulation of ribosome biogenesis can contribute to the pathogenesis of DS and explain the clinical heterogeneity characteristic of the syndrome.

Summary: Biological age (BA) has been proposed to evaluate the aging status instead of chronological age (CA). Our study shows evidence that there might be multiple “clocks” within the whole-body system: systemic aging drivers/clocks overlaid with organ/tissue-specific counterparts. We utilize multi-omics data, including clinical tests, immune repertoire, targeted metabolomic molecules, gut microbiomes, physical fitness examinations, and facial skin examinations, to estimate the BA of different organs (e.g., liver, kidney) and systems (immune and metabolic system). The aging rates of organs/systems are diverse. People’s aging patterns are different. We also demonstrate several applications of organs/systems BA in two independent datasets. Mortality predictions are compared among organs' BA in the dataset of the United States National Health and Nutrition Examination Survey. Polygenic risk score of BAs constructed in the Chinese Longitudinal Healthy Longevity Survey cohort can predict the possibility of becoming centenarian.

Abstract A central clinical question as the world deals with the COVID-19 pandemic is what the long-term sequelae for the millions of individuals will be who recover from the hyperinflammatory state characterizing COVID-19 and in particular for the hundreds of thousands who are ill enough to need hospitalization and in particular ICU care. Even when the pandemic is finally controlled, will COVID-19 survivors face exaggerated internal inflammatory processes, worsening co-morbidities, and increased susceptibility to age-related diseases? Clues for what may happen in post-COVID-19 patients can be elicited from those who recovered from other conditions that lead to similar hyperinflammatory states such as Severe Acute Respiratory Syndrome (SARS), acute respiratory disease syndrome (ARDS), cytokine storm syndrome, and post-ICU syndrome. The short-and long-term sequalae following recovery from each of these conditions suggests that these syndromes lead to an accelerated state of chronic subclinical systemic inflammation often seen in aging (termed inflammaging) resulting in increased and worsening age-related conditions including frailty even in younger individuals.

Abstract Background The cline of human genetic diversity observable across Europe is recapitulated at a micro-geographic scale by variation within the Italian population. Besides resulting from extensive gene flow, this might be ascribable also to local adaptations to diverse ecological contexts evolved by people who anciently spread along the Italian Peninsula. Dissecting the evolutionary history of the ancestors of present-day Italians may thus improve the understanding of demographic and biological processes that contributed to shape the gene pool of European populations. However, previous SNP array-based studies failed to investigate the full spectrum of Italian variation, generally neglecting low-frequency genetic variants and examining a limited set of small effect size alleles, which may represent important determinants of population structure and complex adaptive traits. To overcome these issues, we analyzed 38 high-coverage whole-genome sequences representative of population clusters at the opposite ends of the cline of Italian variation, along with a large panel of modern and ancient Euro-Mediterranean genomes. Results We provided evidence for the early divergence of Italian groups dating back to the Late Glacial and for Neolithic and distinct Bronze Age migrations having further differentiated their gene pools. We inferred adaptive evolution at insulin-related loci in people from Italian regions with a temperate climate, while possible adaptations to pathogens and ultraviolet radiation were observed in Mediterranean Italians. Some of these adaptive events may also have secondarily modulated population disease or longevity predisposition. Conclusions We disentangled the contribution of multiple migratory and adaptive events in shaping the heterogeneous Italian genomic background, which exemplify population dynamics and gene-environment interactions that played significant roles also in the formation of the Continental and Southern European genomic landscapes.

The requirement of blood-circulating sensitive biomarkers for monitoring liver transplant (LT) is currently a necessary step aiming at the reduction of standard invasive protocols, such as liver biopsy. In this respect, the main objective of this study is to assess circulating microRNA (c-miR) changes in recipients’ blood before and after LT and to correlate their blood levels with gold standard biomarkers and with outcomes such as rejection or complications after graft. An miR profile was initially performed; then, the most deregulated miRs were validated by RT-qPCR in 14 recipients pre- and post-LT and compared to a control group of 24 nontransplanted healthy subjects. MiR-122-5p, miR-92a-3p, miR-18a-5p, and miR-30c-5p, identified in the validation phase, were also analyzed considering an additional 19 serum samples collected from LT recipients and focusing on different follow-up (FU) times. The results showed significant, FU-related changes in c-miRs. In particular, miR-122-5p, miR-92a-3p, and miR-18a-5p revealed the same trend after transplantation and an increase in their level was found in patients with complications, independently from FU times. Conversely, the variations in the standard haemato-biochemical parameters for liver function assessment were not significant in the same FU period, confirming the importance of c-miRs as potential noninvasive biomarkers for monitoring patients’ outcomes.

