Frailty In Mice Research Articles

Short senolytic or senostatic interventions rescue progression of radiation-induced frailty and premature ageing in mice.

Cancer survivors suffer from progressive frailty, multimorbidity, and premature morbidity. We hypothesise that therapy-induced senescence and senescence progression via bystander effects are significant causes of this premature ageing phenotype. Accordingly, the study addresses the question whether a short anti-senescence intervention is able to block progression of radiation-induced frailty and disability in a pre-clinical setting. Male mice were sublethally irradiated at 5 months of age and treated (or not) with either a senolytic drug (Navitoclax or dasatinib + quercetin) for 10 days or with the senostatic metformin for 10 weeks. Follow-up was for 1 year. Treatments commencing within a month after irradiation effectively reduced frailty progression (p<0.05) and improved muscle (p<0.01) and liver (p<0.05) function as well as short-term memory (p<0.05) until advanced age with no need for repeated interventions. Senolytic interventions that started late, after radiation-induced premature frailty was manifest, still had beneficial effects on frailty (p<0.05) and short-term memory (p<0.05). Metformin was similarly effective as senolytics. At therapeutically achievable concentrations, metformin acted as a senostatic neither via inhibition of mitochondrial complex I, nor via improvement of mitophagy or mitochondrial function, but by reducing non-mitochondrial reactive oxygen species production via NADPH oxidase 4 inhibition in senescent cells. Our study suggests that the progression of adverse long-term health and quality-of-life effects of radiation exposure, as experienced by cancer survivors, might be rescued by short-term adjuvant anti-senescence interventions.

Preclinical frailty assessments: Phenotype and frailty index identify frailty in different mice and are variably affected by chronic medications.

Use of different objective frailty assessment tools may improve understanding of the biology of frailty and allow evaluation of effects of interventions on frailty. Polypharmacy is associated with increased risk of frailty in epidemiologic studies, regardless of frailty definition, but the pathophysiology of the association is not well understood. This study aims to (1) assess and compare the prevalence of frailty from middle to old age following control, chronic polypharmacy or monotherapy treatment, when measured using the clinical frailty index assessment and the mouse frailty phenotype tools; and (2) to evaluate and compare the effects of chronic polypharmacy regimens with zero, low and high Drug Burden Index (DBI) and monotherapies from middle to old age on the rate of deficit accumulation on the frailty index, mean number of phenotype criteria, odds of being frail assessed by the frailty index or phenotype, and the time to onset of frailty assessed by the frailty index or phenotype. In a longitudinal study, middle-aged (12months) male C57BL/6J(B6) mice were administered non medicated control feed and water, or therapeutic doses of different polypharmacy combinations or monotherapies in feed and/or water. Frailty assessments were performed at 12, 15, 18, 21 and 24months. There was limited overlap between animals identified as frail using different frailty assessments. Polypharmacy has measurable and different effects on each frailty assessment. Long-term chronic administration of some polypharmacy and monotherapy therapeutic drug regimens increased the number of frailty deficits (clinical frailty index: low DBI polypharmacy (15 and 21months), high DBI polypharmacy (15-21months), oxycodone (15-18months), oxybutynin (15-18months), citalopram (15-21months) and metoprolol monotherapy (15months) and modified frailty phenotype assessment (over the whole duration of treatment, low DBI polypharmacy (adjusted Risk Ratio(aRR)=1.97, 95% confidence interval (CI) 1.43-2.72), high DBI polypharmacy (aRR=1.88; 95% CI 1.36-2.60), oxybutynin (aRR=1.48; 95% CI 1.01-2.16) and citalopram monotherapy (aRR=1.96; 95% CI 1.41-2.74), p<0.05) . The odds of developing frailty measured with the clinical frailty index increased with high DBI polypharmacy (adjusted odds ratio (aOR)=3.13; 95% CI 1.01-9.66) and when measured with the frailty phenotype assessment increased with low DBI polypharmacy (aOR=4.38, 95% CI 1.40-13.74), high DBI polypharmacy (aOR=3.43; 95% CI 1.12-10.50) and citalopram monotherapy (aOR=4.63; 95% CI 1.39-15.54)). No treatment affected time to frailty using either frailty assessment. Analysis of the number of deficits on the frailty index or number of positive criteria on the frailty phenotype allows analysis of rate of change and provides greater sensitivity, while the odds of being frail analysis provided a clinically relevant indicator of whether mice had greater chance of reaching a cut-off for becoming frail with medication exposure than without. Our results are consistent with clinical studies, demonstrating that certain polypharmacy regimens induce frailty, with different relationships observed when using different frailty assessments and analyses.

