Viscoelastic Strain Response (ViSR) ultrasound assessment of viscoelastic properties in human duchenne muscular dystrophy, in vivo

2012 ◽  
Author(s):  
Mallory R. Scola ◽  
Melissa C. Caughey ◽  
Diane O. Meyer ◽  
Regina Emmett ◽  
James F. Howard ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Viscoelastic Properties ◽  
Strain Response ◽  
Viscoelastic Strain ◽  
Ultrasound Assessment

scholarly journals In vivo genome editing in mouse restores dystrophin expression in Duchenne muscular dystrophy patient muscle fibers

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Menglong Chen ◽  
Hui Shi ◽  
Shixue Gou ◽  
Xiaomin Wang ◽  
Lei Li ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Mouse Model ◽  
Genome Editing ◽  
Large Scale ◽  
Gene Editing ◽  
High Efficiency ◽  
Muscle Fibers ◽  
Muscle Cells

Abstract Background Mutations in the DMD gene encoding dystrophin—a critical structural element in muscle cells—cause Duchenne muscular dystrophy (DMD), which is the most common fatal genetic disease. Clustered regularly interspaced short palindromic repeat (CRISPR)-mediated gene editing is a promising strategy for permanently curing DMD. Methods In this study, we developed a novel strategy for reframing DMD mutations via CRISPR-mediated large-scale excision of exons 46–54. We compared this approach with other DMD rescue strategies by using DMD patient-derived primary muscle-derived stem cells (DMD-MDSCs). Furthermore, a patient-derived xenograft (PDX) DMD mouse model was established by transplanting DMD-MDSCs into immunodeficient mice. CRISPR gene editing components were intramuscularly delivered into the mouse model by adeno-associated virus vectors. Results Results demonstrated that the large-scale excision of mutant DMD exons showed high efficiency in restoring dystrophin protein expression. We also confirmed that CRISPR from Prevotella and Francisella 1(Cas12a)-mediated genome editing could correct DMD mutation with the same efficiency as CRISPR-associated protein 9 (Cas9). In addition, more than 10% human DMD muscle fibers expressed dystrophin in the PDX DMD mouse model after treated by the large-scale excision strategies. The restored dystrophin in vivo was functional as demonstrated by the expression of the dystrophin glycoprotein complex member β-dystroglycan. Conclusions We demonstrated that the clinically relevant CRISPR/Cas9 could restore dystrophin in human muscle cells in vivo in the PDX DMD mouse model. This study demonstrated an approach for the application of gene therapy to other genetic diseases.

scholarly journals In vivo non-invasive monitoring of dystrophin correction in a new Duchenne muscular dystrophy reporter mouse

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Leonela Amoasii ◽  
Hui Li ◽  
Yu Zhang ◽  
Yi-Li Min ◽  
Efrain Sanchez-Ortiz ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Genetic Disorder ◽  
Dystrophin Gene ◽  
Luciferase Reporter ◽  
Luciferase Expression ◽  
Gene Correction ◽  
Reading Frame ◽  
Non Invasive

Abstract Duchenne muscular dystrophy (DMD) is a fatal genetic disorder caused by mutations in the dystrophin gene. To enable the non-invasive analysis of DMD gene correction strategies in vivo, we introduced a luciferase reporter in-frame with the C-terminus of the dystrophin gene in mice. Expression of this reporter mimics endogenous dystrophin expression and DMD mutations that disrupt the dystrophin open reading frame extinguish luciferase expression. We evaluated the correction of the dystrophin reading frame coupled to luciferase in mice lacking exon 50, a common mutational hotspot, after delivery of CRISPR/Cas9 gene editing machinery with adeno-associated virus. Bioluminescence monitoring revealed efficient and rapid restoration of dystrophin protein expression in affected skeletal muscles and the heart. Our results provide a sensitive non-invasive means of monitoring dystrophin correction in mouse models of DMD and offer a platform for testing different strategies for amelioration of DMD pathogenesis.

