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# The intersection of disuse-induced muscle atrophy and satellite cell content: Reply to Snijders, Nederveen, and Parise

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## Abstract

to the editor: We thank Snijders et al. ([5][1]) for raising important questions worthy of further discussion. In our study, 14 days of bed rest induced robust myofiber atrophy and muscle weakness that was accompanied by a reduction in satellite cell content ([1][2]). Individual changes in satellite

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... The SCs are essential for muscle fiber regeneration and repair and are the source of new myonuclei during muscle fiber hypertrophy (8); thus, preservation and activation of SCs during incidents of short-term atrophy may be beneficial for subsequent recovery. It is, however, debated whether SCs are lost during short-term muscle fiber atrophy (9,10). ...
... Immunohistochemical analysis of SC content revealed that SCs were not lost in the CON leg during hospitalization, and that E-Stim did not increase the SC content. It is important to emphasize that it is not fully clear whether SCs are lost during disuse atrophy (9,10). Without a significant reduction in the SC pool in the CON leg in the present study, it is not possible to conclude whether E-Stim could have preserved the SC pool in these patients. ...
Article
Aim: To examine changes in lean mass during hospitalization in geriatric patients and the effect of muscle activation by neuromuscular electrical stimulation. Methods: Thirteen patients (69-94 years) at a geriatric ward completed tests at hospital admission (day 2-3) and discharge (day 8-10). One leg received daily stimulation of the knee extensors while the other leg served as a control leg. Lean mass was evaluated by DEXA scans and muscle thickness by ultrasound scans. Muscle biopsies were collected from both legs at admission and discharge in 9 patients and analyzed for fibre size, satellite cell number and activation and expression of genes associated with muscle protein synthesis and breakdown, connective tissue and cellular stress. Results: The relative decline in leg lean mass and midthigh region lean mass was larger in the control (-2.8±1.5%) vs. the stimulated leg (-0.5±1.4%, p<0.05). Although there were no changes in fibre size or satellite cell number, the mRNA data revealed that, compared to control, the stimulation resulted in a downregulation of Myostatin (p<0.05) and a similar trend for MAFbx (p=0.099), together with an upregulation of Collagen I (p<0.001), TenascinC (p<0.001), CD68 (p<0.01) and Ki67 (p<0.05) mRNA. Conclusion: These findings demonstrate a moderate decline in leg lean mass during a hospital stay in geriatric patients, while leg lean mass was preserved with daily neuromuscular electrical muscle activation. At the cellular level the stimulation had a clear influence on suppression of atrophy signaling pathways in parallel with a stimulation of connective tissue and cellular remodeling processes.
... The progressive loss of muscle mass and function that occurs with aging is associated with morphological and structural alterations within skeletal muscle. For example, aged muscles have smaller fibers (Arentson-Lantz et al., 2016;Miljkovic et al., 2015;Schiaffino and Reggiani, 2011), a phenomenon that is particularly evident in type II fibers, along with a decrease in capillary content (Gueugneau et al., 2016). The decline in skeletal muscle mass (sarcopenia) advances slowly with healthy aging but can be greatly accelerated by concurrent diseases resulting in mobility limitation, disability, frailty and loss of independence (Beaudart et al., 2015;Cosquéric et al., 2006). ...
Article
ABSTRACT Objectives: Adequate muscle perfusion supports the transport of nutrients, oxygen and hormones into muscle fibers. Aging is associated with a substantial decrease in skeletal muscle capillarization, fiber size and oxidative capacity, which may be improved with regular physical activity. The aim of this study was to investigate the relationship between muscle capillarization and indices of muscle hypertrophy (i.e. lean mass; fiber cross sectional area (CSA)) in older adults before and after 12 weeks of progressive resistance exercise training (RET). Design: Interventional study Setting and Participants: 19 subjects (10 male and 9 female; 71.1±4.3 years; 27.6±3.2 BMI) were enrolled in the study and performed a whole body RET program for 12 weeks. Subjects where then retrospectively divided into a LOW or HIGH group, based on their pre-RET capillary-to-fiber perimeter exchange index (CFPE). Physical activity level, indices of capillarization (capillaries-to-fiber ratio, C:Fi; CFPE index and capillary-to-fiber interface, LC-PF index), muscle hypertrophy, muscle protein turnover and mitochondrial function were assessed before and after RET. Results: Basal capillarization (C:Fi; CFPE and LP-CF index) correlates with daily physical activity level (C:Fi, r=0.57, p=0.019; CFPE index, r=0.55, p=0.024; LC-PF index, r=0.56, p=0.022) and CFPE and LC-PF indices were also positively associated with oxidative capacity (respectively r=0.45, p=0.06; r=0.67, p=0.004). Following RET, subjects in the HIGH group underwent hypertrophy with significant improvements in muscle protein synthesis and muscle fiber CSA (p<0.05). However, RET did not promote muscle hypertrophy in the LOW group, but RET significantly increased muscle capillary density (p<0.05). Conclusion/Implications: Muscle fiber capillarization before starting an exercise training program may be predictive of the muscle hypertrophic response to RET in older adults. Increases in muscle fiber size following RET appear to be blunted when muscle capillarization is low, suggesting that an adequate initial capillarization is critical to achieve a meaningful degree of muscle adaptation to RET.
... This reduced type II fibre-associated SC content in the SR leg compared with healthy controls suggests that inactivity contributed to a contraction of the SC pool. Whether immobilization results in a contraction of the SC pool in younger adults is debatable 35,36 ; however, it appears that older skeletal muscle may be susceptible to inactivity-induced loss of SC. 32 Mechanistically, the factors responsible for the loss of SC with inactivity are not defined but may be related to cell apoptosis. For example, SC isolated from aged muscle display greater rates of apoptosis concomitant with rises in caspase family members and cell death genes. ...
... This reduced type II fibre-associated SC content in the SR leg compared with healthy controls suggests that inactivity contributed to a contraction of the SC pool. Whether immobilization results in a contraction of the SC pool in younger adults is debatable 35,36 ; however, it appears that older skeletal muscle may be susceptible to inactivity-induced loss of SC. 32 Mechanistically, the factors responsible for the loss of SC with inactivity are not defined but may be related to cell apoptosis. For example, SC isolated from aged muscle display greater rates of apoptosis concomitant with rises in caspase family members and cell death genes. ...
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Background Age‐related sarcopenia is accelerated by physical inactivity. Low‐load resistance exercise (LLRE) counters inactivity‐induced muscle atrophy in older adults, but changes in muscle fibre morphology are unstudied. We aimed to determine the impact of LLRE during short‐term inactivity (step‐reduction) on muscle fibre size and capillarity as well as satellite cell (SC) content in older skeletal muscle. Methods Fourteen older (~71 years) male adults underwent 14 days of step reduction (<1500 steps/day) while performing six sessions of LLRE (~30% maximal strength) with one leg (SR + EX) while the contralateral leg served as an untrained control (SR). Seven healthy ambulatory age‐matched male adults (~69 years) served as a comparator group (COM). Muscle biopsies were taken from the vastus lateralis after 14 days, and immunohistochemical analysis was performed to determine muscle fibre cross‐sectional area (CSA), myonuclear content, SC content (PAX7⁺ cells), and total (C:F) and fibre type‐specific (C:Fi) capillary‐to‐fibre ratios. Results Type I and II fibre CSA was greater in SR + EX compared with SR. Whereas there were no differences across fibre types between SR + EX and CON, type II fibre CSA was significantly lower in SR compared with COM. Type II myonuclear domain was greater in SR + EX compared with COM and SR. Pax7⁺ cells associated with type I and II fibres were lower in SR compared with SR + EX. Type II PAX7+ cells were also lower in SR compared with COM with a similar trend for type I fibres. There were trends for a lower C:Fi in SR compared with SR + EX for both fibre types with no differences for each compared with COM. Conclusions Minimal LLRE during a period of decreased physical activity is associated with greater muscle fibre CSA, SC content, and capillarization. These results support the use of LLRE as an effective countermeasure to inactivity‐induced alterations in muscle morphology with age.
