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Progress in Neurobiology Apr 2024Dystrophin loss due to mutations in the Duchenne muscular dystrophy (DMD) gene is associated with a wide spectrum of neurocognitive comorbidities, including an aberrant... (Review)
Review
Dystrophin loss due to mutations in the Duchenne muscular dystrophy (DMD) gene is associated with a wide spectrum of neurocognitive comorbidities, including an aberrant unconditioned fear response to stressful/threat stimuli. Dystrophin-deficient animal models of DMD demonstrate enhanced stress reactivity that manifests as sustained periods of immobility. When the threat is repetitive or severe in nature, dystrophinopathy phenotypes can be exacerbated and even cause sudden death. Thus, it is apparent that enhanced sensitivity to stressful/threat stimuli in dystrophin-deficient vertebrates is a legitimate cause of concern for patients with DMD that could impact neurocognition and pathophysiology. This review discusses our current understanding of the mechanisms and consequences of the hypersensitive fear response in preclinical models of DMD and the potential challenges facing clinical translatability.
Topics: Animals; Humans; Dystrophin; Fear; Muscular Dystrophy, Duchenne; Mutation; Vertebrates
PubMed: 38484964
DOI: 10.1016/j.pneurobio.2024.102590 -
International Journal of Molecular... Feb 2024Dystrophin () gene mutations are associated with skeletal muscle diseases such as Duchenne and Becker Muscular Dystrophy (BMD) and X-linked dilated cardiomyopathy...
Dystrophin () gene mutations are associated with skeletal muscle diseases such as Duchenne and Becker Muscular Dystrophy (BMD) and X-linked dilated cardiomyopathy (XL-DCM). To investigate the molecular basis of DCM in a 37-year-old woman. Clinical and genetic investigations were performed. Genetic testing was performed with whole exome sequencing (WES) using the Illumina platform. According to the standard protocol, a variant found by WES was confirmed in all available members of the family by bi-directional capillary Sanger resequencing. The effect of the variant was investigated by using an in silico prediction of pathogenicity. The index case was a 37-year-old woman diagnosed with DCM at the age of 33. A germline heterozygous A>G transversion at nucleotide 10103 in the gene, leading to an aspartic acid-glycine substitution at the amino acid 3368 of the DMD protein (c.10103A>G p.Asp3368Gly), was identified and confirmed by PCR-based Sanger sequencing of the exon 70. In silico prediction suggests that this variant could have a deleterious impact on protein structure and functionality (CADD = 30). The genetic analysis was extended to the first-degree relatives of the proband (mother, father, and sister) and because of the absence of the variant in both parents, the p.Asp3368Gly substitution was considered as occurring de novo. Then, the direct sequencing analysis of her 8-year-old son identified as hemizygous for the same variant. The young patient did not present any signs or symptoms attributable to DCM, but reported asthenia and presented with bilateral calf hypertrophy at clinical examination. Laboratory testing revealed increased levels of creatinine kinase (maximum value of 19,000 IU/L). We report an early presentation of dilated cardiomyopathy in a 33-year-old woman due to a de novo pathogenic variant of the dystrophin () gene (p.Asp3368Gly). Genetic identification of this variant allowed an early diagnosis of a skeletal muscle disease in her son.
Topics: Humans; Female; Adult; Child; Dystrophin; Cardiomyopathy, Dilated; Muscular Dystrophy, Duchenne; Mothers
PubMed: 38474032
DOI: 10.3390/ijms25052787 -
International Journal of Molecular... Feb 2024The diaphragm muscle is essential for breathing, and its dysfunctions can be fatal. Many disorders affect the diaphragm, including muscular dystrophies. Despite the...
The diaphragm muscle is essential for breathing, and its dysfunctions can be fatal. Many disorders affect the diaphragm, including muscular dystrophies. Despite the clinical relevance of targeting the diaphragm, there have been few studies evaluating diaphragm function following a given experimental treatment, with most of these involving anti-inflammatory drugs or gene therapy. Cell-based therapeutic approaches have shown success promoting muscle regeneration in several mouse models of muscular dystrophy, but these have focused mainly on limb muscles. Here we show that transplantation of as few as 5000 satellite cells directly into the diaphragm results in consistent and robust myofiber engraftment in dystrophin- and fukutin-related protein-mutant dystrophic mice. Transplanted cells also seed the stem cell reservoir, as shown by the presence of donor-derived satellite cells. Force measurements showed enhanced diaphragm strength in engrafted muscles. These findings demonstrate the feasibility of cell transplantation to target the diseased diaphragm and improve its contractility.
Topics: Mice; Animals; Muscular Dystrophy, Duchenne; Diaphragm; Mice, Inbred mdx; Muscle, Skeletal; Cell Transplantation
PubMed: 38473751
DOI: 10.3390/ijms25052503 -
Journal of Neuromuscular Diseases 2024Single exon duplications account for disease in a minority of Duchenne muscular dystrophy patients. Exon skipping in these patients has the potential to be highly...
