Fight Aging! Newsletter, November 4th 2024

Extracellular vesicles (EVs) are vital for cell-to-cell communication, transferring proteins, lipids, and nucleic acids in various physiological and pathological processes. They play crucial roles in immune modulation and tissue regeneration but are also involved in pathogenic conditions like inflammation and degenerative disorders. EVs have heterogeneous populations and cargo, with numerous subpopulations currently under investigations. EV therapy shows promise in stimulating tissue repair and serving as a drug delivery vehicle, offering advantages over cell therapy, such as ease of engineering and minimal risk of tumorigenesis.

Despite the rapid growth in the EV field, much remains to be studied. There are numerous EV classes and subclasses yet to be fully characterized. The heterogeneity within EV classes leads to variability in their effects. Studies isolating EVs from the same cell line report different cargo and mechanisms of action. The effects of EV subpopulations are also not fully understood, with research predominantly focused on exosomes. The roles of microvesicles (MVs) are still controversial, as they can be either therapeutic or pathogenic depending on their source. MVs and other large EVs may be worth exploring further since they can deliver more cargo.

Characterization and purification of EVs are crucial for clinical application. To control the effects of EVs, production methods need to be strictly replicable to avoid heterogeneity, with specific culture and modulation techniques. EVs are involved in multiple pathological processes, such as inflammation, tumorigenesis, and toxic protein spreading. Blocking these pathological EVs is challenging due to a lack of specificity, necessitating more precise techniques for targeting them. Multiple sources of EVs have been studied, but not thoroughly across all domains of treatment. Some cell sources may be better suited for certain roles; for example, stem cells for neuroregeneration, glia for immunomodulation, and endothelial cells for angiogenesis. The targeting ability of EVs is another area that has not been extensively explored. Optimal EV preconditioning, administration regimens, and safety profiles also require further investigation.

A better understanding of EV subtypes and their specific roles will mark a significant milestone in medicine, leading to safer, more effective and disease-tailored EV therapy for a variety of neurological disorders.

We established the Mammalian Methylation Consortium with two primary objectives. The first objective was to develop DNA methylation-based measures to track the passage of time, culminating in the creation of the pan-mammalian methylation clock. The second objective aimed to understand the epigenetic correlates of maximum mammalian lifespan, a goal explored through a trilogy of papers. In the first paper, we developed multivariate predictors of maximum lifespan based on cytosine methylation. This predictor can estimate species-specific characteristics, such as maximum lifespan, from a DNA sample, even if the species is unknown. The second paper characterized individual CpGs and clusters of CpGs (modules) that correlate with maximum lifespan, providing insights into the methylation landscape of long-lived species. Interestingly, this study found only a weak overlap between CpGs that correlate with maximum lifespan and those associated with chronological age. While we understand that a species characteristic like maximum lifespan is genetically hardwired and does not change with the individual's age, this finding challenges the intuitive hypothesis that individual aging (the passage of time) must relate to maximum lifespan (a species characteristic).

Prior studies have linked the rate of change in methylation to maximum lifespan in smaller samples of mammalian species, and have suggested a strong position correlation between slower average rate of change in methylation and increased species maximum lifespan. But this is not always the case. By leveraging the large dataset from our Mammalian Methylation Consortium and a careful mathematical framework we demonstrate that the strong correlation between slower average rate of change in methylation and increased species maximum lifespan is only found in certain chromatin states such as bivalent promoters. In other chromatin states, the correlation is either not present or even reversed. Future studies might delve deeper into which chromatin regions result in the opposite interpretation, where a rapid rate of change correlates with an increased maximum lifespan.

Recent years have witnessed exciting advances relating to methylation clocks and programmatic aging theory. We have highlighted here the convergence of these two paths of progress, and described several new ideas arising from this convergence. Understanding of the biological processes of aging that methylation clocks correspond to has lagged behind the recent rapid progress in clock characterization. Several features of clocks suggest that this hidden biology involves developmental processes.

