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Amyloid fibrils and fibril-like structures are currently estimated to represent many different products of several genes in humans and play a key role in many types of proteinopathies, commonly associated with ageing process. They share the mutual feature of aggregation-prone proteins and the building up of molecular-supramolecular structure, such as inter-neuronal plaques in the brain of Alzheimer's Disease (AD) patients, characterized by an extraordinary strength. Noteworthy, this type of structure has been reported in different organisms, in particular in invertebrates. The aim of the current review is to focus on alpha and beta amyloids i.e., SAAs, SAP and APP, elucidating the structure and function of amyloid proteins in invertebrates (such as nematods, annelids, molluscs, insects, ascidians) and highlighting their striking pattern of functional conservation when compared to human amyloid-like fibrils, thus focusing on possible new studies and applications for innovative therapies, particularly for AD, the most common and worldwide type of dementia.

Atherosclerosis is considered a chronic inflammatory disease of arteries associated with the aging process. Many risk factors have been identified and they are mainly related to life-styles, gene-environment interactions and socioeconomic status. Carotid and coronary artery diseases are the two major atherosclerotic conditions, being the primary cause of stroke and heart attack, respectively. Nevertheless, carotid plaque assumes particular aspects not only for the specific molecular mechanisms, but also for the types of atheroma which may be associated with a better or a worst prognosis. The identification of circulating blood biomarkers able to distinguish carotid plaque types (stable or vulnerable) is a crucial step for the improvement of adequate therapeutic approaches avoiding or delaying endarterectomy in the oldest old individuals (> 80 years), a population predicted to growth in the next years. The review highlights the most recent knowledge on carotid plaque molecular mechanisms, focusing on microRNAs (miRs), as a site-specific accelerated aging within the conceptual framework of Geroscience for new affordable therapies.

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. publishedVersion

Blood circulating microRNAs (c‐miRs) are potential biomarkers to trace aging and longevity trajectories to identify molecular targets for anti‐aging therapies. Based on a cross‐sectional study, a discovery phase was performed on 12 donors divided into four groups: young, old, healthy, and unhealthy centenarians. The identification of healthy and unhealthy phenotype was based on cognitive performance and capabilities to perform daily activities. Small RNA sequencing identified 79 differentially expressed c‐miRs when comparing young, old, healthy centenarians, and unhealthy centenarians. Two miRs, that is, miR‐19a‐3p and miR‐19b‐3p, were found increased at old age but decreased at extreme age, as confirmed by RT‐qPCR in 49 donors of validation phase. The significant decrease of those miR levels in healthy compared to unhealthy centenarians appears to be due to the presence of isomiRs, not detectable with RT‐qPCR, but only with a high‐resolution technique such as deep sequencing. Bioinformatically, three main common targets of miR‐19a/b‐3p were identified, that is, SMAD4, PTEN, and BCL2L11, converging into the FoxO signaling pathway, known to have a significant role in aging mechanisms. For the first time, this study shows the age‐related increase of plasma miR‐19a/b‐3p in old subjects but a decrease in centenarians. This decrease is more pronounced in healthy centenarians and was confirmed by the modified pattern of isomiRs comparing healthy and unhealthy centenarians. Thus, our study paves the way for functional studies using c‐miRs and isomiRs as additional parameter to track the onset of aging and age‐related diseases using new potential biomarkers., Small RNA deep sequencing on plasma‐derived RNA from 12 donors (young and old subjects, healthy, and unhealthy centenarians) was performed: 79 differentially expressed c‐miRs were identified. MiR‐19a/b‐3p increased at old age but decreased at extreme age, as confirmed by RT‐qPCR in 49 subjects. The decrease was more relevant in healthy centenarians, also confirmed by the modified pattern of isomiRs. Three common targets of miR‐19a/b‐3p were identified converging into the FoxO signaling pathway.

Atherosclerosis may starts early in life and each artery has peculiar characteristics likely affecting atherogenesis. The primary objective of the work was to underpin the microRNA (miR)-profiling differences in human normal femoral, abdominal aortic, and carotid arteries. The secondary aim was to investigate if those identified miRs, differently expressed in normal conditions, may also have a role in atherosclerotic arteries at adult ages. MiR-profiles were performed on normal tissues, revealing that aorta and carotid arteries are more similar than femoral arteries. MiRs emerging from profiling comparisons, i.e., miR-155-5p, -27a-5p, and -139-5p, were subjected to validation by RT-qPCR in normal arteries and also in pathological/atheroma counterparts, considering all the available 20 artery specimens. The three miRs were confirmed to be differentially expressed in normal femoral vs aorta/carotid arteries. Differential expression of those miRs was also observed in atherosclerotic arteries, together with some miR-target proteins, such as vimentin, CD44, E-cadherin and an additional marker SLUG. The different expression of miRs and targets/markers suggests that aorta/carotid and femoral arteries differently activate molecular drivers of pathological condition, thus conditioning the morphology of atheroma in adult life and likely suggesting the future use of artery-specific treatment to counteract atherosclerosis.

A redox steady state is important in maintaining vital cellular functions and is therefore homeostatically controlled by a number of antioxidative agents, the most important of which are enzymes. Oxidative Stress (OS) is associated with (or/and caused by) excessive production of damaging reactive oxygen and/or nitrogen species (ROS, RNS), which play a role in many pathologies. Because OS is a risk factor for many diseases, much effort (and money) is devoted to early diagnosis and treatment of OS. The desired benefit of the “identify (OS) and treat (by low molecular weight antioxidants, LMWA)” approach is to enable selective treatment of patients under OS. The present work aims at gaining understanding of the benefit of the antioxidants based on interrelationship between the concentration of different OS biomarkers and LMWA. Both the concentrations of a variety of biomarkers and of LMWA were previously determined and some analyses have been published by the MARK-AGE team. For the sake of simplicity, we assume that the concentration of an OS biomarker is a linear function of the concentration of a LMWA (if the association is due to causal relationship). A negative slope of this dependence (and sign of the correlation coefficient) can be intuitively expected for an antioxidant, a positive slope indicates that the LMWA is pro-oxidative, whereas extrapolation of the OS biomarker to [LMWA] = 0 is an approximation of the concentration of the OS biomarker in the absence of the LMWA. Using this strategy, we studied the effects of 12 LMWA (including tocopherols, carotenoids and ascorbic acid) on the OS status, as observed with 8 biomarkers of oxidative damage (including malondialdehyde, protein carbonyls, 3-nitrotyrosine). The results of this communication show that in a cross-sectional study the LMWA contribute little to the redox state and that different “antioxidants” are very different, so that single LMWA treatment of OS is not scientifically justified assuming our simple model. In view of the difficulty of quantitating the OS and the very different effects of various LMWA, the use of the “identify and treat” approach is questionable.

