Senotherapeutic drug treatment ameliorates chemotherapy-induced cachexia.

Cachexia is a debilitating skeletal muscle wasting condition for which we currently lack effective treatments. In the context of cancer, certain chemotherapeutics cause DNA damage and cellular senescence. Senescent cells exhibit chronic activation of the transcription factor NF-κB, a known mediator of the proinflammatory senescence-associated secretory phenotype (SASP) and skeletal muscle atrophy. Thus, targeting NF-κB represents a logical therapeutic strategy to alleviate unintended consequences of genotoxic drugs. Herein, we show that treatment with the IKK/NF-κB inhibitor SR12343 during a course of chemotherapy reduces markers of cellular senescence and the SASP in liver, skeletal muscle, and circulation and, correspondingly, attenuates features of skeletal muscle pathology. Lastly, we demonstrate that SR12343 mitigates chemotherapy-induced reductions in body weight, lean mass, fat mass, and muscle strength. These findings support senescent cells as a promising druggable target to counteract the SASP and skeletal muscle wasting in the context of chemotherapy.

p21 induces a senescence program and skeletal muscle dysfunction.

Recent work has established associations between elevated p21, the accumulation of senescent cells, and skeletal muscle dysfunction in mice and humans. Using a mouse model of p21 overexpression (p21OE), we examined if p21 mechanistically contributes to cellular senescence and pathological features in skeletal muscle. We show that p21 induces several core properties of cellular senescence in skeletal muscle, including an altered transcriptome, DNA damage, mitochondrial dysfunction, and the senescence-associated secretory phenotype (SASP). Furthermore, p21OE mice exhibit manifestations of skeletal muscle pathology, such as atrophy, fibrosis, and impaired physical function when compared to age-matched controls. These findings suggest p21 alone is sufficient to drive a cellular senescence program and reveal a novel source of skeletal muscle loss and dysfunction.

Characterization of cellular senescence in aging skeletal muscle.

Senescence is a cell fate that contributes to multiple aging-related pathologies. Despite profound age-associated changes in skeletal muscle (SkM), whether its constituent cells are prone to senesce has not been methodically examined. Herein, using single cell and bulk RNA-sequencing and complementary imaging methods on SkM of young and old mice, we demonstrate that a subpopulation of old fibroadipogenic progenitors highly expresses p16 Ink4a together with multiple senescence-related genes and, concomitantly, exhibits DNA damage and chromatin reorganization. Through analysis of isolated myofibers, we also detail a senescence phenotype within a subset of old cells, governed instead by p2 Cip1 . Administration of a senotherapeutic intervention to old mice countered age-related molecular and morphological changes and improved SkM strength. Finally, we found that the senescence phenotype is conserved in SkM from older humans. Collectively, our data provide compelling evidence for cellular senescence as a hallmark and potentially tractable mediator of SkM aging.

Exercise Counters the Age-Related Accumulation of Senescent Cells.

We propose the beneficial effects of exercise are in part mediated through the prevention and elimination of senescent cells. Exercise counters multiple forms of age-related molecular damage that initiate the senescence program and activates immune cells responsible for senescent cell clearance. Preclinical and clinical evidence for exercise as a senescence-targeting therapy and areas needing further investigation are discussed.

Deletion of SA ß-Gal+ cells using senolytics improves muscle regeneration in old mice.

Systemic deletion of senescent cells leads to robust improvements in cognitive, cardiovascular, and whole-body metabolism, but their role in tissue reparative processes is incompletely understood. We hypothesized that senolytic drugs would enhance regeneration in aged skeletal muscle. Young (3 months) and old (20 months) male C57Bl/6J mice were administered the senolytics dasatinib (5 mg/kg) and quercetin (50 mg/kg) or vehicle bi-weekly for 4 months. Tibialis anterior (TA) was then injected with 1.2% BaCl2 or PBS 7- or 28 days prior to euthanization. Senescence-associated ß-Galactosidase positive (SA ß-Gal+) cell abundance was low in muscle from both young and old mice and increased similarly 7 days following injury in both age groups, with no effect of D+Q. Most SA ß-Gal+ cells were also CD11b+ in young and old mice 7- and 14 days following injury, suggesting they are infiltrating immune cells. By 14 days, SA ß-Gal+/CD11b+ cells from old mice expressed senescence genes, whereas those from young mice expressed higher levels of genes characteristic of anti-inflammatory macrophages. SA ß-Gal+ cells remained elevated in old compared to young mice 28 days following injury, which were reduced by D+Q only in the old mice. In D+Q-treated old mice, muscle regenerated following injury to a greater extent compared to vehicle-treated old mice, having larger fiber cross-sectional area after 28 days. Conversely, D+Q blunted regeneration in young mice. In vitro experiments suggested D+Q directly improve myogenic progenitor cell proliferation. Enhanced physical function and improved muscle regeneration demonstrate that senolytics have beneficial effects only in old mice.