Single exon duplications account for disease in a minority of Duchenne muscular dystrophy patients. Exon skipping in these patients has the potential to be highly therapeutic through restoration of full-length dystrophin expression. We conducted a 48-week open label study of casimersen and golodirsen in 3 subjects with an exon 45 or 53 duplication. Two subjects (aged 18 and 23 years) were non-ambulatory at baseline. Upper limb, pulmonary, and cardiac function appeared stable in the 2 subjects in whom they could be evaluated. Dystrophin expression increased from 0.94 % ±0.59% (mean±SD) of normal to 5.1% ±2.9% by western blot. Percent dystrophin positive fibers also rose from 14% ±17% at baseline to 50% ±42% . Our results provide initial evidence that the use of exon-skipping drugs may increase dystrophin levels in patients with single-exon duplications.
Topics: Adolescent; Humans; Male; Young Adult; Dystrophin; Exons; Gene Duplication; Muscular Dystrophy, Duchenne; Oligonucleotides
PubMed: 38461513
DOI: 10.3233/JND-230107 -
EMBO Molecular Medicine Apr 2024Clinical deployment of oligonucleotides requires delivery technologies that improve stability, target tissue accumulation and cellular internalization. Exosomes show...
Clinical deployment of oligonucleotides requires delivery technologies that improve stability, target tissue accumulation and cellular internalization. Exosomes show potential as ideal delivery vehicles. However, an affordable generalizable system for efficient loading of oligonucleotides on exosomes remain lacking. Here, we identified an Exosomal Anchor DNA Aptamer (EAA) via SELEX against exosomes immobilized with our proprietary CP05 peptides. EAA shows high binding affinity to different exosomes and enables efficient loading of nucleic acid drugs on exosomes. Serum stability of thrombin inhibitor NU172 was prolonged by exosome-loading, resulting in increased blood flow after injury in vivo. Importantly, Duchenne Muscular Dystrophy PMO can be readily loaded on exosomes via EAA (EXO). EXO elicited significantly greater muscle cell uptake, tissue accumulation and dystrophin expression than PMO in vitro and in vivo. Systemic administration of EXO elicited therapeutic levels of dystrophin restoration and functional improvements in mdx mice. Altogether, our study demonstrates that EAA enables efficient loading of different nucleic acid drugs on exosomes, thus providing an easy and generalizable strategy for loading nucleic acid therapeutics on exosomes.
Topics: Animals; Mice; Dystrophin; Mice, Inbred mdx; Exosomes; Morpholinos; Muscular Dystrophy, Duchenne; Oligonucleotides
PubMed: 38448545
DOI: 10.1038/s44321-024-00049-7 -
EMBO Molecular Medicine Apr 2024Cell therapy for muscular dystrophy has met with limited success, mainly due to the poor engraftment of donor cells, especially in fibrotic muscle at an advanced stage...
Cell therapy for muscular dystrophy has met with limited success, mainly due to the poor engraftment of donor cells, especially in fibrotic muscle at an advanced stage of the disease. We developed a cell-mediated exon skipping that exploits the multinucleated nature of myofibers to achieve cross-correction of resident, dystrophic nuclei by the U7 small nuclear RNA engineered to skip exon 51 of the dystrophin gene. We observed that co-culture of genetically corrected human DMD myogenic cells (but not of WT cells) with their dystrophic counterparts at a ratio of either 1:10 or 1:30 leads to dystrophin production at a level several folds higher than what predicted by simple dilution. This is due to diffusion of U7 snRNA to neighbouring dystrophic resident nuclei. When transplanted into NSG-mdx-Δ51mice carrying a mutation of exon 51, genetically corrected human myogenic cells produce dystrophin at much higher level than WT cells, well in the therapeutic range, and lead to force recovery even with an engraftment of only 3-5%. This level of dystrophin production is an important step towards clinical efficacy for cell therapy.
Topics: Animals; Humans; Mice; Disease Models, Animal; Dystrophin; Exons; Genetic Vectors; Mice, Inbred mdx; Muscles; Muscular Dystrophy, Duchenne
PubMed: 38438561
DOI: 10.1038/s44321-024-00031-3 -
PLoS Genetics Mar 2024The basement membrane (BM) is an essential structural element of tissues, and its diversification participates in organ morphogenesis. However, the traffic routes...
The basement membrane (BM) is an essential structural element of tissues, and its diversification participates in organ morphogenesis. However, the traffic routes associated with BM formation and the mechanistic modulations explaining its diversification are still poorly understood. Drosophila melanogaster follicular epithelium relies on a BM composed of oriented BM fibrils and a more homogenous matrix. Here, we determined the specific molecular identity and cell exit sites of BM protein secretory routes. First, we found that Rab10 and Rab8 define two parallel routes for BM protein secretion. When both routes were abolished, BM production was fully blocked; however, genetic interactions revealed that these two routes competed. Rab10 promoted lateral and planar-polarized secretion, whereas Rab8 promoted basal secretion, leading to the formation of BM fibrils and homogenous BM, respectively. We also found that the dystrophin-associated protein complex (DAPC) and Rab10 were both present in a planar-polarized tubular compartment containing BM proteins. DAPC was essential for fibril formation and sufficient to reorient secretion towards the Rab10 route. Moreover, we identified a dual function for the exocyst complex in this context. First, the Exo70 subunit directly interacted with dystrophin to limit its planar polarization. Second, the exocyst complex was also required for the Rab8 route. Altogether, these results highlight important mechanistic aspects of BM protein secretion and illustrate how BM diversity can emerge from the spatial control of distinct traffic routes.
