Tissue regeneration declines with ageing but little is known about whether this arises from changes in stem-cell heterogeneity. Here, in homeostatic skeletal muscle, we identify two quiescent ...stem-cell states distinguished by relative CD34 expression: CD34
, with stemness properties (genuine state), and CD34
, committed to myogenic differentiation (primed state). The genuine-quiescent state is unexpectedly preserved into later life, succumbing only in extreme old age due to the acquisition of primed-state traits. Niche-derived IGF1-dependent Akt activation debilitates the genuine stem-cell state by imposing primed-state features via FoxO inhibition. Interventions to neutralize Akt and promote FoxO activity drive a primed-to-genuine state conversion, whereas FoxO inactivation deteriorates the genuine state at a young age, causing regenerative failure of muscle, as occurs in geriatric mice. These findings reveal transcriptional determinants of stem-cell heterogeneity that resist ageing more than previously anticipated and are only lost in extreme old age, with implications for the repair of geriatric muscle.
Assessment of a low skeletal muscle mass (SM) is important for diagnosis of ageing and disease-associated sarcopenia and is hindered by heterogeneous methods and terminologies that lead to ...differences in diagnostic criteria among studies and even among consensus definitions. The aim of this review was to analyze and summarize previously published cut-offs for SM applied in clinical and research settings and to facilitate comparison of results between studies. Multiple published reference values for discrepant parameters of SM were identified from 64 studies and the underlying methodological assumptions and limitations are compared including different concepts for normalization of SM for body size and fat mass (FM). Single computed tomography or magnetic resonance imaging images and appendicular lean soft tissue by dual X-ray absorptiometry (DXA) or bioelectrical impedance analysis (BIA) are taken as a valid substitute of total SM because they show a high correlation with results from whole body imaging in cross-sectional and longitudinal analyses. However, the random error of these methods limits the applicability of these substitutes in the assessment of individual cases and together with the systematic error limits the accurate detection of changes in SM. Adverse effects of obesity on muscle quality and function may lead to an underestimation of sarcopenia in obesity and may justify normalization of SM for FM. In conclusion, results for SM can only be compared with reference values using the same method, BIA- or DXA-device and an appropriate reference population. Limitations of proxies for total SM as well as normalization of SM for FM are important content-related issues that need to be considered in longitudinal studies, populations with obesity or older subjects.
Extensive analyses of mice carrying null mutations in paired box 7 (Pax7) have confirmed the progressive loss of the satellite cell lineage in skeletal muscle, resulting in severe muscle atrophy and ...death. A recent study using floxed alleles and tamoxifen-induced inactivation concluded that after 3 wk of age, Pax7 was entirely dispensable for satellite cell function. Here, we demonstrate that Pax7 is an absolute requirement for satellite cell function in adult skeletal muscle. Following Pax7 deletion, satellite cells and myoblasts exhibit cell-cycle arrest and dysregulation of myogenic regulatory factors. Maintenance of Pax7 deletion through continuous tamoxifen administration prevented regrowth of Pax7-expressing satellite cells and a profound muscle regeneration deficit that resembles the phenotype of skeletal muscle following genetically engineered ablation of satellite cells. Therefore, we conclude that Pax7 is essential for regulating the expansion and differentiation of satellite cells during both neonatal and adult myogenesis.
Skeletal muscle satellite cells are adult stem cells responsible for postnatal skeletal muscle growth and regeneration. Paired-box transcription factor Pax7 plays a central role in satellite cell ...survival, self-renewal, and proliferation. However, how Pax7 is regulated during the transition from proliferating satellite cells to differentiating myogenic progenitor cells is largely unknown. In this study, we find that miR-1 and miR-206 are sharply up-regulated during satellite cell differentiation and down-regulated after muscle injury. We show that miR-1 and miR-206 facilitate satellite cell differentiation by restricting their proliferative potential. We identify Pax7 as one of the direct regulatory targets of miR-1 and miR-206. Inhibition of miR-1 and miR-206 substantially enhances satellite cell proliferation and increases Pax7 protein level in vivo. Conversely, sustained Pax7 expression as a result of the loss of miR-1 and miR-206 repression elements at its 3' untranslated region significantly inhibits myoblast differentiation. Therefore, our experiments suggest that microRNAs participate in a regulatory circuit that allows rapid gene program transitions from proliferation to differentiation.