... Instead, we observed robust myofiber atrophy and depletion of sat- ellite cells in both groups following bed rest; a relationship that was correlated (r = 0.34, p = 0.05) as others have dem- onstrated. 39,52 Myofiber atrophy is consistent in older adults after 4 days of cast immobilization. 51 However, it is impor- tant to point out that several short-term immobilization studies (5 days) 14,26,50 and even some longer bed rest studies (7 days) 53 failed to show atrophy at the fiber level as a result of disuse but rather noticed differences at the whole muscle level using CT and/or DXA. ...
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Short-term bed rest in older adults is characterized by significant loss in leg lean mass and strength posing significant health consequences. The purpose of this study was to determine in healthy older adults if the daily combination of neuromuscular electrical stimulation and protein supplementation (NMES+PRO) would protect muscle mass and function after 5-days of bed rest. Twenty healthy older adults (~70y) were subjected to 5-days of continuous bed rest and were randomized into one of two groups: NMES+PRO (n=10) or CON (n=10). The NMES+PRO group received bilateral NMES to quadriceps (40 min/session, 3x/day (morning, afternoon, evening) followed by an interventional protein supplement (17g). The CON group received an isocaloric equivalent beverage. Before and after bed rest, vastus lateralis biopsies occurred before and after acute essential amino acid (EAA) ingestion for purposes of acutely stimulating mechanistic target of rapamycin (mTORC1) signaling, a major regulator of muscle protein synthesis, in response to bed rest and NMES+PRO. Baseline (pre and post bed rest) muscle samples were also used to assess myofiber characteristics and gene expression of muscle atrophy markers. Thigh lean mass and muscle function were measured before and after bed rest. Five days of bed rest reduced thigh lean mass, muscle function, myofiber CSA, and satellite cell content, blunted EAA-induced mTORC1 signaling and increased Myostatin and MAFbx mRNA expression. Interestingly, NMES+PRO during bed rest maintained thigh lean mass, but not muscle function. Thigh muscle preservation during bed rest with NMES+PRO may partly be explained by attenuation of Myostatin and MAFbx mRNA expression rather than restoration of nutrient-induced mTORC1 signaling. We conclude that the combination of NMES and protein supplementation 3 times a day may be an effective therapeutic tool to use to preserve thigh muscle mass during periods of short-term hospitalization in older adults. However this combined intervention was not effective to prevent the loss in muscle function.
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Bed rest, a ground-based spaceflight analog, induces robust atrophy of skeletal muscle, an effect that is exacerbated with increasing age. We examined the effect of 14 days of bed rest on skeletal muscle satellite cell content and fiber type atrophy in middle-aged adults, an understudied age demographic with few overt signs of muscle aging that is representative of astronauts who perform long-duration spaceflight. Muscle biopsies were obtained from the vastus lateralis of healthy middle-aged adults (n=7 [4M;3F]; age: 51±1 y) before (Pre-BR) and after (Post-BR) 14 days of bed rest. Immunohistochemical analyses were used to quantify myosin heavy chain (MyHC) isoform expression, cross-sectional area (CSA), satellite cell and myonuclear content, and capillary density. Peak oxygen consumption, knee extensor strength, and body composition were also measured Pre- and Post-BR. Post-BR MyHC Type 2a fiber percentage was reduced and mean CSA decreased in all fiber types (-24±5%; P<0.05). Satellite cell content was also reduced Post-BR (-39±9%; P<0.05) and the change in satellite cell content was significantly correlated with the change in mean fiber CSA (R2=0.60; P<0.05). A decline in capillary density was observed Post-BR (-23±6%; P<0.05), and Post-BR capillary content was significantly associated with Post-BR peak aerobic capacity (R2=0.59; P<0.05). A subtle decline in myonuclear content occurred during bed rest (-5±1%; P<0.05). The rapid maladaptation of skeletal muscle to 14 days of mechanical unloading in middle-aged adults emphasizes the need for robust countermeasures to preserve muscle function in astronauts.
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Background: Physical inactivity triggers a rapid loss of muscle mass and function in older adults. Middle-aged adults show few phenotypic signs of aging yet may be more susceptible to inactivity than younger adults. Objective: The aim was to determine whether leucine, a stimulator of translation initiation and skeletal muscle protein synthesis (MPS), can protect skeletal muscle health during bed rest. Design: We used a randomized, double-blind, placebo-controlled trial to assess changes in skeletal MPS, cellular signaling, body composition, and skeletal muscle function in middle-aged adults (n = 19; age ± SEM: 52 ± 1 y) in response to leucine supplementation (LEU group: 0.06 g ∙ kg(-1) ∙ meal(-1)) or an alanine control (CON group) during 14 d of bed rest. Results: Bed rest decreased postabsorptive MPS by 30% ± 9% (CON group) and by 10% ± 10% (LEU group) (main effect for time, P < 0.05), but no differences between groups with respect to pre-post changes (group × time interactions) were detected for MPS or cell signaling. Leucine protected knee extensor peak torque (CON compared with LEU group: -15% ± 2% and -7% ± 3%; group × time interaction, P < 0.05) and endurance (CON compared with LEU: -14% ± 3% and -2% ± 4%; group × time interaction, P < 0.05), prevented an increase in body fat percentage (group × time interaction, P < 0.05), and reduced whole-body lean mass loss after 7 d (CON compared with LEU: -1.5 ± 0.3 and -0.8 ± 0.3 kg; group × time interaction, P < 0.05) but not 14 d (CON compared with LEU: -1.5 ± 0.3 and -1.0 ± 0.3 kg) of bed rest. Leucine also maintained muscle quality (peak torque/kg leg lean mass) after 14 d of bed-rest inactivity (CON compared with LEU: -9% ± 2% and +1% ± 3%; group × time interaction, P < 0.05). Conclusions: Bed rest has a profoundly negative effect on muscle metabolism, mass, and function in middle-aged adults. Leucine supplementation may partially protect muscle health during relatively brief periods of physical inactivity. This trial was registered at clinicaltrials.gov as NCT00968344.
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Skeletal muscle satellite cells are considered to play a crucial role in muscle fiber maintenance, repair and remodeling. Our knowledge of the role of satellite cells in muscle fiber adaptation has traditionally relied on in vitro cell and in vivo animal models. Over the past decade, a genuine effort has been made to translate these results to humans under physiological conditions. Findings from in vivo human studies suggest that satellite cells play a key role in skeletal muscle fiber repair/remodeling in response to exercise. Mounting evidence indicates that aging has a profound impact on the regulation of satellite cells in human skeletal muscle. Yet, the precise role of satellite cells in the development of muscle fiber atrophy with age remains unresolved. This review seeks to integrate recent results from in vivo human studies on satellite cell function in muscle fiber repair/remodeling in the wider context of satellite cell biology whose literature is largely based on animal and cell models.
