Postnatal growth of skeletal muscle largely depends on the expansion and differentiation of resident stem cells, the so-called satellite cells. Here, we demonstrate that postnatal satellite cells ...express components of the bone morphogenetic protein (BMP) signaling machinery. Overexpression of noggin in postnatal mice (to antagonize BMP ligands), satellite cell-specific knockout of
(the gene encoding the BMP transmembrane receptor) or overexpression of inhibitory SMAD6 decreased satellite cell proliferation and accretion during myofiber growth, and ultimately retarded muscle growth. Moreover, reduced BMP signaling diminished the adult satellite cell pool. Abrogation of BMP signaling in satellite cell-derived primary myoblasts strongly diminished cell proliferation and upregulated the expression of cell cycle inhibitors
and
In conclusion, these results show that BMP signaling defines postnatal muscle development by regulating satellite cell-dependent myofiber growth and the generation of the adult muscle stem cell pool.
Cell size is determined by the balance between protein synthesis and degradation. This equilibrium is affected by hormones, nutrients, energy levels, mechanical stress and cytokines. Mutations that ...inactivate myostatin lead to excessive muscle growth in animals and humans, but the signals and pathways responsible for this hypertrophy remain largely unknown. Here we show that bone morphogenetic protein (BMP) signaling, acting through Smad1, Smad5 and Smad8 (Smad1/5/8), is the fundamental hypertrophic signal in mice. Inhibition of BMP signaling causes muscle atrophy, abolishes the hypertrophic phenotype of myostatin-deficient mice and strongly exacerbates the effects of denervation and fasting. BMP-Smad1/5/8 signaling negatively regulates a gene (Fbxo30) that encodes a ubiquitin ligase required for muscle loss, which we named muscle ubiquitin ligase of the SCF complex in atrophy-1 (MUSA1). Collectively, these data identify a critical role for the BMP pathway in adult muscle maintenance, growth and atrophy.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Objective
This study was undertaken to determine whether a low residual quantity of dystrophin protein is associated with delayed clinical milestones in patients with DMD mutations.
Methods
We ...performed a retrospective multicentric cohort study by using molecular and clinical data from patients with DMD mutations registered in the Universal Mutation Database–DMD France database. Patients with intronic, splice site, or nonsense DMD mutations, with available muscle biopsy Western blot data, were included irrespective of whether they presented with severe Duchenne muscular dystrophy (DMD) or milder Becker muscular dystrophy (BMD). Patients were separated into 3 groups based on dystrophin protein levels. Clinical outcomes were ages at appearance of first symptoms; loss of ambulation; fall in vital capacity and left ventricular ejection fraction; interventions such as spinal fusion, tracheostomy, and noninvasive ventilation; and death.
Results
Of 3,880 patients with DMD mutations, 90 with mutations of interest were included. Forty‐two patients expressed no dystrophin (group A), and 31 of 42 (74%) developed DMD. Thirty‐four patients had dystrophin quantities < 5% (group B), and 21 of 34 (61%) developed BMD. Fourteen patients had dystrophin quantities ≥ 5% (group C), and all but 4 who lost ambulation beyond 24 years of age were ambulant. Dystrophin quantities of <5%, as low as <0.5%, were associated with milder phenotype for most of the evaluated clinical outcomes, including age at loss of ambulation (p < 0.001).
Interpretation
Very low residual dystrophin protein quantity can cause a shift in disease phenotype from DMD toward BMD. ANN NEUROL 2021;89:280–292
The mechanism of pattern formation during limb muscle development remains poorly understood. The canonical view holds that naïve limb muscle progenitor cells (MPCs) invade a pre-established pattern ...of muscle connective tissue, thereby forming individual muscles. Here, we show that early murine embryonic limb MPCs highly accumulate pSMAD1/5/9, demonstrating active signaling of bone morphogenetic proteins (BMP) in these cells. Overexpression of inhibitory human SMAD6 (huSMAD6) in limb MPCs abrogated BMP signaling, impaired their migration and proliferation, and accelerated myogenic lineage progression. Fewer primary myofibers developed, causing an aberrant proximodistal muscle pattern. Patterning was not disturbed when huSMAD6 was overexpressed in differentiated muscle, implying that the proximodistal muscle pattern depends on BMP-mediated expansion of MPCs before their differentiation. We show that limb MPCs differentially express Hox genes, and Hox-expressing MPCs displayed active BMP signaling. huSMAD6 overexpression caused loss of HOXA11 in early limb MPCs. In conclusion, our data show that BMP signaling controls expansion of embryonic limb MPCs as a prerequisite for establishing the proximodistal muscle pattern, a process that involves expression of Hox genes.