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., 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.

Tissue-specific effects of 17 beta-estradiol are delivered via both estrogen receptors and microRNAs (miRs). Menopause is known to affect the whole-body fat distribution in women. This investigation aimed at identifying menopause-and hormone replacement therapy (HRT)-associated miR profiles and miR targets in subcutaneous abdominal adipose tissue and serum from the same women. A discovery phase using array technology was performed in 13 women, including monozygotic twin pairs discordant for HRT and premenopausal young controls. Seven miRs, expressed in both adipose tissue and serum, were selected for validation phase in 34 women from a different cohort. An age/menopause-related increase of miRs-16-5p, -451a, -223-3p, -18a-5p, -19a-3p,-486-5p and -363-3p was found in the adipose tissue, but not in serum. MiR-19a-3p, involved in adipocyte development and estrogen signaling, resulted to be higher in HRT users in comparison with non-users. Among the identified targets, AKT1, BCL-2 and BRAF proteins showed lower expression in both HRT and No HRT users in comparison with premenopausal women. Unexpectedly, ESR1 protein expression was not modified although its mRNA was lower in No HRT users compared to premenopausal women and HRT users. Thus, both HRT and menopause appear to affect adipose tissue homeostasis via miR-mediated mechanism.

The gut microbiota (GM) is a complex, evolutionarily molded ecological system, which contributes to a variety of physiological functions. The GM is highly dynamic, being sensitive to environmental stimuli, and its composition changes over the host’s entire lifespan. However, the basic question of how much these changes may be ascribed to variables such as population, diet, genetics and gender, and/or to the aging process per se is still largely unanswered. We argue that comparison among studies on centenarians—the best model of healthy aging and longevity—recruited from different geographical areas/populations (different genetics and dietary habits) can help to disentangle the contribution of aging and non-aging-related variables to GM remodeling with age. The current review focuses on the role of population, gender and host genetics as possible drivers of GM modification along the human aging process. The feedback impact of age-associated GM variation on the GM–brain axis and GM metabolomics is also discussed. We likewise address the role of GM in neurodegenerative diseases such as Parkinson’s and Alzheimer’s, and its possible therapeutic use, taking advantage of the fact that centenarians are characterized by an extreme (healthy) phenotype versus patients suffering from age-related pathologies. Finally, it is argued that longitudinal studies combining metagenomics sequencing and in-depth phylogenetic analysis with a comprehensive phenotypic characterization of centenarians and patients using up-to-date omics (metabolomics, transcriptomics and meta-transcriptomics) are urgently needed.

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. Electronic supplementary material The online version of this article (10.1186/s12967-019-2004-2) contains supplementary material, which is available to authorized users.

Longevity has been described as the result of a complex combination of variables, hailing from genetics, lifestyle and environment. In this context, the intestinal microbiome has been proposed as a possible mediator of healthy aging that preserves host-environment homeostasis by counteracting inflammaging, intestinal permeability, and deterioration of cognitive and bone health. Correlations have been previously found between age-related gut microbiota dysbioses and levels of pro-inflammatory cytokines, hospitalization, poor diet and frailty in the elderly. More recently, the longest human gut microbiota trajectory along aging has been built, by comparing the fecal bacterial taxa from healthy adults and older individuals, also including semi-supercentenarians, i.e. people aged 105 or older. However, the functional changes that occur in the gut microbiome along with age are still largely unexplored. In an attempt to provide some glimpses in this direction and advance our knowledge on whether and how the gut microbiome may support the maintenance of health in extreme aging, here we characterized the fecal microbiome of 62 individuals, with age ranging from 22 to 105 years, by shotgun metagenomics. According to our findings, aging is characterized by an increased number of genes involved in xenobiotics degradation, as well as by rearrangements in metabolic pathways related to carbohydrate, amino acid and lipid metabolism. These microbiome features are even more boosted in semi-supercentenarians, probably representing the result of a life-long adaptive response to progressive changes in diet and lifestyle.

We propose in this review that hormesis, a concept profoundly and systematically addressed by Mark Mattson, has to be considered a sort of comprehensive "contact point" capable of unifying several conceptualizations of the aging process, including those focused on the stress response, oxidative stress and chronic inflammation/inflammaging. A major strength of hormesis and inflammaging is that they have a strong evolutionary basis. Moreover, both hormesis and inflammaging frame the aging process within a lifelong perspective of adaptation to different types of stresses. Such adaptation perspective also suggests that the aging process is malleable, and predicts that effective anti-aging strategies should mimic what evolution did in the course of million years and that we have to learn how to exploit the great potential inherent in the hormetic/inflammatory responses. To this regard, new topics such as the production of mitokines to cope with mitochondrial dysfunction are emerging as possible anti-aging target. This approach opens theoretically the door to the possibility of modulating the individual aging rate and trajectory by adopting the most effective scientifically-based lifestyle regarding fundamentally nutrition and physical activity. In this scenario Mark Mattson's lesson and personal example will permanently enlighten the aging field and the quest for a healthy aging and longevity.

Background and Aim: A state of chronic, subclinical inflammation known as inflammaging is present in elderly people and represents a risk factor for all age-related diseases. Dietary supplementation with ad hoc fortified foods seems an appealing strategy to counteract inflammaging. The purpose of this study was to test the efficacy of elderly-tailored fortified milk on inflammaging and different health parameters. Methods: A double-blind randomized cross-over study was performed on forty-eight volunteers aged 63&ndash, 80 years. The fortified milk was enriched with &omega, 3 polyunsaturated fatty acids (eicosapentaenoic acid, EPA, docosahexaenoic acid, DHA), vitamins (25-hydroxyvitamin D, E, C, B6, B9, B12), and trace elements (zinc, selenium). The two intervention periods lasted for 12 weeks, with a 16-week washout intermission. Results: Compared to placebo, the consumption of fortified milk increased the circulating levels of different micronutrients, including vitamins and the &omega, 3 index of erythrocyte membranes. Conversely, it reduced the amount of arachidonic acid, homocysteine, and &omega, 6/&omega, 3 ratio. Conclusion: Twelve-week daily consumption of adhoc fortified milk has an overall positive impact on different health parameters related to inflammaging in the elderly.

