Fight Aging!

In today's open access paper, researchers report a modest improvement in cerebral blood flow and cognitive performance in a small study of older individuals suffering cognitive impairment as a result of sustained hyperbaric oxygen treatment over a period of months. This seems a compensatory approach to therapy, in that improvements in cerebral blood flow should be expected to improve cognitive function at any age. This is the mechanism by which exercise rapidly improves memory function, for example. A direct comparison of hyperbaric oxygen treatment and exercise would be interesting.

This result might help to inform discussions of the degree to which loss of blood supply to the brain contributes to cognitive decline in patients diagnosed with neurodegenerative conditions. Vascular dementia is an acknowledged and well-researched condition, but to what degree is the impairment of Alzheimer's patients at various stages due to vascular aging and consequent reduced blood flow to the brain, versus the harmful protein aggregation and neuroinflammation characteristic of Alzheimer's? Absent a way to remove just one of these pathologies, it is hard to answer that question.

It is worth noting that this study was conducted and published by the same groups who put together the poor study and accompanying overhyped media materials regarding the effects of hyperbaric oxygen treatment on measures of metabolism related to aging. It is most likely a good idea to treat this and any future work conducted by these researchers with an appropriately greater level of scrutiny and skepticism.

Hyperbaric oxygen therapy alleviates vascular dysfunction and amyloid burden in an Alzheimer's disease mouse model and in elderly patients

Vascular dysfunction is entwined with aging and the pathogenesis of Alzheimer's disease (AD), and contributes to reduced cerebral blood flow (CBF) and consequently, hypoxia. Hyperbaric oxygen therapy (HBOT) is in clinical use for a wide range of medical conditions. In the current study, we exposed 5XFAD mice, a well-studied AD model that presents impaired cognitive abilities, to HBOT and then investigated the therapeutical effects. HBOT increased arteriolar luminal diameter and elevated CBF, thus contributing to reduced hypoxia. Furthermore, HBOT reduced amyloid burden by reducing the volume of pre-existing plaques and attenuating the formation of new ones. This was associated with changes in amyloid precursor protein processing, elevated degradation and clearance of amyloid-ß protein and improved behavior of 5XFAD mice. Hence, our findings are consistent with the effects of HBOT being mediated partially through a persistent structural change in blood vessels that reduces brain hypoxia.

To understand whether the ability of HBOT to change CBF and affect cognitive function also applied to elderly people, we performed a human study in which six elderly patients (age 70.00 ± 2.68 years) with significant memory loss at baseline (memory domain score < 100) were treated with HBOT (60 daily HBOT sessions within 3 months). CBF and cognitive function were evaluated before and after HBOT. CBF was measured by MRI, while cognitive functions were evaluated using computerized cognitive tests. Following HBOT, there were significant CBF increases in several brain areas.

At baseline, patients attained a mean global cognitive score (102.4±7.3) similar to the average score in the general population normalized for age and education level (100), while memory scores were significantly lower (86.6 ± 9.2). Cognitive assessment following HBOT revealed a significant increase in the global cognitive score (102.4 ± 7.3 to 109.5 ± 5.8), where memory, attention and information processing speed domain scores were the most ameliorated. Moreover, post-HBOT mean memory scores improved to the mean score (100.9 ± 7.8), normalized per age and education level (100). The improvements in these scores correlate with improved short and working memory, and reduced times of calculation and response, as well as increased capacity to choose and concentrate on a relevant stimulus.

The analysis of the effects of genetic variants on human life expectancy has employed ever large databases in recent years: more genes, more sequences, more people. As the data grows, the likely size of the effect of genetic variation on human longevity has become smaller. Outside of a few interesting genes, such as those relating to blood cholesterol levels and cardiovascular disease risk, he picture is one of countless variants with small, interacting, environment-dependent effects, different in every study population.

How much of this picture is a true assessment versus a consequence of larger effects being hidden in the interactions between gene variants? Past studies have near all focused on a variant by variant analysis, considering each variant alone - and so this is an interesting question. Interesting or not, it remains the case that there may be no practical application here, however. Old people are still aged, damaged, and increasingly frail, whether or not they carry rare gene variants associated with longevity. Finding ways to emulate survivors to old age is an inherently poor approach to the treatment of aging, at least in comparison to working towards the repair of the underlying molecular damage that causes aging, in order to produce rejuvenation.

A major goal of aging research is identifying genetic targets that could be used to slow or reverse aging - changes in the body and extend limits of human lifespan. However, the majority of genes that showed the anti-aging and pro-survival effects in animal models were not replicated in humans, with few exceptions. Potential reasons for this lack of translation include a highly conditional character of genetic influence on lifespan, and its heterogeneity, meaning that better survival may be result of not only activity of individual genes, but also gene-environment and gene-gene interactions, among other factors.

In this paper, we explored associations of genetic interactions with human lifespan. We selected candidate genes from well-known aging pathways (IGF1/FOXO growth signaling, P53/P16 apoptosis/senescence, and mTOR/SK6 autophagy and survival) that jointly decide on outcomes of cell responses to stress and damage, and so could be prone to interactions. We estimated associations of pairwise statistical epistasis between SNPs in these genes with survival to age 85+ in the Atherosclerosis Risk in Communities study, and found significant effects of interactions between SNPs in IGF1R, TGFBR2, and BCL2 on survival to age 85 and older. We validated these findings in the Cardiovascular Health Study sample, using survival to age 85+, and to the 90th percentile, as outcomes.

Our results show that interactions between SNPs in genes from the aging pathways influence survival more significantly than individual SNPs in the same genes, which may contribute to heterogeneity of lifespan, and to lack of animal to human translation in aging research.

