Dietary restriction impacts health and lifespan of genetically diverse mice.

Caloric restriction extends healthy lifespan in multiple species1. Intermittent fasting, an alternative form of dietary restriction, is potentially more sustainable in humans, but its effectiveness remains largely unexplored2-8. Identifying the most efficacious forms of dietary restriction is key for developing interventions to improve human health and longevity9. Here we performed an extensive assessment of graded levels of caloric restriction (20% and 40%) and intermittent fasting (1 and 2 days fasting per week) on the health and survival of 960 genetically diverse female mice. We show that caloric restriction and intermittent fasting both resulted in lifespan extension in proportion to the degree of restriction. Lifespan was heritable and genetics had a larger influence on lifespan than dietary restriction. The strongest trait associations with lifespan included retention of body weight through periods of handling-an indicator of stress resilience, high lymphocyte proportion, low red blood cell distribution width and high adiposity in late life. Health effects differed between interventions and exhibited inconsistent relationships with lifespan extension. 40% caloric restriction had the strongest lifespan extension effect but led to a loss of lean mass and changes in the immune repertoire that could confer susceptibility to infections. Intermittent fasting did not extend the lifespan of mice with high pre-intervention body weight, and two-day intermittent fasting was associated with disruption of erythroid cell populations. Metabolic responses to dietary restriction, including reduced adiposity and lower fasting glucose, were not associated with increased lifespan, suggesting that dietary restriction does more than just counteract the negative effects of obesity. Our findings indicate that improving health and extending lifespan are not synonymous and raise questions about which end points are the most relevant for evaluating aging interventions in preclinical models and clinical trials.

The freedom to run: developing an autonomous robot matching the needs of visually impaired citizens to technology opportunities.

Purpose: Visual impairment poses significant challenges in daily life, especially when navigating unfamiliar environments, resulting in inequalities and reduced quality of life. This study aimed to gain an in-depth understanding of the needs and perspectives of visually impaired people in sports-related contexts through surveys and focus groups, and to understand whether their needs are being met by current technological solutions.Materials and methods: To accomplish this, opinions gathered from focus groups and interviews were compared to the technological solutions found in the literature. Since many unmet needs were identified, participants from associations and organizations were asked to identify key characteristics for the development of a robot guide. The results underscored the paramount importance of an easy-to-use guide that offers accurate and personalized assistance. Participants expressed a strong desire for advanced features such as object recognition and navigation in complex environments, as well as adaptability to the user's speed while providing the necessary safety features to ensure a high level of autonomy.Results: This research serves as a bridge between technological advances and the needs of the visually impaired, contributing to a more accessible and inclusive society. By addressing the unique challenges faced by the visually impaired individuals and tailoring technology to meet their needs, this study takes a significant step toward reducing disparities and improving the independence and quality of life for this community.Conclusions: As technology continues to advance, it has the potential to be a powerful tool in breaking down barriers and fostering a world where everyone, regardless of their visual ability, can navigate the world with confidence and ease.

Inclusive design: Recognizing the importance of incorporating the unique requirements and perspectives of visually impaired individuals can guide the development of rehabilitation technology and services, ensuring they effectively support daily activities and active participation in sports and physical pursuits.Tailored-assistive technology: Understanding the specific needs of visually impaired individuals with regards to assistive technology, such as dependable robotic guides and essential features, can inform the design and customization of rehabilitation aids to enhance mobility and independence.Promising technologies: Exploring promising technologies like Aira, Be My Eyes, RoboCart, and Wayband can inspire the integration of these innovations into rehabilitation programs, facilitating better orientation, mobility, and accessibility for individuals with visual impairments.Continued research and development: Emphasizing the necessity for ongoing research and development efforts underscores the importance of advancing rehabilitation solutions that effectively address the distinct needs of visually impaired individuals, particularly in navigating unfamiliar environments.

Longitudinal fragility phenotyping contributes to the prediction of lifespan and age-associated morbidity in C57BL/6 and Diversity Outbred mice.