Cytokines have been investigated extensively in elderly people, with conflicting results. We performed a comprehensive analysis of the plasma levels of 62 cytokines and growth factors involved in the regulation of the immune system, in healthy centenarians, and middle-aged controls. We confirmed the previously observed increase in the levels of several pro-inflammatory cytokines, such as TNF-α and IL-6, and found that several other cytokines, directly or indirectly involved in inflammation (such as IFN-α, IL-23, CCL-5), were present at higher levels in centenarians. We did not observe any increase in the levels of anti-inflammatory cytokines, with the notable exception of the Th2-shifting cytokine IL-19. No relevant difference was observed in cytokines regulating T cell immunity. Several growth factors having a role in regulating immunity, such as G-CSF, GM-CSF, EGF, and VEGF, were upregulated in centenarians, too. Principal component analysis of the cytokine dataset showed that pro and anti-inflammatory cytokines were the variables that contributed the most to the variability of the data we observed.

In the present paper, we have analysed the role of age and sex in the fatal outcome of COVID-19, as there are conflicting results in the literature. As such, we have answered three controversial questions regarding this aspect of the COVID-19 pandemic: (1) Have women been more resilient than men? (2) Did centenarians die less than the remaining older people? (3) Were older centenarians more resistant to SARS-CoV-2 than younger centenarians? The literature review demonstrated that: (1) it is women who are more resilient, in agreement with data showing that women live longer than men even during severe famines and epidemics; however, there are conflicting data regarding centenarian men; (2) centenarians overall did not die less than remaining older people, likely linked to their frailty; (3) in the first pandemic wave of 2020, centenarians > 101 years old (i.e., born before 1919), but not “younger centenarians”, have been more resilient to COVID-19 and this may be related to the 1918 Spanish flu epidemic, although it is unclear what the mechanisms might be involved.

Epigenetic clocks were initially developed to track chronological age, but accumulating evidence indicates that they can also predict biological age. They are usually based on the analysis of DNA methylation by genome-wide methods, but targeted approaches, based on the assessment of a small number of CpG sites, are advisable in several settings. In this study, we developed a targeted epigenetic clock purposely optimized for the measurement of biological age. The clock includes six genomic regions mapping in ELOVL2, NHLRC1, AIM2, EDARADD, SIRT7 and TFAP2E genes, selected from a re-analysis of existing microarray data, whose DNA methylation is measured by EpiTYPER assay. In healthy subjects (n = 278), epigenetic age calculated using the targeted clock was highly correlated with chronological age (Spearman correlation = 0.89). Most importantly, and in agreement with previous results from genome-wide clocks, epigenetic age was significantly higher and lower than expected in models of increased (persons with Down syndrome, n = 62) and decreased (centenarians, n = 106; centenarians’ offspring, n = 143; nutritional intervention in elderly, n = 233) biological age, respectively. These results support the potential of our targeted epigenetic clock as a new marker of biological age and open its evaluation in large cohorts to further promote the assessment of biological age in healthcare practice.

We performed a genome-wide association study (GWAS) of human extreme longevity (EL), defined as surviving past the 99th survival percentile, by aggregating data from four centenarian studies. The combined data included 2304 EL cases and 5879 controls. The analysis identified a locus in CDKN2B-AS1 (rs6475609, p = 7.13 × 10−8) that almost reached genome-wide significance and four additional loci that were suggestively significant. Among these, a novel rare variant (rs145265196) on chromosome 11 had much higher longevity allele frequencies in cases of Ashkenazi Jewish and Southern Italian ancestry compared to cases of other European ancestries. We also correlated EL-associated SNPs with serum proteins to link our findings to potential biological mechanisms that may be related to EL and are under genetic regulation. The findings from the proteomic analyses suggested that longevity-promoting alleles of significant genetic variants either provided EL cases with more youthful molecular profiles compared to controls or provided some form of protection from other illnesses, such as Alzheimer’s disease, and disease progressions.

