The mouse Social Frailty Index (mSFI): a novel behavioral assessment for impaired social functioning in aging mice.

Various approaches exist to quantify the aging process and estimate biological age on an individual level. Frailty indices based on an age-related accumulation of physical deficits have been developed for human use and translated into mouse models. However, declines observed in aging are not limited to physical functioning but also involve social capabilities. The concept of "social frailty" has been recently introduced into human literature, but no index of social frailty exists for laboratory mice yet. To fill this gap, we developed a mouse Social Frailty Index (mSFI) consisting of seven distinct assays designed to quantify social functioning which is relatively simple to execute and is minimally invasive. Application of the mSFI in group-housed male C57BL/6 mice demonstrated a progressively elevated levels of social frailty through the lifespan. Conversely, group-housed females C57BL/6 mice manifested social frailty only at a very old age. Female mice also showed significantly lower mSFI score from 10 months of age onward when compared to males. We also applied the mSFI in male C57BL/6 mice under chronic subordination stress and in chronic isolation, both of which induced larger increases in social frailty compared to age-matched group-housed males. Lastly, we show that the mSFI is enhanced in mouse models that show accelerated biological aging such as progeroid Ercc1-/Δ and Xpg-/- mice of both sexes compared to age matched littermate wild types. In summary, the mSFI represents a novel index to quantify trajectories of biological aging in mice and may help elucidate links between impaired social behavior and the aging process.

Acarbose ameliorates Western diet-induced metabolic and cognitive impairments in the 3xTg mouse model of Alzheimer's disease.

Age is the greatest risk factor for Alzheimer's disease (AD) as well as for other disorders that increase the risk of AD such as diabetes and obesity. There is growing interest in determining if interventions that promote metabolic health can prevent or delay AD. Acarbose is an anti-diabetic drug that not only improves glucose homeostasis, but also extends the lifespan of wild-type mice. Here, we test the hypothesis that acarbose will not only preserve metabolic health, but also slow or prevent AD pathology and cognitive deficits in 3xTg mice, a model of AD, fed either a Control diet or a high-fat, high-sucrose Western diet (WD). We find that acarbose decreases the body weight and adiposity of WD-fed 3xTg mice, increasing energy expenditure while also stimulating food consumption, and improves glycemic control. Both male and female WD-fed 3xTg mice have worsened cognitive deficits than Control-fed mice, and these deficits are ameliorated by acarbose treatment. Molecular and histological analysis of tau and amyloid pathology identified sex-specific effects of acarbose which are uncoupled from the dramatic improvements in cognition, suggesting that the benefits of acarbose on AD are largely driven by improved metabolic health. In conclusion, our results suggest that acarbose may be a promising intervention to prevent, delay, or even treat AD, especially in individuals consuming a Western diet.

Dietary isoleucine content defines the metabolic and molecular response to a Western diet.

The amino acid composition of the diet has recently emerged as a critical regulator of metabolic health. Consumption of the branched-chain amino acid isoleucine is positively correlated with body mass index in humans, and reducing dietary levels of isoleucine rapidly improves the metabolic health of diet-induced obese male C57BL/6J mice. However, it is unknown how sex, strain, and dietary isoleucine intake may interact to impact the response to a Western Diet (WD). Here, we find that although the magnitude of the effect varies by sex and strain, reducing dietary levels of isoleucine protects C57BL/6J and DBA/2J mice of both sexes from the deleterious metabolic effects of a WD, while increasing dietary levels of isoleucine impairs aspects of metabolic health. Despite broadly positive responses across all sexes and strains to reduced isoleucine, the molecular response of each sex and strain is highly distinctive. Using a multi-omics approach, we identify a core sex- and strain- independent molecular response to dietary isoleucine, and identify mega-clusters of differentially expressed hepatic genes, metabolites, and lipids associated with each phenotype. Intriguingly, the metabolic effects of reduced isoleucine in mice are not associated with FGF21 - and we find that in humans plasma FGF21 levels are likewise not associated with dietary levels of isoleucine. Finally, we find that foods contain a range of isoleucine levels, and that consumption of dietary isoleucine is lower in humans with healthy eating habits. Our results demonstrate that the dietary level of isoleucine is critical in the metabolic and molecular response to a WD, and suggest that lowering dietary levels of isoleucine may be an innovative and translatable strategy to protect from the negative metabolic consequences of a WD.