Methods for measuring diet composition and quantifying nutrient intake with sufficient validity are essential to study the association between nutrition and health outcomes and risk of diseases. 7-day food records provides a quantification of food actually and currently consumed and is interesting for its use in intervention studies to monitor diet in a short-term period and to guide participants toward changing their intakes. The objective of this study is to analyze the correlation/association between the daily intake of selected nutrients (collected by a 7-day food records plus a mineral/vitamin supplementation questionnaire) and estimates of energy expenditure as well as blood and urine biomarkers of dietary intakes in 1,140 healthy elderly subjects (65–79 years) at baseline of the NU-AGE intervention study (NCT01754012, clinicaltrials.gov). The results show that: the daily intake of energy correlated significantly with predicted total energy expenditure (pTEE) (ρ = 0.459, p < 0.001, and q < 0.001); protein intake correlated significantly with the ratio of 24 h urinary urea to creatinine excretion (ρ = 0.143 for total protein intake, ρ = 0.296 for animal protein intake, and ρ = 0.359 for protein intake/body weight, p < 0.001 and q < 0.001 for each correlation); vitamin B12 and folate intakes correlated significantly with their serum concentrations (ρ = 0.151 and ρ = 0.363, respectively; p < 0.001 and q < 0.001 for each correlation); sodium and potassium intakes correlated significantly with their 24 h urinary excretion (ρ = 0.298 and ρ = 0.123, respectively; p < 0.001 and q < 0.001 for each correlation); vitamin B12 and folate intakes were negatively associated with plasma homocysteine measure (p = 0.001 and p = 0.004, respectively); stratifying subjects by gender, the correlations between energy intake and pTEE and between potassium intake and its 24 h urinary excretion lost their significance in women. Even if the plasma and urinary levels of these nutrients depend on several factors, the significant correlations between daily reported intake of nutrients (protein, vitamin B12, folate, and sodium) and their blood/urinary markers confirmed that the 7-day food records (plus a supplementation questionnaire) provides reliable data to evaluate short-term current dietary intake in European elderly subjects and it can be exploited to guide and monitor NU-AGE participants through the shift of their diet according NU-AGE recommendations.

Cell‐free DNA (cfDNA) is present in the circulating plasma and other body fluids and is known to originate mainly from apoptotic cells. Here, we provide the first in vivo evidence of global and local chromatin changes in human aging by analyzing cfDNA from the blood of individuals of different age groups. Our results show that nucleosome signals inferred from cfDNA are consistent with the redistribution of heterochromatin observed in cellular senescence and aging in other model systems. In addition, we detected a relative cfDNA loss at several genomic locations, such as transcription start and termination sites, 5′UTR of L1HS retrotransposons and dimeric AluY elements with age. Our results also revealed age and deteriorating health status correlate with increased enrichment of signals from cells in different tissues. In conclusion, our results show that the sequencing of circulating cfDNA from human blood plasma can be used as a noninvasive methodology to study age‐associated changes to the epigenome in vivo.

The feasibility of liver transplantation from old healthy donors suggests that this organ is able to preserve its functionality during aging. To explore the biological basis of this phenomenon, we characterized the epigenetic profile of liver biopsies collected from 45 healthy liver donors ranging from 13 to 90 years old using the Infinium HumanMethylation450 BeadChip. The analysis indicates that a large remodeling in DNA methylation patterns occurs, with 8,823 age-associated differentially methylated CpG probes. Notably, these age-associated changes tended to level off after the age of 60, as confirmed by Horvath's clock. Using stringent selection criteria, we further identified a DNA methylation signature of aging liver including 75 genomic regions. We demonstrated that this signature is specific for liver compared to other tissues and that it is able to detect biological age-acceleration effects associated with obesity. Finally, we combined DNA methylation measurements with available expression data. Although the intersection between the two omic characterizations was low, both approaches suggested a previously unappreciated role of epithelial-mesenchymal transition and Wnt-signaling pathways in the aging of human liver.

Deep sequencing technologies are completely revolutionizing the approach to DNA analysis. Mitochondrial DNA (mtDNA) studies entered in the "postgenomic era": the burst in sequenced samples observed in nuclear genomics is expected also in mitochondria, a trend that can already be detected checking complete mtDNA sequences database submission rate. Tools for the analysis of these data are available, but they fail in throughput or in easiness of use. We present here a new pipeline based on previous algorithms, inherited from the "nuclear genomic toolbox," combined with a newly developed algorithm capable of efficiently and easily classify new mtDNA sequences according to PhyloTree nomenclature. Detected mutations are also annotated using data collected from publicly available databases. Thanks to the analysis of all freely available sequences with known haplogroup obtained from GenBank, we were able to produce a PhyloTree-based weighted tree, taking into account each haplogroup pattern conservation. The combination of a highly efficient aligner, coupled with our algorithm and massive usage of asynchronous parallel processing, allowed us to build a high-throughput pipeline for the analysis of mtDNA sequences that can be quickly updated to follow the ever-changing nomenclature. HaploFind is freely accessible at the following Web address: https://haplofind.unibo.it.

The possible genotoxicity of extremely low frequency magnetic field (ELF-MF) exposure is still a controversial topic. The most of the reported data suggests that it alone does not affect DNA integrity, but several recent reports have suggested that sinusoidal ELF-MF may increase the effect of known genotoxic agents. Only a few studies deal with non sinusoidal ELF-MF, including pulsed magnetic field (PMF), which are produced by several devices. The aim of this study is to investigate whether PMF exposure can interfere with DNA damage and repair in the presence of a genotoxic oxidative agent in neuronal type cells. To this purpose gamma-H2AX foci formation, which is a sensitive marker of DNA double strand breaks (DSB), was investigated at different points of time (1, 24, 48, 72. h) after the H2O2 treatment (300. μM for 1. h) under PMF exposure (1. mT, 50. Hz) in human neuroblastoma BE(2)C cells. Moreover, cytotoxicity evaluation, by MTT assay and cell cycle analysis, was performed at various points of time after the treatment. Taken together, results suggest that PMF exposure does not interfere with genotoxicity and cytotoxicity induced by oxidative stress.