Functional improvements to 6 months of physical activity are not related to changes in size or density of multiple lower-extremity muscles in mobility-limited older individuals.

Older adults are encouraged to engage in multicomponent physical activity, which includes aerobic and muscle-strengthening activities. The current work is an extension of the Vitality, Independence, and Vigor in the Elderly 2 (VIVE2) study - a 6-month multicenter, randomized, placebo-controlled trial of physical activity and nutritional supplementation in community dwelling 70-year-old seniors. Here, we examined whether the magnitude of changes in muscle size and quality differed between major lower-extremity muscle groups and related these changes to functional outcomes. We also examined whether daily vitamin-D-enriched protein supplementation could augment the response to structured physical activity. Forty-nine men and women (77 ± 5 yrs) performed brisk walking, muscle-strengthening exercises for the lower limbs, and balance training 3 times weekly for 6 months. Participants were randomized to daily intake of a nutritional supplement (20 g whey protein + 800 IU vitamin D), or a placebo. Muscle cross-sectional area (CSA) and radiological attenuation (RA) were assessed in 8 different muscle groups using single-slice CT scans of the hip, thigh, and calf at baseline and after the intervention. Walking speed and performance in the Short Physical Performance Battery (SPPB) were also measured. For both CSA and RA, there were muscle group × time interactions (P < 0.01). Significant increases in CSA were observed in 2 of the 8 muscles studied, namely the knee extensors (1.9%) and the hip adductors (2.8%). For RA, increases were observed in 4 of 8 muscle groups, namely the hip flexors (1.1 HU), hip adductors (0.9 HU), knee extensors (1.2 HU), and ankle dorsiflexors (0.8 HU). No additive effect of nutritional supplementation was observed. While walking speed (13%) and SPPB performance (38%) improved markedly, multivariate analysis showed that these changes were not associated with the changes in muscle CSA and RA after the intervention. We conclude that this type of multicomponent physical activity program results in significant improvements in physical function despite relatively small changes in muscle size and quality of some, but not all, of the measured lower extremity muscles involved in locomotion.

Skeletal muscle aging, cellular senescence, and senotherapeutics: Current knowledge and future directions.

Cellular senescence is a state of cell cycle arrest induced by several forms of metabolic stress. Senescent cells accumulate with advancing age and have a distinctive phenotype, characterized by profound chromatin alterations and a robust senescence-associated secretory phenotype (SASP) that exerts negative effects on tissue health, both locally and systemically. In preclinical models, pharmacological agents that eliminate senescent cells (senotherapeutics) restore health and youthful properties in multiple tissues. To date, however, very little is understood about the vulnerability of terminally-differentiated skeletal muscle fibers and the resident mononuclear cells that populate the interstitial microenvironment of skeletal muscle to senescence, and their contribution to the onset and progression of skeletal muscle loss and dysfunction with aging. Scientific advances in these areas have the potential to highlight new therapeutic approaches to optimize late-life muscle health. To this end, this review highlights the current evidence and the key questions that need to be addressed to advance the field's understanding of cellular senescence as a mediator of skeletal muscle aging and the potential for emerging senescent cell-targeting therapies to counter age-related deficits in muscle mass, strength, and function. This article is part of the Special Issue - Senolytics - Edited by Joao Passos and Diana Jurk.

Myonuclear transcriptional dynamics in response to exercise following satellite cell depletion.

Skeletal muscle is composed of post-mitotic myofibers that form a syncytium containing hundreds of myonuclei. Using a progressive exercise training model in the mouse and single nucleus RNA-sequencing (snRNA-seq) for high-resolution characterization of myonuclear transcription, we show myonuclear functional specialization in muscle. After 4 weeks of exercise training, snRNA-seq reveals that resident muscle stem cells, or satellite cells, are activated with acute exercise but demonstrate limited lineage progression while contributing to muscle adaptation. In the absence of satellite cells, a portion of nuclei demonstrates divergent transcriptional dynamics associated with mixed-fate identities compared with satellite cell replete muscles. These data provide a compendium of information about how satellite cells influence myonuclear transcription in response to exercise.

The Second Annual Symposium of the Midwest Aging Consortium: The Future of Aging Research in the Midwestern United States.