Aging studies in mammalian models often depend on natural lifespan data as a primary outcome. Tools for lifespan prediction could accelerate these studies and reduce the need for veterinary intervention. Here, we leveraged large-scale longitudinal frailty and lifespan data on two genetically distinct mouse cohorts to evaluate noninvasive strategies to predict life expectancy in mice. We applied a modified frailty assessment, the Fragility Index, derived from existing frailty indices with additional deficits selected by veterinarians. We developed an ensemble machine learning classifier to predict imminent mortality (95% proportion of life lived [95PLL]). Our algorithm represented improvement over previous predictive criteria but fell short of the level of reliability that would be needed to make advanced prediction of lifespan and thus accelerate lifespan studies. Highly sensitive and specific frailty-based predictive endpoint criteria for aged mice remain elusive. While frailty-based prediction falls short as a surrogate for lifespan, it did demonstrate significant predictive power and as such must contain information that could be used to inform the conclusion of aging experiments. We propose a frailty-based measure of healthspan as an alternative target for aging research and demonstrate that lifespan and healthspan criteria reveal distinct aspects of aging in mice.

Depression and Anxiety in Pediatric Patients with Beckwith-Wiedemann Syndrome: A Pilot Study.

The study's aim was to determine the prevalence of depression and anxiety in children with Beckwith-Wiedemann syndrome (BWS) and their effects on social relationships and family acceptance. The Pediatric Symptom Checklist-35 items (PSC-35), Screen for Child Anxiety Related Emotional Disorders (SCARED), and the Vineland Adaptive Behavior Scale Second Edition (VABS-II) were administered to the children. The parental Acceptance Rejection/Control Questionnaire (PARQ/Control) and Zarit Burden Inventory (ZBI) were administered to parents. In total, 6 patients and 10 parents were included. Patients showed a significant presence of internalizing behavior in PSC-35 (mean, 7.66 ± 3.67), anxiety symptoms (SCARED: mean, 46.33 ± 17.50) and socialization difficulties (mean, 90.83 ± 10.09). Parents reported a perceived good acceptance (mean, 56.33 ± 1.03) and a moderate control (mean, 24.17 ± 1.83), but the burden level was ranked moderate to severe (mean, 59.33 ± 16.78). It was found that the severity of the burden level reported by parents was related to internalizing behavior (OR = 2.000; 95% CI = 0.479-3.521; p = 0.022) and anxiety symptoms (SCARED total score: OR = 3.000; 95% CI = 1.479-4.521; p = 0.005) of children. During psychological counseling in the context of BWS treatment, it is important to identify specific resources that can support patients and families in dealing with stress and identify any critical areas that could hinder the adaptation process.

Longitudinal Fragility Phenotyping Predicts Lifespan and Age-Associated Morbidity in C57BL/6 and Diversity Outbred Mice.

Aging studies in mammalian models often depend on natural lifespan data as a primary outcome. Tools for lifespan prediction could accelerate these studies and reduce the need for veterinary intervention. Here, we leveraged large-scale longitudinal frailty and lifespan data on two genetically distinct mouse cohorts to evaluate noninvasive strategies to predict life expectancy in mice. We applied a modified frailty assessment, the Fragility Index, derived from existing frailty indices with additional deficits selected by veterinarians. We developed an ensemble machine learning classifier to predict imminent mortality (95% proportion of life lived [95PLL]). Our algorithm represented improvement over previous predictive criteria but fell short of the level of reliability that would be needed to make advanced prediction of lifespan and thus accelerate lifespan studies. Highly sensitive and specific frailty-based predictive endpoint criteria for aged mice remain elusive. While frailty-based prediction falls short as a surrogate for lifespan, it did demonstrate significant predictive power and as such must contain information that could be used to inform the conclusion of aging experiments. We propose a frailty-based measure of healthspan as an alternative target for aging research and demonstrate that lifespan and healthspan criteria reveal distinct aspects of aging in mice.

Perioperative Management of a Pediatric Patient with Beckwith-Wiedemann Syndrome Undergoing a Partial Glossectomy According to Egyedi/Obwegeser.