Centenarians are rare and exceptional individuals characterized by a peculiar phenotype. They are the best example of healthy aging in humans as most of them have escaped or substantially delayed the onset of major age-related diseases. Within this scenario, the purpose of the present work was to understand if immune status is associated with survival and health status in centenarians. To this aim, 116 centenarians were concomitantly characterized for their immunological, health and functional status, and followed-up for five-year survival. On the basis of previous knowledge we focused on a core of fundamental and basic immune parameters (number of leukocytes, monocytes, total lymphocytes, CD3(+) T lymphocytes, CD4(+) helper T lymphocytes, CD8(+) cytotoxic T lymphocytes, CD19(+) B lymphocytes and plasma levels of IgM), and the most important findings can be summarized as follows: i. a hierarchical cluster analysis was able to define Cluster1 (88 centenarians) and Cluster2 (28 centenarians) characterized by low and high values of all these immune parameters, respectively; ii. centenarians of Cluster2 showed a statistically longer five-year survival and more favorable values of other important immune (naïve, activated/memory and effector/memory T cells) and metabolic (glycemia, insulin and HOMA-IR) parameters, in accord with previous observations that centenarians have a peculiar immune profile, a preserved insulin pathway and a lower incidence of type 2 diabetes; and iii. unexpectedly, parameters related to frailty, as well as functional and cognitive status, did not show any significant correlation with the immune clustering, despite being capable per se of predicting survival. In conclusion, high values of basic immunological parameters and important T cell subsets correlate with five-year survival in centenarians, independent of other phenotypic characteristics. This unexpected biological scenario is compatible with the general hypothesis that in centenarians a progressive disconnection and loss of biological coherence among the different functions of the body occur, where survival/mortality result from the failure of any of these domains which apparently follow an independent age-related trajectory.

Induced pluripotent stem cell (iPSC)-technology is an important platform in medicine and disease modeling. Physiological degeneration and disease onset are common occurrences in the aging population. iPSCs could offer regenerative medical options for age-related degeneration and disease in the elderly. However, reprogramming somatic cells from the elderly is inefficient when successful at all. Perhaps due to their low rates of replication in culture, traditional transduction and reprogramming approaches with centenarian fibroblasts met with little success. A simple and reproducible reprogramming process is reported here which enhances interactions of the cells with the viral vectors that leads to improved iPSC generation. The improved methods efficiently generates fully reprogrammed iPSC lines from 105–107 years old subjects in feeder-free conditions using an episomal, Sendai-Virus (SeV) reprogramming vector expressing four reprogramming factors. In conclusion, dermal fibroblasts from human subjects older than 100 years can be efficiently and reproducibly reprogrammed to fully pluripotent cells with minor modifications to the standard reprogramming procedures. Efficient generation of iPSCs from the elderly may provide a source of cells for the regeneration of tissues and organs with autologous cells as well as cellular models for the study of aging, longevity and age-related diseases.

During the last decades survival has significantly improved and centenarians are becoming a fast-growing group of the population. Human life span is mainly dependent on environmental and genetic factors. Favourable modifications of lifestyle factors (e.g. physical activity, diet and not smoking) and healthcare (e.g. effective vascular disease prevention) have also increased human life span. Genetic factors contribute to the variation of human life span by around 25%, which is believed to be more profound after 85 years of age. It is likely that multiple factors influence life span and we need answers to questions such as: 1) What does it take to reach 100?, 2) Do centenarians have better health during their lifespan compared with contemporaries who died at a younger age?, 3) Do centenarians have protective modifications of body composition, fat distribution and energy expenditure, maintain high physical and cognitive function, and sustained engagement in social and productive activities?, 4) Do centenarians have genes which contribute to longevity?, 5) Do centenarians benefit from epigenetic phenomena?, 6). Is it possible to influence the transgenerational epigenetic inheritance (epigenetic memory) which leads to longevity?, 7) Is the influence of nutrigenomics important for longevity?, 8) Do centenarians benefit more from drug treatment, particularly in primary prevention?, and, 9) Are there any potential goals for drug research? Many definitions of successful ageing have been proposed, but at present there is no consensus definition. Such definitions may need to differentiate between "Longevity Syndrome" and "Exceptional Longevity"

Background Myelofibrosis (MF) is a clonal hemopoietic stem/progenitor cells (HSPCs) disorder with mutations in 3 driver genes (JAK2, MPL, or CALR) and inflammatory microenvironment. Triple negative (TN) patients (pts) do not carry the 3 driver mutations and display a significantly worse survival-rate which can be partially due to a greater mutation complexity. Circulating microvesicles (MVs; 0.1-1 μm), most of megakaryocytes (MKMVs) and platelets (PMVs) origin, play a role in intercellular signalling and are increased in inflammation and cancer including MF. We hypothesized that in TN pts the HSPCs and/or the inflammatory microenvironment show a more aggressive and malignant phenotype. Aims To identify a biomolecular signature of TN pts by comparatively evaluating the circulating HSPCs behaviour and their functional interplay with the inflammatory microenvironment. Methods EDTA-anticoagulated peripheral blood (PB) was collected from 16 MF pts (WHO-2016 criteria-JAK2(V617F) mutated (n=10) and TN (n=6)) and 20 age/sex-matched healthy donors (HD). Driver mutation status was obtained by PCR/NGS. Plasma MVs were isolated by ultracentrifugation (45.000 rpm for 2 hours), counted by Nanosight technology and phenotypically characterized for PMVs (CD61+CD62P+) and MKMVs (CD61+CD62P-) by flow cytometry (size 0.5-0.9 µm). in vitro survival (24 hours), CXCL12-driven migration (12 hours) and clonogenic ability of immunomagnetically isolated CD34+ cells from pts PB or cord blood (CB; n=10) was performed in the presence/absence of the isolated MVs or IL-1β, TNF-α and IL-6 (alone/ in combination). Incorporation of CFSE-labelled MVs into CD34+ cells was analyzed by confocal microscopy. MicroRNA (miRs) profile analysis of the isolated MVs and Gene expression profile (GEP) analysis of CD34+ cells from 3 JAK2(V617F) mutated, 3 TN pts and 3 HD/CB was performed. Plasma inflammatory cytokines levels were evaluated by ELISA. Results We firstly compared the TN and JAK2(V617F) CD34+ cells. TN CD34+ cells showed decreased in vitro survival and clonogenic ability but comparable migration capacity (Fig. 1 A, B, C). Consistently, comparing the GEP, the expression of selected anti-apoptotic (TSPYL5, GFI-1 and FCMR) and pro-apototic (TNFSF10, TP53INP1) genes was significantly down- and up-regulated, respectively, in TN CD34+ cells. We then investigated whether signals from the microenvironment may affect the in vitro behaviour of the TN CD34+ cells. IL-1β, IL-6 and TNF-α plasma levels were significantly increased (p Conclusions Our results demonstrate that TN CD34+ cells show in vitro defective function and are unresponsive to the inflammatory microenvironment. Interestingly, only the autologous circulating MVs promoted the malignant clone, suggesting that MVs, as signals from microenvironment, may have a pathogenetic role in TN MF and may be the target of novel therapeutic approaches. Figure 1. Figure 1. Disclosures Fabbri: Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS: Employment. Cavo:Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Honoraria, Membership on an entity's Board of Directors or advisory committees; GlaxoSmithKline: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Palandri:Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.