Link: https://doi.org/10.3389/fcell.2021.692020

Researchers here report on a broad comparison of protein sequences across many mammalian species, conducted in order to search for small differences between individual proteins that correlate with species life span. They find that humans, as one of the longer-lived mammals, already have most of these differences present across most of the the population. Further, the nature of these differences between proteins, meaning the specific functions of differing proteins in cell metabolism, is argued to support the hypothesis that quality control processes responsible for maintaining protein structure and removing damaged proteins make a sizable contribution to species differences in life span.

A key mechanism that may contribute to differences in lifespan between species is the maintenance of the proteostasis network. Protein stability or proteostasis refers to the capacity to protect protein structures and functions against environmental stressors, including aging. In fact, dysfunction of the protein quality control mechanisms is a hallmark of aging and there is substantial evidence linking proteostasis and longevity. For instance, improved protein stability is determinant for longevity in exceptionally long-lived mollusks and in the naked mole-rat, the longest-living rodent. In addition, interventions that enhance proteome stability can improve health or increase lifespan in model organisms, such as pharmacological chaperones that have been investigated as potential therapeutic targets to reduce the adverse effects of misfolding of aging-related proteins.

A mammalian-wide study of the genomic underpinnings of lifespan has never been carried out with the combined goals of identifying individual mutations linked to longevity; analyzing the functional properties of their genes and the pathways in which they take part; and studying how the stability of proteins coded by these genes may differentiate long- and short-lived species. Here, we performed the largest phylogeny-based genome-phenotype analysis to date, focusing on the detection of individual mutations and genes that underlie the enormous variation of lifespan in mammals. We report the discovery of more than 2,000 longevity-related genes and show that, overall, they present a trend towards increased protein stability in long-lived organisms. In addition, we successfully show that our findings enhance the interpretation of the results of longevity genome-wide association studies that have been carried out in humans.

We discovered a total of 2,737 single amino acid differences (AA) in 2,004 genes that distinguish long- and short-lived mammals, significantly more than expected by chance. These genes belong to pathways involved in regulating lifespan, such as inflammatory response and hemostasis. Among them, a total 1,157 AA showed a significant association with maximum lifespan in a phylogenetic test. Interestingly, most of the detected AA positions do not vary in extant human populations (81.2%) or have allele frequencies below 1% (99.78%). Consequently, almost none of these putatively important variants could have been detected by genome-wide association studies. Additionally, we identified four more genes whose rate of protein evolution correlated with longevity in mammals. Crucially, SNPs located in the detected genes explain a larger fraction of human lifespan heritability than expected, successfully demonstrating for the first time that comparative genomics can be used to enhance interpretation of human genome-wide association studies. Finally, we show that the human longevity-associated proteins are significantly more stable than the orthologous proteins from short-lived mammals, strongly suggesting that general protein stability is linked to increased lifespan.

Link: https://doi.org/10.1093/molbev/msab219

Rejuvenation takes place very early in embryonic development. The germline cells that go into the creation of an embryo are well protected and maintained in comparison to the average somatic cell in the adult body. Nonetheless, there is an accumulation of age-related epigenetic changes and molecular damage. Cells purge themselves of as much of this change and damage as possible, in order to ensure that the young are born with young somatic cells and tissues. This is primarily a resetting of epigenetic controls over gene expression, decorations on the structure of the genome that control shape and access to specific genes by the molecular machinery responsible for producing proteins from genetic blueprints.

A cell is a state machine, largely governed in operation by the matter of which proteins are produced, and in what quantities. Not completely governed: some damage, such as mutations to nuclear DNA, is irreversible. Some molecular waste cannot be managed even by cells in a youthful epigenetic state, and will degrade normal function. In a collection of replicating cells, that waste can be diluted via cell division, or even passed off entirely to a sacrificial daughter cell in a process of asymmetric division. So long as no one cell or small number of cells are vital, even serious mutation can be evaded by replication, provided that mutated cells are rejected. This is how single celled life, such as bacteria, can continue indefinitely. Further, a few lower organisms, such as the hydra, essentially a tiny bundle of stem cells in which every structure is replaceable, use this strategy in order to achieve individual immortality. Higher animals, with complex central nervous systems that include many non-replicating cells that cannot be sacrificed, cannot use this strategy, and so suffer from degenerative aging.

Embryonic rejuvenation is a process that can be understood, induced, and manipulated. The creation of induced pluripotent stem cells from normal adult somatic cells via reprogramming is one example of what becomes possible given sufficient knowledge and technical aptitude. This combines, in the same way as occurs in the early embryo, both an epigenetic reset and loss of somatic cell state, such as the shape and function of a skin cell or a brain cell, producing dedifferentiation into a pluripotent stem cell state. Researchers are presently looking beyond experiments in cell cultures towards the application of reprogramming in living animals. An epigenetic reset is a desirable outcome for somatic tissues throughout the aged body, likely able to reverse to some degree many age-related issues, such as loss of mitochondrial function. Dedifferentiation of somatic cells in an adult individual, on the other hand, is a roadblock and a challenge. It will lead to cancer where it occurs to a lesser degree, and it will cause pathology and death if prevalent. Differentiated cell state is vital to normal tissue function.

Thus an important question currently under investigation is whether or not these two aspects of reprogramming are inseparable. Is there an approach to reprogramming that will produce maximal epigenetic rejuvenation with minimal dedifferentiation? If so, that could prove to the the basis for a very useful approach to the treatment of aging. It likely cannot help much in the case of stochastic nuclear DNA damage leading to somatic mosaicism, and it cannot help with the accumulation of some forms of persistent molecular waste in long-lived cells, but it could nonetheless be beneficial enough to be interesting.