Recently developed programmatic theories offer a framework of ideas within which such developmental aging mechanisms can potentially be understood. Such a framework includes explanations for how such clocks evolve, the programmatic mechanisms in which they act, and how those mechanism contribute to late-life disease. Central to this framework is the hypothesis that epigenetic and developmental changes occurring throughout life history are part and parcel of the same process. According to evolutionary theory, genes exhibiting antagonistic pleiotropy, that are beneficial early in life yet detrimental later on, may be favored by natural selection, causing aging. In the context of programmatic theories, the basic hypothesis is that developmental gene action and processes that are beneficial earlier in life continue later in life, in a futile fashion, becoming detrimental and pathogenic.

Here we explore how recent discoveries about methylation clocks cohere with the developmental theory of aging. We present several new perspectives, questions, and speculations arising from the cross-referencing of these two subjects, and argue for the timeliness of interdisciplinary integration of biogerontology with developmental biology. Central to these ideas is the hypothesis that epigenetic changes occurring throughout life history (including ontogenesis and senescence) are part of wider developmental changes. We have elaborated upon this explanatory framework, introducing new terminology (including the distinction between onto-developmental and maturo-developmental processes). Hypotheses proposed include the regulation by polycomb proteins of a trade-off between developmental fidelity and plasticity, and the presence of a conserved developmental sequence as a major, previously unseen element in the aging process. These suggestions provide examples of the sort of thinking that is possible by combining ideas from evolutionary biology, biogerontology, and developmental biology, in what we have described as a developmental gerontology or devo-gero approach.

While previous studies identified common genetic variants associated with longevity in centenarians, the role of the rare loss-of-function (LOF) mutation burden remains largely unexplored. Here, we investigated the burden of rare LOF mutations in Ashkenazi Jewish individuals from the Longevity Genes Project and LonGenity study cohorts using whole-exome sequencing data.

In this study, we have discovered that centenarians, within the large cohort we examined, possess a significantly lower burden of predicted deleterious LOF variants compared to controls. This finding suggests that a protective genetic background, characterized by the depletion of damaging coding mutations, contributes to the exceptional longevity of centenarians. Notably, we also observed a lower mutation burden in centenarian offspring, although the effect was less pronounced. These findings support the notion of a heritable component to longevity outside of protective and common variants and suggest that the combined genetic background, including protective variants and depletion of damaging variants, may be transmitted across generations to support exceptional longevity.

Our pathway analysis revealed that centenarian exomes are depleted of LOF variants in several pathways related to aging and disease, including Class A/1 (Rhodopsin-like receptors), hyaluronan metabolism, post-translational protein modification, and mitochondrial translation. Class A/1 (Rhodopsin-like) receptors are involved in various physiological processes and have been implicated in age-related diseases, suggesting their potential role in longevity. Hyaluronan is a key component of the extracellular matrix that has been shown to decline with age, and its increase contributes to the extension of lifespan. Variants that maintain hyaluronan homeostasis may, therefore, promote healthy aging in humans. Post-translational protein modifications play crucial roles in protein function and stability, and their dysregulation has been associated with various age-related diseases. Mitochondrial translation has also been linked to lifespan extension in model organisms.

To complement our analysis of rare LOF variants, we also investigated the causal role of identified longevity genes in aging-related traits using Mendelian randomization (MR) analyzes. This approach allows us to infer potential causal relationships between gene expression and phenotypes of interest by using expression quantitative trait loci, eQTLs (common variants that are associated with gene expression) as instrumental variables. Our MR analyzes provided evidence for the causal effects of several longevity-associated genes, including RGP1, PCNX2, and ANO9, on multiple aging-related traits. PCNX2 was identified to be associated with longevity in an independent genome-wide association study, while ANO9 was associated with various cancers. These findings suggest that these genes may directly influence the aging process and contribute to the extended healthspan and lifespan. The consistent causal effect estimates across different aging-related traits further support the robustness of these associations.