Inflamm-aging, that is the age-associated inflammatory status, is considered one of the most striking consequences of immunosenescence, as it is believed to be linked to the majority of age-associated diseases sharing an inflammatory basis. Nevertheless, evidence is emerging that inflamm-aging is at least in part independent from immunological stimuli. Moreover, centenarians who avoided or delayed major inflammatory diseases display markers of inflammation. In this paper we proposed a reappraisal of the concept of inflamm- aging, suggesting that its pathological effects can be independent from the total amount of pro-inflammatory mediators, but they would be rather associated with the anatomical district and type of cells where they are produced and where they primarily act.

The study of the extreme limits of human lifespan may allow a better understanding of how human beings can escape, delay, or survive the most frequent age-related causes of morbidity, a peculiarity shown by long-living individuals. Longevity is a complex trait in which genetics, environment, and stochasticity concur to determine the chance to reach 100 or more years of age [1]. Because of its impact on human metabolism and immunology, the gut microbiome has been proposed as a possible determinant of healthy aging [2, 3]. Indeed, the preservation of host-microbes homeostasis can counteract inflammaging [4], intestinal permeability [5], and decline in bone and cognitive health [6, 7]. Aiming at deepening our knowledge on the relationship between the gut microbiota and a long-living host, we provide for the first time the phylogenetic microbiota analysis of semi-supercentenarians, i.e.,105-109 years old, in comparison to adults, elderly,and centenarians, thus reconstructing the longest available human microbiota trajectory along aging. We highlighted the presence of a core microbiota ofhighly occurring, symbiotic bacterial taxa (mostly belonging to the dominant Ruminococcaceae, Lachnospiraceae, and Bacteroidaceae families), with a cumulative abundance decreasing along withage. Aging is characterized by an increasing abundance of subdominant species, as well as a rearrangementintheir co-occurrence network. These features are maintained in longevity and extreme longevity, but peculiarities emerged, especially in semi-supercentenarians, describing changes that, even accommodating opportunistic and allochthonous bacteria, might possibly supporthealth maintenance during aging, such as an enrichment and/or higher prevalence of health-associated groups (e.g., Akkermansia, Bifidobacterium, and Christensenellaceae).

A typical feature of ageing is a chronic, low-grade inflammation characterized by a general increase in the production of pro-inflammatory cytokines and inflammatory markers ("inflamm-ageing"). This status may slowly damage one or several organs, especially when unfavorable genetic polymorphisms and epigenetic alterations are concomitant, leading to an increased risk of frailty together with the onset of age-related chronic diseases. The contribution of different tissues (adipose tissue, muscle), organs (brain, liver), immune system and ecosystems (gut microbiota) to age-related inflammation ("inflamm-ageing") will be discussed in this review in the context of its onset/progression leading to site-restricted and systemic effects. Moreover, some of the possible strategies and therapies to counteract the different sources of molecular mediators which lead to the age-related inflammatory phenotype will be presented.

In this study, the induction of apoptosis after exposure to 900 MHz radiofrequency radiation (GSM signal) was investigated by assessing caspase 3 activation in exponentially growing Jurkat cells and in quiescent and proliferating human peripheral blood lymphocytes (PBLs). The exposure was carried out at an average specific absorption rate of 1.35 W/kg in a dual wire patch cell exposure system where the temperature of cell cultures was accurately controlled. After 1 h exposure to the radiofrequency field, a slight but statistically significant increase in caspase 3 activity, measured 6 h after exposure, was observed in Jurkat cells (32.4%) and in proliferating human PBLs (22%). In contrast, no effect was detected in quiescent human PBLs. In the same experimental conditions, apoptosis was also evaluated in Jurkat cells by Western blot analysis and in both cell types by flow cytometry. To evaluate late effects due to caspase 3 activity, flow cytometry was also employed to assess apoptosis and viability 24 h after radiofrequency-radiation exposure in both cell types. Neither the former nor the latter was affected. Since in recent years it has been reported that caspases are also involved in processes other than apoptosis, additional cell cycle studies were carried out on proliferating T cells exposed to radiofrequency radiation; however, we found no differences between sham-exposed and exposed cultures. Further studies are warranted to investigate the biological significance of our findings of a dose-response increase in caspase 3 activity after exposure to radiofrequency radiation.

The role of inherited and somatic mutations of mitochondrial DNA (mtDNA) in aging and longevity is complex and highly controversial, owing to its peculiar genetics, including the phenomenon of heteroplasmy. Most of the data on mtDNA and longevity have been obtained on humans and particularly on centenarians, i. e., people who escaped or delayed the major age-related pathologies and reached the extreme limit of human lifespan. In this review we summarize the most recent advances in this field that suggest a consistent role in human longevity of both germ-line inherited and somatically acquired mutations. The particular case of the association with longevity of the somatic C150T mutation is extensively discussed, challenging the tenet that mtDNA mutations are basically detrimental. We also stress several limitations of our present knowledge, regarding the difficulty in extrapolating to humans the results obtained in animal models, owing to a variety of biological differences, including the very limited genetic variability of mtDNA in the strains used in laboratory experiments. The use of high-throughput technologies and the extensive analysis, possibly at the single cell level, of different tissues and cell types derived from the same individual will help in disentangling the complexity of mtDNA in aging and longevity.