The presence of circulating microbiome in blood has been reported in both physiological and pathological conditions, although its origins, identities and function remain to be elucidated. This study aimed to investigate the presence of blood microbiome by quantitative real-time PCRs targeting the 16S rRNA gene. To our knowledge, this is the first study in which the circulating microbiome has been analyzed in such a large sample of individuals since the study was carried out on 1285 Randomly recruited Age-Stratified Individuals from the General population (RASIG). The samples came from several different European countries recruited within the EU Project MARK-AGE in which a series of clinical biochemical parameters were determined. The results obtained reveal an association between microbial DNA copy number and geographic origin. By contrast, no gender and age-related difference emerged, thus demonstrating the role of the environment in influencing the above levels independent of age and gender at least until the age of 75. In addition, a significant positive association was found with Free Fatty Acids (FFA) levels, leukocyte count, insulin, and glucose levels. Since these factors play an essential role in both health and disease conditions, their association with the extent of the blood microbiome leads us to consider the blood microbiome as a potential biomarker of human health.

Genome-wide association studies have only revealed a handful of genetic loci for longevity. Here, in a case–control design based on phenotype definitions of individuals surviving at or beyond the age corresponding to the 90th and 99th survival percentile, the authors report two additional loci located in the APOE locus and near GPR78.

Abstract Background Here, we isolated, expanded and functionally characterized regulatory T cells (Tregs) from patients with end stage kidney and liver disease, waiting for kidney/liver transplantation (KT/LT), with the aim to establish a suitable method to obtain large numbers of immunomodulatory cells for adoptive immunotherapy post-transplantation. Methods We first established a preclinical protocol for expansion/isolation of Tregs from peripheral blood of LT/KT patients. We then scaled up and optimized such protocol according to good manufacturing practice (GMP) to obtain high numbers of purified Tregs which were phenotypically and functionally characterized in vitro and in vivo in a xenogeneic acute graft-versus-host disease (aGVHD) mouse model. Specifically, immunodepressed mice (NOD-SCID-gamma KO mice) received human effector T cells with or without GMP-produced Tregs to prevent the onset of xenogeneic GVHD. Results Our small scale Treg isolation/expansion protocol generated functional Tregs. Interestingly, cryopreservation/thawing did not impair phenotype/function and DNA methylation pattern of FOXP3 gene of the expanded Tregs. Fully functional Tregs were also isolated/expanded from KT and LT patients according to GMP. In the mouse model, GMP Tregs from LT or KT patient proved to be safe and show a trend toward reduced lethality of acute GVHD. Conclusions These data demonstrate that expanded/thawed GMP-Tregs from patients with end-stage organ disease are fully functional in vitro. Moreover, their infusion is safe and results in a trend toward reduced lethality of acute GVHD in vivo, further supporting Tregs-based adoptive immunotherapy in solid organ transplantation.

Abstract Background Perilipin2 (Plin2) belongs to a family of five highly conserved proteins, known for their role in lipid storage. Recent data indicate that Plin2 has an important function in cell metabolism and is involved in several human pathologies, including liver steatosis and Type II diabetes. An association between Plin2 and lower muscle mass and strength has been found in elderly and inactive people, but its function in skeletal muscle is still unclear. Here, we addressed the role of Plin2 in adult muscle by gain and loss of function experiments. Methods By mean of in vivo Plin2 down‐regulation (shPlin2) and overexpression (overPlin2) in murine tibialis anterior muscle, we analysed the effects of Plin2 genetic manipulations on myofiber size and lipid composition. An analysis of skeletal muscle lipid composition was also performed in vastus lateralis samples from young and old patients undergoing hip surgery. Results We found that Plin2 down‐regulation was sufficient to induce a 30% increase of myofiber cross‐sectional area, independently of mTOR pathway. Alterations of lipid content and modulation of genes involved in lipid synthesis occurred in hypertrophic muscles. In particular, we showed a decrease of triglycerides, ceramides, and phosphatidylcoline:phosphatidylethanolamine ratio, a condition known to impact negatively on muscle function. Plin2 overexpression did not change fibre size; however, lipid composition was strongly affected in a way that is similar to that observed in human samples from old patients. Conclusions Altogether these data indicate that Plin2 is a critical mediator for the control of muscle mass, likely, but maybe not exclusively, through its critical role in the regulation of intracellular lipid content and composition.