Here, we report the perioperative management of a clinical case of a 6 year, 5 month old girl suffering from Beckwith-Wiedemann syndrome undergoing a partial glossectomy procedure in a patient with surgical indication for obstructive sleep apnea syndrome (OSAS), difficulty swallowing, feeding, and speech. On surgery day, Clonidine (4 µg/kg) was administered. Following this, a general anesthesia induction was performed by administering Sevoflurane, Fentanyl, continuous intravenous Remifentanil, and lidocaine to the vocal cords, and a rhinotracheal intubation with a size 4.5 tube was carried out. Before starting the procedure, a block of the Lingual Nerve was performed with Levobupivacaine. Analgosedation was maintained with 3% Sevoflurane in air and oxygen (FiO2 of 40%) and Remifentanil in continuous intravenous infusion at a rate of 0.08-0.15 µg/kg/min. The surgical procedure lasted 2 h and 32 min. At the end of the surgery, the patient was under close observation during the first 72 h. In the pediatric patient with Beckwith-Wiedemann syndrome submitted to major maxillofacial surgery, the difficulty in managing the airways in the preoperative phase during intubation and in the post-operative phase during extubation should be considered.

An Automated, Home-Cage, Video Monitoring-based Mouse Frailty Index Detects Age-associated Morbidity in C57BL/6 and Diversity Outbred Mice.

Frailty indexes (FIs) provide quantitative measurements of nonspecific health decline and are particularly useful as longitudinal monitors of morbidity in aging studies. For mouse studies, frailty assessments can be taken noninvasively, but they require handling and direct observation that is labor-intensive to the scientist and stress inducing to the animal. Here, we implement, evaluate, and provide a refined digital FI composed entirely of computational analyses of home-cage video and compare it to manually obtained frailty scores in both C57BL/6 and genetically heterogeneous Diversity Outbred mice. We show that the frailty scores assigned by our digital index correlate with both manually obtained frailty scores and chronological age. Thus, we provide an automated tool for frailty assessment that can be collected reproducibly, at scale, without substantial labor cost.

Age and diet shape the genetic architecture of body weight in diversity outbred mice.

Understanding how genetic variation shapes a complex trait relies on accurately quantifying both the additive genetic and genotype-environment interaction effects in an age-dependent manner. We used a linear mixed model to quantify diet-dependent genetic contributions to body weight measured through adulthood in diversity outbred female mice under five diets. We observed that heritability of body weight declined with age under all diets, except the 40% calorie restriction diet. We identified 14 loci with age-dependent associations and 19 loci with age- and diet-dependent associations, with many diet-dependent loci previously linked to neurological function and behavior in mice or humans. We found their allelic effects to be dynamic with respect to genomic background, age, and diet, identifying several loci where distinct alleles affect body weight at different ages. These results enable us to more fully understand and predict the effectiveness of dietary intervention on overall health throughout age in distinct genetic backgrounds.

Body weight is one trait influenced by genes, age and environmental factors. Both internal and external environmental pressures are known to affect genetic variation over time. However, it is largely unknown how all factors ­ including age ­ interact to shape metabolism and bodyweight. Wright et al. set out to quantify the interactions between genes and diet in ageing mice and found that the effect of genetics on mouse body weight changes with age. In the experiments, Wright et al. weighed 960 female mice with diverse genetic backgrounds, starting at two months of age into adulthood. The animals were randomized to different diets at six months of age. Some mice had unlimited food access, others received 20% or 40% less calories than a typical mouse diet, and some fasted one or two days per week. Variations in their genetic background explained about 80% of differences in mice's weight, but the influence of genetics relative to non-genetic factors decreased as they aged. Mice on the 40% calorie restriction diet were an exception to this rule and genetics accounted for 80% of their weight throughout adulthood, likely due to reduced influence from diet and reduced interactions between diet and genes. Several genes involved in metabolism, neurological function, or behavior, were associated with mouse weight. The experiments highlight the importance of considering interactions between genetics, environment, and age in determining complex traits like body weight. The results and the approaches used by Wright et al. may help other scientists learn more about how the genetic predisposition to disease changes with environmental stimuli and age.

Automated, high-dimensional evaluation of physiological aging and resilience in outbred mice.

