scholarly journals Changes in Myonuclear Number During Postnatal Growth –Implications for AAV Gene Therapy for Muscular Dystrophy

2021 ◽  
pp. 1-8
Author(s):  
Jennifer Morgan ◽  
Francesco Muntoni
Keyword(s):  
Skeletal Muscle ◽  
Gene Therapy ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Muscle Growth ◽  
Postnatal Growth ◽  
Muscle Fibres ◽  
Postnatal Life ◽  
Adult Skeletal Muscle ◽  
Dividing Cells

Adult skeletal muscle is a relatively stable tissue, as the multinucleated muscle fibres contain post-mitotic myonuclei. During early postnatal life, muscle growth occurs by the addition of skeletal muscle stem cells (satellite cells) or their progeny to growing muscle fibres. In Duchenne muscular dystrophy, which we shall use as an example of muscular dystrophies, the muscle fibres lack dystrophin and undergo necrosis. Satellite-cell mediated regeneration occurs, to repair and replace the necrotic muscle fibres, but as the regenerated muscle fibres still lack dystrophin, they undergo further cycles of degeneration and regeneration. AAV gene therapy is a promising approach for treating Duchenne muscular dystrophy. But for a single dose of, for example, AAV coding for dystrophin, to be effective, the treated myonuclei must persist, produce sufficient dystrophin and a sufficient number of nuclei must be targeted. This latter point is crucial as AAV vector remains episomal and does not replicate in dividing cells. Here, we describe and compare the growth of skeletal muscle in rodents and in humans and discuss the evidence that myofibre necrosis and regeneration leads to the loss of viral genomes within skeletal muscle. In addition, muscle growth is expected to lead to the dilution of the transduced nuclei especially in case of very early intervention, but it is not clear if growth could result in insufficient dystrophin to prevent muscle fibre breakdown. This should be the focus of future studies.

scholarly journals Co-Transplantation of Bone Marrow-MSCs and Myogenic Stem/Progenitor Cells from Adult Donors Improves Muscle Function of Patients with Duchenne Muscular Dystrophy

Cells ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 1119
Author(s):  
Aleksandra Klimczak ◽  
Agnieszka Zimna ◽  
Agnieszka Malcher ◽  
Urszula Kozlowska ◽  
Katarzyna Futoma ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Bone Marrow ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Progenitor Cells ◽  
Muscle Function ◽  
Progenitor Cell ◽  
Genetic Disorder ◽  
Cell Populations ◽  
Muscle Fibres

Duchenne muscular dystrophy (DMD) is a genetic disorder associated with a progressive deficiency of dystrophin that leads to skeletal muscle degeneration. In this study, we tested the hypothesis that a co-transplantation of two stem/progenitor cell populations, namely bone marrow-derived mesenchymal stem cells (BM-MSCs) and skeletal muscle-derived stem/progenitor cells (SM-SPCs), directly into the dystrophic muscle can improve the skeletal muscle function of DMD patients. Three patients diagnosed with DMD, confirmed by the dystrophin gene mutation, were enrolled into a study approved by the local Bioethics Committee (no. 79/2015). Stem/progenitor cells collected from bone marrow and skeletal muscles of related healthy donors, based on HLA matched antigens, were expanded in a closed MC3 cell culture system. A simultaneous co-transplantation of BM-MSCs and SM-SPCs was performed directly into the biceps brachii (two patients) and gastrocnemius (one patient). During a six-month follow-up, the patients were examined with electromyography (EMG) and monitored for blood kinase creatine level. Muscle biopsies were examined with histology and assessed for dystrophin at the mRNA and protein level. A panel of 27 cytokines was analysed with multiplex ELISA. We did not observe any adverse effects after the intramuscular administration of cells. The efficacy of BM-MSC and SM-SPC application was confirmed through an EMG assessment by an increase in motor unit parameters, especially in terms of duration, amplitude range, area, and size index. The beneficial effect of cellular therapy was confirmed by a decrease in creatine kinase levels and a normalised profile of pro-inflammatory cytokines. BM-MSCs may support the pro-regenerative potential of SM-SPCs thanks to their trophic, paracrine, and immunomodulatory activity. Both applied cell populations may fuse with degenerating skeletal muscle fibres in situ, facilitating skeletal muscle recovery. However, further studies are required to optimise the dose and timing of stem/progenitor cell delivery.