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Aim: Muscle fibre hypertrophy is accompanied by an increase in myonuclear number, an increase in myonuclear domain size, or both. It has been suggested that increases in myonuclear domain size precede myonuclear accretion and subsequent muscle fibre hypertrophy during prolonged exercise training. In this study we assessed the changes in muscle fibre size, myonuclear and satellite cell content throughout 12 wks of resistance type exercise training in young men. Methods: Twenty-two young men (23±1 y) were assigned to a progressive, 12-wk resistance type exercise training program (3 sessions/wk). Muscle biopsies from the vastus lateralis muscle were taken before and after 2, 4, 8 and 12 wks of exercise training. Muscle fibre size, myonuclear content, myonuclear domain size, and satellite cell content were assessed by immunohistochemistry. Results: Type I and type II muscle fibre size increased gradually throughout the 12 wks of training (type I: 18±5%, type II: 41±6%, P<0.01). Myonuclear content increased significantly over time in both the type I (P<0.01) and type II (P<0.001) muscle fibres. No changes in type I and type II myonuclear domain size were observed at any time point throughout the intervention. Satellite cell content increased significantly over time in both type I and type II muscle fibres (P<0.001). Conclusion: Increases in myonuclear domain size do not appear to drive myonuclear accretion and muscle fibre hypertrophy during prolonged resistance type exercise training in vivo in humans. This article is protected by copyright. All rights reserved.
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Aim: Skeletal muscle satellite cells (SCs) are important for muscle repair and hypertrophy in response mechanical stimuli. Neuron-glial antigen 2 positive (NG2(+)) and alkaline phosphatase positive (ALP(+)) pericytes may provide an alternative source of myogenic progenitors and/or secrete paracrine factors to induce Pax7(+) SC proliferation and differentiation. The purpose of this study was to investigate NG2(+) and ALP(+) cell quantity, as well as SC content and activation in human skeletal muscle following prolonged concentric (Conc) or eccentric (Ecc) resistance training. Methods: Male subjects engaged in unilateral resistance training utilizing isolated Ecc or Conc contractions. After 12 weeks, muscle biopsies were analyzed for NG2(+) and ALP(+) pericytes, total Pax7(+) SCs, activated SCs (Pax7(+)MyoD(+)), and differentiating myogenic cells (Pax7(-) MyoD(+)). Results: NG2(+) cells localized to CD31(+) vessels and the majority co-expressed ALP. NG2(+) pericyte quantity decreased following both Conc and Ecc training (p<0.05). ALP(+) pericyte quantity declined following Conc (p<0.05), but not Ecc training. Conversely, total Pax7(+) SC content was elevated following Conc only (p<0.001), while Pax7(+)MyoD(+) SC content was increased following Conc and Ecc (p<0.001). Follow up analyses demonstrated that CD90(+) and PDGFRα(+) mononuclear cell proliferation was also increased in response to both Conc and Ecc training (p<0.01). Conclusion: Resistance training results in a decline in pericyte quantity and increase in mesenchymal progenitor cell proliferation, and these events likely influence SC pool expansion and increased activation observed post-training.
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Reduced vessel density in adipose tissue and skeletal muscle is associated with obesity and may result in decreased perfusion, decreased oxygen consumption, and insulin resistance. In the presence of VEGFA, Angiopoietin-2 (Angpt2) and Angiopoietin-1 (Angpt1) are central determinants of angiogenesis, with greater Angpt2:Angpt1 ratios promoting angiogenesis. In skeletal muscle, exercise training stimulates angiogenesis and modulates transcription of VEGFA, Angpt1, and Angpt2. However, it remains unknown whether exercise training stimulates vessel growth in human adipose tissue, and it remains unknown whether adipose angiogenesis is mediated by angiopoietin signaling. We sought to determine whether insulin-resistant subjects would display an impaired angiogenic response to aerobic exercise training. Insulin-sensitive (IS, N = 12) and insulin-resistant (IR, N = 14) subjects had subcutaneous adipose and muscle (vastus lateralis) biopsies before and after 12 weeks of cycle ergometer training. In both tissues, we measured vessels and expression of pro-angiogenic genes. Exercise training did not increase insulin sensitivity in IR Subjects. In skeletal muscle, training resulted in increased vessels/muscle fiber and increased Angpt2:Angpt1 ratio in both IR and IS subjects. However, in adipose, exercise training only induced angiogenesis in IS subjects, likely due to chronic suppression of VEGFA expression in IR subjects. These results indicate that skeletal muscle of IR subjects exhibits a normal angiogenic response to exercise training. However, the same training regimen is insufficient to induce angiogenesis in adipose tissue of IR subjects, which may help to explain why we did not observe improved insulin sensitivity following aerobic training. © 2015 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.
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Fully-sedated patients, being treated in the ICU, experience substantial skeletal muscle loss. Consequently, survival rate is reduced and full recovery after awakening is compromised. Neuromuscular electrical stimulation (NMES) represents an effective method to stimulate muscle protein synthesis and alleviate muscle disuse atrophy in healthy subjects. We investigated the efficacy of twice-daily NMES to alleviate muscle loss in six fully-sedated ICU patients admitted for acute critical illness (n=3 males, n=3 females; age 63±6 y; APACHE II disease severity-score: 29±2). One leg was subjected to twice-daily NMES of the quadriceps muscle for a period of 7±1 d while the other leg acted as non-stimulated control (CON). Directly before the first and on the morning after the final NMES session, quadriceps muscle biopsies were collected from both legs to assess muscle fiber-type specific cross-sectional area (CSA). Furthermore, phosphorylation status of key proteins involved in the regulation of muscle protein synthesis was assessed, and mRNA expression of selected genes was measured. In the CON leg, type I and type II muscle fiber CSA decreased by 16±9 and 24±7%, respectively (P<0.05). No muscle atrophy was observed in the stimulated leg. NMES increased mTOR phosphorylation by 19% when compared to baseline (P<0.05), with no changes in the CON leg. Furthermore, mRNA expression of key genes involved in muscle protein breakdown either declined (FOXO1; P<0.05) or remained unchanged (MAFBx and MuRF1), with no differences between legs. In conclusion, NMES represents an effective and feasible interventional strategy to prevent skeletal muscle atrophy in critically ill, comatose patients.
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Context. The exact relationship between the bed rest-induced loss of skeletal muscle and reductions in muscle strength and physical performance in the older individuals is still unclear. Objective. We examined the effect of 10 days of bed rest on changes in regional body composition, muscle strength, and functional status, and the relationship between these variables in older individuals. Design, Participants, and Intervention. Regional body composition was measured using dual energy x-ray absorptiometry. We also determined changes in leg strength and several indices of functional status, including walking speed. Results. Body weight, body mass index, and total and lower extremity lean mass decreased with bed rest. There were also significant reductions in knee extension one repetition maximum, isometric knee extension, knee extension 60° concentric, stair ascent time, stair ascent power, stair descent time, VO2 max, floor transfer test, 5-minute walk time, and chair stand. The overall change in total and lower extremity lean mass was also directly related to bed rest-induced reductions in one repetition maximum knee extension. Conclusions. Bed rest promoted overall declines in muscle mass, muscle strength, and physical function in older individuals. The changes in lean tissue were closely correlated with the bed rest-induced decline of muscle strength.
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Muscle disuse leads to a considerable loss of skeletal muscle mass and strength. However, the cellular mechanisms underlying disuse-induced muscle fibre atrophy remain to be elucidated. Therefore, we assessed the impact of muscle disuse on quadriceps muscle cross-sectional area (CSA), muscle fibre size and satellite cell content, and associated myocellular signalling pathways. Methods: Twelve healthy, young (24±1 y) men were subjected to 2 wks of one-legged knee immobilisation via a full leg cast. Before and immediately after the immobilisation period and after six weeks of natural rehabilitation, muscle strength (one-repetition maximum), muscle cross-sectional area (single slice CT-scan), and muscle fibre type characteristics (muscle biopsies) were assessed. Protein and/or mRNA expression of key genes (i.e. MyoD, myogenin and myostatin) in the satellite cell regulatory pathways were determined using Western blotting and rtPCR analyses, respectively. Results: Quadriceps CSA declined following immobilisation by 8±2% (P<0.05). In line, both type I and type II muscle fibre size decreased by 7±3% and 13±4%, respectively (P<0.05). No changes were observed in satellite cell content following immobilisation in either type I or type II muscle fibres. Muscle myogenin mRNA expression had doubled (P<0.05) while myostatin protein expression had decreased by 30±9% (P<0.05) following immobilisation. Muscle mass and strength returned to baseline values within six weeks of recovery without any specific rehabilitative program. Conclusion: Two weeks of muscle disuse leads to considerable loss of skeletal muscle mass and strength. The loss of muscle mass is attributed to both type I and type II muscle fibre atrophy, and is not accompanied by a decline in satellite cell content.