Bats are widely known for their long lifespan, cancer resistance, and viral tolerance. As highly complex and pleiotropic processes, the genes and mechanisms underlying these phenotypes can be challenging to identify. Here we outline an approach that enables functional comparative biology by generating cell lines from wing punches of wild caught bats for genome assembly, comparative genomics, and functional follow up. Cell lines are generated from minimally-invasive biopsies collected in the field thus avoiding disturbing natural populations. Given the high density of bat species concentrated at single locations world-wide, it is feasible to collect wing punches from a large number of individuals across a wide phylogenetic range; these wing punches can be used to generate cell lines and sequencing libraries for reference genomes in a matter of weeks.

By explicitly modeling the evolution of lifespan separately from body size, we recapitulate the extant relationship between body size and lifespan across mammals in evolutionary time. Contrary to prior work, we show that overall bats exhibit allometric lifespan scaling, comparable to other mammals. However, two bat clades - Myotis and Phyllostomidae - exhibit distinct trends with Myotis demonstrating an increased rate of change in lifespan given body size compared to other mammals. This altered scaling of longevity in Myotis has dramatic consequences for their intrinsic, per-cell cancer risk and for the evolution of tumor-suppressor genes and pathways.

We found a number of genes under selection across multiple longevity-associated pathways, consistent with the pleiotropic nature of the aging process. These include members of canonical longevity pathways such as mTOR-IGF signaling, DNA damage repair, oxidative stress, and the senescence-associated secretory phenotype. We additionally identified selection in various pathways that have likely emerged as a result of the unique biology of bats, including genes at the intersection of immunity and senescence, such as Serpin-family genes; genes in metabolic pathways including amino acid metabolism; and pervasive selection observed in the ferroptosis pathway, which sits at the intersection of bats' extreme oxidative challenges, metabolic demands, immune function, and cancer resistance.

By quantifying the relative contributions of genes under selection to cancer-related pathways at each node, we found significant enrichment of these processes across the phylogeny, especially at nodes undergoing the greatest changes in lifespan and cancer risk. While cancer risk scales linearly with body size, it scales over time as a power law of 6. Unlike other systems where the evolution of cancer resistance has been driven by rapid changes in body size, the body size of Myotis has not significantly changed since their common ancestor. Instead, the rapid and repeated changes in lifespan across an order of magnitude in Myotis lead to some of the most significant changes in intrinsic cancer risk seen across mammals.

While no reports or studies of neoplasia rates have been published in Myotis, the use of in vitro models of carcinogenesis provides a promising avenue for comparative studies of cancer resistance under controlled conditions. In agreement with our results, in vitro and xenograft transplant models have shown that cells of long-lived bats, including M. lucifugus, are more resistant to carcinogenesis than shorter-lived bats and other mammals.

The mechanism that triggers the progressive dysregulation of cell functions, inflammation, and breakdown of tissues during aging is currently unknown. We propose here a previously unknown mechanism due to tissue autodigestion by the digestive enzymes. After synthesis in the pancreas, these powerful enzymes are activated and transported inside the lumen of the small intestine to which they are compartmentalized by the mucin/epithelial barrier. We hypothesize that this barrier leaks active digestive enzymes (e.g. during meals) and leads to their accumulation in tissues outside the gastrointestinal tract.

Using immunohistochemistry we provide evidence in young (4 months) and old (24 months) rats for significant accumulation of pancreatic trypsin, elastase, lipase, and amylase in peripheral organs, including liver, lung, heart, kidney, brain, and skin. The mucin layer density on the small intestine barrier is attenuated in the old and trypsin leaks across the tip region of intestinal villi with depleted mucin. The accumulation of digestive enzymes is accompanied in the same tissues of the old by damage to collagen, as detected with collagen fragment hybridizing peptides. We provide evidence that the hyperglycemia in the old is accompanied by proteolytic cleavage of the extracellular domain of the insulin receptor. Blockade of pancreatic trypsin in the old by a two-week oral treatment with a serine protease inhibitor (tranexamic acid) serves to significantly reduce trypsin accumulation in organs outside the intestine, collagen damage, as well as hyperglycemia and insulin receptor cleavage.

These results support the hypothesis that the breakdown of tissues in aging is due to autodigestion and a side-effect of the fundamental requirement for digestion.

