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Extraocular muscles (EOM) have a strikingly different disease profile than limb skeletal muscles. It has long been known that they are spared in Duchenne (DMD) and other forms of muscular dystrophy. Despite many studies, the cause for this sparing is not understood. We have proposed that differences in myogenic precursor cell properties in EOM maintain normal morphology over the lifetime of individuals with DMD due to either greater proliferative potential or greater resistance to injury. This hypothesis was tested by exposing wild type and mdx:utrophin+/− (het) mouse EOM and limb skeletal muscles to 18 Gy gamma irradiation, a dose known to inhibit satellite cell proliferation in limb muscles. As expected, over time het limb skeletal muscles displayed reduced central nucleation mirrored by a reduction in Pax7-positive cells, demonstrating a significant loss in regenerative potential. In contrast, in the first month post-irradiation in the het EOM, myofiber cross-sectional areas first decreased, then increased, but ultimately returned to normal compared to non-irradiated het EOM. Central nucleation significantly increased in the first post-irradiation month, resembling the dystrophic limb phenotype. This correlated with decreased EECD34 stem cells and a concomitant increase and subsequent return to normalcy of both Pax7 and Pitx2-positive cell density. By two months, normal het EOM morphology returned. It appears that irradiation disrupts the normal method of EOM remodeling, which react paradoxically to produce increased numbers of myogenic precursor cells. This suggests that the EOM contain myogenic precursor cell types resistant to 18 Gy gamma irradiation, allowing return to normal morphology 2 months post-irradiation. This supports our hypothesis that ongoing proliferation of specialized regenerative populations in the het EOM actively maintains normal EOM morphology in DMD. Ongoing studies are working to define the differences in the myogenic precursor cells in EOM as well as the cellular milieu in which they reside. 相似文献
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Cintia Yuri Matsumura Bruno Menezes de Oliveira Madeleine Durbeej Maria Julia Marques 《PloS one》2013,8(6)
Duchenne muscular dystrophy (DMD) is the most common childhood myopathy, characterized by muscle loss and cardiorespiratory failure. While the genetic basis of DMD is well established, secondary mechanisms associated with dystrophic pathophysiology are not fully clarified yet. In order to obtain new insights into the molecular mechanisms of muscle dystrophy during earlier stages of the disease, we performed a comparative proteomic profile of the spared extraocular muscles (EOM) vs. affected diaphragm from the mdx mice, using a label based shotgun proteomic approach. Out of the 857 identified proteins, 42 to 62 proteins had differential abundance of peptide ions. The calcium-handling proteins sarcalumenin and calsequestrin-1 were increased in control EOM compared with control DIA, reinforcing the view that constitutional properties of EOM are important for their protection against myonecrosis. The finding that galectin-1 (muscle regeneration), annexin A1 (anti-inflammatory) and HSP 47 (fibrosis) were increased in dystrophic diaphragm provides novel insights into the mechanisms through which mdx affected muscles are able to counteract dystrophy, during the early stage of the disease. Overall, the shotgun technique proved to be suitable to perform quantitative comparisons between distinct dystrophic muscles and allowed the suggestion of new potential biomarkers and drug targets for dystrophinopaties. 相似文献
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Extraocular muscles (EOM) represent a unique muscle group that controls eye movements and originates from head mesoderm, while the more typically studied body and limb muscles are somite-derived. Aiming to investigate myogenic progenitors (satellite cells) in EOM versus limb and diaphragm of adult mice, we have been using flow cytometry in combination with myogenic-specific Cre-loxP lineage marking for cell isolation. While analyzing cells from the EOM of mice that harbor Myf5Cre-driven GFP expression, we identified in addition to the expected GFP+ myogenic cells (presumably satellite cells), a second dominant GFP+ population distinguished as being Sca1+, non-myogenic, and exhibiting a fibro/adipogenic potential. This unexpected population was not only unique to EOM compared