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The purpose of this study was to explore the possible role of muscle stem cells, also referred to as satellite cells (SCs), in adaptation and remodeling following a nonhypertrophic stimulus in humans. Muscle biopsies were obtained from the vastus lateralis of previously untrained women (n=15; age: 27±8 yr, BMI: 29±6 kg/m(2)) before and after 6 wk of aerobic interval training. The fiber type-specific SC response to training was analyzed using immunofluorescent microscopy of muscle cross sections. Following training, the number of SCs associated with fibers expressing myosin heavy-chain type I and II isoforms (hybrid fibers) increased (pre: 0.062±0.035 SC/hybrid fiber; post: 0.38±0.063 SC/hybrid fiber; P<0.01). In addition, there was a greater number of MyoD(+)/Pax7(-) SCs, indicative of differentiating SCs, associated with hybrid fibers (0.18±0.096 MyoD(+)/Pax7(-) SC/hybrid fiber) compared to type I (0.015±0.00615 MyoD(+)/Pax7(-) SC/type I fiber) or II (0.012±0.00454 MyoD(+)/Pax7(-) SC/type II fiber) fibers (P<0.05). There was also a training-induced increase in the number of hybrid fibers containing centrally located nuclei (15.1%) compared to either type I (3.4%) or II fibers (3.6%) (P<0.01). These data are consistent with the hypothesis that SCs contribute to the remodeling of muscle fibers even in the absence of hypertrophy.-Joanisse, S., Gillen, J. B., Bellamy, L. M., McKay, B. R., Tarnopolsky, M. A., Gibala, M. J., Parise, G. Evidence for the contribution of muscle stem cells to nonhypertrophic skeletal muscle remodeling in humans.
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Recovery of skeletal muscle mass from immobilisation-induced atrophy is faster in young than older individuals, yet the cellular mechanisms remain unknown. We examined the cellular and molecular regulation of muscle recovery in young and old human subjects subsequent to 2 weeks of immobility-induced muscle atrophy. Re-training consisted of 4 weeks of supervised resistive exercise in 9 older (OM: 67.3yrs, range 61-74) and 11 young (YM: 24.4yrs, range 21-30) males. Measures of myofiber area (MFA), Pax7-positive satellite cells (SC) associated with type I and type II muscle fibres, as well as gene expression analysis of key growth and transcription factors associated with local skeletal muscle milieu were performed after 2 weeks immobility (Imm) and following 3 days (+3d) and 4 weeks (+4wks) of re-training. OM demonstrated no detectable gains in MFA (VL muscle) and no increases in number of Pax7-positive SCs following 4 wks re-training, whereas YM increased their MFA (p<0.05), number of Pax7-positive cells, and had more Pax7-positive cells per type II fibre than OM at +3d and +4wks (p<0.05). No age-related differences were observed in mRNA expression of IGF-1Ea, MGF, MyoD1 and HGF with re-training, whereas myostatin expression levels were more down-regulated in YM compared to OM at +3d (p<0.05). In conclusion, the diminished muscle re-growth after immobilisation in elderly humans was associated with lesser response in satellite cell proliferation in combination with an age-specific regulation of myostatin. In contrast, expression of local growth factors did not seem to explain the age related difference in muscle mass recovery.
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Important insights concerning the molecular basis of skeletal muscle disuse-atrophy and aging related muscle loss have been obtained in cell culture and animal models, but these regulatory signaling pathways have not previously been studied in aging human muscle. In the present study, muscle atrophy was induced by immobilization in healthy old and young individuals to study the time-course and transcriptional factors underlying human skeletal muscle atrophy. The results reveal that irrespectively of age, mRNA expression levels of MuRF-1 and Atrogin-1 increased in the very initial phase (2-4 days) of human disuse-muscle atrophy along with a marked reduction in PGC-1α and PGC-1β (1-4 days) and a ∼10% decrease in myofiber size (4 days). Further, an age-specific decrease in Akt and S6 phosphorylation was observed in young muscle within the first days (1-4 days) of immobilization. In contrast, Akt phosphorylation was unchanged in old muscle after 2 days and increased after 4 days of immobilization. Further, an age-specific down-regulation of MuRF-1 and Atrogin-1 expression levels was observed following 2 weeks of immobilization, along with a slowing atrophy response in aged skeletal muscle. Neither the immediate loss of muscle mass, nor the subsequent age-differentiated signaling responses could be explained by changes in inflammatory mediators, apoptosis markers or autophagy indicators. Collectively, these findings indicate that the time-course and regulation of human skeletal muscle atrophy is age dependent, leading to an attenuated loss in aging skeletal muscle when exposed to longer periods of immobility-induced disuse.
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Human aging is accompanied by a progressive loss of muscle mass (sarcopenia). We tested the hypothesis that older males (OMs, 70±4 yr, n=9) would have a blunted myogenic response to a physiological stimulus compared to younger controls (21±3 yr, n=9). Subjects completed an acute bout of intense unilateral muscle loading. Young healthy males matched for body mass and activity level served as the control group. Muscle biopsies and blood were obtained before and at 3, 24, and 48 h after muscle loading. The muscle stem cell response was analyzed using flow cytometry, immunofluorescent microscopy, and standard protein and mRNA analysis. OMs had 35% fewer basal stem cells and a type II fiber-specific impairment in stem cell content and proliferation. Myogenic determination factor staining and cell cycle analysis illustrated a severely blunted progression through the myogenic program. Myostatin protein and mRNA were 2-fold higher in OMs. Stem cell-specific myostatin levels were not different at baseline; however, there were 67% more myostatin-positive type II-associated stem cells in OMs at 24 h. These data illustrate an age-related impairment of stem cell function in a fiber type-specific manner. The greater colocalization of myostatin with stem cells provides a mechanism for the impaired myogenic capacity of aged muscle.
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An important unresolved question in skeletal muscle plasticity is whether satellite cells are necessary for muscle fiber hypertrophy. To address this issue, a novel mouse strain (Pax7-DTA) was created which enabled the conditional ablation of >90% of satellite cells in mature skeletal muscle following tamoxifen administration. To test the hypothesis that satellite cells are necessary for skeletal muscle hypertrophy, the plantaris muscle of adult Pax7-DTA mice was subjected to mechanical overload by surgical removal of the synergist muscle. Following two weeks of overload, satellite cell-depleted muscle showed the same increases in muscle mass (approximately twofold) and fiber cross-sectional area with hypertrophy as observed in the vehicle-treated group. The typical increase in myonuclei with hypertrophy was absent in satellite cell-depleted fibers, resulting in expansion of the myonuclear domain. Consistent with lack of nuclear addition to enlarged fibers, long-term BrdU labeling showed a significant reduction in the number of BrdU-positive myonuclei in satellite cell-depleted muscle compared with vehicle-treated muscle. Single fiber functional analyses showed no difference in specific force, Ca(2+) sensitivity, rate of cross-bridge cycling and cooperativity between hypertrophied fibers from vehicle and tamoxifen-treated groups. Although a small component of the hypertrophic response, both fiber hyperplasia and regeneration were significantly blunted following satellite cell depletion, indicating a distinct requirement for satellite cells during these processes. These results provide convincing evidence that skeletal muscle fibers are capable of mounting a robust hypertrophic response to mechanical overload that is not dependent on satellite cells.