Behavior and physiology are essential readouts in many studies but have not benefited from the high-dimensional data revolution that has transformed molecular and cellular phenotyping. To address this, we developed an approach that combines commercially available automated phenotyping hardware with a systems biology analysis pipeline to generate a high-dimensional readout of mouse behavior/physiology, as well as intuitive and health-relevant summary statistics (resilience and biological age). We used this platform to longitudinally evaluate aging in hundreds of outbred mice across an age range from 3 months to 3.4 years. In contrast to the assumption that aging can only be measured at the limits of animal ability via challenge-based tasks, we observed widespread physiological and behavioral aging starting in early life. Using network connectivity analysis, we found that organism-level resilience exhibited an accelerating decline with age that was distinct from the trajectory of individual phenotypes. We developed a method, Combined Aging and Survival Prediction of Aging Rate (CASPAR), for jointly predicting chronological age and survival time and showed that the resulting model is able to predict both variables simultaneously, a behavior that is not captured by separate age and mortality prediction models. This study provides a uniquely high-resolution view of physiological aging in mice and demonstrates that systems-level analysis of physiology provides insights not captured by individual phenotypes. The approach described here allows aging, and other processes that affect behavior and physiology, to be studied with improved throughput, resolution, and phenotypic scope.

Sex- and strain-specific effects of mitochondrial uncoupling on age-related metabolic diseases in high-fat diet-fed mice.

Mild uncoupling of oxidative phosphorylation is an intrinsic property of all mitochondria and may have evolved to protect cells against the production of damaging reactive oxygen species. Therefore, compounds that enhance mitochondrial uncoupling are potentially attractive anti-aging therapies; however, chronic ingestion is associated with a number of unwanted side effects. We have previously developed a controlled-release mitochondrial protonophore (CRMP) that is functionally liver-directed and promotes oxidation of hepatic triglycerides by causing a subtle sustained increase in hepatic mitochondrial inefficiency. Here, we sought to leverage the higher therapeutic index of CRMP to test whether mild mitochondrial uncoupling in a liver-directed fashion could reduce oxidative damage and improve age-related metabolic disease and lifespan in diet-induced obese mice. Oral administration of CRMP (20 mg/[kg-day] × 4 weeks) reduced hepatic lipid content, protein kinase C epsilon activation, and hepatic insulin resistance in aged (74-week-old) high-fat diet (HFD)-fed C57BL/6J male mice, independently of changes in body weight, whole-body energy expenditure, food intake, or markers of hepatic mitochondrial biogenesis. CRMP treatment was also associated with a significant reduction in hepatic lipid peroxidation, protein carbonylation, and inflammation. Importantly, long-term (49 weeks) hepatic mitochondrial uncoupling initiated late in life (94-104 weeks), in conjugation with HFD feeding, protected mice against neoplastic disorders, including hepatocellular carcinoma (HCC), in a strain and sex-specific manner. Taken together, these studies illustrate the complex variation of aging and provide important proof-of-concept data to support further studies investigating the use of liver-directed mitochondrial uncouplers to promote healthy aging in humans.

Impact of Large Granular Lymphocyte Leukemia on Blood DNA Methylation and Epigenetic Clock Modeling in Fischer 344 Rats.

Age-dependent differences in methylation at specific cytosine-guanine (CpG) sites have been used in "epigenetic clock" formulas to predict age. Deviations of epigenetic age from chronological age are informative of health status and are associated with adverse health outcomes, including mortality. In most cases, epigenetic clocks are performed on methylation from DNA extracted from circulating blood cells. However, the effect of neoplastic cells in the circulation on estimation and interpretation of epigenetic clocks is not well understood. Here, we explored this using Fischer 344 (F344) rats, a strain that often develops large granular lymphocyte leukemia (LGLL). We found clear histological markers of LGLL pathology in the spleens and livers of 27 out of 61 rats aged 17-27 months. We assessed DNA methylation by reduced representation bisulfite sequencing with coverage of 3 million cytosine residues. Although LGLL broadly increased DNA methylation variability, it did not change epigenetic aging. Despite this, the inclusion of rats with LGLL in clock training sets significantly altered predictor selection probability at 83 of 121 commonly utilized CpG sites. Furthermore, models trained on rat samples that included individuals with LGLL had greater absolute age error than those trained exclusively rats free of LGLL (39% increase; p < .0001). We conclude that the epigenetic signals for aging and LGLL are distinct, such that LGLL assessment is not necessary for valid measures of epigenetic age in F344 rats. The precision and architecture of constructed epigenetic clock formulas, however, can be influenced by the presence of neoplastic hematopoietic cells in training set populations.