Extreme longevity is the paradigm of healthy aging as individuals who reached the extreme decades of human life avoided or largely postponed all major age-related diseases. In this study, we sequenced at high coverage (90X) the whole genome of 81 semi-supercentenarians and supercentenarians [105+/110+] (mean age: 106.6 ± 1.6) and of 36 healthy unrelated geographically matched controls (mean age 68.0 ± 5.9) recruited in Italy. The results showed that 105+/110+ are characterized by a peculiar genetic background associated with efficient DNA repair mechanisms, as evidenced by both germline data (common and rare variants) and somatic mutations patterns (lower mutation load if compared to younger healthy controls). Results were replicated in a second independent cohort of 333 Italian centenarians and 358 geographically matched controls. The genetics of 105+/110+ identified DNA repair and clonal haematopoiesis as crucial players for healthy aging and for the protection from cardiovascular events.

Alzheimer's disease (AD) is characterized by specific alterations of brain DNA methylation (DNAm) patterns. Age and sex, two major risk factors for AD, are also known to largely affect the epigenetic profiles in brain, but their contribution to AD-associated DNAm changes has been poorly investigated. In this study we considered publicly available DNAm datasets of four brain regions (temporal, frontal, entorhinal cortex, and cerebellum) from healthy adult subjects and AD patients, and performed a meta-analysis to identify sex-, age-, and AD-associated epigenetic profiles. In one of these datasets it was also possible to distinguish 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) profiles. We showed that DNAm differences between males and females tend to be shared between the four brain regions, while aging differently affects cortical regions compared to cerebellum. We found that the proportion of sex-dependent probes whose methylation is modified also during aging is higher than expected, but that differences between males and females tend to be maintained, with only a few probes showing age-by-sex interaction. We did not find significant overlaps between AD- and sex-associated probes, nor disease-by-sex interaction effects. On the contrary, we found that AD-related epigenetic modifications are significantly enriched in probes whose DNAm varies with age and that there is a high concordance between the direction of changes (hyper or hypo-methylation) in aging and AD, supporting accelerated epigenetic aging in the disease. In summary, our results suggest that age-associated DNAm patterns concur to the epigenetic deregulation observed in AD, providing new insights on how advanced age enables neurodegeneration.

Aging is accompanied by physiological changes affecting body composition and functionality, including accumulation of fat mass at the expense of muscle mass, with effects upon morbidity and quality of life. The gut microbiome has recently emerged as a key environmental modifier of human health that can modulate healthy aging and possibly longevity. However, its associations with adiposity in old age are still poorly understood. Here we profiled the gut microbiota in a well-characterized cohort of 201 Italian elderly subjects from the NU-AGE study, by 16S rRNA amplicon sequencing. We then tested for association with body composition from dual-energy X-ray absorptiometry (DXA), with a focus on visceral and subcutaneous adipose tissue. Dietary patterns, serum metabolome and other health-related parameters were also assessed. This study identified distinct compositional structures of the elderly gut microbiota associated with DXA parameters, diet, metabolic profiles and cardio-metabolic risk factors.

Organismal ageing is associated with many physiological changes, including differences in the immune system of most animals. These differences are often considered to be a key cause of age-associated diseases as well as decreased vaccine responses in humans. The most often cited vaccine failure is seasonal influenza, but, while it is usually the case that the efficiency of this vaccine is lower in older than younger adults, this is not always true, and the reasons for the differential responses are manifold. Undoubtedly, changes in the innate and adaptive immune response with ageing are associated with failure to respond to the influenza vaccine, but the cause is unclear. Moreover, recent advances in vaccine formulations and adjuvants, as well as in our understanding of immune changes with ageing, have contributed to the development of vaccines, such as those against herpes zoster and SARS-CoV-2, that can protect against serious disease in older adults just as well as in younger people. In the present article, we discuss the reasons why it is a myth that vaccines inevitably protect less well in older individuals, and that vaccines represent one of the most powerful means to protect the health and ensure the quality of life of older adults.