Epidemiological studies have revealed a progressive increase in the prevalence of diabetes mellitus in the elderly. Numerous factors are responsible for this trend, among them there are (a) the long-lasting disease due the improved therapeutic remedial (pharmacological, dietary treatments and physical activity), (b) the increased life span expectancy. The prevalence of diabetes mellitus in long living subjects is lower than in elderly people (subjects aged from 65 to 84). Senile diabetes is prevalent in long living people, and usually begins after 90 years. The incidence of neodiagnosed diabetes is higher in the oldest old than in the elderly people. Based on the results, diabetes mellitus is a negative factor for survival, and does not usually allow to achieve very old age, i.e. centenarian.

Within the MARK-AGE project, a population study (3337 subjects) was conducted to identify a set of biomarkers of ageing which, as a combination of parameters with appropriate weighting, would measure biological age better than any single marker. The MARK-AGE project involves 14 European countries and a total of 26 research centres. In such a study, standard operating procedures (SOPs) are an essential task, which are binding for all MARK-AGE Beneficiaries. The SOPs cover all aspects of subject's recruitment, collection, shipment and distribution of biological samples (blood and its components, buccal mucosa cells or BMC and urine) as well as the anthropometric measurements and questionnaires. published

Recent studies on "nonthermal" effects of mobile phone radiofrequency (RF) suggest that RF can interact with cellular functions and molecular pathways. To study the possible RF effects on human lymphocyte activation, we analyzed CD25, CD95, CD28 molecules in unstimulated and stimulated CD4+ e CD8+ T cells in vitro. Peripheral blood mononuclear cells (PBMCs) from young and elderly donors were exposed or sham-exposed to RF (1,800 MHz, Specific Absorption Rate 2 W/kg) with or without mitogenic stimulation. No significant changes in the percentage of these cell subsets were found between exposed and sham-exposed lymphocytes in both young and elderly donors. Nevertheless, after RF exposure we observed a slight, but significant, downregulation of CD95 expression in stimulated CD4+ T lymphocytes from elderly, but not from young donors. This age-related result is noteworthy given the importance of a such molecule in regulation of the immune response.

In mammals, the thymus is the primary central organ of the lymphoid system; after birth, it progressively diminishes in size, undergoing gradual atrophy. Physiological maturation and/or involution of the thymus may be accelerated by endogenous or exogenous factors. Exposure to extremely low frequency EMF seems to interfere with thymic cell death. Data suggest that, in the rat model, a prolonged exposure to 50 Hz electric and magnetic fields, independently from field strength, seems to affect thymic cell death and possibly thymic physiology, since alterations in the balance of cell death and other parameters such as mitoses might interfere with the positive and negative selection of thymocytes and with the immunosurveillance properties of the thymus.

In this paper, we review data of recent literature on the distribution in centenarians of candidate germ-line polymorphisms that likely affect the individual chance to reach the extreme limit of human life. On the basis of previous observations on the immunology, endocrinology and cellular biology of centenarians we focused on genes that regulate immune responses and inflammation (IL-6, IL-1 cluster, IL-10), genes involved in the insulin/IGF-I signalling pathway and genes that counteract oxidative stress (PON1). On the whole, data indicate that polymorphisms of these genes likely contribute to human longevity, in accord with observations emerging from a variety of animal models, and suggest that a common core of master genes and metabolic pathways are responsible for aging and longevity across animal species. Moreover, in the concern of our plan to discover new genetic factors related to longevity, we explored the possibility to by-pass the need of an a-priori choice of candidate genes, extending the search to genes and genomic regions of still unknown function. Alu sequences may be considered as good markers of highly variable and potentially unstable loci in functionally important genomic regions. We extensively screened Alu-rich genomic sites and found a new genomic region associated with longevity.

In the last 30 years, an increasing public concern about the possible harmful effects of electromagnetic fields generated by power lines and domestic appliances has pushed the scientific community to search for a correct and comprehensive answer to this problem. In this work the effects of exposure to 50 Hz sinusoidal magnetic fields, with a magnetic flux density of 0.05 mT and 2.5 mT (peak values), were studied on human peripheral blood mononuclear cells (PBMCs) collected from healthy young and elderly donors. Cell activation and proliferation were investigated by using flow cytometry techniques and 3H-TdR incorporation assays, respectively. The results obtained indicated that exposure to the fields altered neither DNA synthesis nor the capacity of lymphocytes to enter the activation phase and progress into the cell cycle. Thus, the conclusions are that two important functional phases of human lymphocytes, such as activation and proliferation, are not affected by exposures to 50 Hz magnetic fields similar to those found under power lines.

Syncytium formation in cells that express herpes simplex virus glycoprotein B (gB), gD, gH, and gL is blocked by gK (E. Avitabile, G. Lombardi, and G. Campadelli-Fiume, J. Virol. 77: 6836-6844, 2003). Here, we report the results of two series of experiments. First, UL20 protein (UL20p) expression weakly inhibited cell-cell fusion. Coexpression of UL20p and gK drastically reduced fusion in a cell-line-dependent manner, with the highest inhibition in BHK cells. Singly expressed UL20p and gK localized at the endoplasmic reticulum and nuclear membranes. When they were coexpressed, both proteins relocalized to the Golgi apparatus. Remarkably, in cells that coexpressed UL20p and gK, the antifusion activity correlated with a downmodulation of gD, gB, gH, and gL cell surface expression. Second, gB Δ867 has a partial deletion in the cytoplasmic tail that removed endocytosis motifs. Whereas wild-type (wt) gB was internalized in vesicles lined with the endosomal marker Rab5, gB Δ867 was not internalized, exhibited enhanced cell surface expression, and was more efficient in mediating cell-cell fusion than wt gB. The antifusion activity of UL20p and gK was also exerted when gB Δ867 replaced wt gB in the cell fusion assay. These studies show that the gB C tail carries a functional endocytosis motif(s) and that the removal of the motif correlated with increased gB surface expression and increased fusion activity. We conclude that cell-cell fusion in wt-virus-infected cells is negatively controlled by at least two mechanisms. The novel mechanism described here involves the concerted action of UL20p and gK and correlates with a moderate but consistent reduction in the cell surface expression of the fusion glycoproteins. This mechanism is independent of the one exerted through endocytosis-mediated downmodulation of gB from the plasma membrane.