A Novel Trimethylamine Oxide-Induced Model Implicates Gut Microbiota-Related Mechanisms in Frailty.

Frailty is a complicated syndrome that occurs at various ages, with highest incidence in aged populations, suggesting associations between the pathogenesis of frailty and age-related changes. Gut microbiota (GM) diversity and abundance change with age, accompanied by increased levels of trimethylamine oxide (TMAO), a systemic inflammation-inducing GM metabolite. Thus, we hypothesized that TMAO may be involved in the development of frailty. We successfully established and verified a novel model of frailty in adult mice based on a 4-week intraperitoneal injection regime of TMAO followed by LPS challenge. The frailty index significantly increased in TMAO-treated mice after LPS challenge. TMAO also decreased claudin-1 immunofluorescent staining intensity in the jejunum, ileum, and colon, indicating that the destruction of intestinal wall integrity may increase vulnerability to exogenous pathogens and invoke frailty. 16S sequencing showed that TMAO significantly reduced the GM Firmicutes/Bacteroidetes (F/B) ratio, but not α-diversity. Interestingly, after LPS challenge, more genera of bacterial taxa were differently altered in the control mice than in the TMAO-treated mice. We infer that a variety of GM participate in the maintenance of homeostasis, whereas TMAO could blunt the GM and impair the ability to recover from pathogens, which may explain the continuous increase in the frailty index in TMAO-treated mice after LPS challenge. TMAO also significantly increased serum imidazole metabolites, and led to different patterns of change in serum peptide and phenylpropanoid metabolites after LPS stimulation. These changes indicate that glucose metabolism may be one mechanism by which GM inactivation causes frailty. In conclusion, TMAO leads to frailty by destroying intestinal barrier integrity and blunting the GM response.

Both Vitamin D Supplementation and HIIT Boost Muscle VDR Expression, Which May Underlie Benefits for Frailty

Frailty is a condition of poor response to stressors that increases susceptibility to adverse outcomes - including disability and death. Declining physical function is an important hallmark of frailty, and we previously published that long term vitamin D insufficiency from young to middle-age leads to declines in endurance and gait disturbances. Furthermore, we report that aged mice (24-months) made vitamin D insufficient for 4 months exhibit increased frailty, whereas those made hyper-sufficient do not. Exercise, including short session high intensity interval training (HIIT - 10 minutes/3x-week), also reverses frailty in aged mice. Here we investigate the impacts of aging, vitamin D, and exercise on underlying muscle quality and muscle stem cell activity. Our preliminary data reveal muscle vitamin D receptor (VDR) expression is lower in aged mice (24-28 months) relative to young mice (6 months). Yet HIIT, either one hour after a single session or following 6 weeks, increases VDR expression. HIIT also increases myonuclear accretion in muscle fibers – an indicator of in vivo stem cell activity – and stimulates progenitor cells proliferation ex vivo. Likewise, we observe that vitamin D hyper supplementation alone also increases muscle VDR expression and the number of satellite cells. These data indicate that both vitamin D supplementation and HIIT independently enhance VDR expression in skeletal muscle with associated greater satellite and muscle progenitor cell activity. These data critically link vitamin D physiology and HIIT in muscle, and thus provide a mechanistic basis for their benefits for muscle quality, function, and health during aging.

Glucose 6-P dehydrogenase delays the onset of frailty by protecting against muscle damage.