While the average human life span continues to increase, there is little evidence that this is leading to a contemporaneous increase in "healthy years" experienced by our aging population. Consequently, many scientists focus their research on understanding the process of aging and trialing interventions that can promote healthspan. The 2021 Midwest Aging Consortium consensus statement is to develop and further the understanding of aging and age-related disease using the wealth of expertise across universities in the Midwestern United States. This report summarizes the cutting-edge research covered in a virtual symposium held by a consortium of researchers in the Midwestern United States, spanning topics such as senescence biomarkers, serotonin-induced DNA protection, immune system development, multisystem impacts of aging, neural decline following severe infection, the unique transcriptional impact of calorie restriction of different fat depots, the pivotal role of fasting in calorie restriction, the impact of peroxisome dysfunction, and the influence of early life trauma on health. The symposium speakers presented data from studies conducted in a variety of common laboratory animals as well as less-common species, including Caenorhabditis elegans, Drosophila, mice, rhesus macaques, elephants, and humans. The consensus of the symposium speakers is that this consortium highlights the strength of aging research in the Midwestern United States as well as the benefits of a collaborative and diverse approach to geroscience.

Exercise reduces circulating biomarkers of cellular senescence in humans.

Cellular senescence has emerged as a significant and potentially tractable mechanism of aging and multiple aging-related conditions. Biomarkers of senescent cell burden, including molecular signals in circulating immune cells and the abundance of circulating senescence-related proteins, have been associated with chronological age and clinical parameters of biological age in humans. The extent to which senescence biomarkers are affected by interventions that enhance health and function has not yet been examined. Here, we report that a 12-week structured exercise program drives significant improvements in several performance-based and self-reported measures of physical function in older adults. Impressively, the expression of key markers of the senescence program, including p16, p21, cGAS, and TNFα, were significantly lowered in CD3+ T cells in response to the intervention, as were the circulating concentrations of multiple senescence-related proteins. Moreover, partial least squares discriminant analysis showed levels of senescence-related proteins at baseline were predictive of changes in physical function in response to the exercise intervention. Our study provides first-in-human evidence that biomarkers of senescent cell burden are significantly lowered by a structured exercise program and predictive of the adaptive response to exercise.

Satellite Cell Depletion Disrupts Transcriptional Coordination and Muscle Adaptation to Exercise.

Satellite cells are required for postnatal development, skeletal muscle regeneration across the lifespan, and skeletal muscle hypertrophy prior to maturity. Our group has aimed to address whether satellite cells are required for hypertrophic growth in mature skeletal muscle. Here, we generated a comprehensive characterization and transcriptome-wide profiling of skeletal muscle during adaptation to exercise in the presence or absence of satellite cells in order to identify distinct phenotypes and gene networks influenced by satellite cell content. We administered vehicle or tamoxifen to adult Pax7-DTA mice and subjected them to progressive weighted wheel running (PoWeR). We then performed immunohistochemical analysis and whole-muscle RNA-seq of vehicle (SC+) and tamoxifen-treated (SC-) mice. Further, we performed single myonuclear RNA-seq to provide detailed information on how satellite cell fusion affects myonuclear transcription. We show that while skeletal muscle can mount a robust hypertrophic response to PoWeR in the absence of satellite cells, growth, and adaptation are ultimately blunted. Transcriptional profiling reveals several gene networks key to muscle adaptation are altered in the absence of satellite cells.

Depletion of resident muscle stem cells negatively impacts running volume, physical function, and muscle fiber hypertrophy in response to lifelong physical activity.

To date, studies that have aimed to investigate the role of satellite cells during adult skeletal muscle adaptation and hypertrophy have utilized a nontranslational stimulus and/or have been performed over a relatively short time frame. Although it has been shown that satellite cell depletion throughout adulthood does not drive skeletal muscle loss in sedentary mice, it remains unknown how satellite cells participate in skeletal muscle adaptation to long-term physical activity. The current study was designed to determine whether reduced satellite cell content throughout adulthood would influence the transcriptome-wide response to physical activity and diminish the adaptive response of skeletal muscle. We administered vehicle or tamoxifen to adult Pax7-diphtheria toxin A (DTA) mice to deplete satellite cells and assigned them to sedentary or wheel-running conditions for 13 mo. Satellite cell depletion throughout adulthood reduced balance and coordination, overall running volume, and the size of muscle proprioceptors (spindle fibers). Furthermore, satellite cell participation was necessary for optimal muscle fiber hypertrophy but not adaptations in fiber type distribution in response to lifelong physical activity. Transcriptome-wide analysis of the plantaris and soleus revealed that satellite cell function is muscle type specific; satellite cell-dependent myonuclear accretion was apparent in oxidative muscles, whereas initiation of G protein-coupled receptor (GPCR) signaling in the glycolytic plantaris may require satellite cells to induce optimal adaptations to long-term physical activity. These findings suggest that satellite cells play a role in preserving physical function during aging and influence muscle adaptation during sustained periods of physical activity.