Distinct cell populations with regenerative capacity have been reported to contribute to myofibres after skeletal muscle injury, including non-satellite cells as well as myogenic satellite cells. ...However, the relative contribution of these distinct cell types to skeletal muscle repair and homeostasis and the identity of adult muscle stem cells remain unknown. We generated a model for the conditional depletion of satellite cells by expressing a human diphtheria toxin receptor under control of the murine Pax7 locus. Intramuscular injection of diphtheria toxin during muscle homeostasis, or combined with muscle injury caused by myotoxins or exercise, led to a marked loss of muscle tissue and failure to regenerate skeletal muscle. Moreover, the muscle tissue became infiltrated by inflammatory cells and adipocytes. This localised loss of satellite cells was not compensated for endogenously by other cell types, but muscle regeneration was rescued after transplantation of adult Pax7(+) satellite cells alone. These findings indicate that other cell types with regenerative potential depend on the presence of the satellite cell population, and these observations have important implications for myopathic conditions and stem cell-based therapeutic approaches.
Skeletal muscle contraction increases intracellular ATP turnover, calcium flux, and mechanical stress, initiating signal transduction pathways that modulate peroxisome proliferator‐activated receptor ...γ coactivator‐1α (PGC‐1α)‐dependent transcriptional programmes. The purpose of this study was to determine if the intensity of exercise regulates PGC‐1α expression in human skeletal muscle, coincident with activation of signalling cascades known to regulate PGC‐1α transcription. Eight sedentary males expended 400 kcal (1674 kj) during a single bout of cycle ergometer exercise on two separate occasions at either 40% (LO) or 80% (HI) of . Skeletal muscle biopsies from the m. vastus lateralis were taken at rest and at +0, +3 and +19 h after exercise. Energy expenditure during exercise was similar between trials, but the high intensity bout was shorter in duration (LO, 69.9 ± 4.0 min; HI, 36.0 ± 2.2 min, P < 0.05) and had a higher rate of glycogen utilization (P < 0.05). PGC‐1α mRNA abundance increased in an intensity‐dependent manner +3 h after exercise (LO, 3.8‐fold; HI, 10.2‐fold, P < 0.05). AMP‐activated protein kinase (AMPK) (2.8‐fold, P < 0.05) and calcium/calmodulin‐dependent protein kinase II (CaMKII) phosphorylation (84%, P < 0.05) increased immediately after HI but not LO. p38 mitogen‐activated protein kinase (MAPK) phosphorylation increased after both trials (∼2.0‐fold, P < 0.05), but phosphorylation of the downstream transcription factor, activating transcription factor‐2 (ATF‐2), increased only after HI (2.4‐fold, P < 0.05). Cyclic‐AMP response element binding protein (CREB) phosphorylation was elevated at +3 h after both trials (∼80%, P < 0.05) and class IIa histone deacetylase (HDAC) phosphorylation increased only after HI (2.0‐fold, P < 0.05). In conclusion, exercise intensity regulates PGC‐1α mRNA abundance in human skeletal muscle in response to a single bout of exercise. This effect is mediated by differential activation of multiple signalling pathways, with ATF‐2 and HDAC phosphorylation proposed as key intensity‐dependent mediators.
In skeletal muscle, the nature of adaptation to repeated sessions of exercise (exercise training) is determined by the intensity, duration and frequency of the individual exercise sessions. We compared the molecular response in human skeletal muscle to a single session of either high or low intensity cycling exercise where the total energy expended (number of calories) was similar. We show that high intensity exercise, but of shorter duration, results in greater activation of key regulatory pathways controlling skeletal muscle gene expression compared to low intensity, longer duration exercise. High intensity exercise training may, consequently, be more time‐efficient in promoting training adaptations. This study increases our understanding of the molecular basis for the intensity‐dependent adaptation to exercise.