BackgroundFrailty is a major age‐associated syndrome leading to disability. Oxidative damage plays a significant role in the promotion of frailty. The cellular antioxidant system relies on reduced nicotinamide adenine dinucleotide phosphate (NADPH) that is highly dependent on glucose 6‐P dehydrogenase (G6PD). The G6PD‐overexpressing mouse (G6PD‐Tg) is protected against metabolic stresses. Our aim was to examine whether this protection delays frailty.MethodsOld wild‐type (WT) and G6PD‐Tg mice were evaluated longitudinally in terms of frailty. Indirect calorimetry, transcriptomic profile, and different skeletal muscle quality markers and muscle regenerative capacity were also investigated.ResultsThe percentage of frail mice was significantly lower in the G6PD‐Tg than in the WT genotype, especially in 26‐month‐old mice where 50% of the WT were frail vs. only 13% of the Tg ones (P < 0.001). Skeletal muscle transcriptomic analysis showed an up‐regulation of respiratory chain and oxidative phosphorylation (P = 0.009) as well as glutathione metabolism (P = 0.035) pathways in the G6PD‐Tg mice. Accordingly, the Tg animals exhibited an increase in reduced glutathione (34.5%, P < 0.01) and a decrease on its oxidized form (−69%, P < 0.05) and in lipid peroxidation (4‐HNE: −20.5%, P < 0.05). The G6PD‐Tg mice also showed reduced apoptosis (BAX/Bcl2: −25.5%, P < 0.05; and Bcl‐xL: −20.5%, P < 0.05), lower levels of the intramuscular adipocyte marker FABP4 (−54.7%, P < 0.05), and increased markers of mitochondrial content (COX IV: 89.7%, P < 0.05; Grp75: 37.8%, P < 0.05) and mitochondrial OXPHOS complexes (CII: 81.25%, P < 0.01; CIII: 52.5%, P < 0.01; and CV: 37.2%, P < 0.05). Energy expenditure (−4.29%, P < 0.001) and the respiratory exchange ratio were lower (−13.4%, P < 0.0001) while the locomotor activity was higher (43.4%, P < 0.0001) in the 20‐month‐old Tg, indicating a major energetic advantage in these mice. Short‐term exercise training in young C57BL76J mice induced a robust activation of G6PD in skeletal muscle (203.4%, P < 0.05), similar to that achieved in the G6PD‐Tg mice (142.3%, P < 0.01).ConclusionsGlucose 6‐P dehydrogenase deficiency can be an underestimated risk factor for several human pathologies and even frailty. By overexpressing G6PD, we provide the first molecular model of robustness. Because G6PD is regulated by pharmacological and physiological interventions like exercise, our results provide molecular bases for interventions that by increasing G6PD will delay the onset of frailty.

Sex Differences in the Relation Between Frailty and Endothelial Dysfunction in Old Mice.

Vascular endothelial function declines with age on average, but there is high variability in the magnitude of this decline within populations. Measurements of frailty, known as frailty index (FI), can be used as surrogates for biological age, but it is unknown if frailty relates to the age-related decline in vascular function. To examine this relation, we studied young (4-9 months) and old (23-32 months) C57BL6 mice of both sexes. We found that FI was greater in old compared with young mice, but did not differ between old male and female mice. Middle cerebral artery (MCA) and mesenteric artery endothelium-dependent dilation (EDD) also did not differ between old male and female mice; however, there were sex differences in the relations between FI and EDD. For the MCA, FI was inversely related to EDD among old female mice, but not old male mice. In contrast, for the mesenteric artery, FI was inversely related to EDD among old male mice, but not old female mice. A higher FI was related to a greater improvement in EDD with the superoxide scavenger 4-hydroxy-2,2,6,6-tetramethylpiperidin-1-oxyl in the MCAs for old female mice and in the mesenteric arteries for old male mice. FI related to mesenteric artery gene expression negatively for extracellular superoxide dismutase (Sod3) and positively for interleukin-1β (Il1b). In summary, we found that the relation between frailty and endothelial function is dependent on sex and the artery examined. Arterial oxidative stress and proinflammatory signaling are potential mediators of the relations of frailty and endothelial function.