Duchenne muscular dystrophy gene therapy in the canine model

2018 ◽  
Author(s):  
◽  
Kasun Kodippili
Keyword(s):  
Skeletal Muscle ◽  
Gene Therapy ◽  
Animal Model ◽  
Duchenne Muscular Dystrophy ◽  
Sarcoplasmic Reticulum ◽  
Muscular Dystrophy ◽  
Canine Model ◽  
Disease Pathogenesis ◽  
Preclinical Treatment ◽  
Expression And Activity

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] Duchenne muscular dystrophy (DMD) is a devastating muscle wasting disease caused by loss of function mutations in the dystrophin gene, resulting in the absence of dystrophin, a structural protein in muscle. DMD is the most common form of inherited muscle disease in childhood, with an incidence of 1 in 5000 live male births worldwide. The dystrophin-null mdx mouse has been the most widely used animal model for DMD research over the last 30 years. Dystrophin-deficient DMD dogs have also gained prominence as a highly relevant preclinical animal model due to their high phenotypic homology to human DMD patients. Preclinical treatment studies in these dogs are expected to better inform and guide clinical trials in human patients. However, there are still significant gaps in our understanding of the disease pathogenesis and gene therapy in the canine model. The goals of my dissertation work were to establish reagents and methodologies to study preclinical treatment in the canine model, and subsequently characterize the disease pathogenesis and gene therapy in DMD dogs. To this end, I first characterized 65 epitope-specific human dystrophin monoclonal antibodies for their reactivity in canine skeletal and cardiac muscle by both immunofluorescence (IF) staining and western blot. I found species-specific, tissue-specific and assay-specific patterns of reactivity in these antibodies. Importantly, out of the 65 antibodies that I characterized, I recognized 15 antibodies that worked well for canine tissue on both IF staining and western blot, which are recommended for DMD research in the canine model. ... Dystrophin-independent gene therapy for DMD takes advantage of disease-modifying genes that are either structural and/or functional homologues of dystrophin, or alternative targets that are involved in disease pathogenesis. One such alternative target gene is the sarcoplasmic reticulum calcium ATPase 2a (SERCA2a), a pump that transports calcium ions from the cytoplasm into the sarcoplasmic reticulum. I show that SERCA2a expression and activity are impaired, and that calcium homeostasis is dysregulated in DMD dog skeletal muscle. Furthermore, gene therapy with human SERCA2a restored expression and activity of the pump, and improved several aspects of muscle function and histopathology in DMD dog skeletal muscle. In summary, this dissertation work advances our knowledge of the disease pathogenesis and gene therapy prospects in the canine model of DMD, a highly relevant and valuable preclinical DMD model.

scholarly journals The influence of undernutrition during gestation on skeletal muscle cellularity and on the expression of genes that control muscle growth

2004 ◽  
Vol 91 (3) ◽  
pp. 331-339 ◽  
Author(s):  
Stéphanie Bayol ◽  
Doiran Jones ◽  
Geoffrey Goldspink ◽  
Neil C. Stickland
Keyword(s):  
Skeletal Muscle ◽  
Growth Rate ◽  
Muscle Growth ◽  
Postnatal Growth ◽  
Nuclear Antigen ◽  
Control Group ◽  
Muscle Fibres ◽  
Control Muscle ◽  
Expression Of Genes ◽  
Ad Libitum