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Skeletal muscle tissue provides mechanical force for locomotion of all vertebrate animals. It is prone to damage from acute physical trauma and physiological stress. To cope with this, it possesses a tremendous capacity for rapid and effective repair that is widely held to be accomplished by the satellite cells lying between the muscle fiber plasmalemma and the basement membrane. Cell transplantation and lineage-tracing studies have demonstrated that Pax7-expressing (Pax7(+)) satellite cells can repair damaged muscle tissue repeatedly after several bouts of acute injury. These findings provided evidence that Pax7(+) cells are muscle stem cells. However, stem cells from a variety of other origins are also reported to contribute to myofibers upon engraftment into muscles, questioning whether satellite cells are the only stem cell source for muscle regeneration. Here, we have engineered genetic ablation of Pax7(+) cells to test whether there is any significant contribution to muscle regeneration after acute injury from cells other than this source. We find that such elimination of Pax7(+) cells completely blocks regenerative myogenesis either following injury to the tibialis anterior (TA) muscle or after transplantation of extensor digitorum longus (EDL) muscles into nude mice. As Pax7 is specifically expressed in satellite cells, we conclude that they are essential for acute injury-induced muscle regeneration. It remains to be established whether there is any significant role for stem cells of other origins. The implications of our results for muscle stem cell-based therapy are discussed.
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Muscle regeneration requires the coordinated interaction of multiple cell types. Satellite cells have been implicated as the primary stem cell responsible for regenerating muscle, yet the necessity of these cells for regeneration has not been tested. Connective tissue fibroblasts also are likely to play a role in regeneration, as connective tissue fibrosis is a hallmark of regenerating muscle. However, the lack of molecular markers for these fibroblasts has precluded an investigation of their role. Using Tcf4, a newly identified fibroblast marker, and Pax7, a satellite cell marker, we found that after injury satellite cells and fibroblasts rapidly proliferate in close proximity to one another. To test the role of satellite cells and fibroblasts in muscle regeneration in vivo, we created Pax7(CreERT2) and Tcf4(CreERT2) mice and crossed these to R26R(DTA) mice to genetically ablate satellite cells and fibroblasts. Ablation of satellite cells resulted in a complete loss of regenerated muscle, as well as misregulation of fibroblasts and a dramatic increase in connective tissue. Ablation of fibroblasts altered the dynamics of satellite cells, leading to premature satellite cell differentiation, depletion of the early pool of satellite cells, and smaller regenerated myofibers. Thus, we provide direct, genetic evidence that satellite cells are required for muscle regeneration and also identify resident fibroblasts as a novel and vital component of the niche regulating satellite cell expansion during regeneration. Furthermore, we demonstrate that reciprocal interactions between fibroblasts and satellite cells contribute significantly to efficient, effective muscle regeneration.
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Effects of previous strength training can be long-lived, even after prolonged subsequent inactivity, and retraining is facilitated by a previous training episode. Traditionally, such "muscle memory" has been attributed to neural factors in the absence of any identified local memory mechanism in the muscle tissue. We have used in vivo imaging techniques to study live myonuclei belonging to distinct muscle fibers and observe that new myonuclei are added before any major increase in size during overload. The old and newly acquired nuclei are retained during severe atrophy caused by subsequent denervation lasting for a considerable period of the animal's lifespan. The myonuclei seem to be protected from the high apoptotic activity found in inactive muscle tissue. A hypertrophy episode leading to a lasting elevated number of myonuclei retarded disuse atrophy, and the nuclei could serve as a cell biological substrate for such memory. Because the ability to create myonuclei is impaired in the elderly, individuals may benefit from strength training at an early age, and because anabolic steroids facilitate more myonuclei, nuclear permanency may also have implications for exclusion periods after a doping offense.
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To highlight the losses in muscle mass, strength, power, and functional capacity incurred in older adults during bed rest-mediated inactivity and to provide practical recommendations for both the prevention and rehabilitation of these losses. In addition to sarcopenic muscle loss, older adults lose lean tissue more rapidly than the young during prolonged periods of physical inactivity. Amino acid or protein supplementation has the potential to maintain muscle protein synthesis and may reduce inactivity-induced muscle loss, but should ideally be part of an integrated countermeasure regimen consisting of nutrition, exercise, and, when appropriate, pharmacologic interventions. In accordance with recent mechanistic advances, we recommend an applied, broad-based two-phase approach to limit inactivity-mediated losses of muscle mass and function in older adults: (i) Lifestyle: consume a moderate amount (25-30 g) of high-quality protein with each meal and incorporate habitual exercise in close temporal proximity to protein-containing meals; (ii) Crises: react aggressively to combat the accelerated loss of muscle mass and function during acute catabolic crises and periods of reduced physical activity. As a base strategy, this should include nutritional support such as targeted protein or amino acid supplementation and integrated physical therapy.
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to the editor: Arentson-Lantz et al. ([1][1]) report that 14 days of bed rest (BR) induces a profound decline in muscle strength (−13%) and fiber cross-sectional area (CSA) (−24%) in middle-aged adults. More importantly, they report that short-term disuse-induced muscle fiber atrophy is
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Studies in humans and animal models provide compelling evidence for age-related skeletal muscle denervation, which may contribute to muscle fiber atrophy and loss. Skeletal muscle denervation seems relentless; however, long-term, high-intensity physical activity appears to promote muscle reinnervation. Whether 5-month resistance training (RT) enhances skeletal muscle innervation in obese older adults is unknown. This study found that neural cell-adhesion molecule, NCAM+ muscle area decreased with RT and was inversely correlated with muscle strength. NCAM1 and RUNX1 gene transcripts significantly decreased with the intervention. Type I and type II fiber grouping in the vastus lateralis did not change significantly but increases in leg press and knee extensor strength inversely correlated with type I, but not with type II, fiber grouping. RT did not modify the total number of satellite cells, their number per area, or the number associated with specific fiber subtypes or innervated/denervated fibers. Our results suggest that RT has a beneficial impact on skeletal innervation, even when started late in life by sedentary obese older adults.
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Skeletal muscle mass, function, and repair capacity all progressively decline with aging, restricting mobility, voluntary function, and quality of life. Skeletal muscle repair is facilitated by a population of dedicated muscle stem cells (MuSCs), also known as satellite cells, that reside in anatomically defined niches within muscle tissues. In adult tissues, MuSCs are retained in a quiescent state until they are primed to regenerate damaged muscle through cycles of self-renewal divisions. With aging, muscle tissue homeostasis is progressively disrupted and the ability of MuSCs to repair injured muscle markedly declines. Until recently, this decline has been largely attributed to extrinsic age-related alterations in the microenvironment to which MuSCs are exposed. However, as highlighted in this Perspective, recent reports show that MuSCs also progressively undergo cell-intrinsic alterations that profoundly affect stem cell regenerative function with aging. A more comprehensive understanding of the interplay of stem cell–intrinsic and extrinsic factors will set the stage for improving cell therapies capable of restoring tissue homeostasis and enhancing muscle repair in the aged.