Atrial Fibrillation in Aging and Frail Mice: Modulation by Natriuretic Peptide Receptor C.

[Figure: see text].

Neuroprotective Metabolites of Hericium erinaceus Promote Neuro-Healthy Aging.

Frailty is a geriatric syndrome associated with both locomotor and cognitive decline, typically linked to chronic systemic inflammation, i.e., inflammaging. In the current study, we investigated the effect of a two-month oral supplementation with standardized extracts of H. erinaceus, containing a known amount of Erinacine A, Hericenone C, Hericenone D, and L-ergothioneine, on locomotor frailty and cerebellum of aged mice. Locomotor performances were monitored comparing healthy aging and frail mice. Cerebellar volume and cytoarchitecture, together with inflammatory and oxidative stress pathways, were assessed focusing on senescent frail animals. H. erinaceus partially recovered the aged-related decline of locomotor performances. Histopathological analyses paralleled by immunocytochemical evaluation of specific molecules strengthened the neuroprotective role of H. erinaceus able to ameliorate cerebellar alterations, i.e., milder volume reduction, slighter molecular layer thickness decrease and minor percentage of shrunken Purkinje neurons, also diminishing inflammation and oxidative stress in frail mice while increasing a key longevity regulator and a neuroprotective molecule. Thus, our present findings demonstrated the efficacy of a non-pharmacological approach, based on the dietary supplementation using H. erinaceus extract, which represent a promising adjuvant therapy to be associated with conventional geriatric treatments.

Sex Differences in the Relation Between Frailty and Endothelial Function in Old Mice

Frailty is an accumulation of deficits that stem from impairments in multiple organ systems, potentially including the vasculature. Older women typically have higher frailty scores than older men, but it is unknown if there are sex differences in the relation of frailty to physiological function. Endothelial function declines with age on average, but there is a high variability in the amount of decline. Thus, we sought to determine sex differences in the relation of frailty to endothelial dysfunction among old mice. In young male (n= 24, 8.5±0.27 mo), young female (n=20, 4.9±0.01 mo), old male (n=32, 28±0.56 mo) and old female (n= 17, 25.8±0.47 mo) C57BL/6 mice, we evaluated frailty by a 31-item frailty index (FI) based on established signs of deterioration. We assessed endothelial function by endothelium-dependent dilation (EDD, maximal response to acetylcholine) ex vivo in isolated, pressurized mesentery arteries and middle cerebral arteries (MCAs). We also assessed EDD after incubation with the superoxide scavenger TEMPOL and endothelium-independent dilation (EID, maximal response to sodium nitroprusside). We quantified gene expression for pro-inflammatory interleukin-1β (Il1b), pro-oxidant NADPH oxidase 2 (Nox2), and antioxidants superoxide dismutases (Sod1, Sod2, and Sod3) in mesentery arteries. Old mice had a greater FI compared with young mice (old: 5.53±2.3, young: 1.01±0.67, p=0.003), but FI did not differ between old male and female mice (p=0.41). When specific FI components were examined, old female mice had higher scores for alopecia, rectal prolapse, and cataracts but lower scores for vaginal/penile prolapse and grimace scale compared with old male mice (p<0.05). Old mice had a 25% impairment in MCA EDD and 16% impairment in mesentery artery EDD compared with young mice (p<0.05). While EDD did not differ between arteries from old male and female mice (p>0.05), there were sex differences in the relations between FI and EDD. For the MCA, FI was inversely related to EDD among old female (r=-0.49, p=0.02), but not old male mice (r=0.15, p=0.24). In contrast, for the mesentery artery, FI was inversely related to EDD among old males (r=-0.42, p=0.01), but not old female mice (r=0.37, p=0.07). EID did not differ with sex or age group (p>0.05). A higher FI was related to a greater improvement in EDD with TEMPOL in the MCA in only old female mice and in the mesentery arteries in only old male mice (p<0.05). In mesentery arteries from old compared with young mice, gene expression was 42% higher for Il1b, 34% lower for Sod2 and 54% lower for Sod3 (p<0.05), but not different between ages for Sod1 or Nox2 (p>0.05). Among old mice, Il1b expression was23% higher in mesenteries from old male compared with female mice (p=0.001), but sex differences were not present for the other genes (p>0.05). In summary, we found no sex differences for composite FI scores or endothelial impairment among old mice. However, there are sex differences in the individual components of FI and in the relations of frailty to endothelial function, and these may be mediated by sex differences in arterial oxidative stress and pro-inflammatory signaling.