to the other muscles but also specific to the Myf5Cre-driven reporter when compared to the MyoDCre driver. Histological studies of periocular tissue preparations demonstrated the presence of Myf5Cre-driven GFP+ cells in connective tissue locations adjacent to the muscle masses, including cells in the vasculature wall. These vasculature-associated GFP+ cells were further identified as mural cells based on the presence of the specific XLacZ4 transgene. Unlike the EOM satellite cells that originate from a Pax3-negative lineage, these non-myogenic Myf5Cre-driven GFP+ cells appear to be related to cells of a Pax3-expressing origin, presumably derived from the neural crest. In all, our lineage tracing based on multiple reporter lines has demonstrated that regardless of common ancestral expression of Myf5, there is a clear distinction between periocular myogenic and non-myogenic cell lineages according to their mutually exclusive antecedence of MyoD and Pax3 gene activity. 相似文献
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Le Grand F Auda-Boucher G Levitsky D Rouaud T Fontaine-Pérus J Gardahaut MF 《Experimental cell research》2004,301(2):232-241
We investigated whether the vessel-associated or endothelial cells within mouse embryo muscles can be a source of myogenic progenitors. Immunodetection of the stem cell surface markers, CD34 and Flk1, which are known to characterize the endothelial lineage, was done throughout the course of embryo muscle development. Both markers appeared to be restricted to the vessel-associated cells. On the basis of CD34 labeling, the reactive cells were purified by magnetic-bead selection from the limb muscles of 17-dpc desmin+/-LacZ mouse embryos and characterized by fluorescence-activated cell sorting. The cells in the selected CD34(+) population appeared to be approximately 95% positive for Flk1, but usually negative for CD45. We demonstrated that in vitro the CD34(+)/Flk1(+) population differentiated into endothelial cells and skeletal myofibers. When transplanted into mdx mouse muscle, this population displayed a high propensity to disperse within the recipient muscle, fuse with the host myofibers, and restore dystrophin expression. The marked ability of the embryonic muscle endothelial cells to activate myogenic program could be related to their somitic origin. 相似文献
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Alexander Nystr?m Johanna Holmblad Fatima Pedrosa-Domell?f Takako Sasaki Madeleine Durbeej 《Matrix biology》2006,25(6):382-385
Mutations in the gene encoding laminin (LM) alpha2 chain cause congenital muscular dystrophy. Here, we show that extraocular muscle (EOM) is spared upon complete LMalpha2 chain absence. The major LM chains in limb muscle basement membranes are alpha2, beta1, beta2 and gamma1 whereas alpha2, alpha4, beta1, beta2 and gamma1 chains are expressed in EOM. Expression of LMalpha4 chain mRNA is further increased in LMalpha2 chain deficient EOM. Mainly integrin alpha7X1 subunit, which binds to laminin-411, is expressed in EOM and in contrast to dystrophic limb muscle, sustained integrin alpha7B expression is seen in LMalpha2 chain deficient EOM. We propose that LMalpha4 chain, possibly by binding to integrin alpha7BX1beta1D, protects EOM in LMalpha2 chain deficient muscular dystrophy. 相似文献
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Nakae Y Hirasaka K Goto J Nikawa T Shono M Yoshida M Stoward PJ 《Histochemistry and cell biology》2008,129(4):489-501
Dystrophic muscles suffer from enhanced oxidative stress. We have investigated whether administration of an antioxidant, epigallocatechin-3-gallate
(EGCG), a component of green tea, reduces their oxidative stress and pathophysiology in mdx mice, a mild phenotype model of human Duchenne-type muscular dystrophy. EGCG (5 mg/kg body weight in saline) was injected
subcutaneously 4× a week into the backs of C57 normal and dystrophin-deficient mdx mice for 8 weeks after birth. Saline was injected into normal and mdx controls. EGCG had almost no observable effects on normal mice or on the body weights of mdx mice. In contrast, it produced the following improvements in the blood chemistry, muscle histology, and electrophysiology
of the treated mdx mice. First, the activities of serum creatine kinase were reduced to normal levels. Second, the numbers of fluorescent lipofuscin
granules per unit volume of soleus and diaphragm muscles were significantly decreased by about 50% compared to the numbers