scholarly journals Applications of CRISPR/Cas9 for the Treatment of Duchenne Muscular Dystrophy

2018 ◽  
Vol 8 (4) ◽  
pp. 38 ◽  
Author(s):  
Kenji Lim ◽  
Chantal Yoon ◽  
Toshifumi Yokota
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Wide Spectrum ◽  
Membrane Integrity ◽  
Dystrophin Gene ◽  
Muscle Membrane ◽  
Reading Frame ◽  
Long Term Treatment

Duchenne muscular dystrophy (DMD) is a fatal X-linked recessive neuromuscular disease prevalent in 1 in 3500 to 5000 males worldwide. As a result of mutations that interrupt the reading frame of the dystrophin gene (DMD), DMD is characterized by a loss of dystrophin protein that leads to decreased muscle membrane integrity, which increases susceptibility to degeneration. CRISPR/Cas9 technology has garnered interest as an avenue for DMD therapy due to its potential for permanent exon skipping, which can restore the disrupted DMD reading frame in DMD and lead to dystrophin restoration. An RNA-guided DNA endonuclease system, CRISPR/Cas9 allows for the targeted editing of specific sequences in the genome. The efficacy and safety of CRISPR/Cas9 as a therapy for DMD has been evaluated by numerous studies in vitro and in vivo, with varying rates of success. Despite the potential of CRISPR/Cas9-mediated gene editing for the long-term treatment of DMD, its translation into the clinic is currently challenged by issues such as off-targeting, immune response activation, and sub-optimal in vivo delivery. Its nature as being mostly a personalized form of therapy also limits applicability to DMD patients, who exhibit a wide spectrum of mutations. This review summarizes the various CRISPR/Cas9 strategies that have been tested in vitro and in vivo for the treatment of DMD. Perspectives on the approach will be provided, and the challenges faced by CRISPR/Cas9 in its road to the clinic will be briefly discussed.

Nintedanib as a new therapeutic agent for Duchenne muscular dystrophy: preclinical in vitro and in vivo studies

2017 ◽  
Vol 27 ◽  
pp. S191
Author(s):  
P. Piñol ◽  
E. Fernández-Simón ◽  
X. Suárez ◽  
N. de Luna ◽  
A. Molins ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Therapeutic Agent ◽  
In Vivo Studies

scholarly journals Microtubules that form the stationary lattice of muscle fibers are dynamic and nucleated at Golgi elements

2013 ◽  
Vol 203 (2) ◽  
pp. 205-213 ◽  
Author(s):  
Sarah Oddoux ◽  
Kristien J. Zaal ◽  
Victoria Tate ◽  
Aster Kenea ◽  
Shuktika A. Nandkeolyar ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Mouse Muscle ◽  
Muscle Diseases ◽  
Superresolution Microscopy ◽  
Nuclear Membranes ◽  
Organizing Center ◽  
Static Images

Skeletal muscle microtubules (MTs) form a nonclassic grid-like network, which has so far been documented in static images only. We have now observed and analyzed dynamics of GFP constructs of MT and Golgi markers in single live fibers and in the whole mouse muscle in vivo. Using confocal, intravital, and superresolution microscopy, we find that muscle MTs are dynamic, growing at the typical speed of ∼9 µm/min, and forming small bundles that build a durable network. We also show that static Golgi elements, associated with the MT-organizing center proteins γ-tubulin and pericentrin, are major sites of muscle MT nucleation, in addition to the previously identified sites (i.e., nuclear membranes). These data give us a framework for understanding how muscle MTs organize and how they contribute to the pathology of muscle diseases such as Duchenne muscular dystrophy.

scholarly journals Soluble guanylate cyclase stimulation mitigates skeletal and cardiac muscle dysfunction in a mdx model of Duchenne muscular dystrophy

2021 ◽  
Author(s):  
Ling Zhang ◽  
Yuanyuan Xu ◽  
Keyvan Yousefi ◽  
Camila I. Irion ◽  
Roger A. Alvarez ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Guanylate Cyclase ◽  
Soluble Guanylate Cyclase ◽  
Signal Propagation ◽  
Guanosine Monophosphate ◽  
Muscle Pathology ◽  
Mdx Mice

