Mechanics of dystrophin deficient skeletal muscles in very young mice and effects of age

2021 ◽  
Author(s):  
Michael A. Lopez ◽  
Sherina Bontiff ◽  
Mary Adeyeye ◽  
Aziz I Shaibani ◽  
Matthew S. Alexander ◽  
...  
Keyword(s):  
Skeletal Muscles ◽  
Force Production ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Fiber Direction ◽  
Transverse Fiber ◽  
Muscle Necrosis ◽  
Relaxation Curves ◽  
Passive Mechanics

The MDX mouse is an animal model of Duchenne muscular dystrophy, a human disease marked by an absence of the cytoskeletal protein, dystrophin. We hypothesized that (1) dystrophin serves a complex mechanical role in skeletal muscles by contributing to passive compliance, viscoelastic properties, and contractile force production and (2) age is a modulator of passive mechanics of skeletal muscles of the MDX mouse. Using an in vitro biaxial mechanical testing apparatus, we measured passive length-tension relationships in the muscle fiber direction as well as transverse to the fibers, viscoelastic stress-relaxation curves, and isometric contractile properties. To avoid confounding secondary effects of muscle necrosis, inflammation, and fibrosis, we used very young 3-week-old mice whose muscles reflected the pre-fibrotic and pre-necrotic state. Compared to controls, 1) muscle extensibility and compliance were greater in both along fiber direction and transverse to fiber direction in MDX mice and 2) the relaxed elastic modulus was greater in dystrophin-deficient diaphragms. Furthermore, isometric contractile muscle stress was reduced in the presence and absence of transverse fiber passive stress. We also examined the effect of age on the diaphragm length-tension relationships and found that diaphragm muscles from 9-months old MDX mice were significantly less compliant and less extensible than those of muscles from very young MDX mice. Our data suggest that the age of the MDX mouse is a determinant of the passive mechanics of the diaphragm; in the pre-fibrotic/pre-necrotic stage, muscle extensibility and compliance, as well as viscoelasticity, and muscle contractility are altered by loss of dystrophin.

Muscle genome-wide expression profiling during disease evolution in mdx mice

2009 ◽  
Vol 37 (2) ◽  
pp. 119-132 ◽  
Author(s):  
Mario Marotta ◽  
Claudia Ruiz-Roig ◽  
Yaris Sarria ◽  
Jose Luis Peiro ◽  
Fatima Nuñez ◽  
...  
Keyword(s):  
Candidate Genes ◽  
Genetic Defect ◽  
Strong Relationship ◽  
Pcr Analysis ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Matrix Remodeling ◽  
Disease Evolution ◽  
Genome Wide ◽  
Muscle Necrosis

Mdx mice show a milder phenotype than Duchenne patients despite bearing an analogous genetic defect. Our aim was to sort out genes, differentially expressed during the evolution of skeletal muscle mdx mouse disease, to elucidate the mechanisms by which these animals overcome the lack of dystrophin. Genome-wide microarray-based gene expression analysis was carried out at 3 wk and 1.5 and 3 mo of life. Candidate genes were selected by comparing: 1) mdx vs. controls at each point in time, and 2) mdx mice and 3) control mice among the three points in time. The first analysis showed a strong upregulation (96%) of inflammation-related genes and in >75% of genes related to cell adhesion, muscle structure/regeneration, and extracellular matrix remodeling during mdx disease evolution. Lgals3, Postn, Ctss, and Sln genes showed the strongest variations. The analysis performed among points in time demonstrated significant changes in Ecm1, Spon1, Thbs1, Csrp3, Myo10, Pde4b, and Adamts-5 exclusively during mdx mice lifespan. RT-PCR analysis of Postn, Sln, Ctss, Thbs1, Ecm1, and Adamts-5 expression from 3 wk to 9 mo, confirmed microarray data and demonstrated variations beyond 3 mo of age. A high-confidence functional network analysis demonstrated a strong relationship between them and showed two main subnetworks, having Dmd- Utrn- Myo10 and Adamts5- Thbs1- Spon1-Postn as principal nodes, which are functionally linked to Abca1, Actn4, Crebbp, Csrp3, Lama1, Lama3, Mical2, Mical3, Myf6, Pxn, and Sparc genes. Candidate genes may participate in the decline of muscle necrosis in mdx mice and could be considered potential therapeutic targets for Duchenne patients.