Cellular reprogramming and epigenetic rejuvenation

A recent addition to the anti-ageing strategies being developed comes from cellular reprogramming approaches. Induced pluripotency studies provided evidence that age-related cellular phenotypes such as mitochondrial morphology, function and number, as well as nuclear envelope integrity, are not irreversible. However, developmental cellular reprogramming turns a cell to a pluripotent state, where it has the potential to generate any somatic cell type. This process is not appropriate for an anti-ageing therapy in vivo because it requires not only the loss of the original cellular identity, but also the re-establishment of self-renewal capabilities. Therefore, induction of pluripotency or the direct injection of pluripotent cells in vivo, invariably lead to cancer in mice. For a cellular reprogramming-based intervention to be considered rejuvenative (turning an old cell into a younger cell), we need to uncouple its effects from dedifferentiation (loss of somatic cell identity).

Cellular reprogramming has demonstrated potential not only in regenerative medicine, but also in the ageing field through the amelioration of both physiological and cellular ageing hallmarks. While partial reprogramming might be used as a catch-all term to describe this type of rejuvenation, it does not reflect the fact that the described interrupted cellular reprogramming techniques are applied with the aim of (epigenetic) rejuvenation as opposed to inducing pluripotency (loss of cell identity). Reprogramming-induced rejuvenation (RIR) is a better term, capturing the nature of the utilised process and final aim of the interventions.

RIR has shown promise as a treatment to safely reverse ageing whilst retaining the ability to revert to or maintain original cell identity, both in vivo and in vitro. However, the precise nature of RIR still needs to be fully understood before it can be safely implemented as an anti-ageing treatment. For example, tracking any traces of pluripotency in partially reprogrammed cells (particularly in vivo) is a necessary precaution to minimise long-term cancer risk. Additionally, can rejuvenated partially reprogrammed cells be cultured long-term? The rejuvenated phenotype of some OSKM-treated cells lasts at least four weeks, but does this phenotype remain stable or eventually start to deteriorate at a rate faster than normal ageing?

Other important RIR safety concerns include how the reprogramming factors are introduced in vivo. Retroviruses are commonly used to integrate reprogramming factors into the genome. However, this method bears risks, such as insertional mutagenesis, residual expression and re-activation of reprogramming factors, and retrotransposon activation, all of which could increase cancer risk in vivo. Non-integrative delivery methods, such as transient transfection, non-integrating viral vectors, and RNA transfection are safer alternatives. For example, researchers have successfully used mRNA transfection to non-integratively conduct RIR. Another safe alternative is chemical-based reprogramming, which involves direct conversion of a somatic cell to a pluripotent state by use of small molecules and growth factors. It is conceivable that, in the future, chemical-based reprogramming could be adapted to achieve rejuvenation, however, this reprogramming approach currently only works for mice.

While RIR applied to skeletal muscle stem cells appears effective in improving regenerative capacity and muscle function in immunocompromised mice, further analysis is required regarding the somatic mosaicism of partially reprogrammed stem cells. Somatic variants at a stem or early progenitor cell level in turn can cause lineage bias, reduced stem cell function, and increased risk of developing haematologic cancer (e.g. age-related clonal haematopoesis). This can lead to the development of pre-malignant cells, which have a higher propensity to transform to a malignant state, the effect of which could be attenuated or exacerbated by RIR.

It also remains to be further explored whether and how RIR would work on post-mitotic terminally differentiated cells, such as neurons, cardiomyocytes, or adipocytes, but also other non-dividing cells such as quiescent or senescent cells. Pilot work has been done in the latter two states, demonstrating that a rejuvenated phenotype is achievable after restoration of cell division. These results may point to a scenario where proliferation is an essential requirement for rejuvenation. Indeed, induced pluripotency of postnatal neurons was only possible after forced cell proliferation via p53 expression. Coincidentally, the natural rejuvenation event in the early mouse embryo spans over stages of very active cell proliferation.

Overall, RIR is currently the best prospect to achieve epigenetic rejuvenation. Further studies are required to fully determine its limitations and efficacy.

Loss of hair cells in the inner ear is thought to be the primary mechanism behind the progression of age-related hearing loss, though there is some debate over whether it is in fact loss of cells versus loss of the connections that link hair cells to the brain. For some years, the research community has investigated whether or not it is possible to generate new hair cells in a living animal, bypassing the usual inability to replace losses in this cell population. Various approaches to signaling and cell therapy have been attempted, but despite interesting technology demonstrations, there is as yet little progress towards clinical translation of this research.

Various mechanisms can cause sensorineural hearing loss, among which irreversible damage to inner ear hair cells is the main cause. Although the commonly used hearing aids and cochlear implants in clinical practice improve the hearing of patients, their effect depends on the quantity and quality of residual hair cells and spiral neurons. Therefore, the ideal way to treat sensorineural hearing loss is to regenerate hair cells, through the use of stem cells to repair the structure and function of the cochlea, so as to fundamentally restore hearing.

Stem cell therapy in the auditory field has been a research hotspot in recent years. Although some progress has been made, almost all are results at the animal level, and there is still a long way to go before clinical transformation. The microenvironment of inner ear stem cells and the interaction with neighboring cells are very important for inner ear stem cells or sensory precursor cells to induce differentiation into mature inner ear hair cells. In the reported studies, the efficiency of differentiation of inner ear stem cells or sensory precursor cells into hair cells is still low. An insufficient number of new hair cells, immature new hair cells without the function of mature hair cells, and long-term survival of new hair cells are all key problems and difficulties that need to be solved urgently.

These results indicate that it is more difficult to regulate a single signal pathway to regenerate functional hair cells, and it may require coordinated regulation of multiple genes to effectively promote hair cell regeneration and the functional maturity and survival of new hair cells. Still, inducing the committed differentiation of stem cells into hair cells or nerve cells, the exploration of the methods of stem cell transplantation into the inner ear, and the safety research of stem cell transplantation have collectively laid the foundation for the transplantation of stem cells in vivo.