AbstractThe impairment of neuronal nitric oxide synthase (nNOS) signaling contributes to disease pathology in the muscle wasting disorder Duchenne muscular dystrophy (DMD). nNOS signal propagation occurs through nitric oxide sensitive soluble guanylate cyclase (sGC), a critical source of cyclic guanosine monophosphate (cGMP) in muscle. Although both nNOS and sGC activity are impaired in DMD patients, little is known about sGC as a therapeutic target. In this study, we tested the hypothesis that stimulating sGC activity with the allosteric agonist BAY41-8543 mitigates striated muscle pathology in the mdx4cv mouse model of DMD. In contrast to DMD patients, mdx mice exhibited greater basal sGC activity than wild type controls with preservation of cGMP levels due partly to upregulation of sGC in some muscles. Stimulating sGC activity in mdx mice with BAY41-8543 substantially reduced skeletal muscle damage, macrophage densities and inflammation and significantly increased resistance to contraction-induced fatigue. BAY41-8543 also enhanced in vivo diaphragm function while reducing breathing irregularities suggesting improved respiratory function. BAY41-8543 attenuated cardiac hypertrophic remodeling, fibrosis and diastolic dysfunction including left atrium enlargement in aged mdx mice. Overall, sGC stimulation significantly mitigated skeletal and cardio-respiratory dysfunction in mdx4cv mice. Importantly, this study provides compelling pre-clinical evidence supporting sGC as a novel target in DMD and the repurposing of FDA-approved sGC stimulators, such as riociguat and veraciguat, as a novel therapeutic approach for DMD.

scholarly journals Ultrasonography validation for early alteration of diaphragm echodensity and function in the mdx mouse model of Duchenne muscular dystrophy

PLoS ONE ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. e0245397
Author(s):  
Antonietta Mele ◽  
Paola Mantuano ◽  
Adriano Fonzino ◽  
Francesco Rana ◽  
Roberta Francesca Capogrosso ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Mouse Model ◽  
Ex Vivo ◽  
Eccentric Contraction ◽  
Invasive Technique ◽  
Mdx Mouse ◽  
Early Stages ◽  
Over Time

The mdx mouse model of Duchenne muscular dystrophy is characterized by functional and structural alterations of the diaphragm since early stages of pathology, closely resembling patients’ condition. In recent years, ultrasonography has been proposed as a useful longitudinal non-invasive technique to assess mdx diaphragm dysfunction and evaluate drug efficacy over time. To date, only a few preclinical studies have been conducted. Therefore, an independent validation of this method by different laboratories is needed to increase results reliability and reduce biases. Here, we performed diaphragm ultrasonography in 3- and 6-month-old mdx mice, the preferred age-window for pharmacology studies. The alteration of diaphragm function over time was measured as diaphragm ultrasound movement amplitude. At the same time points, a first-time assessment of diaphragm echodensity was performed, as an experimental index of progressive loss of contractile tissue. A parallel evaluation of other in vivo and ex vivo dystrophy-relevant readouts was carried out. Both 3- and 6-month-old mdx mice showed a significant decrease in diaphragm amplitude compared to wild type (wt) mice. This index was well-correlated either with in vivo running performance or ex vivo isometric tetanic force of isolated diaphragm. In addition, diaphragms from 6-month-old dystrophic mice were also highly susceptible to eccentric contraction ex vivo. Importantly, we disclosed an age-dependent increase in echodensity in mdx mice not observed in wt animals, which was independent from abdominal wall thickness. This was accompanied by a notable increase of pro-fibrotic TGF-β1 levels in the mdx diaphragm and of non-muscle tissue amount in diaphragm sections stained by hematoxylin & eosin. Our findings corroborate the usefulness of diaphragm ultrasonography in preclinical drug studies as a powerful tool to monitor mdx pathology progression since early stages.

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