Protein restriction slows the development and progression of pathology in a mouse model of Alzheimer's disease.

Dietary protein is a critical regulator of metabolic health and aging. Low protein diets are associated with healthy aging in humans, and dietary protein restriction extends the lifespan and healthspan of mice. In this study, we examined the effect of protein restriction (PR) on metabolic health and the development and progression of Alzheimer's disease (AD) in the 3xTg mouse model of AD. Here, we show that PR promotes leanness and glycemic control in 3xTg mice, specifically rescuing the glucose intolerance of 3xTg females. PR induces sex-specific alterations in circulating and brain metabolites, downregulating sphingolipid subclasses in 3xTg females. PR also reduces AD pathology and mTORC1 activity, increases autophagy, and improves the cognition of 3xTg mice. Finally, PR improves the survival of 3xTg mice. Our results suggest that PR or pharmaceutical interventions that mimic the effects of this diet may hold promise as a treatment for AD.

Quantification of healthspan in aging mice: introducing FAMY and GRAIL.

The population around the world is graying, and as many of these individuals will spend years suffering from the burdens of age associated diseases, understanding how to increase healthspan, defined as the period of life free from disease and disability, is an urgent priority of geroscience research. The lack of agreed-upon quantitative metrics for measuring healthspan in aging mice has slowed progress in identifying interventions that do not simply increase lifespan, but also healthspan. Here, we define FAMY (Frailty-Adjusted Mouse Years) and GRAIL (Gauging Robust Aging when Increasing Lifespan) as new summary statistics for quantifying healthspan in mice. FAMY integrates lifespan data with longitudinal measurements of a widely utilized clinical frailty index, while GRAIL incorporates these measures and also adds information from widely utilized healthspan assays and the hallmarks of aging. Both metrics are conceptually similar to quality-adjusted life years (QALY), a widely utilized measure of disease burden in humans, and can be readily calculated from data acquired during longitudinal and cross-sectional studies of mouse aging. We find that interventions generally thought to promote health, including calorie restriction, robustly improve healthspan as measured by FAMY and GRAIL. Finally, we show that the use of GRAIL provides new insights, and identify dietary restriction of protein or isoleucine as interventions that robustly promote healthspan but not longevity in female HET3 mice. We suggest that the routine integration of these measures into studies of aging in mice will allow the identification and development of interventions that promote healthy aging even in the absence of increased lifespan.

Dietary lipid is largely deposited in skin and rapidly affects insulating properties.

Skin has been shown to be a regulatory hub for energy expenditure and metabolism: mutations of skin lipid metabolism enzymes can change the rate of thermogenesis and susceptibility to diet-induced obesity. However, little is known about the physiological basis for this function. Here we show that the thermal properties of skin are highly reactive to diet: within three days, a high fat diet reduces heat transfer through skin. In contrast, a dietary manipulation that prevents obesity accelerates energy loss through skins. We found that skin was the largest target in a mouse body for dietary fat delivery, and that fat was assimilated both by epidermis and by dermal white adipose tissue. Dietary triglyceride acyl groups persist in skin for weeks after feeding. Using multi-modal lipid profiling, we have implicated both keratinocytes and sebocytes in the altered lipids which correlate with thermal function. In response to high fat feeding, wax diesters and ceramides accumulate, and triglycerides become more saturated. In contrast, in response to the dramatic loss of adipose tissue that accompanies restriction of the branched chain amino acid isoleucine, skin becomes highly heat-permeable: skins shows limited uptake of dietary lipids and editing of wax esters, and acquires a signature of depleted signaling lipids, which include the acyl carnitines and lipid ethers. We propose that skin should be routinely included in physiological studies of lipid metabolism, given the size of the skin lipid reservoir and its adaptable functionality.