Link: https://doi.org/10.3389/fncel.2021.732507

Researchers here note that reduction in the calorie content in the diet is the important trigger for the benefits of calorie restriction, not reductions in the quantity of food ingested. The practice of calorie restriction, reducing calorie intake while still obtaining optimal micronutrient intake, has been shown to extend life span in near all species and lineages tested to date. Firm data on human life span has yet to be obtained, and is expected to be modest, on the order of a few years only, but short-term beneficial changes to the operation of metabolism are quite similar between mice and humans. Research into calorie restriction has given rise to a broad field of development of calorie restriction mimetic drugs, targeting many of the same cellular stress response mechanisms that are triggered by a low calorie intake. One shouldn't expect miracles from this line of work: we know the limits of calorie restriction in humans, and while it is certainly beneficial, it doesn't greatly change the duration of a human life.

Although calorie restriction has been reported to extend lifespan in several organisms, animals subjected to calorie restriction consume not only fewer calories but also smaller quantities of food. Whether it is the overall restriction of calories or the coincidental reduction in the quantity of food consumed that mediates the anti-aging effects is unclear. Here, we subjected mice to five dietary interventions. We showed that both calorie and quantity restriction could improve early survival, but no maximum lifespan extension was observed in the mice fed isocaloric diet in which food quantity was reduced.

Mice fed isoquant diet with fewer calories showed maximum lifespan extension and improved health among all the groups, suggesting that calorie intake rather than food quantity consumed is the key factor for the anti-aging effect of calorie restriction. Midlife liver gene expression correlations with lifespan revealed that calorie restriction raised fatty acid biosynthesis and metabolism and biosynthesis of amino acids but inhibited carbon metabolism, indicating different effects on fatty acid metabolism and carbohydrate metabolism. Our data illustrate the effects of calories and food quantity on the lifespan extension by calorie restriction and their potential mechanisms, which will provide guidance on the application of calorie restriction to humans.

Link: https://doi.org/10.18632/aging.203493

The processes of autophagy act to remove damaged molecular machinery and structures in the cell. Autophagy becomes dysfunctional with age, however. This is likely downstream of underlying causes of aging that cause changes in gene expression that degrade the function of autophagic processes in one way or another. For example mitophagy, the clearance of damaged mitochondria by autophagy, is indirectly negatively impacted by changes in mitochondrial dynamics resulting from altered gene expression. Equally, age-related changes in gene expression produce defects in the formation of autophagosomes, and this affects all aspects of autophagy.

Many of the known interventions that slow aging in animal models appear to improve the efficiency of autophagy, and functional autophagy is required for the extension of healthy life span via calorie restriction to take place. While improvement of autophagy has been a goal in the research community for quite some time, surprisingly little concrete progress has been made towards the development of therapies that specifically target dysfunction in autophagic processes.

Calorie restriction mimetics such as mTOR inhibitors improve autophagy, and mitochondrially targeted antioxidants and NAD+ upregulation may act to restore mitophagy. These were not designed with the enhancement of autophagy in mind; rather, it has been found to be one of their outcomes. The research and development communities have yet to see success in the development of narrowly targeted means of restoring a youthful function of autophagy in old tissues, though a few groups, such as the startup Selphagy Therapeutics that emerged from work on LAMP2A upregulation in the liver, are working in that area.

Selective Autophagy as a Potential Therapeutic Target in Age-Associated Pathologies

Cellular garbage disposal is critical for recycling defective cell constituents, such as proteins and organelles, towards the maintenance of cellular homeostasis. One of the main degradative molecule pathways is autophagy, which is a physiological catabolic process shared by all eukaryotes. Derived from the Greek words 'auto' meaning self, and 'phagy', meaning eating, autophagy, it was initially considered to be a bulk degradation process, while now its highly selective nature is increasingly appreciated. This self-digestive mechanism relieves the cell from proteotoxic, genotoxic, oxidative, and nutrient stress. It is accomplished in an intricate stepwise manner, which leads to clearance of damaged cell constituents, in the degradative organelle, the lysosome. Failure to complete this procedure has been implicated in many age-related diseases.

Homeostatic mechanisms that respond to mitochondrial damage are less efficient during aging. Mitophagy is a physiological eukaryotic pathway, which involves the degradation of superfluous or damaged mitochondria. When perturbed, normal mitochondrial function is hindered, resulting in the production of excessive reactive oxygen species (ROS). This ultimately leads to cellular dysfunction and tissue damage. Defective mitophagy is evident in a variety of age-related pathologies such as neurodegeneration, metabolic syndromes, and myopathies.

Aggrephagy degrades aggregation-prone proteins via targeted macroautophagy, in addition to chaperone-mediated autophagy and the proteasomal pathway. These proteins typically form aggresomes near the nucleus, which are surrounded by intermediate filament cytoskeleton, and are further processed to be degraded by autophagy. Protein aggregation usually occurs due to misfolding and can cause, among others, dysregulation of calcium homeostasis, inflammation, neurotoxicity.

Recycling of peroxisomes is also regulated by autophagy. These small dynamic single membrane organelles regulate fatty acid oxidation, production of bile acid and other lipids, while also producing ROS, which is neutralized by catalase. Moreover, peroxisomes interact with a multitude of other cellular constituents, such as lipids, the endoplasmic reticulum (ER), and mitochondria. Pexophagy and peroxisome biogenesis have recently been implicated with disease. During aging, peroxisomal targeting signal 1 (PTS1) protein import deteriorates and catalase function is diminished. Peroxisomes become more abundant and PEX5 accumulates on their membranes. This causes increased production of ROS, which further blocks peroxisomal protein import and contributes to aging.