Infections have been associated with the incidence of Alzheimer's disease and related dementias, but the mechanisms responsible for these associations remain unclear. Using a multicohort approach, we found that influenza, viral, respiratory, and skin, and subcutaneous infections were associated with increased long-term dementia risk. These infections were also associated with region-specific brain volume loss, most commonly in the temporal lobe.

We identified 260 out of 942 immunologically relevant proteins in plasma that were differentially expressed in individuals with an infection history. Of the infection-related proteins, 35 predicted volumetric changes in brain regions vulnerable to infection-specific atrophy. Several of these proteins, including PIK3CG, PACSIN2, and PRKCB, were related to cognitive decline and plasma biomarkers of dementia (Aβ42/40, GFAP, NfL, pTau-181). Genetic variants that influenced expression of immunologically relevant infection-related proteins, including ITGB6 and TLR5, predicted brain volume loss. Our findings support the role of infections in dementia risk and identify molecular mediators by which infections may contribute to neurodegeneration.

"In prior work, we developed a new 3D bioprinting method, known as "sacrificial writing in functional tissue" (SWIFT), for patterning hollow channels within a living cellular matrix. Here, building on this method, we introduce coaxial SWIFT (co-SWIFT) that recapitulates the multilayer architecture found in native blood vessels, making it easier to form an interconnected endothelium and more robust to withstand the internal pressure of blood flow." The key innovation developed by the team was a unique core-shell nozzle with two independently controllable fluid channels for the "inks" that make up the printed vessels: a collagen-based shell ink and a gelatin-based core ink. The interior core chamber of the nozzle extends slightly beyond the shell chamber so that the nozzle can fully puncture a previously printed vessel to create interconnected branching networks for sufficient oxygenation of human tissues and organs via perfusion. The size of the vessels can be varied during printing by changing either the printing speed or the ink flow rates.

The team used a shell ink that was infused with smooth muscle cells (SMCs), which comprise the outer layer of human blood vessels. After melting out the gelatin core ink, they then perfused endothelial cells (ECs), which form the inner layer of human blood vessels, into their vasculature. After seven days of perfusion, both the SMCs and the ECs were alive and functioning as vessel walls - there was a three-fold decrease in the permeability of the vessels compared to those without ECs.

Finally, they were ready to test their method inside living human tissue. They constructed hundreds of thousands of cardiac organ building blocks (OBBs) - tiny spheres of beating human heart cells, which are compressed into a dense cellular matrix. Next, using co-SWIFT, they printed a biomimetic vessel network into the cardiac tissue. Finally, they removed the sacrificial core ink and seeded the inner surface of their SMC-laden vessels with ECs via perfusion and evaluated their performance. Not only did these printed biomimetic vessels display the characteristic double-layer structure of human blood vessels, but after five days of perfusion with a blood-mimicking fluid, the cardiac OBBs started to beat synchronously - indicative of healthy and functional heart tissue.

A prospective cohort study was performed on 9,349 adults aged 40 to 79 years from the population-based European Prospective Investigation into Cancer in Norfolk Study, with repeated measurements of physical activity (PA) and diet (from 1993 till 2004) and subsequent follow-up till 2022 (median follow-up 18.8 years). Validated questionnaires were used to derive physical activity energy expenditure (PAEE) as a proxy of total PA and adherence to the Mediterranean diet score (MDS, range 0-15 points) as an indicator of overall diet quality, and their changes over time (∆PAEE and ∆MDS).

Over 149,681 person-years of follow-up, there were 3,534 deaths. In adjusted models, for each 1 standard deviation difference in baseline PAEE (4.64 kJ/kg/day), ∆PAEE (0.65 kJ/kg/day per year), baseline MDS (1.30 points) and ∆MDS (0.32 points per year), hazard ratios (HRs) for all-cause mortality were 0.90, 0.89, 0.95, and 0.93, respectively. Compared with participants with sustained low PAEE (under 5 kJ/kg/day) and low MDS (less than 8.5 points), those with sustained high PAEE and high MDS had lower all-cause mortality (HR 0.78), as did those who improved both PAEE and MDS (HR 0.60). There was no evidence of interaction between PA and diet quality exposures on mortality risk. Population impact estimates suggested that if all participants had maintained high levels of PA and diet quality consistently, cumulative adjusted mortality rate would have been 8.8% lower.