Late-life isoleucine restriction promotes physiological and molecular signatures of healthy aging.

In defiance of the paradigm that calories from all sources are equivalent, we and others have shown that dietary protein is a dominant regulator of healthy aging. The restriction of protein or the branched-chain amino acid isoleucine promotes healthspan and extends lifespan when initiated in young or adult mice. However, many interventions are less efficacious or even deleterious when initiated in aged animals. Here, we investigate the physiological, metabolic, and molecular consequences of consuming a diet with a 67% reduction of all amino acids (Low AA), or of isoleucine alone (Low Ile), in male and female C57BL/6J.Nia mice starting at 20 months of age. We find that both diet regimens effectively reduce adiposity and improve glucose tolerance, which were benefits that were not mediated by reduced calorie intake. Both diets improve specific aspects of frailty, slow multiple molecular indicators of aging rate, and rejuvenate the aging heart and liver at the molecular level. These results demonstrate that Low AA and Low Ile diets can drive youthful physiological and molecular signatures, and support the possibility that these dietary interventions could help to promote healthy aging in older adults.

Quantification of healthspan in aging mice: Introducing FAMY and GRAIL.

The population around the world is graying, and as many of these individuals will spend years suffering from the burdens of age associated diseases, understanding how to increase healthspan, defined as the period of life free from disease and disability, is an urgent priority of geroscience research. The lack of agreed-upon quantitative metrics for measuring healthspan in aging mice has slowed progress in identifying interventions that do not simply increase lifespan, but also healthspan. Here, we define FAMY (Frailty-Adjusted Mouse Years) and GRAIL (Gauging Robust Aging when Increasing Lifespan) as new summary statistics for quantifying healthspan in mice. FAMY integrates lifespan data with longitudinal measurements of a widely utilized clinical frailty index, while GRAIL incorporates these measures and also adds information from widely utilized healthspan assays and the hallmarks of aging. Both metrics are conceptually similar to quality-adjusted life years (QALY), a widely-utilized measure of disease burden in humans, and can be readily calculated from data acquired during longitudinal and cross-sectional studies of mouse aging. We find that interventions generally thought to promote health, including calorie restriction, robustly improve healthspan as measured by FAMY and GRAIL. Finally, we show that the use of GRAIL provides new insights, and identify dietary restriction of protein or isoleucine as interventions that robustly promote healthspan but not longevity in female HET3 mice. We suggest that the routine integration of these measures into studies of aging in mice will allow the identification and development of interventions that promote healthy aging even in the absence of increased lifespan.

Geroprotector drugs and exercise: friends or foes on healthy longevity?

Physical activity and several pharmacological approaches individually combat age-associated conditions and extend healthy longevity in model systems. It is tantalizing to extrapolate that combining geroprotector drugs with exercise could extend healthy longevity beyond any individual treatment. However, the current dogma suggests that taking leading geroprotector drugs on the same day as exercise may limit several health benefits. Here, we review leading candidate geroprotector drugs and their interactions with exercise and highlight salient gaps in knowledge that need to be addressed to identify if geroprotector drugs can have a harmonious relationship with exercise.

Dietary restriction of individual amino acids stimulates unique molecular responses in mouse liver.