Immunosenescence is marked by a systemic process named inflammaging along with a series of defects in the immunological activity that results in poor responses to infectious agents and to vaccination. Inflammaging, a state of low-grade chronic inflammation, usually leads to chronic inflammatory diseases and frailty in the elderly. However, some elderly escape from frailty and reach advanced age free of the consequences of inflammaging. This process has been called immunological remodeling, and it is the hallmark of healthy aging as described in the studies of centenarians in Italy. The biological markers of healthy aging are still a matter of debate, and the studies on the topic have focused on inflammatory versus remodeling processes and molecules. The sub-clinical inflammatory status associated with aging might be a deleterious event for populations living in countries where chronic infectious diseases are not prevalent. Nevertheless, in other parts of the world where they are, two possibilities may occur. Inflammatory responses may have a protective effect against these infectious agents. At the same time, the long-term consequences of protective immune responses during chronic infections may result in accelerated immunosenescence in these individuals. Therefore, the biological markers of healthy aging can vary according to environmental, cultural, and geographical settings that reflect worldwide, and in a non-biased, non-westernized perspective, the changes that we experience regarding our contacts with microorganisms and the outcomes of such contacts.

Cold climates represent one of the major environmental challenges that anatomically modern humans faced during their dispersal out of Africa. The related adaptive traits have been achieved by modulation of thermogenesis and thermoregulation processes where nuclear (nuc) and mitochondrial (mt) genes play a major role. In human populations, mitonuclear genetic interactions are the result of both the peculiar genetic history of each human group and the different environments they have long occupied. This study aims to investigate mitonuclear genetic interactions by considering all the mitochondrial genes and 28 nuclear genes involved in brown adipose tissue metabolism, which have been previously hypothesized to be crucial for cold adaptation. For this purpose, we focused on three human populations (i.e., Finnish, British, and Central Italian people) of European ancestry from different biogeographical and climatic areas, and we used a machine learning approach to identify relevant nucDNA–mtDNA interactions that characterized each population. The obtained results are twofold: (i) at the methodological level, we demonstrated that a machine learning approach is able to detect patterns of genetic structure among human groups from different latitudes both at single genes and by considering combinations of mtDNA and nucDNA loci; (ii) at the biological level, the analysis identified population-specific nuclear genes and variants that likely play a relevant biological role in association with a mitochondrial gene (such as the “obesity gene” FTO in Finnish people). Further studies are needed to fully elucidate the evolutionary dynamics (e.g., migration, admixture, and/or local adaptation) that shaped these nucDNA–mtDNA interactions and their functional role.

Growth differentiation factor 15 (GDF15) is a stress molecule produced in response to mitochondrial, metabolic and inflammatory stress with a number of beneficial effects on metabolism. However, at the level of skeletal muscle it is still unclear whether GDF15 is beneficial or detrimental. The aim of the study was to analyse the levels of circulating GDF15 in people of different age, characterized by different level of physical activity and to seek for correlation with hematological parameters related to inflammation. The plasma concentration of GDF15 was determined in a total of 228 subjects in the age range from 18 to 83 years. These subjects were recruited and divided into three different groups based on the level of physical activity: inactive patients with lower limb mobility impairment, active subjects represented by amateur endurance cyclists, and healthy controls taken from the general population. Cyclists were sampled before and after a strenuous physical bout (long distance cycling race). The plasma levels of GDF15 increase with age and are inversely associated with active lifestyle. In particular, at any age, circulating GDF15 is significantly higher in inactive patients and significantly lower in active people, such as cyclists before the race, with respect to control subjects. However, the strenuous physical exercise causes in cyclists a dramatic increase of GDF15 plasma levels, that after the race are similar to that of patients. Moreover, GDF15 plasma levels significantly correlate with quadriceps torque in patients and with the number of total leukocytes, neutrophils and lymphocytes in both cyclists (before and after race) and patients. Taken together, our data indicate that GDF15 is associated with decreased muscle performance and increased inflammation.