in the corresponding saline-treated controls. Third, in sections of diaphragm and soleus muscles, the relative area occupied
by histologically normal muscle fibres increased significantly 1.5- to 2-fold whereas the relative areas of connective tissue
and necrotic muscle fibres were substantially reduced. Fourth, the times for the maximum tetanic force of soleus muscles to
fall by a half increased to almost normal values. Fifth, the amount of utrophin in diaphragm muscles increased significantly
by 17%, partially compensating for the lack of dystrophin expression. 相似文献
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Anna Bodanovsky Noga Guttman Hila Barzilai-Tutsch Ola Genin Oshrat Levy Mark Pines Orna Halevy 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》2014
Halofuginone has been shown to prevent fibrosis via the transforming growth factor-β/Smad3 pathway in muscular dystrophies. We hypothesized that halofuginone would reduce apoptosis—the presumed cause of satellite-cell depletion during muscle degradation—in the mdx mouse model of Duchenne muscular dystrophy. Six-week-old mdx mouse diaphragm exhibited fourfold higher numbers of apoptotic nuclei compared with wild-type mice as determined by a TUNEL assay. Apoptotic nuclei were found in macrophages and in Pax7-expressing cells; some were located in centrally-nucleated regenerating myofibers. Halofuginone treatment of mdx mice reduced the apoptotic nuclei number in the diaphragm, together with reduction in Bax and induction in Bcl2 levels in myofibers isolated from these mice. A similar effect was observed when halofuginone was added to cultured myofibers. No apparent effect of halofuginone was observed in wild-type mice. Inhibition of apoptosis or staurosporine-induced apoptosis by halofuginone in mdx primary myoblasts and C2 myogenic cell line, respectively, was reflected by less pyknotic/apoptotic cells and reduced Bax expression. This reduction was reversed by a phosphinositide-3-kinase and mitogen-activated protein kinase/extracellular signal-regulated protein kinase inhibitors, suggesting involvement of these pathways in mediating halofuginone's effects on apoptosis. Halofuginone increased apoptosis in α smooth muscle actin- and prolyl 4-hydroxylase β-expressing cells in mdx diaphragm and in myofibroblasts, the major source of extracellular matrix. The data suggest an additional mechanism by which halofuginone improves muscle pathology and function in muscular dystrophies. 相似文献
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Intraarterial injection of muscle-derived CD34(+)Sca-1(+) stem cells restores dystrophin in mdx mice 总被引:22,自引:0,他引:22
Torrente Y Tremblay JP Pisati F Belicchi M Rossi B Sironi M Fortunato F El Fahime M D'Angelo MG Caron NJ Constantin G Paulin D Scarlato G Bresolin N 《The Journal of cell biology》2001,152(2):335-348
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Judith N. Haslett Peter B. Kang Mei Han Alvin T. Kho Despina Sanoudou Jay M. Volinski Alan H. Beggs Isaac S. Kohane Louis M. Kunkel 《Mammalian genome》2005,16(10):739-748
The phenotypic differences among Duchenne muscular dystrophy patients, mdx mice, and mdx5cv mice suggest that despite the common etiology of dystrophin deficiency, secondary mechanisms have a substantial influence
on phenotypic severity. The differential response of various skeletal muscles to dystrophin deficiency supports this hypothesis.
To explore these differences, gene expression profiles were generated from duplicate RNA targets extracted from six different
skeletal muscles (diaphragm, soleus, gastrocnemius, quadriceps, tibialis anterior, and extensor digitorum longus) from wild-type,
mdx, and mdx5cv mice, resulting in 36 data sets for 18 muscle samples. The data sets were compared in three different ways: (1) among wild-type
samples only, (2) among all 36 data sets, and (3) between strains for each muscle type. The molecular profiles of soleus and
diaphragm separate significantly from the other four muscle types and from each other. Fiber-type proportions can explain
some of these differences. These variations in wild-type gene expression profiles may also reflect biomechanical differences
known to exist among skeletal muscles. Further exploration of the genes that most distinguish these muscles may help explain
the origins of the biomechanical differences and the reasons why some muscles are more resistant than others to dystrophin
deficiency.
Electronic Supplementary Material Electronic Supplementary material is available for this article at
and accessible for authorised users.