Phosphorylation of eukaryotic initiation factor 4E is dispensable for skeletal muscle hypertrophy.

The eukaryotic initiation factor 4E (eIF4E) is a major mRNA cap-binding protein that has a central role in translation initiation. Ser209 is the single phosphorylation site within eIF4E and modulates its activity in response to MAPK pathway activation. It has been reported that phosphorylation of eIF4E at Ser209 promotes translation of key mRNAs, such as cyclin D1, that regulate ribosome biogenesis. We hypothesized that phosphorylation at Ser209 is required for skeletal muscle growth in response to a hypertrophic stimulus by promoting ribosome biogenesis. To test this hypothesis, wild-type (WT) and eIF4E knocked-in (KI) mice were subjected to synergist ablation to induce muscle hypertrophy of the plantaris muscle as the result of mechanical overload; in the KI mouse, Ser209 of eIF4E was replaced with a nonphosphorylatable alanine. Contrary to our hypothesis, we observed no difference in the magnitude of hypertrophy between WT and KI groups in response to 14 days of mechanical overload induced by synergist ablation. Similarly, the increases in cyclin D1 protein levels, ribosome biogenesis, and translational capacity did not differ between WT and KI groups. Based on these findings, we conclude that phosphorylation of eIF4E at Ser209 is dispensable for skeletal muscle hypertrophy in response to mechanical overload.

Resident muscle stem cells are not required for testosterone-induced skeletal muscle hypertrophy.

It is postulated that testosterone-induced skeletal muscle hypertrophy is driven by myonuclear accretion as the result of satellite cell fusion. To directly test this hypothesis, we utilized the Pax7-DTA mouse model to deplete satellite cells in skeletal muscle followed by testosterone administration. Pax7-DTA mice (6 mo of age) were treated for 5 days with either vehicle [satellite cell replete (SC+)] or tamoxifen [satellite cell depleted (SC-)]. Following a washout period, a testosterone propionate or sham pellet was implanted for 21 days. Testosterone administration caused a significant increase in muscle fiber cross-sectional area in SC+ and SC- mice in both oxidative (soleus) and glycolytic (plantaris and extensor digitorum longus) muscles. In SC+ mice treated with testosterone, there was a significant increase in both satellite cell abundance and myonuclei that was completely absent in testosterone-treated SC- mice. These findings provide direct evidence that testosterone-induced muscle fiber hypertrophy does not require an increase in satellite cell abundance or myonuclear accretion.Listen to a podcast about this Rapid Report with senior author E. E. Dupont-Versteegden (https://ajpcell.podbean.com/e/podcast-on-paper-that-shows-testosterone-induced-skeletal-muscle-hypertrophy-does-not-need-muscle-stem-cells/).

Elevated myonuclear density during skeletal muscle hypertrophy in response to training is reversed during detraining.

Myonuclei gained during exercise-induced skeletal muscle hypertrophy may be long-lasting and could facilitate future muscle adaptability after deconditioning, a concept colloquially termed "muscle memory." The evidence for this is limited, mostly due to the lack of a murine exercise-training paradigm that is nonsurgical and reversible. To address this limitation, we developed a novel progressive weighted-wheel-running (PoWeR) model of murine exercise training to test whether myonuclei gained during exercise persist after detraining. We hypothesized that myonuclei acquired during training-induced hypertrophy would remain following loss of muscle mass with detraining. Singly housed female C57BL/6J mice performed 8 wk of PoWeR, while another group performed 8 wk of PoWeR followed by 12 wk of detraining. Age-matched sedentary cage-dwelling mice served as untrained controls. Eight weeks of PoWeR yielded significant plantaris muscle fiber hypertrophy, a shift to a more oxidative phenotype, and greater myonuclear density than untrained mice. After 12 wk of detraining, the plantaris muscle returned to an untrained phenotype with fewer myonuclei. A finding of fewer myonuclei simultaneously with plantaris deconditioning argues against a muscle memory mechanism mediated by elevated myonuclear density in primarily fast-twitch muscle. PoWeR is a novel, practical, and easy-to-deploy approach for eliciting robust hypertrophy in mice, and our findings can inform future research on the mechanisms underlying skeletal muscle adaptive potential and muscle memory.