These findings suggest that adopting and maintaining higher levels of PA and diet quality are associated with lower mortality. Significant public health benefits could be realised by enabling active living and healthy eating through adulthood.

Aging is a complex and time-dependent decline in physiological function that affects most organisms, leading to increased risk of age-related diseases. Investigating the molecular underpinnings of aging is crucial to identify geroprotectors, precisely quantify biological age, and propose healthy longevity approaches. This review explores pathways that are currently being investigated as intervention targets and aging biomarkers spanning molecular, cellular, and systemic dimensions. Interventions that target these hallmarks may ameliorate the aging process, with some progressing to clinical trials.

Biomarkers of these hallmarks are used to estimate biological aging and risk of aging-associated disease. Utilizing aging biomarkers, biological aging clocks can be constructed that predict a state of abnormal aging, age-related diseases, and increased mortality. Biological age estimation can therefore provide the basis for a fine-grained risk stratification by predicting all-cause mortality well ahead of the onset of specific diseases, thus offering a window for intervention. Yet, despite technological advancements, challenges persist due to individual variability and the dynamic nature of these biomarkers. Addressing this requires longitudinal studies for robust biomarker identification. Overall, utilizing the hallmarks of aging to discover new drug targets and develop new biomarkers opens new frontiers in medicine. Prospects involve multi-omics integration, machine learning, and personalized approaches for targeted interventions, promising a healthier aging population.

Metabolic compromise is crucial in aggravating age-associated chronic inflammation, oxidative stress, mitochondrial damage, increased LDL and triglycerides, and elevated blood pressure. Excessive adiposity, hyperglycemia, and insulin resistance due to aging are associated with elevated levels of damaging free radicals, inducing a proinflammatory state and hampering immune cell activity, leading to a malfunctioning cardiometabolic condition. The age-associated oxidative load and redox imbalance are contributing factors for cardiometabolic morbidities via vascular remodelling and endothelial damage.

Recent evidence has claimed the importance of gut microbiota in maintaining regular metabolic activity, which declines with chronological aging and cardiometabolic comorbidities. Genetic mutations, polymorphic changes, and environmental factors strongly correlate with increased vulnerability to aberrant cardiometabolic changes by affecting key physiological pathways. Numerous studies have reported a robust link between biological aging and cardiometabolic dysfunction. This review outlines the scientific evidence exploring potential mechanisms behind the onset and development of cardiovascular and metabolic issues, particularly exacerbated with aging.

Emerging preclinical evidence suggests that semaglutide, a glucagon-like peptide receptor agonist (GLP-1RA) for type 2 diabetes mellitus (T2DM) and obesity, protects against neurodegeneration and neuroinflammation. We conducted emulation target trials based on a nationwide database of electronic health records (EHRs) of 116 million US patients. Seven target trials were emulated among 1,094,761 eligible patients with T2DM who had no prior Alzheimer's disease (AD) diagnosis by comparing semaglutide with seven other antidiabetic medications.

Semaglutide was associated with significantly reduced risk for first-time AD diagnosis, most strongly compared with insulin (hazard ratio [HR], 0.33) and most weakly compared with other GLP-1RAs (HR, 0.59). Similar results were seen across obesity status, gender, and age groups. Semaglutide was associated with significantly lower AD-related medication prescriptions. Our findings provide real-world evidence supporting the potential clinical benefits of semaglutide in mitigating AD initiation and development in patients with T2DM.

Within a species, larger individuals often have shorter lives and higher rates of age-related disease. Despite this well-known link, we still know little about underlying age-related epigenetic differences, which could help us better understand inter-individual variation in aging and the etiology, onset, and progression of age-associated disease. Dogs exhibit this negative correlation between size, health, and longevity and thus represent an excellent system in which to test the underlying mechanisms. Here, we quantified genome-wide DNA methylation in a cohort of 864 dogs in the Dog Aging Project.