IL-6 can singlehandedly drive many features of frailty in mice.

Frailty is a geriatric syndrome characterized by age-related declines in function and reserve resulting in increased vulnerability to stressors. The most consistent laboratory finding in frail subjects is elevation of serum IL-6, but it is unclear whether IL-6 is a causal driver of frailty. Here, we characterize a new mouse model of inducible IL-6 expression (IL-6TET-ON/+ mice) following administration of doxycycline (Dox) in food. In this model, IL-6 induction was Dox dose-dependent. The Dox dose that increased IL-6 levels to those observed in frail old mice directly led to an increase in frailty index, decrease in grip strength, and disrupted muscle mitochondrial homeostasis. Littermate mice lacking the knock-in construct failed to exhibit frailty after Dox feeding. Both naturally old mice and young Dox-induced IL-6TET-ON/+ mice exhibited increased IL-6 levels in sera and spleen homogenates but not in other tissues. Moreover, Dox-induced IL-6TET-ON/+ mice exhibited selective elevation in IL-6 but not in other cytokines. Finally, bone marrow chimera and splenectomy experiments demonstrated that non-hematopoietic cells are the key source of IL-6 in our model. We conclude that elevated IL-6 serum levels directly drive age-related frailty, possibly via mitochondrial mechanisms.

Pinpointing a Role for Vitamin D in Frailty: A Time for Animal Models?

Frailty is a condition marked by greater susceptibility to adverse outcomes, including disability and mortality, which affects up to 50% of those 80 years of age and older. Concurrently, serum vitamin D insufficiency and deficiency, for which as many as 70% of older adults may be at risk, potentially play an important role in frailty onset and progression. Large population driven studies have uncovered associations between low serum vitamin D levels and higher incidence of frailty. However, attempts to apply vitamin D therapeutically to treat and/or prevent frailty have not yielded consistent support for benefits. Given the complexity and inconsistency arising from human studies involving vitamin D, our research group has recently published on animal models of vitamin D insufficiency. Combining our model with the emerging development of animal frailty assessment, we identified that higher than standard levels of vitamin D supplementation may delay frailty in mice. In this viewpoint article, we will discuss current knowledge regarding the importance of vitamin D in frailty progression, the emerging significance of animal models in addressing these relationships, and the future for pre-clinical and clinical research.

Lifelong restriction of dietary branched-chain amino acids has sex-specific benefits for frailty and lifespan in mice.

Protein restricted (PR) diets promote health and longevity in many species. While the precise components of a PR diet that mediate the beneficial effects to longevity have not been defined, we recently showed that many metabolic effects of PR can be attributed to reduced dietary levels of the branched-chain amino acids (BCAAs) leucine, isoleucine, and valine. Here, we demonstrate that restricting dietary BCAAs increases the survival of two different progeroid mouse models, delays frailty and promotes the metabolic health of wild-type C57BL/6J mice when started in midlife, and leads to a 30% increase in lifespan and a reduction in frailty in male, but not female, wild-type mice when fed lifelong. Our results demonstrate that restricting dietary BCAAs can increase healthspan and longevity in mice, and suggest that reducing dietary BCAAs may hold potential as a translatable intervention to promote healthy aging.

Effects of long-term in vivo micro-CT imagingon hallmarks of osteopenia and frailty in aging mice.