Judith N. Haslett, Peter B. Kang These authors contributed equally to this work. 相似文献
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Thibaut Larcher Aude Lafoux Laurent Tesson Séverine Remy Virginie Thepenier Virginie Fran?ois Caroline Le Guiner Helicia Goubin Maéva Dutilleul Lydie Guigand Gilles Toumaniantz Anne De Cian Charlotte Boix Jean-Baptiste Renaud Yan Cherel Carine Giovannangeli Jean-Paul Concordet Ignacio Anegon Corinne Huchet 《PloS one》2014,9(10)
A few animal models of Duchenne muscular dystrophy (DMD) are available, large ones such as pigs or dogs being expensive and difficult to handle. Mdx (X-linked muscular dystrophy) mice only partially mimic the human disease, with limited chronic muscular lesions and muscle weakness. Their small size also imposes limitations on analyses. A rat model could represent a useful alternative since rats are small animals but 10 times bigger than mice and could better reflect the lesions and functional abnormalities observed in DMD patients. Two lines of Dmd mutated-rats (Dmdmdx) were generated using TALENs targeting exon 23. Muscles of animals of both lines showed undetectable levels of dystrophin by western blot and less than 5% of dystrophin positive fibers by immunohistochemistry. At 3 months, limb and diaphragm muscles from Dmdmdx rats displayed severe necrosis and regeneration. At 7 months, these muscles also showed severe fibrosis and some adipose tissue infiltration. Dmdmdx rats showed significant reduction in muscle strength and a decrease in spontaneous motor activity. Furthermore, heart morphology was indicative of dilated cardiomyopathy associated histologically with necrotic and fibrotic changes. Echocardiography showed significant concentric remodeling and alteration of diastolic function. In conclusion, Dmdmdx rats represent a new faithful small animal model of DMD. 相似文献
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Bigot A Jacquemin V Debacq-Chainiaux F Butler-Browne GS Toussaint O Furling D Mouly V 《Biology of the cell / under the auspices of the European Cell Biology Organization》2008,100(3):189-199
Background information. Aging of human skeletal muscle results in a decline in muscle mass and force, and excessive turnover of muscle fibres, such as in muscular dystrophies, further increases this decline. Although it has been shown in rodents, by cross‐age transplantation of whole muscles, that the environment plays an important role in this process, the implication of proliferating aging of the muscle progenitors has been poorly investigated, particularly in humans, since the regulation of cell proliferation differs between rodents and humans. The myogenic differentiation of human myoblasts is regulated by the muscle‐specific regulatory factors. Cross‐talk between the muscle‐specific regulatory factors and the cell cycle regulators is essential for differentiation. The aim of the present study was to determine the effects of replicative senescence on the myogenic programme of human myoblasts. Results. We showed that senescent myoblasts, which could not re‐enter the cell cycle, are still able to differentiate and form multinucleated myotubes. However, these myotubes are significantly smaller. The expression of muscle‐specific regulatory factors and cell cycle regulators was analysed in proliferating myoblasts and compared with senescent cells. We have observed a delay and a decrease in the muscle‐specific regulatory factors and the cyclin‐dependent kinase inhibitor p57 during the early step of differentiation in senescent myoblasts, as well as an increase in the fibroblastic markers. Conclusions. Our results demonstrate that replicative senescence alters the expression of the factors triggering muscle differentiation in human myoblasts and could play a role in the regenerative defects observed in muscular diseases and during normal skeletal‐muscle aging. 相似文献
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The mdx mutant is a murine homologous model of Duchenne muscular dystrophy (DMD). Fibre types determined by the myosin-ATPase technique in soleus muscles were compared in C57BL/10 control and mdx mice from 3 to 52 weeks of age. The control strain continuously presented 70% of type 2 fibres whereas the mdx strain showed an increase to 80% at 6 weeks and a subsequent decline. In mdx muscles, necrosis which begins at 3 weeks of age did not affect specifically one type of fibre. Type grouping was never observed when muscle regeneration occurred. Fibres of intermediate type (1C and 2C) were found continuously up to 52 weeks of age in the mdx mutant. The foci of small immature regenerating fibres were of type 2C but never 1C. A few mature fibres were either of type 2C or 1C. We suggest that the presence of intermediate type fibres could result from the co-expression of type 1 and 2 myosin heavy chains, indicating a transition from type 2 to type 1 in regenerating fibres. 相似文献
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Muscular dystrophy is a genetic disorder of skeletal muscle characterized by progressive muscle weakness. Here we assessed whether muscle wasting affects cell viability and mechanical properties of extensor digitorum longus (EDL) and of tibialis anterior (TA) tendons from mdx dystrophic mice compared to wild type (WT) mice. mdx mice represent the classical animal model for human Duchenne muscular dystrophy, and show several signs of the pathology, including a decrease in specific force and an increase of fibrotic index. Cell viability of tendons was evaluated by histological analysis, and viscoelastic properties have been assessed by a rapid measurement protocol that allowed us to compute, at the same time, tissue complex compliance for all the frequencies of interest. Confocal microscopy and mechanical properties measurements revealed that mdx tendons, compared to WT ones, have an increase in the number of dead cells and a significant reduction in tissue elasticity for all the frequencies that were tested. These findings indicate a reduced quality of the tissue. Moreover, mdx tendons have an increase in the viscous response, indicating that during dynamic loading, they dissipate more energy compared to WT. Our results demonstrate that muscular dystrophy involves not only muscle wasting, but also alteration in the viscoelastic properties of tendons, suggesting a paracrine effect of altered skeletal muscle on tendinous tissue. 相似文献