Dietary protein and essential amino acid (EAA) restriction promotes favorable metabolic reprogramming, ultimately resulting in improvements to both health and lifespan. However, as individual EAAs have distinct catabolites and engage diverse downstream signaling pathways, it remains unclear to what extent shared or AA-specific molecular mechanisms promote diet-associated phenotypes. Here, we investigated the physiological and molecular effects of restricting either dietary methionine, leucine, or isoleucine (Met-R, Leu-R, and Ile-R) for 3 weeks in C57BL/6J male mice. While all 3 AA-depleted diets promoted fat and lean mass loss and slightly improved glucose tolerance, the molecular responses were more diverse; while hepatic metabolites altered by Met-R and Leu-R were highly similar, Ile-R led to dramatic changes in metabolites, including a 3-fold reduction in the oncometabolite 2-hydroxyglutarate. Pathways regulated in an EAA-specific manner included glycolysis, the pentose phosphate pathway (PPP), nucleotide metabolism, the TCA cycle and amino acid metabolism. Transcriptiome analysis and global profiling of histone post-translational modifications (PTMs) revealed different patterns of responses to each diet, although Met-R and Leu-R again shared similar transcriptional responses. While the pattern of global histone PTMs were largely unique for each dietary intervention, Met-R and Ile-R had similar changes in histone-3 methylation/acetylation PTMs at lysine-9. Few similarities were observed between the physiological or molecular responses to EAA restriction and treatment with rapamycin, an inhibitor of the mTORC1 AA-responsive protein kinase, indicating the response to EAA restriction may be largely independent of mTORC1. Together, these results demonstrate that dietary restriction of individual EAAs has unique, EAA-specific effects on the hepatic metabolome, epigenome, and transcriptome, and suggests that the specific EAAs present in dietary protein may play a key role at regulating health at the molecular level.

What approaches are needed to understand human development and disease?

Researchers are leveraging what we have learned from model organisms to understand if the same principles arise in human physiology, development, and disease. In this collection of Voices, we asked researchers from different fields to discuss what tools and insights they are using to answer fundamental questions in human biology.

Dietary restriction of isoleucine increases healthspan and lifespan of genetically heterogeneous mice.

Low-protein diets promote health and longevity in diverse species. Restriction of the branched-chain amino acids (BCAAs) leucine, isoleucine, and valine recapitulates many of these benefits in young C57BL/6J mice. Restriction of dietary isoleucine (IleR) is sufficient to promote metabolic health and is required for many benefits of a low-protein diet in C57BL/6J males. Here, we test the hypothesis that IleR will promote healthy aging in genetically heterogeneous adult UM-HET3 mice. We find that IleR improves metabolic health in young and old HET3 mice, promoting leanness and glycemic control in both sexes, and reprograms hepatic metabolism in a sex-specific manner. IleR reduces frailty and extends the lifespan of male and female mice, but to a greater degree in males. Our results demonstrate that IleR increases healthspan and longevity in genetically diverse mice and suggests that IleR, or pharmaceuticals that mimic this effect, may have potential as a geroprotective intervention.

Resistance exercise protects mice from protein-induced fat accretion.

Low-protein (LP) diets extend the lifespan of diverse species and are associated with improved metabolic health in both rodents and humans. Paradoxically, many athletes and bodybuilders consume high-protein (HP) diets and protein supplements, yet are both fit and metabolically healthy. Here, we examine this paradox using weight pulling, a validated progressive resistance exercise training regimen, in mice fed either an LP diet or an isocaloric HP diet. We find that despite having lower food consumption than the LP group, HP-fed mice gain significantly more fat mass than LP-fed mice when not exercising, while weight pulling protected HP-fed mice from this excess fat accretion. The HP diet augmented exercise-induced hypertrophy of the forearm flexor complex, and weight pulling ability increased more rapidly in the exercised HP-fed mice. Surprisingly, exercise did not protect from HP-induced changes in glycemic control. Our results confirm that HP diets can augment muscle hypertrophy and accelerate strength gain induced by resistance exercise without negative effects on fat mass, and also demonstrate that LP diets may be advantageous in the sedentary. Our results highlight the need to consider both dietary composition and activity, not simply calories, when taking a precision nutrition approach to health.

Protein restriction slows the development and progression of Alzheimer's disease in mice.