Centenarians are people who escaped from major common diseases, including cancer, and reached the extreme limits of human life-span. The analysis of demographic data indicates that cancer incidence and mortality show a levelling off around the age of 85–90 years, and suggests that oldest old people and centenarians are protected from cancer onset and progression. In this paper, we review data of recent literature on the distribution in centenarians of germ-line polymorphisms, which are supposed to affect the individual susceptibility to cancer (p53, HRAS1, BRCA1, glutathione transferases, cytochrome oxidases, steroid-5 alpha-reductase enzyme type II). Moreover, we add new data on two p53 polymorphisms in a total of 1086 people of different age, including 307 centenarians. In addition, we put forth the hypothesis that the remodelling of the immune system occurring with age is capable of creating a hostile environment for the growth of cancer cells in these exceptional individuals. We conclude that future studies on centenarians regarding the germ-line variability of genes involved in the control of the immune response, including apoptosis (ApoJ), are likely to be of fundamental importance in understanding the basic mechanisms for cancer, aging and their complex relationship.

Owing to organ shortage, livers from old donors are increasingly used for transplantation. The function and duration of such transplanted livers are apparently comparable to those from young donors, suggesting that, despite some morphological and structural age-related changes, no major functional changes do occur in liver with age. We tested this hypothesis by performing a comprehensive study on proteasomes, major cell organelles responsible for proteostasis, in liver biopsies from heart-beating donors. Oxidized and poly-ubiquitin conjugated proteins did not accumulate with age and the three major proteasome proteolytic activities were similar in livers from young and old donors. Analysis of proteasomes composition showed an age-related increased of β5i/α4 ratio, suggesting a shift toward proteasomes containing inducible subunits and a decreased content of PA28α subunit, mainly in the cytosol of hepatocytes. Thus our data suggest that, proteasomes activity is well preserved in livers from aged donors, concomitantly with subtle changes in proteasome subunit composition which might reflect the occurrence of a functional remodelling to maintain an efficient proteostasis. Gender differences are emerging and they deserve further investigations owing to the different aging trajectories between men and women. Finally, our data support the safe use of livers from old donors for transplantation. Owing to organ shortage, livers from old donors are increasingly used for transplantation. The function and duration of such transplanted livers are apparently comparable to those from young donors, suggesting that, despite some morphological and structural age-related changes, no major functional changes do occur in liver with age. We tested this hypothesis by performing a comprehensive study on proteasomes, major cell organelles responsible for proteostasis, in liver biopsies from heart-beating donors. Oxidized and poly-ubiquitin conjugated proteins did not accumulate with age and the three major proteasome proteolytic activities were similar in livers from young and old donors. Analysis of proteasomes composition showed an age-related increased of β5i/α4 ratio, suggesting a shift toward proteasomes containing inducible subunits and a decreased content of PA28α subunit, mainly in the cytosol of hepatocytes. Thus our data suggest that, proteasomes activity is well preserved in livers from aged donors, concomitantly with subtle changes in proteasome subunit composition which might reflect the occurrence of a functional remodelling to maintain an efficient proteostasis. Gender differences are emerging and they deserve further investigations owing to the different aging trajectories between men and women. Finally, our data support the safe use of livers from old donors for transplantation.

Biological aging is associated with physiological deteriorations and its’ remodeling, which are partly due to changes in the hormonal profile. MicroRNAs are known to post-transcriptionally regulate various cellular processes associated with cell senescence; differentiation, replication and apoptosis. Measured from the serum, microRNAs have the potential to serve as noninvasive markers for diagnostics/prognostics and therapeutic targets. We analysed the association of estrogen-based hormone replacement therapy (HRT) with selected microRNAs and inflammation markers from the serum, leukocytes and muscle tissue biopsy samples obtained from 54-62 year-old postmenopausal monozygotic twins (n=11 pairs) discordant for the use of HRT. Premenopausal 30-35 year-old women (n=8) were used as young controls. We focused on the hormonal aging and more specifically, on the interaction between HRT use and the modulation of inflammation associated microRNAs, miR-21 and miR-146a, and classical inflammation markers. Fas-ligand (FasL) was analysed since it functions in both apoptosis and inflammation. The inflammatory profile is healthier among the premenopausal women compared to the older, postmenopausal twins. The serum miR-21 and miR-146a expression levels and FasL concentrations were lower in the HRT users when compared to their non-using co-twins, demonstrating their responsiveness to HRT. Based on the pairwise FasL analysis, the FasL serum concentration is likely to be genetically controlled. Overall, we suggest that postmenopausal estrogen deficiency sustains the development of “inflamm-aging” in women. Estrogen sensitive, specific circulating microRNAs could be potential, early biomarkers for age-associated physiological deteriorations. Highlights:  Unique study design of postmenopausal MZ twins discordant for HRT  Serum miR-21 and miR-146a expressions are lower in HRT users compared to non-users  FasL serum concentrations are lower in HRT users and possibly genetically regulated  Postmenopausal systemic estrogen deficiency partly contributes to the “inflamm-aging”  Serum miR-21/-146a early indicators of age-associated physiological deteriorations peerReviewed

The susceptibility to undergo apoptosis of fresh human peripheral blood mononuclear cells (PBMCs) from three groups of healthy donors of different ages: young people (19-40 years), old people (65-85 years) and centenarians was assessed. Apoptosis was induced by 2-deoxy-D-ribose (dRib), an agent which induces apoptosis in quiescent PBMCs by interfering with cell redox status and mitochondrial membrane potential (MMP). Our major finding is that an inverse correlation emerged between the age of the donors and the propensity of their PBMCs to undergo dRib-induced apoptosis. PBMCs from old people and centenarians also showed an increased resistance to dRib-induced glutathione depletion and a decreased tendency to lose MMP. The anti-apoptotic molecule Bcl-2 was similarly expressed in PBMCs from the three age groups. Moreover, the plasma level of the stable product of transglutaminase, epsilon(gamma-glutamyl)lysine isodipeptide, a marker of total body apoptotic rate, was decreased in centenarians compared to young and elderly people. On the whole, these findings suggest that physiological aging is characterised by a decreased tendency to undergo apoptosis, a phenomenon likely resulting from adaptation to lifelong exposure to damaging agents, such as reactive oxygen species, and may contribute to one of the major phenomena of immunosenescence, i.e. the progressive accumulation of memory/effector T cells.