Topics: Animals; Basement Membrane; Drosophila melanogaster; Dystrophin; Cytoplasm; Epithelium; GTP Phosphohydrolases; Drosophila Proteins
PubMed: 38437244
DOI: 10.1371/journal.pgen.1011169 -
The Journal of International Medical... Mar 2024Extraocular muscles have complex development processes. The present study aimed to analyze the presence of myosin, dystrophin, and collagen IV in the strabismus-affected... (Observational Study)
Observational Study
OBJECTIVE
Extraocular muscles have complex development processes. The present study aimed to analyze the presence of myosin, dystrophin, and collagen IV in the strabismus-affected extraocular muscle.
METHODS
This research was an observational case-control study. Myosin, dystrophin, and collagen IV were detected by histological and immunohistochemical analyses of extraocular muscle samples from concomitant strabismus patients and controls. A semi-quantitative grading method and statistical analysis were used.
RESULTS
In the strabismus-affected extraocular muscle, morphological analysis demonstrated different-sized muscle fibers. Immature muscle fibers and an increased amount of connective tissue were also noted. Strong positive correlations were identified between myosin and collagen IV and between dystrophin and collagen IV.
CONCLUSIONS
The presence of newly formed muscle fibers, increased connective tissue, and variable diameters of skeletal striated muscle fibers indicate the decreased quality of extraocular muscles in strabismus cases. Reduced levels of myosin and dystrophin and a near absence of collagen IV in strabismus-affected skeletal striated muscle fibers characterized the muscular dystrophy of strabismus. Adjuvant therapy aimed at normalizing the metabolism of these muscles may be appropriate alongside concomitant strabismus treatment.
Topics: Humans; Case-Control Studies; Collagen; Dystrophin; Myosins; Oculomotor Muscles; Strabismus
PubMed: 38436252
DOI: 10.1177/03000605241233521 -
Nature Communications Mar 2024In muscular dystrophies, muscle fibers loose integrity and die, causing significant suffering and premature death. Strikingly, the extraocular muscles (EOMs) are spared,...
In muscular dystrophies, muscle fibers loose integrity and die, causing significant suffering and premature death. Strikingly, the extraocular muscles (EOMs) are spared, functioning well despite the disease progression. Although EOMs have been shown to differ from body musculature, the mechanisms underlying this inherent resistance to muscle dystrophies remain unknown. Here, we demonstrate important differences in gene expression as a response to muscle dystrophies between the EOMs and trunk muscles in zebrafish via transcriptomic profiling. We show that the LIM-protein Fhl2 is increased in response to the knockout of desmin, plectin and obscurin, cytoskeletal proteins whose knockout causes different muscle dystrophies, and contributes to disease protection of the EOMs. Moreover, we show that ectopic expression of fhl2b can partially rescue the muscle phenotype in the zebrafish Duchenne muscular dystrophy model sapje, significantly improving their survival. Therefore, Fhl2 is a protective agent and a candidate target gene for therapy of muscular dystrophies.
Topics: Animals; Cytoskeletal Proteins; Dystrophin; Ectopic Gene Expression; Muscular Dystrophy, Duchenne; Oculomotor Muscles; Zebrafish; Muscle Proteins; LIM Domain Proteins
PubMed: 38431640
DOI: 10.1038/s41467-024-46187-x -
Stem Cell Research Apr 2024Duchenne muscular dystrophy (DMD) is a fatal X-linked recessive disorder, which is caused mostly by frame-disrupting, out-of-frame variation in the dystrophin (DMD)...
Duchenne muscular dystrophy (DMD) is a fatal X-linked recessive disorder, which is caused mostly by frame-disrupting, out-of-frame variation in the dystrophin (DMD) gene. Loss-of- function mutations are the most common type of mutation in DMD, accounting for approximately 60-90% of all DMD variations. In this study, we used adenine base editing to generate a human embryonic stem cell line with splice-site mutations to mimic exon deletion variants in clinical Duchenne muscular dystrophy patients. This cell line has differentiation potential and a normal karyotypic.
Topics: Humans; Dystrophin; Muscular Dystrophy, Duchenne; Gene Editing; CRISPR-Cas Systems; Clustered Regularly Interspaced Short Palindromic Repeats; Human Embryonic Stem Cells; Exons; Cell Line; Mutation
PubMed: 38428348
DOI: 10.1016/j.scr.2024.103343