Over the last decade, it has become evident that dietary protein is a critical regulator of metabolic health and aging. Low protein diets are associated with healthy aging in humans, and we and others have shown that dietary protein restriction (PR) extends the lifespan and healthspan of mice. Here, we examined the effect of PR on metabolic health and the development and progression of Alzheimer's disease (AD) in the 3xTg mouse model of AD. We found that PR has metabolic benefits for 3xTg mice and non-transgenic controls of both sexes, promoting leanness and glycemic control in 3xTg mice. We found that PR induces sex-specific alterations in circulating metabolites and in the brain lipidome, downregulating sphingolipid subclasses including ceramides, glucosylceramides, and sphingomyelins in 3xTg females. Consumption of a PR diet starting at 6 months of age reduced AD pathology in conjunction with reduced mTORC1 activity, increased autophagy, and had cognitive benefits for 3xTg mice. Finally, PR improved the survival of 3xTg mice. Our results demonstrate that PR slows the progression of AD at molecular and pathological levels, preserves cognition in this mouse model of AD, and suggests that PR or pharmaceutical interventions that mimic the effects of this diet may hold promise as a treatment for AD.

Blazing a trail for the clinical use of rapamycin as a geroprotecTOR.

Treatment with rapamycin, an inhibitor of the mechanistic Target Of Rapamycin Complex One (mTORC1) protein kinase, has been repeatedly demonstrated to extend lifespan and prevent or delay age-related diseases in diverse model systems. Concerns over the risk of potentially serious side effects in humans, including immunosuppression and metabolic disruptions, have cautiously limited the translation of rapamycin and its analogs as a treatment for aging associated conditions. During the last decade, we and others have developed a working model that suggests that while inhibition of mTORC1 promotes healthy aging, many of the negative side effects of rapamycin are associated with "off-target" inhibition of a second mTOR complex, mTORC2. Differences in the kinetics and molecular mechanisms by which rapamycin inhibits mTORC1 and mTORC2 suggest that a therapeutic window for rapamycin could be exploited using intermittent dosing schedules or alternative rapalogs that may enable more selective inhibition of mTORC1. However, the optimal dosing schedules and the long-term efficacy of such interventions in humans are unknown. Here, we highlight ongoing or upcoming clinical trials that will address outstanding questions regarding the safety, pharmacokinetics, pharmacodynamics, and efficacy of rapamycin and rapalogs on several clinically oriented outcomes. Results from these early phase studies will help guide the design of phase 3 clinical trials to determine whether rapamycin can be used safely to inhibit mTORC1 for the treatment and prevention of age-related diseases in humans.

When a calorie is not just a calorie: Diet quality and timing as mediators of metabolism and healthy aging.

An epidemic of obesity has affected large portions of the world, increasing the risk of developing many different age-associated diseases, including cancer, cardiovascular disease, and diabetes. In contrast with the prevailing notion that "a calorie is just a calorie," there are clear differences, within and between individuals, in the metabolic response to different macronutrient sources. Recent findings challenge this oversimplification; calories from different macronutrient sources or consumed at different times of day have metabolic effects beyond their value as fuel. Here, we summarize discussions conducted at a recent NIH workshop that brought together experts in calorie restriction, macronutrient composition, and time-restricted feeding to discuss how dietary composition and feeding schedule impact whole-body metabolism, longevity, and healthspan. These discussions may provide insights into the long-sought molecular mechanisms engaged by calorie restriction to extend lifespan, lead to novel therapies, and potentially inform the development of a personalized food-as-medicine approach to healthy aging.

Targeting the biology of aging with mTOR inhibitors.