With regard to therapeutic intervention, several pharmacological compounds have been shown to activate mitophagy and alleviate symptoms of age-related diseases, dependent on dysfunctional mitochondria. Rapamycin activates AMPK, while blocking mTOR, maintaining energetic demands and preventing neurological symptoms, such as neuroinflammation. Metformin and pifithrin induce Parkin by inhibiting p53 activity and alleviating diabetic phenotypes. Resveratrol, mainly found in grape skin, as well as, NAD+ precursors found in natural compounds activate mitophagy and mitochondrial biogenesis through the sirtuin 1 (SIRT1)-PGC-1α axis. Urolithin A, an intestinal microbiome-derived metabolite from dietary intake, induces both mitochondrial degradation and biogenesis, and increases health span of model organisms such as C. elegans and mice.

Selective autophagic induction by genetic intervention or chemical compound administration is currently being investigated in multiple diseases as potential therapeutic approach, although no drug has reached the clinic yet. Indeed, clinical studies concerning druggable autophagy targets remains limited. This highlights the complexity and intricacies of selective autophagic pathways, which in humans, cannot be easily targeted due to context-dependence and extensive crosstalk with other functional networks. Thus, initial optimism has subsided, with research now focusing on specific compounds that could target specific aspects of selective autophagy. An important objective of the collective efforts of the research community and pharmaceutical companies is to achieve targeting selective autophagy mediators, while not affecting other cellular processes.

Are there many strategies that can reverse cellular senescence? There are certainly strategies that can lower levels of cellular senescence over time, both in cell cultures and in living animals, but very few are actually reprogramming senescent cells into normal cells. It isn't clear that this reversal of the senescent state is a good idea, given that there is usually a good reason for at least some of such cells to be senescent, such as potentially cancerous mutations. The strategy described here is probably not causing senescent cells to become normal cells in any great number, but rather lowering the rate at which cells become senescent or encouraging senescent cells to self-destruct more rapidly, as well as encouraging normal cells to replicate more rapidly, thus diluting the senescent fraction of the population.

Cellular rejuvenation occurs naturally in embryonic development when sperm and egg (each having a certain chronological age) fuse to each other to form an embryo of age zero. Similarly, reprogramming of somatic cells to pluripotency, producing induced pluripotent stem cells (iPSCs), resets their biological clock as well. At this stage, a core network of transcription factors including NANOG, OCT4, and SOX2 maintains pluripotency in embryonic stem cells (ESCs) and iPSCs. In particular, the pluripotency factor NANOG is essential for maintaining the self-renewal of ESCs over many population doublings.

Although overexpression of NANOG does not confer pluripotency to somatic cells, it has been shown to restore several cellular functions that are compromised by aging including proliferation and differentiation of senescent fibroblasts and mesenchymal stem cells. In vivo endogenous expression of this transcription factor in stratified epithelia of adult mice showed that systemic overexpression of NANOG induces hyperplasia without initiating tumors.

Recently, we discovered that expression of NANOG in myoblasts restored their myogenic differentiation potential, as evidenced by expression of myogenic regulatory factors and the ability to form myotubes, which was impaired by replicative senescence. This result prompted us to investigate the anti-aging effects of NANOG on primary human myoblasts and in skeletal muscle tissue in vivo. Here, we show that overexpression of NANOG reversed the hallmarks of cellular senescence in muscle progenitors in vitro and restored the satellite cell abundance in the skeletal muscle of progeroid mice.

Link: https://doi.org/10.1126/sciadv.abe5671

Researchers here demonstrate that introducing an antioxidant into the diet, one that accumulates in cell lysosomes, helps to prevent macrophage dysfunction and thus reverse atherosclerotic plaque in an animal model of atherosclerosis. The hypothesis is that oxidized LDL particles, ingested and carried to lysosomes for degradation, are an important component of dysfunction in the macrophage cells responsible for clearing out lipid accumulations in blood vessel walls. Macrophages function well in youth, but are challenged and made dysfunctional in later life by the age-related increase in levels of oxidation of lipids and lipid carriers such as LDL particles. Strategies in clinical use to slow atherosclerosis have so far not directly targeted this challenge of oxidation and macrophage function, which may well be why they are of only limited benefit.

Multiple studies suggest that the presence of lysosomal cholesterol accumulation in macrophages, and not the total amount of intracellular lipids, is critical for the observed inflammatory response. We have shown that lysosomes in macrophages are a site of low-density lipoprotein (LDL) oxidation. Seven days after taking up mechanically aggregated LDL or sphingomyelinase aggregated LDL, mouse or human macrophage-like cells and human monocyte-derived macrophages generated ceroid in their lysosomes. Ceroid (lipofuscin) is a polymerized product of lipid oxidation found within foam cells in atherosclerotic lesions.

The lysosomal oxidation of LDL is catalyzed by oxidation-reduction active iron present in the lysosomes of macrophages through the generation of hydroperoxyl radicals at the lysosomal pH of 4.5. This oxidation is inhibited by cysteamine (2-aminoethanethiol), an antioxidant that accumulates in lysosomes. Cysteamine is used in patients for the lysosomal storage disease cystinosis, caused by the absence of the lysosomal cystine transporter cystinosin. Recently, we have shown that cysteamine reduces atherogenic conditions caused by lysosomal LDL oxidation, such as lysosomal dysfunction, cellular senescence, and secretion of various proinflammatory cytokines, such as interleukin-1β, TNF-α, and interleukin-6, and chemokines, such as CCL2, in human macrophages.

LDL receptor-deficient mice were fed a high-fat diet to induce atherosclerotic lesions. They were then reared on chow diet and drinking water containing cysteamine or plain drinking water. Aortic atherosclerosis was assessed, and samples of liver and skeletal muscle were analyzed. There was no regression of atherosclerosis in the control mice, but cysteamine caused regression of between 32% and 56% compared with the control group, depending on the site of the lesions. Cysteamine substantially increased markers of lesion stability, decreased ceroid, and greatly decreased oxidized phospholipids in the lesions. The liver lipid levels and expression of cluster of differentiation 68, acetyl-coenzyme A acetyltransferase 2, cytochromes P450 (CYP)27, and proinflammatory cytokines and chemokines were decreased by cysteamine. Skeletal muscle function and oxidative fibers were increased by cysteamine. There were no changes in the plasma total cholesterol, LDL cholesterol, high-density lipoprotein cholesterol, or triacylglycerol concentrations attributable to cysteamine.