We examined the effects of two levels of gestational undernutrition (50% and 40% of ad libitum) on postnatal growth rate, skeletal muscle cellularity and the expression of genes that control muscle growth, in the offspring at weaning. The results showed that the rat pups born to mothers fed the 50% diet during gestation and a control diet during lactation had an increased postnatal growth rate compared with the pups fed the more restricted diet (40% of ad libitum). Surprisingly, the growth rate of the control group (ad libitum) was intermediate between the 50% and 40% groups. The restricted diets did not alter the number of muscle fibres in the semitendinosus muscle of the offspring but the number of muscle nuclei was reduced by 16% in the 40% group compared with the control group. In the 50% group, the lightest pups at birth (L) had elevated muscle insulin-like growth factor (IGF)-1, IGF binding protein (BP)-5 and proliferating cell nuclear antigen (PCNA) mRNA compared with the L pups from both the control and 40% groups. The heaviest pups at birth (H) in the 50% group had increased levels of IGFBP-4, PCNA and M-cadherin mRNA compared with both the control and 40% groups. Levels of IGF-1 receptor, myostatin and MyoD mRNA did not correlate with postnatal growth. Both H and L pups from the 40% group had reduced muscle IGF-1 mRNA but all other transcripts examined were similar to control levels. The results suggest that the increased postnatal growth rate, which accompanied milder fetal undernutrition (50%), may be due to a more active local muscle IGF system and increased muscle-cell proliferation.

Caveolin-3 in skeletal muscle fibres of Duchenne muscular dystrophy and mdx mouse

1997 ◽  
Vol 7 (6-7) ◽  
pp. 436
Author(s):  
Y. Hagiwara ◽  
Y. Nishina ◽  
M. Imamura ◽  
M. Yoshida ◽  
T. Kikuchi ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Mdx Mouse ◽  
Muscle Fibres ◽  
Skeletal Muscle Fibres

scholarly journals Molecular regulation of skeletal muscle tissue formation and development

2018 ◽  
Vol 63 (No. 11) ◽  
pp. 489-499
Author(s):  
M. Nesvadbova ◽  
G. Borilova
Keyword(s):  
Skeletal Muscle ◽  
Molecular Genetic ◽  
Muscle Growth ◽  
Postnatal Growth ◽  
The Body ◽  
Meat Production ◽  
Muscle Fibres ◽  
Tissue Formation ◽  
Fibre Growth ◽  
Myogenic Factors

This article provides a complex overview of the different stages of myogenesis with an emphasis on the molecular, genetic and cellular bases for skeletal muscle growth. Animals with higher number of medium-sized muscle fibres produce meat of higher quality and in higher quantity. The number of muscle fibres that are created in the body is largely decided during the process of myogenesis. This review describes the main stages of embryonic skeletal myogenesis and the myogenic factors that control myogenesis in epaxial and hypaxial somites, limbs, the head and neck as well as postnatal muscle fibre growth and regeneration. An understanding of the molecular and genetic factors influencing the prenatal and postnatal growth of skeletal muscle is essential for the development of the new strategies and practical approaches to meat production.

scholarly journals In situ localisation of single-stranded DNA breaks in nuclei of a subpopulation of cells within regenerating skeletal muscle of the dystrophic mdx mouse

1992 ◽  
Vol 102 (3) ◽  
pp. 653-662 ◽  
Author(s):  
G.R. Coulton ◽  
B. Rogers ◽  
P. Strutt ◽  
M.J. Skynner ◽  
D.J. Watt
Keyword(s):  
Skeletal Muscle ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Dna Polymerase ◽  
Muscle Regeneration ◽  
Regenerative Process ◽  
Muscle Fibres ◽  
Dna Breaks ◽  
Expression Of Genes ◽  
Muscle Lesions