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We evaluated their circadian rhythms using data from electrocardiographic records and examined the change in circadian period related to normal RR intervals for astronauts who completed a long-term (≥6-month) mission in space. The examinees were seven astronauts, five men and two women, from 2009 to 2010. Their mean ± SD age was 52.0 ± 4.2 years (47-59 yr). Each stayed in space for more than 160 days; their average length of stay was 172.6 ± 14.6 days (163-199 days). We conducted a 24-h Holter electrocardiography before launch (Pre), at one month after launch (DF1), at two months after launch (DF2), at two weeks before return (DF3), and at three months after landing (Post), comparing each index of frequency-domain analysis and 24-h biological rhythms of the NN intervals (normal RR intervals). Results show that the mean period of Normal Sinus (NN) intervals was within 24 ± 4 h at each examination. Inter-individual variability differed among the stages, being significantly smaller at DF3 (Pre versus DF1 versus DF3 versus Post = 22.36 ± 2.50 versus 25.46 ± 4.37 versus 22.46 ± 1.75 versus 26.16 ± 7.18 h, p < 0.0001). The HF component increased in 2 of 7 astronauts, whereas it decreased in 3 of 7 astronauts and 1 was remained almost unchanged at DF1. During DF3, about 6 months after their stay in space, the HF component of 5 of 7 astronauts recovered from the decrease after launch, with prominent improvement to over 20% in 3 astronauts. Although autonomic nervous functions and circadian rhythms were disturbed until one month had passed in space, well-scheduled sleep and wake rhythms and meal times served as synchronizers.
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Determination of muscle fiber composition in human skeletal muscle biopsies is often performed using immunohistochemistry, a method that tends to be both time consuming, technically challenging, and complicated by limited availability of tissue. Here, we introduce quantitative reverse transcriptase polymerase chain reaction (qRT-PCR)-based Gene-family profiling (GeneFam) of myosin heavy chain (MyHC) mRNA expression as a high-throughput, sensitive, and reliable alternative. We show that GeneFam and immunohistochemistry result in similar disclosures of alterations in muscle fiber composition in biopsies from musculus vastus lateralis and musculus biceps brachii of previously untrained young women after 12 weeks of progressive strength training. The adaptations were evident as (a) consistent increases in MyHC2A abundance; (b) consistent decreases in MyHC2X abundance; and (c) consistently stable MyHC1 abundance, and were not found using traditional reference gene-based qRT-PCR analyses. Furthermore, muscle fiber composition found using each of the two approaches was correlated with each other (r = 0.50, 0.74, and 0.78 for MyHC1, A, and X, respectively), suggesting that GeneFam may be suitable for ranking of individual muscle phenotype, particularly for MyHC2 fibers. In summary, GeneFam of MyHC mRNA resulted in reliable assessment of alterations in muscle fiber composition in skeletal muscle of previously untrained women after 12 weeks of strength training.
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Abnormal body compositions such as high adiposity (HA), low muscle mass (LM), or a combination of the 2 [high adiposity with low muscle mass (HA-LM)] are relevant phenotypes, but data on their prevalence and impact on health are still limited. This is largely because of a lack of a consensus definition for these conditions. Of particular interest is the HA-LM phenotype, also termed "sarcopenic obesity," which may confer greater health risk. We propose a new approach for operationalizing abnormal body-composition phenotypes in a representative adult population. Whole-body dual-energy X-ray absorptiometry data obtained from the 1999-2004 NHANES were analyzed for 13,236 subjects aged ≥18 y (maximum weight and height of 136 kg and 1.96 m, respectively). Sex- and body mass index (BMI)-specific decile groups of appendicular skeletal muscle index (ASMI; kg/m(2)) and fat mass index (FMI; kg/m(2)) were developed. Cutoffs for HA and LM were incorporated into a diagnostic framework to characterize 4 specific body-composition phenotypes-low adiposity with high muscle mass, high adiposity with high muscle mass, low adiposity with low muscle mass, and HA-LM-and a subclassification of the phenotypes into class I, II, and III. Abnormal phenotypes were prevalent across the age spectrum and BMI categories. The association between ASMI or FMI and age was modified by sex and BMI. The prevalence of HA-LM in the whole sample was 10.3% in women and 15.2% in men. The prevalence of all subclasses of HA-LM in obese women and men was 14.7% and 22.9%, respectively. HA-LM class III was more prevalent in obese men (2.3%) than in obese women (0.3%). We developed sex- and BMI-specific reference curves to harmonize the classification of body-composition phenotypes. The application of this classification will be particularly useful in the identification of cases of sarcopenic obesity. The association of these phenotypes with metabolic deregulation and increased disease risk awaits verification.
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In the current study, we sought to determine the effect of a traditional, 12-week aerobic training protocol on skeletal muscle fiber type distribution and satellite cell content in sedentary subjects. Muscle biopsies were obtained from the vastus lateralis (n=23 [6M; 17F]; BMI: 30.7±1.2 kg/m2) before and after 12 weeks of aerobic training performed on a cycle ergometer. Immunohistochemical analyses were used to quantify myosin heavy chain (MyHC) isoform expression, cross-sectional area (CSA) and satellite cell and myonuclear content. Following training, a decrease in MyHC hybrid type IIa/IIx fiber frequency occurred, with a concomitant increase in pure MyHC type IIa fibers. Pre-training fiber type correlated with BMI, and the change in fiber type following training was associated with improvements in maximal oxygen consumption. Twelve weeks of aerobic training also induced increases in mean CSA in both MyHC type I and IIa fibers. Satellite cell content was also increased following training, specifically in MyHC type I fibers, with no change in the number of satellite cells associated with MyHC type II fibers. With the increased satellite cell content following training, an increase in myonuclear number per fiber also occurred in MyHC type I fibers. Hypertrophy of MyHC type II fibers occurred without detectable myonuclear addition, suggesting that mechanisms underlying growth in fast and slow fibers differ. These data provide intriguing evidence for a fiber type-specific role of satellite cells in muscle adaptation following aerobic training.
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Sarcopenia has been indicated as a reliable marker of frailty and poor prognosis among the oldest individuals. There are only few data on sarcopenia in healthy general population. We evaluated the prevalence of sarcopenia and its association with functional and clinical status in a population of healthy ambulatory subjects over 45 years living at home, in Paris (France). This study was conducted selecting all ambulatory participants (n = 1,445) aged 45 years and older from October 2008 to September 2011, consulting in the Institute of Physiology (Institut de Jaeger) from Paris (France) for a functional and muscular evaluation, and did not have limitations to moderate physical exercise. All were healthy people. All subjects performed a medical examination, associated with evaluation of muscle mass (body composition assessment using dual-energy X-ray absorptiometry) and of muscle function (by hand grip strength). Diagnosis of sarcopenia required the documentation of low muscle mass with low muscle strength according to the current international consensus definition of sarcopenia. From 1,421 participants (553 males and 868 females) definitively enrolled, 221 subjects (135 females and 86 males) (15.5 %) were identified as sarcopenic. Results from multivariate logistic regression models showed that sarcopenia was inversely associated with BMI with those participants with BMI higher than 22 kg/m(2) showing a lower risk of sarcopenia relative to those with BMI less than 21 kg/m(2) (OR 0.72; 95 % CI 0.60-0.91). Similarly, probability of sarcopenia was lower among subjects involved in leisure physical activities for 3 h or more per week (OR 0.45; 95 % CI 0.24-0.93). According to the category of age [45-54; 55-64; 65-74; 75-84 and 85 years or more], the prevalence of sarcopenia in women increase from 9.1; 12.7; 14.5; 19.4; to 33.3 %, respectively. For the men, the percentage of sarcopenia increase with aging from 8.6; 15.6; 13.6; 63.8 to 45.5 %, respectively. The present study suggests that among healthy ambulatory subjects over 45 years living at home, sarcopenia is frequent, even to the youngest subjects of the studied population, taking place from 9 % from 45 years, until 64.3 % for the subjects over 85 years. Our findings support the hypothesis that muscle mass and function are associated with BMI and physical activity, whatever the age of the subject.