Dystrophin is essential for muscle health: its sarcolemmal absence causes the fatal, X-linked condition, Duchenne muscular dystrophy (DMD). However, its normal, spatial organization remains poorly ...understood, which hinders the interpretation of efficacy of its therapeutic restoration. Using female reporter mice heterozygous for fluorescently tagged dystrophin ( Dmd EGFP ), we here reveal that dystrophin distribution is unexpectedly compartmentalized, being restricted to myonuclear-defined sarcolemmal territories extending ~80 µm, which we called “basal sarcolemmal dystrophin units (BSDUs).” These territories were further specialized at myotendinous junctions, where both Dmd transcripts and dystrophin protein were enriched. Genome-level correction in X-linked muscular dystrophy mice via CRISPR/Cas9 gene editing restored a mosaic of separated dystrophin domains, whereas transcript-level Dmd correction, following treatment with tricyclo-DNA antisense oligonucleotides, restored dystrophin initially at junctions before extending along the entire fiber—with levels ~2% sufficient to moderate the dystrophic process. We conclude that widespread restoration of fiber dystrophin is likely critical for therapeutic success in DMD, perhaps most importantly, at muscle–tendon junctions.
Dystrophin is essential for muscle health: its sarcolemmal absence causes the fatal, X-linked condition, Duchenne muscular dystrophy (DMD). However, its normal, spatial organization remains poorly ...understood, which hinders the interpretation of efficacy of its therapeutic restoration. Using female reporter mice heterozygous for fluorescently tagged dystrophin (
), we here reveal that dystrophin distribution is unexpectedly compartmentalized, being restricted to myonuclear-defined sarcolemmal territories extending ~80 µm, which we called "basal sarcolemmal dystrophin units (BSDUs)." These territories were further specialized at myotendinous junctions, where both
transcripts and dystrophin protein were enriched. Genome-level correction in X-linked muscular dystrophy mice via CRISPR/Cas9 gene editing restored a mosaic of separated dystrophin domains, whereas transcript-level
correction, following treatment with tricyclo-DNA antisense oligonucleotides, restored dystrophin initially at junctions before extending along the entire fiber-with levels ~2% sufficient to moderate the dystrophic process. We conclude that widespread restoration of fiber dystrophin is likely critical for therapeutic success in DMD, perhaps most importantly, at muscle-tendon junctions.