Talin, vinculin and DRP (utrophin) concentrations are increased at mdx myotendinous junctions following onset of necrosis

1994 ◽  
Vol 107 (6) ◽  
pp. 1477-1483 ◽  
Author(s):  
D.J. Law ◽  
D.L. Allen ◽  
J.G. Tidball
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Cytoskeletal Proteins ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Myotendinous Junction ◽  
Glycoprotein Complex ◽  
Transmembrane Glycoprotein ◽  
Muscle Necrosis ◽  
Myotendinous Junctions

Duchenne muscular dystrophy (DMD) and the myopathy seen in the mdx mouse both result from absence of the protein dystrophin. Structural similarities between dystrophin and other cytoskeletal proteins, its enrichment at myotendinous junctions, and its indirect association with laminin mediated by a transmembrane glycoprotein complex suggest that one of dystrophin's functions in normal muscle is to form one of the links between the actin cytoskeleton and the extracellular matrix. Unlike Duchenne muscular dystrophy patients, mdx mice suffer only transient muscle necrosis, and are able to regenerate damaged muscle tissue. The present study tests the hypothesis that mdx mice partially compensate for dystrophin's absence by upregulating one or more dystrophin-independent mechanisms of cytoskeleton-membrane association. Quantitative analysis of immunoblots of adult mdx muscle samples showed an increase of approximately 200% for vinculin and talin, cytoskeletal proteins that mediate thin filament-membrane interactions at myotendinous junctions. Blots also showed an increase (143%) in the dystrophin-related protein called utrophin, another myotendinous junction constituent, which may be able to substitute for dystrophin directly. Muscle samples from 2-week-old animals, a period immediately preceding the onset of muscle necrosis, showed no significant differences in protein concentration between mdx and controls. Quantitative analyses of confocal images of myotendinous junctions from mdx and control muscles show significantly higher concentrations of talin and vinculin at the myotendinous junctions of mdx muscle. These findings indicate that mdx mice may compensate in part for the absence of dystrophin by increased expression of other molecules that subsume dystrophin's mechanical function.

scholarly journals ProNGF/p75NTR Axis Drives Fiber Type Specification by Inducing the Fast-Glycolytic Phenotype in Mouse Skeletal Muscle Cells

Cells ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 2232
Author(s):  
Valentina Pallottini ◽  
Mayra Colardo ◽  
Claudia Tonini ◽  
Noemi Martella ◽  
Georgios Strimpakos ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Myogenic Differentiation ◽  
Fiber Type ◽  
Pcr Analysis ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Muscle Physiology

Despite its undisputable role in the homeostatic regulation of the nervous system, the nerve growth factor (NGF) also governs the relevant cellular processes in other tissues and organs. In this study, we aimed at assessing the expression and the putative involvement of NGF signaling in skeletal muscle physiology. To reach this objective, we employed satellite cell-derived myoblasts as an in vitro culture model. In vivo experiments were performed on Tibialis anterior from wild-type mice and an mdx mouse model of Duchenne muscular dystrophy. Targets of interest were mainly assessed by means of morphological, Western blot and qRT-PCR analysis. The results show that proNGF is involved in myogenic differentiation. Importantly, the proNGF/p75NTR pathway orchestrates a slow-to-fast fiber type transition by counteracting the expression of slow myosin heavy chain and that of oxidative markers. Concurrently, proNGF/p75NTR activation facilitates the induction of fast myosin heavy chain and of fast/glycolytic markers. Furthermore, we also provided evidence that the oxidative metabolism is impaired in mdx mice, and that these alterations are paralleled by a prominent buildup of proNGF and p75NTR. These findings underline that the proNGF/p75NTR pathway may play a crucial role in fiber type determination and suggest its prospective modulation as an innovative therapeutic approach to counteract muscle disorders.

scholarly journals In vitro and in vivo analysis of human fibroblast reprogramming and multipotency

2015 ◽  
Vol 20 (3) ◽  
Author(s):  
Rongqing Pang ◽  
Xiangqing Zhu ◽  
Jia Geng ◽  
Yongyun Zhang ◽  
Qiang Wang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Human Fibroblasts ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Therapeutic Effects ◽  
Tail Vein ◽  
Tail Vein Injection ◽  
Activity Measurements ◽  
Human Dystrophin