Inhibition of the protein kinase mechanistic target of rapamycin (mTOR) with the Food and Drug Administration (FDA)-approved therapeutic rapamycin promotes health and longevity in diverse model organisms. More recently, specific inhibition of mTORC1 to treat aging-related conditions has become the goal of basic and translational scientists, clinicians and biotechnology companies. Here, we review the effects of rapamycin on the longevity and survival of both wild-type mice and mouse models of human diseases. We discuss recent clinical trials that have explored whether existing mTOR inhibitors can safely prevent, delay or treat multiple diseases of aging. Finally, we discuss how new molecules may provide routes to the safer and more selective inhibition of mTOR complex 1 (mTORC1) in the decade ahead. We conclude by discussing what work remains to be done and the questions that will need to be addressed to make mTOR inhibitors part of the standard of care for diseases of aging.

Geroprotective interventions in the 3xTg mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is an age-associated neurodegenerative disease. As the population ages, the increasing prevalence of AD threatens massive healthcare costs in the coming decades. Unfortunately, traditional drug development efforts for AD have proven largely unsuccessful. A geroscience approach to AD suggests that since aging is the main driver of AD, targeting aging itself may be an effective way to prevent or treat AD. Here, we discuss the effectiveness of geroprotective interventions on AD pathology and cognition in the widely utilized triple-transgenic mouse model of AD (3xTg-AD) which develops both ß-amyloid and tau pathologies characteristic of human AD, as well as cognitive deficits. We discuss the beneficial impacts of calorie restriction (CR), the gold standard for geroprotective interventions, and the effects of other dietary interventions including protein restriction. We also discuss the promising preclinical results of geroprotective pharmaceuticals, including rapamycin and medications for type 2 diabetes. Though these interventions and treatments have beneficial effects in the 3xTg-AD model, there is no guarantee that they will be as effective in humans, and we discuss the need to examine these interventions in additional animal models as well as the urgent need to test if some of these approaches can be translated from the lab to the bedside for the treatment of humans with AD.

Strength in diversity: Intra-cellular metabolite sharing enhances longevity.

Much of our foundational knowledge of cellular biology comes from studies in budding yeast, often described as a simple unicellular eukaryotic model. In this issue of Cell, Correia-Melo et al. describe an unappreciated feature of yeast biology involving intra-cellular metabolite exchange, where cells adapt and respond as part of a community, and go on to show that sharing of resources linked to methionine metabolism enhances longevity of cooperating cells.

Regulation of metabolic health by dietary histidine in mice.

Low-protein (LP) diets are associated with a decreased risk of diabetes in humans, and promote leanness and glycaemic control in both rodents and humans. While the effects of an LP diet on glycaemic control are mediated by reduced levels of the branched-chain amino acids, we have observed that reducing dietary levels of the other six essential amino acids leads to changes in body composition. Here, we find that dietary histidine plays a key role in the response to an LP diet in male C57BL/6J mice. Specifically reducing dietary levels of histidine by 67% reduces the weight gain of young, lean male mice, reducing both adipose and lean mass without altering glucose metabolism, and rapidly reverses diet-induced obesity and hepatic steatosis in diet-induced obese male mice, increasing insulin sensitivity. This normalization of metabolic health was associated not with caloric restriction or increased activity, but with increased energy expenditure. Surprisingly, the effects of histidine restriction do not require the energy balance hormone Fgf21. Histidine restriction that was started in midlife promoted leanness and glucose tolerance in aged males but not females, but did not affect frailty or lifespan in either sex. Finally, we demonstrate that variation in dietary histidine levels helps to explain body mass index differences in humans. Overall, our findings demonstrate that dietary histidine is a key regulator of weight and body composition in male mice and in humans, and suggest that reducing dietary histidine may be a translatable option for the treatment of obesity. KEY POINTS: Protein restriction (PR) promotes metabolic health in rodents and humans and extends rodent lifespan. Restriction of specific individual essential amino acids can recapitulate the benefits of PR. Reduced histidine promotes leanness and increased energy expenditure in male mice. Reduced histidine does not extend the lifespan of mice when begun in midlife. Dietary levels of histidine are positively associated with body mass index in humans.