In conclusion, inhibiting the lysosomal oxidation of LDL in atherosclerotic lesions by antioxidants targeted at lysosomes causes the regression of atherosclerosis and improves liver and muscle characteristics in mice and might be a promising novel therapy for atherosclerosis in patients.

Link: https://doi.org/10.1161/JAHA.120.017524

The scientific community is very broad, and there are many groups within that community whose members intermittently produce studies that are either poorly designed, poorly conducted, or poorly presented and explained. Or all three, for all of the usual reasons. Constraints of time and funding, institutional pressure to publish, the involvement of external interests, and so forth. Bad papers do get published, provided that the authors are subtle enough. This does tend to be a self-correcting problem, when considered over a sufficiently long span of time to allow errant individuals and institutions to blacken their reputations with the community at large. Still, at any given moment, one should expect to see that some small fraction of published scientific papers are problematic, rather than merely incorrect.

The problematic paper for today's discussion was published last year, reporting on a study of the effects of hyperbaric oxygen treatment on areas of metabolism that are connected to the study of aging. At the time, claims of reversal of aging were circulating in the media. The paper itself was of poor quality, but far less offensive than the related and entirely unfounded hype. It was the usual circus of ignorant commentary, yes, but also a matter of hyperbaric oxygen treatment providers pushing claims that were completely unsupported by the evidence. Serious researchers will think twice about working with anyone who was involved in this exercise. I talked about this a little at the time, focusing as much on the ridiculous claims being made by institutions involved in the work, and by the media at large, as on issues with the study and interpretation of data. Relatedly, I see that the SENS Research Foundation team have chosen to pick apart the scientific details in a recent article. A little more shaming can't hurt in this case!

Hyperbolic Hyperbaric "Age Reversal"

Lower-quality, clickbait-hungry media outlets love sensationalist claims, but one does expect better from the public relations department of an internationally-respected research university. And it was an easy jump from the already-overstated "In First, Aging Stopped in Humans" and "treatments can reverse two processes associated with aging and its illnesses" to saying that a treatment "can reverse aging process" - and to then land in a mud-pit of self-parody with "Human ageing reversed in 'Holy Grail' study, scientists say."

The actual findings of a recent study on hyperbaric oxygen treatment (HBOT) were much more limited. Despite some intriguing indicators, the actual impact of HBOT on aging based on this study is entirely unclear, quite plausibly negligible, and in any case objectively less impressive than that of (say) regular exercise, which certainly does not "reverse aging."

The actual details of the study show that even the narrow claims of the study abstract aren't fully justified. It's not clear that blood-cell telomeres were lengthened any more than they would have been without HBOT; it's not clear that "senescent" T-cells were reduced in numbers, let alone actually destroyed; and if "senescent" T-cells had been destroyed, it would not demonstrate a senolytic effect of HBOT. Despite the fact that it's standard terminology in the immunology world, "senescent" T-cells aren't actually "senescent cells" in the sense usually used in the geroscience world. Jumping from post-HBOT reductions in the number of these "senescent" T-cells to potential effects on classical senescent cells is really just a misunderstanding of what kinds of cells are involved in each case.

Even if the study had robustly demonstrated that every one of the points above really did occur, it would not constitute "reversing aging" - or even justify the more restrained claims that "blood cells actually grow younger as the treatments progress" or "that the aging process can in fact be reversed at the basic cellular-molecular level."

Aging has comparatively simple root causes, forms of cell and tissue damage that accumulate as a side-effect of the normal operation of metabolism. These comparatively simple causes take effect on a very, very complex system, however. The result is an intricate web of interacting consequences, and ultimately a dysfunctional, failing mess in which it is very hard to pinpoint which of the countless observed mechanisms are actually important. The complexity of the outcome is a result of the complexity of a living organism, not of the complexity of the root causes of aging. Metabolism is incompletely understood, and for so long as that is the case, inspecting the progression of aging will continue to reveal new subtleties. This is why interventions should focus on the causes of aging, far better understood at the present time, and not on manipulating later stages of the process, much of which remains a dark forest.

Researchers have made a surprising discovery about the connection between protein shape and mitochondrial health, providing a piece of evidence for yet another theme in aging research: it's always more complicated than we thought. Proteins within the mitochondria are intricately involved in mitochondrial function, and are protected by the mitochondrial unfolded protein response (UPRmt). When proteins misfold in the mitochondria, which can be caused by external threats like pathogens or mitochondrial toxins, the UPRmt gets activated which helps restore protein shape and function. Past research on the microscopic worm C. elegans has demonstrated that boosting the UPRmt during development contributes to better mitochondrial health and a longer lifespan for the worms.

Consistently, pharmacologically boosting UPRmt has been shown to slow down diseases with mitochondrial dysfunction, such as Alzheimer's. The new research has found that activating the UPRmt in adult worms has the opposite effect: adult worms with a boosted unfolded protein response have worse health and a shorter lifespan. Digging into the details of this surprising outcome led the team to examine the mitochondrial permeability transition pore. Most of the time this pore is closed, keeping the interior of the mitochondria separate from the rest of the cell. Under stress, though, it opens to release calcium into the rest of the cell, signaling that it's time to cut its losses and induce cell death. It turns out that methods to boost the UPRmt in adult C. elegans are caused indirectly - the UPRmt is initiated in response to the opening of the transition pore. While the UPRmt is busy trying to clean things up, the signals coming from the opened pore are too strong for the cell to ignore and result in cell death. Researchers think this is what contributes to the early death of the adult worms.