ABSTRACT
Introduction: The effect of constitutive inactivation of the gene encoding myostatin on the gain in muscle performance during postnatal growth has not been well characterized. Methods: We ...analyzed 2 murine myostatin knockout (KO) models, (i) the Lee model (KOLee) and (ii) the Grobet model (KOGrobet), and measured the contraction of tibialis anterior muscle in situ. Results: Absolute maximal isometric force was increased in 6‐month‐old KOLee and KOGrobet mice, as compared to wild‐type mice. Similarly, absolute maximal power was increased in 6‐month‐old KOLee mice. In contrast, specific maximal force (relative maximal force per unit of muscle mass was decreased in all 6‐month‐old male and female KO mice, except in 6‐month‐old female KOGrobet mice, whereas specific maximal power was reduced only in male KOLee mice. Conclusions: Genetic inactivation of myostatin increases maximal force and power, but in return it reduces muscle quality, particularly in male mice. Muscle Nerve 55: 254–261, 2017
To better define the role of male and female gonad-related factors (MGRF, presumably testosterone, and FGRF, presumably estradiol, respectively) on mouse hindlimb skeletal muscle contractile ...performance/function gain during postnatal development, we analyzed the effect of castration initiated before puberty in male and female mice. We found that muscle absolute and specific (normalized to muscle weight) maximal forces were decreased in 6-mo-old male and female castrated mice compared with age- and sex-matched intact mice, without alteration in neuromuscular transmission. Moreover, castration decreased absolute and specific maximal powers, another important aspect of muscle performance, in 6-mo-old males, but not in females. Absolute maximal force was similarly reduced by castration in 3-mo-old muscle fiber androgen receptor (AR)-deficient and wild-type male mice, indicating that the effect of MGRF was muscle fiber AR independent. Castration reduced the muscle weight gain in 3-mo mice of both sexes and in 6-mo females but not in males. We also found that bone morphogenetic protein signaling through Smad1/5/9 was not altered by castration in atrophic muscle of 3-mo-old mice of both sexes. Moreover, castration decreased the sexual dimorphism regarding muscle performance. Together, these results demonstrated that in the long term, MGRF and FGRF promote muscle performance gain in mice during postnatal development, independently of muscle growth in males, largely via improving muscle contractile quality (force and power normalized), and that MGFR and FGRF also contribute to sexual dimorphism. However, the mechanisms underlying MGFR and FGRF actions remain to be determined.
Background
Dmdmdx, harbouring the c.2983C>T nonsense mutation in Dmd exon 23, is a mouse model for Duchenne muscular dystrophy (DMD), frequently used to test therapies aimed at dystrophin ...restoration. Current translational research is methodologically hampered by the lack of a reporter mouse model, which would allow direct visualization of dystrophin expression as well as longitudinal in vivo studies.
Methods
We generated a DmdEGFP‐mdx reporter allele carrying in cis the mdx‐23 mutation and a C‐terminal EGFP‐tag. This mouse model allows direct visualization of spontaneously and therapeutically restored dystrophin‐EGFP fusion protein either after natural fibre reversion, or for example, after splice modulation using tricyclo‐DNA to skip Dmd exon 23, or after gene editing using AAV‐encoded CRISPR/Cas9 for Dmd exon 23 excision.
Results
Intravital microscopy in anaesthetized mice allowed live‐imaging of sarcolemmal dystrophin‐EGFP fusion protein of revertant fibres as well as following therapeutic restoration. Dystrophin‐EGFP‐fluorescence persisted ex vivo, allowing live‐imaging of revertant and therapeutically restored dystrophin in isolated fibres ex vivo. Expression of the shorter dystrophin‐EGFP isoforms Dp71 in the brain, Dp260 in the retina, and Dp116 in the peripheral nerve remained unabated by the mdx‐23 mutation.
Conclusion
Intravital imaging of DmdEGFP‐mdx muscle permits novel experimental approaches such as the study of revertant and therapeutically restored dystrophin in vivo and ex vivo.
The mechanism of pattern formation during limb muscle development remains poorly understood. The canonical view holds that naïve limb muscle progenitor cells (MPCs) invade a pre-established pattern ...of muscle connective tissue, thereby forming individual muscles. Here we show that early murine embryonic limb MPCs highly accumulate pSMAD1/5/9, demonstrating active signaling of bone morphogenetic proteins (BMP) in these cells. Overexpression of inhibitory SMAD6 in limb MPCs abrogated BMP signaling, impaired their migration and proliferation, and accelerated myogenic lineage progression. Fewer primary myofibers developed, causing an aberrant proximodistal muscle pattern. Patterning was not disturbed when SMAD6 was overexpressed in differentiated muscle, implying that the proximodistal muscle pattern depends on BMP-mediated expansion of MPCs prior to their differentiation. We show that limb MPCs differentially express Hox genes, and Hox-expressing MPCs displayed active BMP signaling. SMAD6 overexpression caused loss of HOXA11 in early limb MPCs. In conclusion, our data show that BMP signaling controls expansion of embryonic limb MPC as a prerequisite for establishing the proximodistal muscle pattern, a process that involves expression of Hox genes.
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