In vivo micro-CT has already been used to monitor microstructural changes of bone in mice of different ages and in models of age-related diseases such as osteoporosis. However, as aging is accompanied by frailty and subsequent increased sensitivity to external stimuli such as handling and anesthesia, the extent to which longitudinal imaging can be applied in aging studies remains unclear. Consequently, the potential of monitoring individual mice during the entire aging process–from healthy to frail status–has not yet been exploited. In this study, we assessed the effects of long-term in vivo micro-CT imaging—consisting of 11 imaging sessions over 20 weeks—on hallmarks of aging both on a local (i.e., static and dynamic bone morphometry) and systemic (i.e., frailty index (FI) and body weight) level at various stages of the aging process. Furthermore, using a premature aging model (PolgA(D257A/D257A)), we assessed whether these effects differ between genotypes. The 6th caudal vertebrae of 4 groups of mice (PolgA(D257A/D257A) and PolgA(+/+)) were monitored by in vivo micro-CT every 2 weeks. One group was subjected to 11 scans between weeks 20 and 40 of age, whereas the other groups were subjected to 5 scans between weeks 26–34, 32–40 and 40–46, respectively. The long-term monitoring approach showed small but significant changes in the static bone morphometric parameters compared to the other groups. However, no interaction effect between groups and genotype was found, suggesting that PolgA mutation does not render bone more or less susceptible to long-term micro-CT imaging. The differences between groups observed in the static morphometric parameters were less pronounced in the dynamic morphometric parameters. Moreover, the body weight and FI were not affected by more frequent imaging sessions. Finally, we observed that longitudinal designs including baseline measurements at young adult age are more powerful at detecting effects of in vivo micro-CT imaging on hallmarks of aging than cross-sectional comparisons between multiple groups of aged mice subjected to fewer imaging sessions.

Alpha-Ketoglutarate, the Metabolite that Regulates Aging in Mice

In this issue of Cell Metabolism, Asadi Shahmirzadi etal. (2020) demonstrate that late-onset dietary supplementation with calcium alpha-ketoglutarate results in increased survival, compressed morbidity, and reduced frailty in mice. The study provides further evidence for critical links between metabolism, inflammation, and aging.

Maladaptive Changes Associated With Cardiac Aging Are Sex-Specific and Graded by Frailty and Inflammation in C57BL/6 Mice

We investigated whether late-life changes in cardiac structure and function were related to high levels of frailty and inflammation in male and female mice. Frailty (frailty index), ventricular structure/function (echocardiography), and serum cytokines (multiplex immunoassay) were measured in 16- and 23-month-old mice. Left ventricular (LV) mass and septal wall thickness increased with age in both sexes. Ejection fraction increased with age in males (60.4 ± 1.4 vs 68.9 ± 1.8%; p < .05) but not females (58.8 ± 2.5 vs 62.6 ± 2.4%). E/A ratios declined with age in males (1.6 ± 0.1 vs 1.3 ± 0.1; p < .05) but not females (1.4 ± 0.1 vs 1.3 ± 0.1) and this was accompanied by increased ventricular collagen levels in males. These changes in ejection fraction (r = 0.52; p = .01), septal wall thickness (r = 0.59; p = .002), E/A ratios (r = −0.49; p = .04), and fibrosis (r = 0.82; p = .002) were closely graded by frailty scores in males. Only septal wall thickness and LV mass increased with frailty in females. Serum cytokines changed modestly with age in both sexes. Nonetheless, in males, E/A ratios, LV mass, LV posterior wall thickness, and septal wall thickness increased as serum cytokines increased (eg, IL-6, IL-3, IL-1α, IL-1β, tumor necrosis factor-α, eotaxin, and macrophage inflammatory protein-1α), while ejection fraction declined with increasing IL-3 and granulocyte-macrophage colony stimulating factor. Cardiac outcomes were not correlated with inflammatory cytokines in females. Thus, changes in cardiac structure and function in late life are closely graded by both frailty and markers of inflammation, but this occurs primarily in males. This suggests poor overall health and inflammation drive maladaptive changes in older male hearts, while older females may be resistant to these adverse effects of frailty.

Targeted Deletion of Interleukin-6 in a Mouse Model of Chronic Inflammation Demonstrates Opposing Roles in Aging: Benefit and Harm.