Research in C. elegans forms the basis of much aging research, but what does this mean for efforts to boost health and prevent disease in people? While the mitochondrial permeability transition pore is already implicated in conditions like stroke and heart attack, the role of the UPRmt is not as well understood. Researchers liken the UPRmt to inflammation, which has a specific purpose and is useful under some conditions, but causes damage under others. One possibility is that, in a stressed cell, the UPRmt uses valuable cellular resources, hastening the already inevitable cell death.

Link: https://www.buckinstitute.org/news/aging-its-more-complicated-than-we-thought/

Immunotherapies that have successfully targeted amyloid-β have failed to help Alzheimer's patients to any meaningful degree. This may be because amyloid-β is only relevant in the earliest stages of the condition, or because the most visible amyloid-β aggregation outside cells is a side-effect of neurodegeneration rather than a core disease process. The research community has in recent years turned increasing attention to immunotherapies that target tau aggregation, characteristic of the later stages of Alzheimer's disease. There appears to be a bidirectional relationship between neuroinflammation and the accumulation of toxic, altered forms of tau. As noted here, targeting tau in human trials is starting to produce data that is more suggestive of patient benefits. Still, this is a painfully incremental process, and the results are still only a marginal improvement. We can hope that targeting inflammatory processes, such as those connected to senescent supporting cells in the brain, may produce better outcomes.

In a first for the field, there is now a hint that a tau immunotherapy may have slightly benefited people with Alzheimer's disease (AD). Semorinemab, a monoclonal antibody specific for tau's N-terminus, stemmed cognitive decline by almost half among people with mild to moderate AD, according to topline results from the Phase 2 LAURIET trial. The findings are a welcome reprieve after most tau immunotherapies thus far, including semorinemab itself, have come up short in trials. In a previous Phase 2 study, called TAURIEL, semorinemab brought no cognitive or functional benefit to people with prodromal AD or mild cognitive impairment. Despite this negative result among people in earlier stages of the disease, the companies moved forward with LAURIET, which enrolled participants in the mild to moderate stages of AD.

LAURIET enrolled 272 participants whose Mini-Mental State Examination (MMSE) scores were between 16 and 21 and who had brain amyloid at baseline. After receiving three doses spaced two weeks apart, participants received monthly intravenous infusions of semorinemab, or placebo, for the remainder of the trial. The study enrolled two cohorts, which received treatment for either 48 or 60 weeks. For both enrolled cohorts, between baseline and 49 weeks, those in the semorinemab groups declined 43.6 percent less on the ADAS-Cog11 than did those in the placebo groups, satisfying one primary outcome. The same was not true for the other primary endpoint, the Alzheimer's Disease Cooperative Study-Activities of Daily Living. Both groups declined similarly on the ADCS-ADL, in which an appointed caregiver scores the participant on how they perform a variety of tasks.

Why a possible efficacy signal in LAURIET, but not a peep in TAURIEL? Researchers think that the difference could come down to which species of tau predominate in different stages of the disease. Perhaps specific hyperphosphorylated forms drive the earlier stages of disease, and semorinemab might not bind them.

Link: https://www.alzforum.org/news/research-news/first-cognitive-signal-tau-immunotherapy-works

The Longevity.technology team has been publishing notes on the recent Aging Research and Drug Discovery (ARDD) conference, one of the few events at the end of this year to be held in person again after the long pandemic hiatus. It was a challenge for conference organizers to look into the crystal ball six to twelve months in advance and commit to a late 2021 event, but some did. The 2022 conference season will no doubt be interesting, as restrictions relax sufficiently for reliable travel and advance scheduling, and a few years of the suppressed urge to network is finally unleashed. The ARDD conferences are more focused on academic science and the established pharmaceutical industry than on investment and entrepreneurship, at least in comparison to say, the Undoing Aging conference series, but it can nonetheless be an interesting event for both investors and entrepreneurs.

For my part, I feel that there is too much of a focus on incremental, unambitious programs and interventions, such as metformin and other calorie restriction mimetics. These unambitious programs may turn out to provide sufficient proof of principle, in animal studies and preclinical drug discovery, for those leaders in the pharmaceutical industry who still find the idea of treating aging a novelty. The outcomes in the clinic will be modest to the point of pointlessness, however. No-one should be spending billions on the development of drugs that cannot in principle do any more for health than exercise and or a reduction in calorie intake. The opportunity cost here is vast, a loss of attention and effort that should have gone towards the development of true rejuvenation therapies with potentially sizable effects on health and life span. This is a complaint that can be directed at the field as a whole, and ARDD only happens to be a representative sampling of that field.

ARDD 2021 kicks off: sessions to cover longevity topics from AI and senolytics to mitochondria and stem cell rejuvenation.

With the goal of building better connections between pharma companies and the longevity field, ARDD's organisers are conscious of the importance of showcasing the most credible research and development in aging. "People want to believe in longevity, but it's important to maintain a stringent a scientific focus as possible. We haven't shown a single molecule working in humans and, while we've been able to slow aging down in mammals, we have never been able to stop it. But I'm very enthusiastic and positive about some of these challenges that we're facing. Years ago, the aging field was many disconnected fields. But the industry and the science has started taking shape, and the conference has evolved through the integration of multiple fields with key hallmarks of aging."

Lots to learn about longevity at ARDD 2021: from pharma and startups to physicians and high schoolers.

The theme of education continues throughout this year's conference, which has its roots in bridging the gap between the longevity field and major pharmaceutical companies. "From the very beginning, we have tried to make this conference very friendly towards the pharmaceutical industry, which needs to buy into this field. There has been a lot of activity in early stage drug discovery in the pharmaceutical industry, specifically on aging and age-associated biology that can be purposed towards multiple diseases."