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It has been previously shown that transiently denervated, neonatal dystrophic muscle fails to undergo the degeneration–regeneration cycle characteristic of murine dystrophy (Moschella and Ontell, 1987). Thus, the myosatellite cells (myogenic stem cells) in these muscles have been spared the mitotic challenge to which dystrophic myosatellite cells are normally subjected early in the time course of the disease. By in vitro evaluation of the proliferative capacity of myosatellite cells derived from extensor digitorum longus (EDL) muscles of 100-day-old genetically normal (+/+) and genetically dystrophic [dy/dy (129ReJdy/dy)] mice and from muscles of age-matched mice that had been neonatally denervated (by sciaticotomy) and allowed to reinnervate, it has been possible to directly determine whether the cessation of spontaneous regeneration in older dy/dy muscles in vivo, is due to an innate defect in the proliferative capacity of the myosatellite cells or exhaustion of the myosatellite cells' mitotic activity during the regenerative phase of the disease. This study demonstrates that transient neonatal denervation of dystrophic muscle (Den.dy/dy) increases the number of muscle colony-forming cells (MCFs) permilligram of wet weight muscle tissue, increases the plating efficiency, and significantly increases the in vitro mitotic activity of dystrophic myosatellite cells toward normal values. The increased mitotic capability of myosatellite cells derived from Den.dy/dy muscle as compared to unoperated dy/dy muscle suggests that there is no innate defect in the proliferative capacity of the myosatellite cells of dy/dy muscles and that the cessation of spontaneous regeneration in the dy/dy muscles is related to the exhaustion of their myosatellite cells' mitotic capability. © 1992 John Wiley & Sons, Inc. 相似文献
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Yusuke Echigoya Joshua Lee Merryl Rodrigues Tetsuya Nagata Jun Tanihata Ashkan Nozohourmehrabad Dharminder Panesar Bailey Miskew Yoshitsugu Aoki Toshifumi Yokota 《PloS one》2013,8(7)
Duchenne muscular dystrophy (DMD), one of the most common and lethal genetic disorders, and the mdx mouse myopathies are caused by a lack of dystrophin protein. These dystrophic muscles contain sporadic clusters of dystrophin-expressing revertant fibers (RFs), as detected by immunohistochemistry. RFs are known to arise from muscle precursor cells with spontaneous exon skipping (alternative splicing) and clonally expand in size with increasing age through the process of muscle degeneration/regeneration. The expansion of revertant clusters is thought to represent the cumulative history of muscle regeneration and proliferation of such precursor cells. However, the precise mechanisms by which RFs arise and expand are poorly understood. Here, to test the effects of mutation types and aging on RF expansion and muscle regeneration, we examined the number of RFs in mdx mice (containing a nonsense mutation in exon 23) and mdx52 mice (containing deletion mutation of exon 52) with the same C57BL/6 background at 2, 6, 12, and 18months of age. Mdx mice displayed a significantly higher number of RFs compared to mdx52 mice in all age groups, suggesting that revertant fiber expansion largely depends on the type of mutation and/or location in the gene. A significant increase in the expression and clustering levels of RFs was found beginning at 6months of age in mdx mice compared with mdx52 mice. In contrast to the significant expansion of RFs with increasing age, the number of centrally nucleated fibers and embryonic myosin heavy chain-positive fibers (indicative of cumulative and current muscle regeneration, respectively) decreased with age in both mouse strains. These results suggest that mutation types and aging differently affect revertant fiber expansion in mdx and mdx52 mice. 相似文献
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Stéphanie Daval Chantal Rocher Yan Cherel Elisabeth Le Rumeur 《Cellular & molecular biology letters》2010,15(1):134-152
The dystrophin-glycoprotein complex (DGC) is a large trans-sarcolemmal complex that provides a linkage between the subsarcolemmal
cytoskeleton and the extracellular matrix. In skeletal muscle, it consists of the dystroglycan, sarcoglycan and cytoplasmic
complexes, with dystrophin forming the core protein. The DGC has been described as being absent or greatly reduced in dystrophin-deficient
muscles, and this lack is considered to be involved in the dystrophic phenotype. Such a decrease in the DGC content was observed
in dystrophin-deficient muscle from humans with muscular dystrophy and in mice with X-linked muscular dystrophy (mdx mice). These deficits were observed in total muscle homogenates and in partially membrane-purified muscle fractions, the
so-called KCl-washed microsomes. Here, we report that most of the proteins of the DGC are actually present at normal levels
in the mdx mouse muscle plasma membrane. The proteins are detected in dystrophic animal muscles when the immunoblot assay is performed
with crude surface membrane fractions instead of the usually employed KCl-washed microsomes. We propose that these proteins
form SDS-insoluble membrane complexes when dystrophin is absent. 相似文献