Progressive Resistance Training Improves Torque Capacity and Strength in Mobility-Limited Older Adults.

BACKGROUND: Progressive resistance training (PRT) is consistently shown to improve muscle strength in older adults. The efficacy of PRT to improve muscle fatigue in older adults with demonstrated mobility limitations remains unclear. METHODS: Mobility-limited (Short Physical Performance Battery [SPPB] ≤ 9) older adults (age 70-92 years) were recruited for this study and randomized to either PRT or home-based flexibility (FLEX) 3 d/wk for 12 weeks. Muscle fatigue and strength outcomes were assessed at baseline and 12 weeks. The primary outcome was torque capacity, a composite measure of strength and fatigue, defined as the sum of peak torques from an isokinetic fatigue test. RESULTS: Seventy participants were randomized (mean [SD] age 78.9 [5.4] years; 60% female; mean [SD] SPPB 7.5 [1.6]). At follow-up, the PRT group improved significantly in torque capacity, mean between-group difference (95% confidence interval) 466.19 (138.4, 793.97) Nm (p = .006), and maximal strength 127.3 (60.96, 193.61) Nm (p = .0003), when compared with FLEX group. Neither group demonstrated significant changes in muscle fatigue or torque variability. CONCLUSION: Twelve weeks of PRT improved torque capacity, as well as strength in mobility-limited older adults. These results demonstrate PRT improves multiple age-related muscular impairments.

Translating the Lifestyle Interventions and Independence for Elders Clinical Trial to Older Adults in a Real-World Community-Based Setting.

BACKGROUND: The Lifestyle Interventions and Independence for Elders (LIFE) clinical trial demonstrated that a structured program of physical activity (PA) reduced mobility-disability in older adults by up to 28%. It remains unknown whether the benefits of LIFE PA can be translated to older adults at risk for mobility-disability in real-world community-based settings. To address this knowledge gap, we conducted the ENhancing independence using Group-based community interventions for healthy AGing in Elders (ENGAGE) pilot study and examined the safety, feasibility, and preliminary effectiveness of translating LIFE PA to a community-based senior center. METHODS: Forty older adults with severe lower extremity functional limitations (age: 76.9 ± 7.3 years; body mass index: 32.7 ± 8 kg/m2; 85% female; short physical performance battery score: 6.3 ± 2.2) were randomized to 24 weeks of PA or a health education control intervention. RESULTS: Community-based PA was safe (serious adverse events: PA vs health education, 0:2; nonserious adverse events: PA vs health education, 3:1) and participants successfully adhered to the PA intervention (65.2%). Compared to health education, PA participants who attended ≥25% of scheduled visits had meaningful and sustained short physical performance battery improvements at follow-up (between group short physical performance battery score differences: ~0.7 units). CONCLUSIONS: ENGAGE has demonstrated the preliminary safety, feasibility, and effectiveness of LIFE PA in a real-world community-based setting. Larger-scale translational studies are needed to further disseminate the benefits of LIFE PA to vulnerable older adults in a variety of community-based settings.

A novel tetracycline-responsive transgenic mouse strain for skeletal muscle-specific gene expression.

BACKGROUND: The tetracycline-responsive system (Tet-ON/OFF) has proven to be a valuable tool for manipulating gene expression in an inducible, temporal, and tissue-specific manner. The purpose of this study was to create and characterize a new transgenic mouse strain utilizing the human skeletal muscle α-actin (HSA) promoter to drive skeletal muscle-specific expression of the reverse tetracycline transactivator (rtTA) gene which we have designated as the HSA-rtTA mouse. METHODS: To confirm the HSA-rtTA mouse was capable of driving skeletal muscle-specific expression, we crossed the HSA-rtTA mouse with the tetracycline-responsive histone H2B-green fluorescent protein (H2B-GFP) transgenic mouse in order to label myonuclei. RESULTS: Reverse transcription-PCR confirmed skeletal muscle-specific expression of rtTA mRNA, while single-fiber analysis showed highly effective GFP labeling of myonuclei in both fast- and slow-twitch skeletal muscles. Pax7 immunohistochemistry of skeletal muscle cross-sections revealed no appreciable GFP expression in satellite cells. CONCLUSIONS: The HSA-rtTA transgenic mouse allows for robust, specific, and inducible gene expression across muscles of different fiber types. The HSA-rtTA mouse provides a powerful tool to manipulate gene expression in skeletal muscle.