Biological aging is a complex process involving multiple biological processes. These can be understood theoretically though considering them as individual networks—e.g., epigenetic networks, cell-cell networks (such as astroglial networks), and population genetics. Mathematical modeling allows the combination of such networks so that they may be studied in unison, to better understand how the so-called “seven pillars of aging” combine and to generate hypothesis for treating aging as a condition at relatively early biological ages. In this review, we consider how recent progression in mathematical modeling can be utilized to investigate aging, particularly in, but not exclusive to, the context of degenerative neuronal disease. We also consider how the latest techniques for generating biomarker models for disease prediction, such as longitudinal analysis and parenclitic analysis can be applied to as both biomarker platforms for aging, as well as to better understand the inescapable condition. This review is written by a highly diverse and multi-disciplinary team of scientists from across the globe and calls for greater collaboration between diverse fields of research.

ABSTRACT The gut microbiome of long-lived people display an increasing abundance of subdominant species, as well as a rearrangement in health-associated bacteria, but less is known about microbiome functions. In order to disentangle the contribution of the gut microbiome to the complex trait of human longevity, we here describe the metagenomic change of the human gut microbiome along with aging in subjects with up to extreme longevity, including centenarians (aged 99 to 104 years) and semisupercentenarians (aged 105 to 109 years), i.e., demographically very uncommon subjects who reach the extreme limit of the human life span. According to our findings, the gut microbiome of centenarians and semisupercentenarians is more suited for xenobiotic degradation and shows a rearrangement in metabolic pathways related to carbohydrate, amino acid, and lipid metabolism. Collectively, our data go beyond the relationship between intestinal bacteria and physiological changes that occur with aging by detailing the shifts in the potential metagenomic functions of the gut microbiome of centenarians and semisupercentenarians as a response to progressive dietary and lifestyle modifications. IMPORTANCE The study of longevity may help us understand how human beings can delay or survive the most frequent age-related diseases and morbidities. In this scenario, the gut microbiome has been proposed as one of the variables to monitor and possibly support healthy aging. Indeed, the disruption of host-gut microbiome homeostasis has been associated with inflammation and intestinal permeability as well as a general decline in bone and cognitive health. Here, we performed a metagenomic assessment of fecal samples from semisupercentenarians, i.e., 105 to 109 years old, in comparison to young adults, the elderly, and centenarians, shedding light on the longest compositional and functional trajectory of the human gut microbiome with aging. In addition to providing a fine taxonomic resolution down to the species level, our study emphasizes the progressive age-related increase in degradation pathways of pervasive xenobiotics in Western societies, possibly as a result of a supportive process within the molecular continuum characterizing aging.

Hypertension is a major global health challenge, as it represents the main risk factor for stroke and cardiovascular disease. It is a multifactorial clinical condition characterized by high and sustained levels of blood pressure, likely resulting from a complex interplay of endogenous and environmental factors. The gut microbiota has been strongly supposed to be involved but its role in hypertension is still poorly understood. In an attempt to fill this gap, here we characterized the microbial composition of fecal samples from 48 hypertensive and 32 normotensive Brazilian individuals by next-generation sequencing of the 16S rRNA gene. In addition, the cytokine production of peripheral blood samples was investigated to build an immunological profile of these individuals. We identified a dysbiosis of the intestinal microbiota in hypertensive subjects, featured by reduced biodiversity and distinct bacterial signatures compared with the normotensive counterpart. Along with a reduction in Bacteroidetes members, hypertensive individuals were indeed mainly characterized by increased proportions of Lactobacillus and Akkermansia while decreased relative abundances of well-known butyrate-producing commensals, including Roseburia and Faecalibacterium within the Lachnospiraceae and Ruminococcaceae families. We also observed an inflamed immune profile in hypertensive individuals with an increase in TNF/IFN-γ ratio, and in TNF and IL-6 production when compared to normotensive ones. Our work provides the first evidence of association of hypertension with altered gut microbiota and inflammation in a Brazilian population. While lending support to the existence of potential microbial signatures of hypertension, likely to be robust to age and geography, our findings point to largely neglected bacteria as potential contributors to intestinal homeostasis loss and emphasize the high vulnerability of hypertensive individuals to inflammation-related disorders.