AbstractMultipotent stem cells have potential therapeutic roles in the treatment of Duchenne muscular dystrophy (DMD). However, the limited access to stem cell sources restricts their clinical application. To address this issue, we established a simple in vitro epigenetic reprogramming technique in which skin fibroblasts are induced to dedifferentiate into multipotent cells. In this study, human fibroblasts were isolated from circumcised adult foreskin and were reprogrammed by co-culture for 72 h with fish oocyte extract (FOE) in serum-free medium. The cells were then observed and analyzed by immunofluorescence staining, flow cytometry and in vitro differentiation assays. Then FOE-treated human fibroblasts were transplanted by tail vein injection into irradiated mdx mice, an animal model of DMD. Two months after injection, the therapeutic effects of FOE-treated fibroblasts on mdx skeletal muscle were evaluated by serum creatine kinase (CK) activity measurements and by immunostaining and RT-PCR of human dystrophin expression. The results indicated that the reprogrammed fibroblasts expressed higher levels of the pluripotent antigen markers SSEA-4, Nanog and Oct-4, and were able to differentiate in vitro into adipogenic cells, osteoblastic cells, and myotube-like cells. Tail vein injection of FOE-treated fibroblasts into irradiated mdx mice slightly reduced serum CK activity and the percentage of centrally nucleated myofibers two months after cell transplantation. Furthermore, we confirmed human dystrophin protein and mRNA expression in mdx mouse skeletal muscle. These data demonstrated that FOE-treated fibroblasts were multipotent and could integrate into mdx mouse myofibers through the vasculature.

Systemic administration of IGF-I enhances oxidative status and reduces contraction-induced injury in skeletal muscles of mdx dystrophic mice

2006 ◽  
Vol 291 (3) ◽  
pp. E499-E505 ◽  
Author(s):  
Jonathan D. Schertzer ◽  
James G. Ryall ◽  
Gordon S. Lynch
Keyword(s):  
Tibialis Anterior ◽  
Skeletal Muscles ◽  
Force Production ◽  
Mdx Mice ◽  
Cross Sectional ◽  
Myosin Heavy Chain Isoform ◽  
Glycoprotein Complex ◽  
Igf I ◽  
Dystrophin Glycoprotein Complex ◽  
Endocrine Factor

The absence of dystrophin and resultant disruption of the dystrophin glycoprotein complex renders skeletal muscles of dystrophic patients and dystrophic mdx mice susceptible to contraction-induced injury. Strategies to reduce contraction-induced injury are of critical importance, because this mode of damage contributes to the etiology of myofiber breakdown in the dystrophic pathology. Transgenic overexpression of insulin-like growth factor-I (IGF-I) causes myofiber hypertrophy, increases force production, and can improve the dystrophic pathology in mdx mice. In contrast, the predominant effect of continuous exogenous administration of IGF-I to mdx mice at a low dose (1.0–1.5 mg·kg−1·day−1) is a shift in muscle phenotype from fast glycolytic toward a more oxidative, fatigue-resistant, slow muscle without alterations in myofiber cross-sectional area, muscle mass, or maximum force-producing capacity. We found that exogenous administration of IGF-I to mdx mice increased myofiber succinate dehydrogenase activity, shifted the overall myosin heavy chain isoform composition toward a slower phenotype, and, most importantly, reduced contraction-induced damage in tibialis anterior muscles. The deficit in force-producing capacity after two damaging lengthening contractions was reduced significantly in tibialis anterior muscles of IGF-I-treated (53 ± 4%) compared with untreated mdx mice (70 ± 5%, P < 0.05). The results provide further evidence that IGF-I administration can enhance the functional properties of dystrophic skeletal muscle and, compared with results in transgenic mice or virus-mediated overexpression, highlight the disparities in different models of endocrine factor delivery.

Tubulyzine®, a novel tri-substituted triazine, prevents the early cell death of transplanted myogenic cells and improves transplantation success

2003 ◽  
Vol 81 (2) ◽  
pp. 81-90 ◽  
Author(s):  
E El Fahime ◽  
M Bouchentouf ◽  
B F Benabdallah ◽  
D Skuk ◽  
J F Lafreniere ◽  
...  
Keyword(s):  
Cell Death ◽  
Annexin V ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Limiting Factor ◽  
Matrix Remodeling ◽  
Tissue Implantation ◽  
Myoblast Transplantation