Chronic inflammation (CI) in older adults is associated with reduced health span and life span. Interleukin-6 (IL-6) is one CI marker that is strongly associated with adverse health outcomes and mortality in aging. We have previously characterized a mouse model of frailty and chronic inflammatory pathway activation (IL-10tm/tm, IL-10 KO) that demonstrates the upregulation of numerous proinflammatory cytokines, including IL-6. We sought to identify a more specific role for IL-6 within the context of CI and aging and developed a mouse with targeted deletion of both IL-10 and IL-6 (IL-10tm/tm/IL-6tm/tm, DKO). Phenotypic characteristics, cytokine measurements, cardiac myocardial oxygen consumption, physical function, and survival were measured in DKO mice and compared to age- and gender-matched IL-10 KO and wild-type mice. Our findings demonstrate that selective knockdown of IL-6 in a frail mouse with CI resulted in the reversal of some of the CI-associated changes. We observed increased protective mitochondrial-associated lipid metabolites, decreased cardiac oxaloacetic acid, improved myocardial oxidative metabolism, and better short-term functional performance in DKO mice. However, the DKO mice also demonstrated higher mortality. This work shows the pleiotropic effects of IL-6 on aging and frailty.

Relevance of oxidative stress and inflammation in frailty based on human studies and mouse models.

Frailty represents a state of vulnerability and increases the risk of negative health outcomes, which is becoming an important public health problem. Over recent years, multiple independent studies have attempted to identify biomarkers that can predict, diagnose, and monitor frailty at the biological level. Among them, several promising candidates have been associated with frailty status including antioxidants and free radicals, and also inflammatory response biomarkers. In this review, we will summarize the more recent advances in this field. Moreover, the identification of scales and measurements to detect and quantify frailty in aged mice, as well as the generation of mouse models, have started to unravel the underlying biological and molecular mechanisms of frailty. We will discuss them here with an emphasis on murine models with overexpression of glucose-6-phosphate dehydrogenase and loss of function of superoxide dismutase and interleukin 10, which reveal that altered oxidative stress and inflammation pathways are involved in the physiopathology of frailty. In summary, we provide the current available evidence, from both human cohorts and experimental animal models, that highlights oxidative damage and inflammation as relevant biomarkers and drivers of frailty.

The role of Interleukin-6 in age-related frailty syndrome

Abstract Aging is accompanied by chronic low-grade inflammation – a term which relates to increase in serum levels of cytokines such as interleukin 6 (IL-6), TNF-α and acute phase proteins such as C-reactive protein. Multiple studies in humans have shown that IL-6 most reliably increases with aging. Frailty is a geriatric syndrome characterized by decrease in physical ability and increased vulnerability to stressors. One of the most consistent findings in frail subjects is an elevation of serum IL-6 but we lack research which delineates is this a causational relationship. We developed a mouse model with inducible IL-6 expression (IL-6TET-ON/+ mice) that enables overexpression of IL-6 following stimulation with doxycycline (dox) administrated in food. IL-6 induction was dose dependent and led to increase in frailty as measured by 30-point murine clinical frailty index. Mice induced to express high levels of IL-6 quickly displayed an increase in frailty index, decrease in muscle grip strength and loss of fat. Mice induced to express 3–4 fold increase in IL-6 similar to frail humans displayed similar changes after months of induction. We measured IL-6 levels in serum and various tissues (gut, muscle, adipose, spleen) of aged (28-month-old) frail mice and adult controls as well as dox fed IL-6TET-ON/+ mice. We observed that IL-6 levels were increased in serums and spleen homogenates of aged mice and IL-6TET-ON/+ mice but not in other tissues. We determined that neutrophils were the main producers of IL-6 and their numbers were higher in spleens of aged and IL-6TET-ON/+ mice compared to adult controls. We conclude that elevated IL-6 serum levels are directly associated with age-related frailty and that spleen neutrophils are likely the main producers of IL-6.