Structural and phenotypic modifications of rat thymocytes from birth up to one year of age, i.e. during maturation and at the beginning of the involutive process of the thymus are described. Since the biological significance and the mechanisms of thymic involution are still a matter of debate, this study aims at clarifying the complexity of the compensatory events occurring during this relatively neglected period of time. Thymuses from Sprague-Dawley rats were analyzed morphologically and morphometrically by light and electron microscopy. At the same time, thymocyte subsets, isolated from the same animals, were characterized by flow cytometry according to physical parameters and phenotypic markers. Results indicate that major changes occur during the first month from birth and from six months onward. In particular, already during the first weeks after birth, thymocytes undergo a slight reduction of mitoses associated with an increased number of apoptoses. Moreover, during the same period of time, flow cytometry revealed an expansion of small thymocytes and changes in thymocyte subsets such as increase of CD4+CD8+ and CD5+alpha(beta)TCR- and a decrease of CD4-CD8-, CD4-CD8+ cells. The thymus of adult rats was characterized by time-dependent decrease of both mitoses and apoptoses, progressive physical disconnection among cells, increase of necrotic areas and fibrosis. Around one year of age tissue changes were associated with a dramatic reduction of the population of large thymocytes and the rise of numerous small thymocytes that were unexpectedly negative for all tested markers. By contrast, medium-size thymocytes exhibited a marked decrease of CD4+CD8+ and CD5+alpha(beta)TCR- subsets. In conclusion, our data indicate that thymus undergoes, with time, a complex remodeling and suggest that thymic involution is not only a simple shrinkage of the organ but rather the result of a series of compensatory mechanisms among different cell populations in a setting of progressive involution.

Inflamm-aging, that is the age-associated inflammatory status, is considered one of the most striking consequences of immunosenescence, as it is believed to be linked to the majority of age-associated diseases sharing an inflammatory basis. Nevertheless, evidence is emerging that inflamm-aging is at least in part independent from immunological stimuli. Moreover, centenarians who avoided or delayed major inflammatory diseases display markers of inflammation. In this paper we proposed a reappraisal of the concept of inflamm-aging, suggesting that its pathological effects can be independent from the total amount of pro-inflammatory mediators, but they would be rather associated with the anatomical district and type of cells where they are produced and where they primarily act.

3-Aminobenzamide (3-ABA) is an inhibitor of poly(ADP-ribose)polymerase (PARP), an enzyme involved in several cellular processes, and exerts its effects by acting at the cytoskeleton level. Here we show that 3-ABA has an antiproliferative effect on the human carcinoma cell line A431, as measured by different assays. 3-ABA was capable of inhibiting cell growth as well as colony formation, this inhibitory effect is reversible. Morphological analyses showed a series of cellular alterations, such as a remarkable increase of dendritic-like protrusions, quite unusual in epithelial cells, and suggestive of a differentiative triggering. Immunocytochemical studies suggested that a major target of 3-ABA was indeed the cytoskeleton. These data, together with those of the literature, indicate that 3-ABA, depending on cell histotype and drug concentration, is a versatile drug capable of exerting antiproliferative and cytostatic effects as well as cytotoxic and antiapoptotic effects, processes sharing an important involvement of cytoskeleton. These unique characteristics of 3-ABA may be of interest for cancer research.

In recent decades, major theoretical and technological advances have been achieved in the field of immunology. These have allowed the scientific community to analyse the immune system in a much more sophisticated manner than was possible even 20 years ago. Moreover, great theoretical changes have also occurred in gerontology - in particular, the hypothesis has been put forward that ageing and diseases are two different phenomena, and that successful ageing, i.e. ageing in good psychophysical conditions, is really possible for most humans and animals. Immunosenescence was then carefully investigated, either in selected healthy people of advanced age or in the oldest old people, such as healthy centenarians. The main results showed that most immune parameters are indeed well preserved even at this far advanced age. This paper deals with some of the most important theoretical problems of immunosenescence. An immunological tenet was that the most important phenomenon of immunosenescence is the involution of the thymus. In most textbooks and papers it is taken for granted that the thymus starts its involution immediately after puberty. When people aged 60-65 were considered old, it was not difficult to think that they could live for the rest of their life with a fully involuted thymus. The findings on centenarians challenge this tenet, as they have only a small reduction of T lymphocytes, and a relatively normal number of virgin and memory T cells, together with a functional T cell repertoire. Other observations reported here on centenarians, concerning the activity of B lymphocytes and the cytokine network, as well as those on the well-preserved innate immunity and the cells' capability of undergoing proliferation after appropriate stimuli, suggest that complex immune changes occur with age, but also indicate that we have to modify our attitude, to grasp the new scenario which is emerging. Immunosenescence can no longer be considered as a unidirectional deterioration, and this complex phenomenon is much better described by terms such as 'remodelling', 'reshaping' or 'retuning'.

Clinical evidence demonstrates that ubiquinol-10, the reduced active form of coenzyme Q10 (CoQ10H₂), improves endothelial function through its antioxidant and probably its anti-inflammatory properties. We previously reported that a biomarker combination including miR-146a, its target protein IL-1 receptor-associated kinase (IRAK-1), and released interleukin (IL)-6, here collectively designated as MIRAKIL, indicates senescence-associated secretory phenotype (SASP) acquisition by primary human umbilical vein endothelial cells (HUVECs). We explore the ability of short- and long-term CoQ10H₂ supplementation to affect MIRAKIL in HUVECs, used as a model of vascular aging, during replicative senescence in the absence/presence of lipopolysaccharide (LPS), a proinflammatory stimulus. Senescent HUVECs had the same ability as young cells to internalize CoQ10 and exhibit an improved oxidative status. LPS-induced NF-κB activation diminished after CoQ10H₂ pretreatment in both young and senescent cells. However, short-term CoQ10H₂ supplementation attenuated LPS-induced MIRAKIL changes in young cells; in senescent cells CoQ10H₂ supplementation significantly attenuated LPS-induced miR-146a and IRAK-1 modulation but failed to curb IL-6 release. Similar results were obtained with long-term CoQ10H₂ incubation. These findings provide new insights into the molecular mechanisms by which CoQ10H₂ stems endothelial cell inflammatory responses and delays SASP acquisition. These phenomena may play a role in preventing the endothelial dysfunction associated with major age-related diseases.