In this work we propose an index to estimate the gut microbiota biodiversity using a modeling approach with the aim of describing its relationship with health and aging. The gut microbiota, a complex ecosystem that links nutrition and metabolism, has a pervasive effect on all body organs and systems, undergoes profound changes with age and life-style, and substantially contributes to the pathogenesis of age-related diseases. For these reasons, the gut microbiota is a suitable candidate for assessing and quantifying healthy aging, i.e. the capability of individuals to reach an advanced age, avoiding or postponing major age-related diseases. The importance of the gut microbiota in health and aging has been proven to be related not only to its taxonomic composition, but also to its ecological properties, namely its biodiversity. Following an ecological approach, here we intended to characterize the relationship between the gut microbiota biodiversity and healthy aging through the development a parsimonious model of gut microbiota from which biodiversity can be estimated. We analysed publicly available metagenomic data relative to subjects of different ages, countries, nutritional habits and health status and we showed that a hybrid niche-neutral model well describes the observed patterns of bacterial relative abundance. Moreover, starting from such ecological modeling, we derived an estimate of the gut microbiota biodiversity that is consistent with classical indices, while having a higher statistical power. This allowed us to unveil an increase of the gut microbiota biodiversity during aging and to provide a good predictor of health status in old age, dependent on life-style and aging disorders.

The role of epigenetics in endothelial cell senescence is a cutting-edge topic in ageing research. However, little is known of the relative contribution to pro-senescence signal propagation provided by microRNAs shuttled by extracellular vesicles (EVs) released from senescent cells. Analysis of microRNA and DNA methylation profiles in non-senescent (control) and senescent (SEN) human umbilical vein endothelial cells (HUVECs), and microRNA profiling of their cognate small EVs (sEVs) and large EVs demonstrated that SEN cells released a significantly greater sEV number than control cells. sEVs were enriched in miR-21-5p and miR-217, which target DNMT1 and SIRT1. Treatment of control cells with SEN sEVs induced a miR-21/miR-217-related impairment of DNMT1-SIRT1 expression, the reduction of proliferation markers, the acquisition of a senescent phenotype and a partial demethylation of the locus encoding for miR-21. MicroRNA profiling of sEVs from plasma of healthy subjects aged 40–100 years showed an inverse U-shaped age-related trend for miR-21-5p, consistent with senescence-associated biomarker profiles. Our findings suggest that miR-21-5p/miR-217 carried by SEN sEVs spread pro-senescence signals, affecting DNA methylation and cell replication.

The aging process is characterized by the chronic inflammatory status called “inflammaging”, which shares major molecular and cellular features with the metabolism-induced inflammation called “metaflammation.” Metaflammation is mainly driven by overnutrition and nutrient excess, but other contributing factors are metabolic modifications related to the specific body composition (BC) changes occurring with age. The aging process is indeed characterized by an increase in body total fat mass and a concomitant decrease in lean mass and bone density, that are independent from general and physiological fluctuations in weight and body mass index (BMI). Body adiposity is also re-distributed with age, resulting in a general increase in trunk fat (mainly abdominal fat) and a reduction in appendicular fat (mainly subcutaneous fat). Moreover, the accumulation of fat infiltration in organs such as liver and muscles also increases in elderly, while subcutaneous fat mass tends to decrease. These specific variations in BC are considered risk factors for the major age-related diseases, such as cardiovascular diseases, type 2 diabetes, sarcopenia and osteoporosis, and can predispose to disabilities. Thus, the maintenance of a balance rate of fat, muscle and bone is crucial to preserve metabolic homeostasis and a health status, positively contributing to a successful aging. For this reason, a detailed assessment of BC in elderly is critical and could be an additional preventive personalized strategy for age-related diseases. Despite BMI and other clinical measures, such as waist circumference measurement, waist-hip ratio, underwater weighing and bioelectrical impedance, are widely used as a surrogate measure for body adiposity, they barely reflect the distribution of body fat. Because of the great advantages offered by imaging tools in research and clinics, the attention of clinicians is now moving to powerful imaging techniques such as computed tomography, magnetic resonance imaging, dual-energy X-ray absorptiometry and ultrasound to obtain a more accurate estimation of BC. The aim of this review is to present the state of the art of the imaging techniques that are currently available to measure BC and that can be applied to the study of BC changes in the elderly, outlining advantages and disadvantages of each technique.