A redox-mediated conformational change in NQO1 controls binding to microtubules and α-tubulin acetylation.

The localization of NQO1 near acetylated microtubules has led to the hypothesis that NQO1 may work in concert with the NAD+-dependent deacetylase SIRT2 to regulate acetyl α-tubulin (K40) levels on microtubules. NQO1 catalyzes the oxidation of NADH to NAD+ and may supplement levels of NAD+ near microtubules to aid SIRT2 deacetylase activity. While HDAC6 has been shown to regulate the majority of microtubule acetylation at K40, SIRT2 is also known to modulate microtubule acetylation (K40) in the perinuclear region. In this study we examined the potential roles NQO1 may play in modulating acetyl α-tubulin levels. Knock-out or knock-down of NQO1 or SIRT2 did not change the levels of acetyl α-tubulin in 16HBE human bronchial epithelial cells and 3T3-L1 fibroblasts; however, treatment with a mechanism-based inhibitor of NQO1 (MI2321) led to a short-lived temporal increase in acetyl α-tubulin levels in both cell lines without impacting the intracellular pools of NADH or NAD+. Inactivation of NQO1 by MI2321 resulted in lower levels of NQO1 immunostaining on microtubules, consistent with redox-dependent changes in NQO1 conformation as evidenced by the use of redox-specific, anti-NQO1 antibodies in immunoprecipitation studies. Given the highly dynamic nature of acetylation-deacetylation reactions at α-tubulin K40 and the crowded protein environment surrounding this site, disruption in the binding of NQO1 to microtubules may temporally disturb the physical interactions of enzymes responsible for maintaining the microtubule acetylome.

NQO1 protects obese mice through improvements in glucose and lipid metabolism.

Chronic nutrient excess leads to metabolic disorders and insulin resistance. Activation of stress-responsive pathways via Nrf2 activation contributes to energy metabolism regulation. Here, inducible activation of Nrf2 in mice and transgenesis of the Nrf2 target, NQO1, conferred protection from diet-induced metabolic defects through preservation of glucose homeostasis, insulin sensitivity, and lipid handling with improved physiological outcomes. NQO1-RNA interaction mediated the association with and inhibition of the translational machinery in skeletal muscle of NQO1 transgenic mice. NQO1-Tg mice on high-fat diet had lower adipose tissue macrophages and enhanced expression of lipogenic enzymes coincident with reduction in circulating and hepatic lipids. Metabolomics data revealed a systemic metabolic signature of improved glucose handling, cellular redox, and NAD+ metabolism while label-free quantitative mass spectrometry in skeletal muscle uncovered a distinct diet- and genotype-dependent acetylation pattern of SIRT3 targets across the core of intermediary metabolism. Thus, under nutritional excess, NQO1 transgenesis preserves healthful benefits.

A rat epigenetic clock recapitulates phenotypic aging and co-localizes with heterochromatin.

Robust biomarkers of aging have been developed from DNA methylation in humans and more recently, in mice. This study aimed to generate a novel epigenetic clock in rats-a model with unique physical, physiological, and biochemical advantages-by incorporating behavioral data, unsupervised machine learning, and network analysis to identify epigenetic signals that not only track with age, but also relates to phenotypic aging. Reduced representation bisulfite sequencing (RRBS) data was used to train an epigenetic age (DNAmAge) measure in Fischer 344 CDF (F344) rats. This measure correlated with age at (r = 0.93) in an independent sample, and related to physical functioning (p=5.9e-3), after adjusting for age and cell counts. DNAmAge was also found to correlate with age in male C57BL/6 mice (r = 0.79), and was decreased in response to caloric restriction. Our signatures driven by CpGs in intergenic regions that showed substantial overlap with H3K9me3, H3K27me3, and E2F1 transcriptional factor binding.

Disulfiram Treatment Normalizes Body Weight in Obese Mice.