Myoblast transplantation (MT) is a potential therapeutic approach for several muscular dystrophies. A major limiting factor is that only a low percentage of the transplanted myoblasts survives the procedure. Recent advances regarding how and when the myoblasts die indicate that events preceding actual tissue implantation and during the first days after the transplantation are crucial. Myoseverin, a recently identified tri-substituted purine, was shown to induce in vitro the fission of multinucleated myotubes and affect the expression of a variety of growth factors, and immunomodulation, extracellular matrix-remodeling, and stress response genes. Since the effects of myoseverin are consistent with the activation of pathways involved in wound healing and tissue regeneration, we have investigated whether pretreatment and co-injection of myoblasts with Tubulyzine® (microtubule lysing triazine), an optimized myoseverin-like molecule recently identified from a triazine library, could reduce myoblast cell death following their transplantation and consequently improves the success of myoblast transplantation. In vitro, using annexin-V labeling, we showed that Tubulyzine (5 µM) prevents normal myoblasts from apoptosis induced by staurosporine (1 µM). In vivo, the pretreatment and co-injection of immortal and normal myoblasts with Tubulyzine reduced significantly cell death (assessed by the radio-labeled thymidine of donor DNA) and increased survival of myoblasts transplanted in Tibialis anterior (TA) muscles of mdx mice, thus giving rise to more hybrid myofibers compared to transplanted untreated cells. Our results suggest that Tubulyzine can be used as an in vivo survival factor to improve the myoblast-mediated gene transfer approach.Key words: myoblast survival, mdx mouse, myoblast transplantation, microtubule-binding molecule, cell death.

scholarly journals Lengthening-contractions in isolated myocardium impact force development and worsen cardiac contractile function in the mdx mouse model of muscular dystrophy

2011 ◽  
Vol 110 (2) ◽  
pp. 512-519 ◽  
Author(s):  
Ying Xu ◽  
Dawn A. Delfín ◽  
Jill A. Rafael-Fortney ◽  
Paul M. L. Janssen
Keyword(s):  
Contractile Function ◽  
Wild Type Mouse ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Wild Type ◽  
Cardiac Contractile Function ◽  
Lengthening Contractions ◽  
Peak Tension ◽  
The Impact

Lengthening-contractions exert eccentric stress on myofibers in normal myocardium. In congestive heart failure caused by a variety of diseases, the impact of lengthening-contractions of myocardium likely becomes more prevalent and severe. The present study introduces a method to investigate the role of stretching imposed by repetitive lengthening-contractions in myocardium under near-physiological conditions. By exerting various stretch-release ramps while the muscle is contracting, consecutive lengthening-contractions and their potential detrimental effect on cardiac function can be studied. We tested our model and hypothesis in age-matched (young and adult) mdx and wild-type mouse right ventricular trabeculae. These linear and ultrathin muscles possess all major cardiac cell types, and their contractile behavior very closely mimics that of the whole myocardium. In the first group of experiments, 10 lengthening-contractions at various magnitudes of stretch were performed in trabeculae from 10-wk-old mdx and wild-type mice. In the second group, 100 lengthening-contractions at various magnitudes were conducted in trabeculae from 10- and 20-wk-old mice. The peak isometric active developed tension (Fdev, in mN/mm2) and kinetic parameters time to peak tension (TTP, in ms) and time from peak tension to half-relaxation (RT50, in ms) were measured. Our results indicate lengthening-contractions significantly impact contractile behavior, and that dystrophin-deficient myocardium in mdx mice is significantly more susceptible to these damaging lengthening-contractions. The results indicate that lengthening-contractions in intact myocardium can be used in vitro to study this emerging contributor to cardiomyopathy.

Quantitative assessment of muscle damage in the mdx mouse model of Duchenne muscular dystrophy using polarization-sensitive optical coherence tomography

2013 ◽  
Vol 115 (9) ◽  
pp. 1393-1401 ◽  
Author(s):  
Xiaojie Yang ◽  
Lixin Chin ◽  
Blake R. Klyen ◽  
Tea Shavlakadze ◽  
Robert A. McLaughlin ◽  
...  
Keyword(s):  
Optical Coherence Tomography ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Muscle Tissue ◽  
Volume Fraction ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Optical Coherence ◽  
Histological Sections ◽  
Muscle Necrosis

Minimally invasive, high-resolution imaging of muscle necrosis has the potential to aid in the assessment of diseases such as Duchenne muscular dystrophy. Undamaged muscle tissue possesses high levels of optical birefringence due to its anisotropic ultrastructure, and this birefringence decreases when the tissue undergoes necrosis. In this study, we present a novel technique to image muscle necrosis using polarization-sensitive optical coherence tomography (PS-OCT). From PS-OCT scans, our technique is able to quantify the birefringence in muscle tissue, generating an image indicative of the tissue ultrastructure, with areas of abnormally low birefringence indicating necrosis. The technique is demonstrated on excised skeletal muscles from exercised dystrophic mdx mice and control C57BL/10ScSn mice with the resulting images validated against colocated histological sections. The technique additionally gives a measure of the proportion (volume fraction) of necrotic tissue within the three-dimensional imaging field of view. The percentage necrosis assessed by this technique is compared against the percentage necrosis obtained from manual assessment of histological sections, and the difference between the two methods is found to be comparable to the interobserver variability of the histological assessment. This is the first published demonstration of PS-OCT to provide automated assessment of muscle necrosis.