Focus on frailty over aging

Anne-Ulrike Trendelenburg from the Novartis Institutes for Biomedical Research gave a brilliant presentation on pathways that should be targeted to treat age-related diseases, and how her research is shifting away from aging and towards defining and targeting frailty. Novartis' research has highlighted five recommendations to guide the future of age-related disease targeting. Firstly, focusing on healthspan as opposed to lifespan, which will have the biggest impact on patient lives, adding quality of life to years, over adding years to life. Trendelenburg also highlighted targeting multimorbidity, rather than continuing to focus on treating each individual disease, which is said to be ineffective and expensive. The team from Novartis want to develop better multimorbidity models in order to progress another of their recommendations, which is to target frailty over age through clinical trials.

ARDD 2021: DNA repair, mitochondrial enhancement, gene editing, and how a new era for longevity will help us beat age-related disease.

The day showcased a variety of longevity research, we heard more about the importance of focussing on mitochondrial targets in relation to finding age-related therapies. Karl Lenhard Rudolph from the Leibniz Institute on Aging spoke on mitochondrial enhancement and its relationship with late life dietary restriction, enabling improvements in stem cells and lifespan. While recognising the positive effects seen with dietary restriction across organisms, Rudolph addressed the worrying data that saw this method losing efficacy when restriction is started late in life. When looking at reduction of mortality rates lifelong dietary restriction shows a reduction in mortality rate, however late life dietary restriction has very little effect.

The combination of dasatinib and quercetin was the first practical senolytic therapy explored in mice and human trials. Treatments tended to be one-time (a few doses a few days apart) or weekly over a period of a few months. Here, researchers try a longer term approach, weekly intervals for much of the adult life of mice.

Senolytic therapies produce rejuvenation in animal studies by selectively destroying senescent cells, which cause pathology as they accumulate in tissues. Present thinking is that this accumulation is more a matter of increased creation and slowed clearance rather than individual senescent cells lingering for the long term. Is the best dosing strategy a frequent one or an infrequent one. Or does it not much matter, so long as cells are periodically cleared?

That the study here shows greater benefits in terms of slowed disc degeneration when the senolytic treatement is started earlier in adult life suggests a few things. Firstly that some forms of structural damage do not tend to recover, even when their causes are removed. Secondly that senescent cells are causing some degree of harm earlier in adult life than we might otherwise have suspected. Assessments of the burden of cellular senescence by age do exist, but are not yet robust. Results like this might cause some reassessment of the ideal strategy for those who would like to take advantage of the existence of readily available senolytic drugs.

Studies of human tissues and mouse models have shown an increased incidence of senescent cells during intervertebral disc aging and degeneration. Intervertebral disc degeneration is highly prevalent within the elderly population and is a leading cause of chronic back pain and disability. Due to the link between disc degeneration and senescence, we explored the ability of the Dasatinib and Quercetin drug combination (D + Q) to prevent an age-dependent progression of disc degeneration in mice.

We treated C57BL/6 mice beginning at 6, 14, and 18 months of age, and analyzed them at 23 months of age. Interestingly, 6- and 14-month D + Q cohorts show lower incidences of degeneration, and the treatment results in a significant decrease in senescence markers p16INK4a, p19ARF, and SASP molecules IL-6 and MMP13. Treatment also preserves cell viability, phenotype, and matrix content. Although transcriptomic analysis shows disc compartment-specific effects of the treatment, cell death and cytokine response pathways are commonly modulated across tissue types.

Our results show that the D + Q combination could target senescence in the mouse disc, and these results provide proof of principle that senolytics may be useful in mitigating age-dependent disc degeneration by decreasing local senescence status, fibrosis and matrix degradation, while promoting cell viability, healthy matrix deposition and lower levels of systemic inflammation.

Link: https://doi.org/10.1038/s41467-021-25453-2

Much of the communication between cells is carried in extracellular vesicles, membrane-wrapped packages of signaling molecules. Vesicles are classified by size at present, though the nomenclature is often used confusingly and inconsistently. Exosomes are one class of smaller and frequently studied vesicle. Since it is now comparatively cheap to analyze the contents of vesicles obtained from blood samples, there exists a wealth of data related to changes in vesicle sizes, types, and contents that take place with age. It remains to be seen as to what can be achieved with this data beyond the construction of biomarkers to measure biological age.

Almost every cell, including stem cells, naturally release extracellular vesicles (EVs) responsible for cell-to-cell communication. They are split into three categories: microvesicles or ectosomes are submicron vesicles with a diameter of 100nm-1000nm, distinguished by biogenesis mechanisms including cytoskeleton remodeling and phosphatidylserine externalization. These microvesicles are formed by the outward budding and fission of the plasma membrane after cell stimulation or stress. Exosomes are the most common EVs studied and the smallest ones with a diameter of 30nm-100nm.

Exosomes are considered key regulators of many biological settings and are present in several extracellular fluids to mediate cellular communication. Recently, they were suggested as biomarkers for several diseases to set up diagnosis and disease progression. Their characteristics hold a great interest in designing therapeutic purposes in metabolic and genetic disorders, neurodegenerative and cardiovascular diseases, and cancer.

The first signature of human aging is the decrease of tissue regeneration and repair, leading to the accumulation of senescent cells. These cells have been described to release more exosomes with different compositions than a normal cell. A cellular transcriptional program is induced whereby the number and composition of exosomes are changed, consequently reflecting the current parent cell profile. Much evidence has increasingly involved exosomes and exosome-derived miRNAs in both normal and pathological aging processes. Evidence has also increasingly involved exosome-derived miRNAs in aging-associated diseases. In this work, we review exosome biogenesis and its involvement in the mechanisms related to aging with a focus on the different pathways described for their secreted miRNAs.

Link: https://doi.org/10.3390/biomedicines9080968