Muscle satellite cells contribute to muscle regeneration. We have used a Pax3superscript GFP/+ mouse line to directly isolate (Pax3)(green fluorescent protein)-expressing muscle satellite cells, by ...flow cytometry from adult skeletal muscles, as a homogeneous population of small, nongranular, Pax7+, CD34+, CD45-, Sca1- cells. The flow cytometry parameters thus established enabled us to isolate satellite cells from wild-type muscles. Such cells, grafted into muscles of mdx nu/nu mice, contributed both to fiber repair and to the muscle satellite cell compartment. Expansion of these cells in culture before engraftment reduced their regenerative capacity.
Adult tissue repair and regeneration require stem-progenitor cells that can self-renew and generate differentiated progeny. Skeletal muscle regenerative capacity relies on muscle satellite cells ...(MuSCs) and their interplay with different cell types within the niche. However, our understanding of skeletal muscle tissue cellular composition is limited. Here, using a combined approach of single-cell RNA sequencing and mass cytometry, we precisely mapped 10 different mononuclear cell types in adult mouse muscle. We also characterized gene signatures and determined key discriminating markers of each cell type. We identified two previously understudied cell populations in the interstitial compartment. One expresses the transcription factor scleraxis and generated tenocytes in vitro. The second expresses markers of smooth muscle and mesenchymal cells (SMMCs) and, while distinct from MuSCs, exhibited myogenic potential and promoted MuSC engraftment following transplantation. The blueprint presented here yields crucial insights into muscle-resident cell-type identities and can be exploited to study muscle diseases.
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•Combination of single-cell RNA sequencing and mass cytometry•Construction of a cell atlas of adult skeletal muscle•Skeletal muscle is composed of 10 mononucleated cell types and myofibers•Skeletal muscle contains interstitial tenocytes and smooth muscle-mesenchymal cells
A strong characterization of cell types present in adult skeletal muscle is needed. Giordani et al. use both single-cell transcriptomics and mass cytometry to build a single-cell survey of adult skeletal muscle tissue and reveal understudied cell populations.
Satellite cells, the quintessential skeletal muscle stem cells, reside in a specialized local environment whose anatomy changes dynamically during tissue regeneration. The plasticity of this niche is ...attributable to regulation by the stem cells themselves and to a multitude of functionally diverse cell types. In particular, immune cells, fibrogenic cells, vessel‐associated cells and committed and differentiated cells of the myogenic lineage have emerged as important constituents of the satellite cell niche. Here, we discuss the cellular dynamics during muscle regeneration and how disease can lead to perturbation of these mechanisms. To define the role of cellular components in the muscle stem cell niche is imperative for the development of cell‐based therapies, as well as to better understand the pathobiology of degenerative conditions of the skeletal musculature.
Multiple, functionally diverse cell types have been shown to contribute to skeletal muscle regeneration. This Review discusses the cellular dynamics and the roles of immune, fibrogenic, vessel‐associated and myogenic cells in the response of the satellite cell niche to muscle injury and disease.
Skeletal muscle stem cells are regulated by Pax3/7. During development, Pax3 is required for the maintenance of these cells in the somite and their migration to sites of myogenesis; high levels of ...Pax3 interfere with muscle cell differentiation, both in the embryo and in the adult. Quantitative fine-tuning of Pax3 is critical, and microRNAs provide a potential mechanism. We identify microRNA-27b (miR-27b), which directly targets the 3'-UTR of Pax3 mRNA, as such a regulator. miR-27b is expressed in the differentiating skeletal muscle of the embryonic myotome and in activated satellite cells of adult muscle. In vivo overexpression of a miR-27b transgene in Pax3-positive cells in the embryo leads to down-regulation of Pax3, resulting in interference with progenitor cell migration and in premature differentiation. In a complementary experiment, miR-27b inhibitors were transfected into cultures of adult muscle satellite cells that normally express miR-27b at the onset of differentiation, when Pax3 protein levels undergo rapid down-regulation. Interference with miR-27b function results in continuing Pax3 expression leading to more proliferation and a delay in the onset of differentiation. Pax7 levels are not affected. Introduction of miR-27b antagomirs at a site of muscle injury in vivo also affects Pax3 expression and regeneration in vivo. We therefore conclude that miR-27b regulates Pax3 protein levels and this down-regulation ensures rapid and robust entry into the myogenic differentiation program.
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