Age strongly patterned the dog epigenome, with the majority (66% of age-associated loci) of regions associating age-related loss of methylation. These age effects were non-randomly distributed in the genome and differed depending on genomic context. We found the LINE1 (long interspersed elements) class of TEs (transposable elements) were the most frequently hypomethylated with age. This LINE1 pattern differed in magnitude across breeds of different sizes - the largest dogs lost 0.26% more LINE1 methylation per year than the smallest dogs. This suggests that epigenetic regulation of TEs, particularly LINE1s, may contribute to accelerated age and disease phenotypes within a species.

Since our study focused on the methylome of immune cells, we looked at LINE1 methylation changes in golden retrievers, a breed highly susceptible to hematopoietic cancers, and found they have accelerated age-related LINE1 hypomethylation compared to other breeds. We also found many of the LINE1s hypomethylated with age are located on the X chromosome and are, when considering X chromosome inactivation, counter-intuitively more methylated in males. These results have revealed the demethylation of LINE1 transposons as a potential driver of inter-species, demographic-dependent aging variation.

Serious scientists have championed the diabetes drug metformin as a potential longevity therapeutic for more than two decades, but more careful scientific testing of the hypothesis over the last ten years has soured many critical thinkers on this use of the drug. Notable evidence against the idea include that metformin failed to extend lifespan in rodents in the National Institute on Aging (NIA)'s Interventions Testing Program and other well-done lifespan studies, and the debunking of the human epidemiological study that appeared to show that people with diabetes who took metformin lived longer than nondiabetics who didn't.

But interest was rekindled by a new study of metformin in aging cynomolgus monkeys. The authors claim to have shown that metformin preserved brain structure, "enhanced" cognitive function, and slowed biological aging by more than six years after less than three and a half years of treatment, as measured on a new species-specific aging clock. So like gamblers who have lost their shirt but can't stop themselves from playing, the conviction that this time it's different has set into the longevity community.

The monkeys in the metformin trial began the study at a body mass index (BMI) of 28, which is in the "overweight" category, and remarkably, the control animals ballooned to BMI 32 (obese) just three months later. By contrast, the monkeys on metformin remained 3 to 5 BMI points lighter than the controls through most of the experiment, up until the control animals began their late-life decline. That's a quite profound weight-protective effect, comparable to the new GLP-1 receptor agonist drugs.

So the first two interrelated problems with the study are that the control animals were obese throughout almost the entire study and that metformin had such a profound protective effect against weight gain in these animals. Why would metformin have made these monkeys so much lighter than their untreated labmates? Metformin is known to cause modest weight loss in humans, especially when it is "enhanced" by common side effects like indigestion or diarrhea. But the weight-buffering effect in these monkeys is much greater than you'll typically see in humans on the drug. Whatever the mechanism, the protective effect against the substantial weight gain seen in the control animals seems highly likely to be responsible for much or all of the benefits in the metformin-treated monkeys, for reasons that have nothing to do with a direct effect on aging processes.

Yamanaka factors (YFs) can reverse some aging features in mammalian tissues, but their effects on the brain remain largely unexplored. Here, we employed a controlled spatiotemporal induction of YFs in the mouse brain across two distinct scenarios: during brain development and in adult stages within the context of neurodegeneration. Our focus on the impact of YFs on neurogenesis during development was influenced by recent findings that a subset of these factors is expressed in various neural progenitors early in this phase.

Here, we report that transient, low-level expression of YFs increased proliferation, resulting in an augmented output of neurons and glia, which led to an enlarged neocortex. This expansion was functionally reflected in enhanced motor and social behavior in adult mice. Because this induction protocol enhanced cognitive skills, we hypothesized that it could exert a similar effect in the context of a neurodegenerative disorder. Thus, we expressed YFs only in mature hippocampal neurons, using the 5xFAD mouse model of Alzheimer's disease. We show that these neurons tolerate intermittent YF expression while preserving their identity. This safe approach led to cognitive, molecular, and histological improvements in the 5xFAD mice.

Our results establish transient YF induction as a powerful tool for modulating neural proliferation, and it may open new therapeutic strategies for brain disorders.