Association between a non‐invasive assessment of frailty and vascular dysfunction in old mice

Advancing age is characterized by not only an increased risk for cardiovascular diseases (CVDs), but also a decline in functional reserve and impaired adaptive capacity across multiple physiologic systems, also known as frailty. Impaired vascular function is a known contributor to CVDs and potentially has a role in increased frailty. In patients with overt disease, measures of frailty are related to vascular endothelial cell dysfunction. However, the relation between vascular endothelial function and frailty in a non‐disease population is unknown. It is also unknown if dysfunction of a particular vascular bed is more closely related to frailty.To examine the relation between vascular function and frailty in the context of similar genetics and environment, we studied wildtype C57BL/6 mice. In young (9 mo, n=7) and old (23–30 mo, n=27) male and female mice, we assessed endothelial‐dependent dilation (EDD) ex vivo in isolated, pressurized mesentery arteries and middle cerebral arteries (MCAs) by measuring the dilation to increasing doses of acetylcholine (ACh, 10−9 to 10−4 M) after pre‐constriction (by phenylephrine, 2 μM). Endothelium‐independent dilation (EID) was assessed by the dose response to sodium nitroprusside (SNP, 10−10 to 10−4 M) after pre‐constriction.Mouse frailty was assessed using a previously established non‐invasive 31‐item frailty index based on clinical signs of deterioration. Clinical assessments included evaluation of the integumentary, musculoskeletal, vestibulocochlear/auditory, ocular, nasal, digestive, urogenital, and respiratory systems as well as body weight, body surface temperature, and signs of discomfort. The severity of each deficit was assessed by two independent observers and assigned a value between 0–1, with a higher score indicating more severe frailty.Frailty index was ~6 fold greater in old compared with young mice (p&lt;0.001). Maximal EDD was 16.1% lower in mesentery arteries (p=0.04) and 11.3% lower in MCAs (p=0.12) for old compared with young mice. Among the old mice, frailty index was correlated with mesentery artery maximal EDD (r=−0.53 p=0.002), and remained significant after controlling for age (partial correlation: r=−0.38, p=0.03). In contrast, there was no correlation between frailty index and MCA maximal EDD among old mice (r=0.16, p=0.21). Among old mice, there was no correlation between frailty index and maximal EID in mesentery arteries or MCAs (p&gt;0.05).This study demonstrates that frailty, independent of age, is associated with mesentery artery endothelial dysfunction in wildtype, non‐diseased mice. However, it remains unknown whether the mesentery artery dysfunction is a cause or consequence of the greater frailty. Interestingly, we found a lack of association between frailty and cerebral artery endothelial function. Future research is needed to determine the mechanisms linking frailty and mesentery artery dysfunction.Support or Funding InformationFunded by NIH R56AG064016

LAV-BPIFB4 associates with reduced frailty in humans and its transfer prevents frailty progression in old mice

Background: There is an increasing concern about age-related frailty because of the growing number of elderly people in the general population. The Longevity-Associated Variant (LAV) of the human BPIFB4 gene was found to correct endothelial dysfunction, one of the mechanisms underlying frailty, in aging mice whereas the RV-BPIFB4 variant induced opposite effects. Thus, we newly hypothesize that, besides being associated with life expectancy, BPIFB4 polymorphisms can predict frailty.Aim and Results: Here we investigated if the BPIFB4 haplotypes, LAV, wild-type (WT) and RV, differentially associate with frailty in a cohort of 237 elderly subjects from Calabria region in Southern Italy. Moreover, we studied the effect of systemic adeno-associated viral vector-mediated LAV-BPIFB4 gene transfer on the progression of frailty in aging mice. We found an inverse correlation of the homozygous LAV-BPIFB4 haplotype with frailty in elderly subjects. Conversely, carriers of the RV-BPIFB4 haplotype showed an increase in the frailty status and risk of death. Moreover, in old mice, LAV-BPIFB4 gene transfer delayed frailty progression.Conclusions: These data indicate that specific BPIFB4 haplotypes could represent useful genetic markers of frailty. In addition, horizontal transfer of a healthy gene variant can attenuate frailty in aging organisms.