Contractile properties of diaphragm muscle segments from old mdx and old transgenic mdx mice

1997 ◽  
Vol 272 (6) ◽  
pp. C2063-C2068 ◽  
Author(s):  
G. S. Lynch ◽  
J. A. Rafael ◽  
R. T. Hinkle ◽  
N. M. Cole ◽  
J. S. Chamberlain ◽  
...  
Keyword(s):  
Life Span ◽  
Functional Properties ◽  
Transgenic Animal ◽  
Mouse Embryos ◽  
Diaphragm Muscle ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Dystrophin Expression ◽  
Structural And Functional Properties

Diaphragm muscles of young (4- to 6-mo-old) mdx mice show severe fiber necrosis and have normalized forces and powers 60 and 46% of the values for control C57BL/10 mice. In contrast, microinjection of mdx mouse embryos with a truncated dystrophin minigene has produced young transgenic mdx (tg-mdx) mice with a level of dystrophin expression and structural and functional properties of diaphragm muscle strips measured in vitro not different from those of control mice. Whether dystrophin expression and functional corrections persist for the life span of these animals is not know. We tested the null hypothesis that, in old (24 mo) tg-mdx mice, dystrophin expression is adequate and diaphragm muscle strips have forces and powers not different from values for diaphragm muscle strips from young tg-mdx mice or control mice. Compared with control values, diaphragm muscle strips from old mdx mice had normalized forces and powers of 48 and 31%, respectively. Expression of dystrophin persisted in diaphragm muscles of old tg-mdx mice, and functional properties were not different from diaphragm muscles of young tg-mdx or young or old control mice. These results suggest that, with a transgenic animal approach, dystrophin expression and functional corrections persist for the life span of the animals.

Hindlimb immobilization applied to 21-day-oldmdx mice prevents the occurrence of muscle degeneration

1999 ◽  
Vol 86 (3) ◽  
pp. 924-931 ◽  
Author(s):  
Asghar Mokhtarian ◽  
Jean Pascal Lefaucheur ◽  
Patrick C. Even ◽  
Alain Sebille
Keyword(s):  
Skeletal Muscle ◽  
Motor Activity ◽  
Skeletal Muscles ◽  
Extensor Digitorum Longus ◽  
Contralateral Side ◽  
Mdx Mouse ◽  
Mdx Mice ◽  
Muscle Degeneration ◽  
Limb Immobilization ◽  
Old Mice

Dystrophin-deficient skeletal muscles of mdx mice undergo their first rounds of degeneration-regeneration at the age of 14–28 days. This feature is thought to result from an increase in motor activity at weaning. In this study, we hypothesize that if the muscle is prevented from contracting, it will avoid the degenerative changes that normally occur. For this purpose, we developed a procedure of mechanical hindlimb immobilization in 3-wk-old mice to restrain soleus (Sol) and extensor digitorum longus (EDL) muscles in the stretched or shortened position. After a 14-day period of immobilization, the striking feature was the low percentage of regenerated (centronucleated) myofibers in Sol and EDL muscles, regardless of the length at which they were fixed, compared with those on the contralateral side (stretched Sol: 8.4 ± 6.5 vs. 46.6 ± 10.3%, P = 0.0008; shortened Sol: 1.2 ± 1.6 vs. 50.4 ± 16.4%, P = 0.0008; stretched EDL: 05 ± 0.5 vs. 32.9 ± 17.5%, P = 0.002; shortened EDL: 3.3 ± 3.1 vs. 34.7 ± 11.1%, P = 0.002). Total numbers of myofibers did not change with immobilization. This study shows that limb immobilization prevents the occurrence of the first round of myofiber necrosis in mdx mice and suggests that muscle contractions play a role in the skeletal muscle degeneration of dystrophin-deficient mdx mouse muscles.

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