Obesity is a top public health concern, and a molecule that safely treats obesity is urgently needed. Disulfiram (known commercially as Antabuse), an FDA-approved treatment for chronic alcohol addiction, exhibits anti-inflammatory properties and helps protect against certain types of cancer. Here, we show that in mice disulfiram treatment prevented body weight gain and abrogated the adverse impact of an obesogenic diet on insulin responsiveness while mitigating liver steatosis and pancreatic islet hypertrophy. Additionally, disulfiram treatment reversed established diet-induced obesity and metabolic dysfunctions in middle-aged mice. Reductions in feeding efficiency and increases in energy expenditure were associated with body weight regulation in response to long-term disulfiram treatment. Loss of fat tissue and an increase in liver fenestrations were also observed in rats on disulfiram. Given the potent anti-obesogenic effects in rodents, repurposing disulfiram in the clinic could represent a new strategy to treat obesity and its metabolic comorbidities.

Iron-Dependent Trafficking of 5-Lipoxygenase and Impact on Human Macrophage Activation.

5-lipoxygenase (5-LOX) is a non-heme iron-containing dioxygenase expressed in immune cells that catalyzes the two initial steps in the biosynthesis of leukotrienes. It is well known that 5-LOX activation in innate immunity cells is related to different iron-associated pro-inflammatory disorders, including cancer, neurodegenerative diseases, and atherosclerosis. However, the molecular and cellular mechanism(s) underlying the interplay between iron and 5-LOX activation are largely unexplored. In this study, we investigated whether iron (in the form of Fe3+ and hemin) might modulate 5-LOX influencing its membrane binding, subcellular distribution, and functional activity. We proved by fluorescence resonance energy transfer approach that metal removal from the recombinant human 5-LOX, not only altered the catalytic activity of the enzyme, but also impaired its membrane-binding. To ascertain whether iron can modulate the subcellular distribution of 5-LOX in immune cells, we exposed THP-1 macrophages and human primary macrophages to exogenous iron. Cells exposed to increasing amounts of Fe3+ showed a redistribution (ranging from ~45 to 75%) of the cytosolic 5-LOX to the nuclear fraction. Accordingly, confocal microscopy revealed that acute exposure to extracellular Fe3+, as well as hemin, caused an overt increase in the nuclear fluorescence of 5-LOX, accompanied by a co-localization with the 5-LOX activating protein (FLAP) both in THP-1 macrophages and human macrophages. The functional relevance of iron overloading was demonstrated by a marked induction of the expression of interleukin-6 in iron-treated macrophages. Importantly, pre-treatment of cells with the iron-chelating agent deferoxamine completely abolished the hemin-dependent translocation of 5-LOX to the nuclear fraction, and significantly reverted its effect on interleukin-6 overexpression. These results suggest that exogenous iron modulates the biological activity of 5-LOX in macrophages by increasing its ability to bind to nuclear membranes, further supporting a role for iron in inflammation-based diseases where its homeostasis is altered and suggesting further evidence of risks related to iron overload.

Benefits of Caloric Restriction in Longevity and Chemical-Induced Tumorigenesis Are Transmitted Independent of NQO1.

Caloric restriction (CR) is the most potent nonpharmacological intervention known to both protect against carcinogenesis and delay aging in laboratory animals. There is a growing number of anticarcinogens and CR mimetics that activate NAD(P)H:quinone oxidoreductase 1 (NQO1). We have previously shown that NQO1, an antioxidant enzyme that acts as an energy sensor through modulation of intracellular redox and metabolic state, is upregulated by CR. Here, we used NQO1-knockout (KO) mice to investigate the role of NQO1 in both the aging process and tumor susceptibility, specifically in the context of CR. We found that NQO1 is not essential for the beneficial effects of CR on glucose homeostasis, physical performance, metabolic flexibility, life-span extension, and (unlike our previously observation with Nrf2) chemical-induced tumorigenesis.

A time to fast.

Nutrient composition and caloric intake have traditionally been used to devise optimized diets for various phases of life. Adjustment of meal size and frequency have emerged as powerful tools to ameliorate and postpone the onset of disease and delay aging, whereas periods of fasting, with or without reduced energy intake, can have profound health benefits. The underlying physiological processes involve periodic shifts of metabolic fuel sources, promotion of repair mechanisms, and the optimization of energy utilization for cellular and organismal health. Future research endeavors should be directed to the integration of a balanced nutritious diet with controlled meal size and patterns and periods of fasting to develop better strategies to prevent, postpone, and treat the socioeconomical burden of chronic diseases associated with aging.