Degeneration of muscle fibres during the early stages of Duchenne Muscular Dystrophy (DMD) is accompanied by muscle fibre regeneration where cell division and myoblast fusion to form multinucleate myotubes within the lesions appear to recapitulate the events of normal muscle development. The mechanisms that govern the expression of genes regulating differentiation of myoblasts in regenerating skeletal muscle are of great interest for the development of future therapies designed to stimulate muscle regeneration. We show here that single-stranded breaks in DNA are localised in nuclei, using an exogenously applied medium containing labelled deoxynucleotides and the Klenow fragment of DNA polymerase I. The nuclei of a sub-population of cells lying in the inflammatory infiltrate of lesions in the skeletal muscle of the muscular dystrophic mouse (mdx), a genetic homologue of DMD, were labelled in this fashion. By contrast, labelled cells were completely absent from the muscles of normal non-myopathic animals (C57BL/10) and non-lesioned areas of mdx muscles. Cells expressing the muscle-specific regulatory gene, myogenin, were also found within mononucleate cells and myotubes within similar mdx muscle lesions. While we cannot yet say that the cells labelled by the DNA polymerase reaction are in fact differentiating, they were found only in significant numbers within mdx muscle lesions where new muscle fibres appear, providing strong circumstantial evidence that they are intimately associated with the regenerative process. Using a range of nucleases and different DNA polymerases, we show that the DNA polymerase-labelling reaction observed was DNA-dependent and most probably due to infilling of naturally occurring single-stranded gaps in DNA. Since the regenerative process in human Duchenne Muscular Dystrophy is apparently less effective than that seen in mdx mice, continued study of single-stranded DNA breaks may help to elucidate further the mechanisms controlling the expression of genes that characterise the myogenic process during skeletal muscle regeneration. Such findings might be applied in the development of future therapies designed to stimulate muscle regeneration in human dystrophies.

Immunological hurdles in the path to gene therapy for Duchenne muscular dystrophy

2002 ◽  
Vol 4 (23) ◽  
pp. 1-23 ◽  
Author(s):  
Dominic J. Wells ◽  
Aurora Ferrer ◽  
Kim E. Wells
Keyword(s):  
Gene Therapy ◽  
Immune Response ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Gene Transfer ◽  
Immune Responses ◽  
Muscle Fibres ◽  
Dystrophic Muscle ◽  
Western Blots ◽  
Muscle Wasting Disease

Patients with Duchenne muscular dystrophy (DMD), an X-linked lethal muscle-wasting disease, have abnormal expression of the protein dystrophin within their muscle fibres. In the mdx mouse model of this condition, both germline and neonatal somatic gene transfers of dystrophin cDNAs have demonstrated the potential of gene therapy in treating DMD. However, in many DMD patients, there appears to be no dystrophin expression when muscle biopsies are immunostained or western blots are performed. This raises the possibility that the expression of dystrophin following gene transfer might trigger a destructive immune response against this ‘neoantigen’. Immune responses can also be generated against the gene transfer vector used to transfect the dystrophic muscle, and the combined immune response could further damage the already inflamed muscle. These problems are now beginning to be investigated in immunocompetent mdx mice. Although much work remains to be done, there are promising indications that these immune responses might not prove as much of a concern as originally envisaged.

scholarly journals Gene Therapy for Duchenne Muscular Dystrophy

2021 ◽  
pp. 1-14
Author(s):  
Nertiyan Elangkovan ◽  
George Dickson
Keyword(s):  
Gene Therapy ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Gene Transfer ◽  
Viral Vector ◽  
Cardiac Complications ◽  
Muscle Fibres ◽  
Large Animal ◽  
Large Animal Models ◽  
Advanced Therapy

Duchenne muscular dystrophy (DMD) is an X-linked, muscle wasting disease that affects 1 in 5000 males. Affected individuals become wheelchair bound by the age of twelve and eventually die in their third decade due to respiratory and cardiac complications. The disease is caused by mutations in the DMD gene that codes for dystrophin. Dystrophin is a structural protein that maintains the integrity of muscle fibres and protects them from contraction-induced damage. The absence of dystrophin compromises the stability and function of the muscle fibres, eventually leading to muscle degeneration. So far, there is no effective treatment for deteriorating muscle function in DMD patients. A promising approach for treating this life-threatening disease is gene transfer to restore dystrophin expression using a safe, non-pathogenic viral vector called adeno-associated viral (AAV) vector. Whilst microdystrophin gene transfer using AAV vectors shows extremely impressive therapeutic success so far in large animal models of DMD, translating this advanced therapy medicinal product from bench to bedside still offers scope for many optimization steps. In this paper, the authors review the current progress of AAV-microdystrophin gene therapy for DMD and other treatment strategies that may apply to a subset of DMD patients depending on the mutations they carry.

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