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Introduction: Both sarcopenia and spinal cord injury (SCI) are characterized by the loss of skeletal muscle mass and function. Despite obvious similarities in atrophy between both models, differences in muscle fiber size and satellite cell content may exist on a muscle fiber type-specific level. Methods: In the present study, we compared skeletal muscle fiber characteristics between wheelchair-dependent young males with SCI (n = 8, 32 ± 4 yr), healthy elderly males (n = 8, 75 ± 2 yr), and young controls (n = 8, 31 ± 3 yr). Muscle biopsies were collected to determine skeletal muscle fiber type composition, fiber size, and satellite cell content. Results: Severe atrophy and a shift toward approximately 90% Type II muscle fibers were observed in muscle obtained from males with SCI. Muscle fiber size was substantially smaller in both the SCI (Types I and II fibers) and elderly subjects (Type II fibers) when compared with the controls. Satellite cell content was substantially lower in the wheelchair-dependent SCI subjects in both the Types I and II muscle fibers (0.049 ± 0.019 and 0.050 ± 0.005 satellite cells per fiber, respectively) when compared with the young controls (0.104 ± 0.011 and 0.117 ± 0.009 satellite cells per fiber, respectively). In the elderly, the number of satellite cells was lower in the Type II muscle fibers only (0.042 ± 0.005 vs 0.117 ± 0.009 satellite cells per fiber in the elderly vs young controls, respectively). Conclusion: This is the first study to show that muscle fiber atrophy as observed with SCI (Types I and II fibers) and aging (Type II fibers) is accompanied by a muscle fiber type-specific reduction in satellite cell content in humans.
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Ulex europaeus I agglutinin, a lectin specific for some alpha-L-fucose-containing glycocompounds, was used in fluorescence microscopy to stain cryostat sections of human tissues. The endothelium of vessels of all sizes was stained ubiquitously in all tissues studied as judged by double staining with a known endothelial marker, antibodies against human clotting factor VIII. Cultured human umbilical vein endothelial cells, but not fibroblasts, also bound Ulex lectin. The staining was not affected by the blood group type of the tissue donor. In some tissues Ulex lectin presented additional binding to epithelial structures. Also, this was independent on the blood group or the ability of the tissue donor to secrete soluble blood group substances. Lotus tetragonolobus agglutinin, another lectin specific for some alpha-L-fucose-containing moieties failed to react with endothelial cells. Our results suggest that Ulex europaeus I agglutinin is a good histologic marker for endothelium in human tissues.
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To investigate the relationship between skeletal muscle fiber type-specific characteristics, circulating hormone concentrations, and skeletal muscle mass and strength in older men. Cross-sectional analyses. University research center. Forty-one community dwelling elderly men (≥ 65). Leg strength (1-repetition maximum, 1RM) and whole-body and limb muscle mass were determined, and muscle fiber type composition, cross-sectional area (CSA), myonuclear content, and satellite cell (SC) content were assessed in skeletal muscle biopsy samples. In addition, blood samples were collected to determine serum testosterone, sex hormone-binding globulin, insulinlike growth factor (IGF)-1, and IGF binding protein-3 concentrations. Muscle mass correlated with muscle strength (0.41 ≤ correlation coefficient (r) ≤ 0.72; P < .01). Muscle fiber CSA, myonuclear content, and SC content were significantly lower in type II than in type I muscle fibers. Myonuclear and SC content were positively correlated with muscle fiber CSA. Furthermore, greater muscle fiber CSA (type I and II) was associated with greater thigh muscle area and muscle strength (0.30 ≤ r ≤ 0.45; P < .05). Testosterone concentration was positively correlated with muscle mass and muscle fiber CSA. Regression analysis showed that SC content, myonuclear content, and testosterone concentration are predictive of muscle fiber CSA. Furthermore, muscle mass and type II muscle fiber CSA are predictive of muscle strength. Skeletal muscle mass and strength in elderly men are positively correlated with muscle fiber type-specific CSA, myonuclear content, and SC content. These findings support the assumption that a decline in SC content plays an important role in age-related decline in muscle mass and strength.
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Spaceflight and bed rest (BR) lead to muscle atrophy. This study assessed the effect of essential amino acid (EAA) supplementation and resistance training with decreased energy intake on molecular changes in skeletal muscle after 28-day BR and 14-day recovery. Thirty-one men (31-55 years) subjected to an 8 ± 6% energy deficit were randomized to receive EAA without resistance training (AA, n = 7), or EAA 3 h after (RT, n = 12) or 5 min before (AART, n = 12) resistance training. During BR, myostatin transcript levels increased twofold in the AA group. During recovery, insulin-like growth factor-1 (IGF-1) mRNA increased in all groups, whereas Pax7, MyoD, myogenin, and MRF4 transcripts increased in AA only (all P < 0.05). MAFbx transcripts decreased twofold with AA and RT. Satellite cells did not change during BR or recovery. This suggests that EAA alone is the least protective countermeasure to muscle loss, and several molecular mechanisms are proposed by which exercise attenuates muscle atrophy during BR with energy deficit.
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Myosatellite cells were examined and quantified at the fine structural level of resolution during aging of skeletal muscles in mice and rats. Satellite cells in the soleus and gastrocnemius muscles of animals between eight and 30 months of age appeared, according to morphological criteria, metabolically less active than those examined in immature muscles. In the soleus muscle of the mouse, satellite cells decreased in number from 4.6% at eight months of age to 2.4% at 30 months. This decrease appeared to be due to the passage of some satellite cells into the interstitial space as a result of the formation of external lamina material around the entire satellite cell surface.
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This work reports on the muscle loss and recovery after 17 wk of continuous bed rest and 8 wk of reambulation in eight normal male volunteers. Muscle changes were assessed by urinary levels of 3-methylhistidine (3-MeH), nitrogen balance, dual-photon absorptiometry (DPA), magnetic resonance imaging (MRI), and isokinetic muscle performance. The total body lean tissue loss during bed rest calculated from nitrogen balance was 3.9 +/- 2.1 (SD) kg (P < 0.05). Although the total loss is minimal, DPA scans showed that nearly all of the lean tissue loss occurred in the lower limbs. Similarly, MRI muscle volume measurements showed greater percent loss in the limbs relative to the back muscles. MRI, DPA, and nitrogen balance suggest that muscle atrophy continued throughout bed rest with rapid recovery after reambulation. Isokinetic muscle strength decreased significantly (P < 0.05) in the thigh and calf with no loss in the arms and with rapid recovery during reambulation. We conclude that there is great variability in the degree and location of muscle loss in bed rest and that the lower limb muscles are primarily affected.
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1. Quadriceps muscle protein turnover was assessed in the post-absorptive state in six men immediately after the end of unilateral leg immobilization (37 ± 4 days) in a plaster cast after tibial fracture. A primed-constant intravenous infusion of l-[1-13C]leucine was administered over 7 h. Quadriceps needle biopsies, taken bilaterally at the end of the infusion, were analysed for muscle protein leucine enrichment with 13C. 2. Quadriceps muscle protein synthetic rate, calculated from the fractional incorporation of [13C]leucine into protein compared with the average enrichment of blood α-ketoisocaproate, was 0.046 ±0.012%/h in the uninjured leg, but was only 0.034 ±0.007%/h in the quadriceps of the previously fractured leg (P > 0.05, means ± sd). 3. Muscle RNA activity (i.e. protein synthetic rate per RNA) fell from 0.27 ±0.08 μg of protein synthesized h−1 μg−1 of RNA in the control leg to 0.14 ±0.03 μg of protein synthesized h−1 μg−1 of RNA in the immobilized leg (P > 0.02). 4. Immobilization was associated with a significant atrophy of type I muscle fibres (mean diameter 69.5 ±21 μm immobilized, 81.1 ±18 μm control, P > 0.05), but no significant change occurred in type II fibre diameter. Mean quadriceps fibre volume calculated from the values for fibre diameter and percentage of each fibre type, was smaller in the injured leg by 10.6%; this value was near to the calculated difference in muscle thigh volume (calculated from thigh circumference and skin-fold thickness) which was less by 8.3%. 5. From estimated mean daily values for quadriceps protein synthetic rate (1.65 ±0.44%/day in the control legs and 1.22±0.28%/day in the injured legs) and change in fibre volume, mean daily muscle protein breakdown rates were calculated as 1.65%/ day and 1.53%/day respectively, suggesting that muscle protein breakdown was not enhanced and may have fallen. 6. The results suggest a decrease in muscle protein turnover during limb immobilization in man, with the decrement in muscle mass being due mainly to a substantial (25%) depression of muscle protein synthesis.
Article
The effects of long-term hindlimb unweighting by tail suspension on postnatal growth of 20-day rat extensor digitorum longus (EDL) and soleus muscles were studied. Morphological assay indicated that radial growth of soleus myofibers was completely inhibited between 3 and 10 days of suspension and reduced thereafter, leading to a severe attenuation (-76% from control) over the total experimental period. Longitudinal growth rate, however, was accelerated 40% over weight-bearing controls. In addition, myofibers were arranged parallel to the long axis of the muscle, an orientation associated with chronologically younger muscles, suggesting morphological maturation of the soleus muscle had been delayed by suspension. In contrast, radial and longitudinal growth of EDL myofibers were minimally affected under similar conditions and remained within approximately 5% of control at all times. Suspension also influenced the normal changes that occur in satellite cell and myonuclear populations during postnatal growth. Both the number and proliferative activity of satellite cells were severely reduced in individual myofibers after only 3 days in both soleus and EDL muscles. The reduced number of satellite cells within 3 days of initiating hindlimb suspension appeared to be the result of their incorporation into myofibers while the long-lasting reduction appeared to be the added effects of decreased proliferative activity. In the soleus, this reduction in number and proliferation of satellite cells persisted throughout the experimental period and resulted in an overall 43% fewer myonuclei and 45% fewer satellite cells than control at 50 days of age. In contrast, both the total number and mitotic activity of satellite cells in the EDL rapidly returned to weight-bearing control levels by day 10 of suspension, resulting in no overall reduction in myonuclear accretion.
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Percutaneous needle biopsy samples were obtained from the vastus lateralis and soleus muscles before and after 30 d of 6 degree head-down bedrest to determine the influence of this model of simulated microgravity on human skeletal muscle. Fiber atrophy was evident in both muscles with both fast-twitch and slow-twitch fiber cross-sectional areas decreasing. Predominant atrophy of slow-twitch fibers was not evident. The soleus had a greater proportion of slow-twitch fibers than the vastus lateralis before bedrest. Neither muscle showed a change in fiber type percentage with bedrest. Phosphofructokinase and lactate dehydrogenase activities in the soleus and vastus lateralis muscles were similar before and after bedrest. The activities of beta-hydroxyacyl-CoA dehydrogenase and citrate synthase, however, were reduced during bedrest with these responses being somewhat greater in the soleus. While the ultrastructure of most of the fibers of the soleus and vastus lateralis appeared normal after bedrest, evidence of remodeling was present in both muscles. The proliferation of core/targetoid lesions, honeycomb networks, regenerating satellite cells, necrotic foci and myofibrillar disorganization after bedrest indicates that force development is an important factor in determining the organization of the fine structure of muscle. The results indicate that short-duration exposure to simulated microgravity decreases fiber size and the capacity for aerobic energy supply of human skeletal muscle. Moreover, disorganization of the contractile machinery occurs. Thus, it appears that bedrest alters the "normal" load-time constraints imposed on skeletal muscle sufficiently to change its inherent structural and metabolic characteristics.
Article
The source of the new nuclei appearing during the growth of muscle fibers was examined in the tibialis anterior muscle of young Sherman rats (14–17 days of age) using radioautography at various intervals after a single injection of a small, non-toxic dose of 3H-thymidine (2 μCi/g body weight). Two techniques were employed: (1) labeled nuclei were detected in 1 μ thick radioautographs examined in the light microscope, and identified by simultaneous electron microscope examination of an adjacent section. The nuclei were then classified either as “true” muscle nuclei (within the plasmalemma of the fibers) or as belonging to “satellite cells” (which are mononucleated cells with scanty cytoplasm wedged between plasmalemma and basement membrane). (2) Muscle fibers freed by collagenase digestion were radioautographed one hour after 3H-thymidine injection in order to determine the total number of labeled nuclei (true muscle nuclei plus those of satellite cells) per unit length of fiber.Certain nuclei within the basement membrane of muscle fibers are labeled one hour after 3H-thymidine and, therefore, synthesize DNA. The electron microscope demonstrates that these nuclei invariably belong to satellite cells, never to true muscle nuclei. Furthermore, the total number of labeled nuclei per unit length of fiber doubles between 1 and 24 hours; and, therefore, the labeled satellite cell nuclei undergo mitosis.Following mitosis, half of the daughters of satellite cells are incorporated into the fibers to become true muscle nuclei. The remaining half divides again later; and half of their daughter cells are incorporated. Thus, satellite cells in young rats divide repeatedly and function as a source of true muscle nuclei.
Article
The mechanisms responsible for the decrease in exercise capacity after bed rest were assessed in 12 apparently healthy men aged 50 +/- 4 years who underwent equilibrium gated blood pool scintigraphy during supine and upright multistage bicycle ergometry before and after 10 days of bed rest. After bed rest, echocardiographically measured supine resting left ventricular end-diastolic volume decreased by 16% (p less than 0.05). Peak oxygen uptake during supine effort after bed rest was diminished by 6% (p = not significant [NS]), whereas peak oxygen uptake during upright effort declined by 15% (p less than 0.05). After bed rest, increases in heart rate were also greater during exercise in the upright than in the supine position (p less than 0.05). Values of left ventricular ejection fraction increased normally during both supine and upright effort after bed rest and were higher than corresponding values before bed rest (p less than 0.05). After bed rest, increased left ventricular ejection fraction and heart rate largely compensated for the reduced cardiac volume during supine effort, but these mechanisms were insufficient to maintain oxygen transport capacity at levels during upright effort before bed rest. These results indicate that orthostatically induced cardiac underfilling, not physical deconditioning or left ventricular dysfunction, is the major cause of reduced effort tolerance after 10 days of bed rest in normal middle-aged men.
Article
Muscle fiber area, capillary density, and capillary to fiber ratio were determined in needle biopsy samples ofM. vastus lateralis processed for electron microscopy from 9 untrained men (average$$\dot V_{{\text{O}}_{\text{2}} }$$max = 61.3 ml/min kg), 3 untrained women ($$\dot V_{{\text{O}}_{\text{2}} }$$max = 43.7 ml/min kg) and 5 well trained orienteers ($$\dot V_{{\text{O}}_{\text{2}} }$$max = 76.1 ml/min kg). The volume density of mitochondria was previously reported to be significantly higher in orienteers (7.32%) than in untrained men (5.19%) and in untrained women (4.08%) in the same material (Hoppeler et al. 1973). The mean fiber area was found to be somewhat larger in orienteers (6,410 μm2 than in untrained men (5,350 μm2, not significant), but significantly smaller values were found in untrained women (3,390 μm2). The capillary density was almost identical in the three experimental groups, it ranged from 416 to 431 mm−2. The absolute values reported for both of the above variables are subjected to a bias due to fiber shrinkage, fiber shortening and fiber kinking occurring during the needle biopsy procedure and the subsequent steps of tissue preparation. The capillary to fiber ratio is much less affected by tissue handling and significantly higher values were found in orienteers (2.70) than in untrained men (2.07) and in untrained women (1.37). There was a significant linear relationship between the capillary to fiber ratio and both$$\dot V_{{\text{O}}_{\text{2}} }$$max and the volume density of mitochondria for all subjects analysed, suggesting an adjustment of the capillary supply to the maximal aerobic capacity as well as to the mitochondrial content inM. vastus lateralis.
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