Journal of Neuromuscular Diseases - Volume 3, issue 2

Authors: Castets, Perrine | Frank, Stephan | Sinnreich, Michael | Rüegg, Markus A.

Article Type: Review Article

Abstract: Recent research has revealed that autophagy, a major catabolic process in cells, is dysregulated in several neuromuscular diseases and contributes to the muscle wasting caused by non-muscle disorders (e.g. cancer cachexia) or during aging (i.e. sarcopenia). From there, the idea arose to interfere with autophagy or manipulate its regulatory signalling to help restore muscle homeostasis and attenuate disease progression. The major difficulty for the development of therapeutic strategies is to restore a balanced autophagic flux, due to the dynamic nature of autophagy. Thus, it is essential to better understand the mechanisms and identify the signalling pathways at play in the …control of autophagy in skeletal muscle. A comprehensive analysis of the autophagic flux and of the causes of its dysregulation is required to assess the pathogenic role of autophagy in diseased muscle. Furthermore, it is essential that experiments distinguish between primary dysregulation of autophagy (prior to disease onset) and impairments as a consequence of the pathology. Of note, in most muscle disorders, autophagy perturbation is not caused by genetic modification of an autophagy-related protein, but rather through indirect alteration of regulatory signalling or lysosomal function. In this review, we will present the mechanisms involved in autophagy, and those ensuring its tight regulation in skeletal muscle. We will then discuss as to how autophagy dysregulation contributes to the pathogenesis of neuromuscular disorders and possible ways to interfere with this process to limit disease progression. Show more

Keywords: Autophagy, skeletal muscle, MAP1LC3, p62/SQSTM1, vacuole, myopathy, dystrophy, mTORC1, FoxO, mitophagy

DOI: 10.3233/JND-160153

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 127-155, 2016

Authors: Goyenvalle, Aurelie | Leumann, Christian | Garcia, Luis

Article Type: Review Article

Abstract: Oligonucleotide therapeutics hold great promise for the treatment of various diseases and the antisense field is constantly gaining interest due to the development of more potent and nuclease resistant chemistries. Despite a rather low success rate with only three antisense drugs being clinically approved, the frontiers of AON therapeutic applications have increased over the past three decades and continue to expand thanks to a steady increase in understanding the mechanisms of action of these molecules, progress in chemical modification and delivery. In this review, we will examine the recent advances obtained with the tricyclo-DNA chemistry which displays unique pharmacological …properties and unprecedented uptake in many tissues after systemic administration. We will review their specific properties and their therapeutic applications mainly for neuromuscular disorders, including exon-skipping for Duchenne muscular dystrophy and exon-inclusion for spinal muscular atrophy, but also aberrant splicing correction for Pompe disease. Finally, we will discuss their advantages and potential limitations, with a focus on the need for careful toxicological screen early in the process of AON drug development. Show more

Keywords: Antisense oligonucleotides, splicing correction, exon-skipping, exon-reinclusion, nanoparticles, delivery

DOI: 10.3233/JND-160146

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 157-167, 2016

Authors: Buchkremer, Stephan | González Coraspe, José Andrés | Weis, Joachim | Roos, Andreas

Article Type: Research Article

Abstract: Chaperone dysfunction leading to the build-up of misfolded proteins could frequently be linked to clinical manifestations also affecting the nervous system and the skeletal muscle. In addition, increase in chaperone function is beneficial to antagonize protein aggregation and thus represents a promising target for therapeutic concepts for many genetic and acquired chaperonopathies. However, little is known on the precise molecular mechanisms defining the cell and tissue abnormalities in the case of impaired chaperone function as well as on underlying effects in the case of compensatory up-regulation of chaperones. This scarcity of knowledge often arises from a lack of appropriate animal …models that mimic closely the human molecular, cellular, and histological characteristics. Here, we introduce the Sil1 -mutant woozy mouse as a suitable model to investigate molecular and cellular mechanisms of impaired ER-chaperone function affecting the integrity of nervous system and skeletal muscle. The overlapping clinical findings in man and mouse indicate that woozy is a good copy of a human phenotype called Marinesco-Sjögren syndrome. We confirm the presence of ER-stress and expand the biochemical knowledge of altered nuclear envelope in muscle, a hallmark of SIL1-disease. In addition, our data suggest that impaired excitation-contraction coupling might be part of the SIL1-pathophysiology. Our results moreover indicate that divergent expression of pro- and anti-survival proteins is decisive for Purkinje cell survival. By summarizing the current knowledge of woozy, we focus on the suitability of this animal model to study neuroprotective co-chaperone function and to investigate the involvement of co-chaperones in the predisposition of other disorders such as diabetic neuropathy. Show more

Keywords: BiP, cerebellar ataxia, chaperone, vacuolar myopathy, woozy mouse

DOI: 10.3233/JND-160152

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 169-181, 2016

Authors: Bouhy, Delphine | Geuens, Thomas | De Winter, Vicky | Almeida-Souza, Leonardo | Katona, Istvan | Weis, Joachim | Hochepied, Tino | Goossens, Steven | Haigh, Jody J. | Janssens, Sophie | Timmerman, Vincent

Article Type: Research Article

Abstract: Background: Charcot-Marie-Tooth (CMT) and associated neuropathies, the most common inherited diseases of the peripheral nervous system, remain so far incurable. Three existing murine models of Charcot-Marie-Tooth type 2F (CMT2F) and/or distal hereditary motor neuropathy type IIb (dHMNIIb), caused by mutations in the small heat shock protein B1 gene (HSPB1 /HSP27 ), partially recapitulate the hallmarks of peripheral neuropathy. Because these models overexpress the HSPB1 mutant proteins they differ from the patients’ situation. Objective: To overcome the possible bias induced by overexpression, we generated and characterized a transgenic model in which the wild type or mutant HSPB1 protein …was expressed at a moderate, more physiologically relevant level. Methods: We generated a new transgenic mouse model in which a human wild type (hHSPB1WT ) or mutant (hHSPB1R127W ; hHSPB1P182L ) HSPB1 transgene was integrated in the mouse ROSA26 locus. The motor and sensory functions of the mice was assessed at 3, 6, 9, 12 and 18 month. Results: However, the mice expressing the mutant hHSPB1 do not develop motor or sensory deficits and do not show any sign of axonal degeneration, even at late age. Quantitative PCR analyses reveal contrasting tissue-specific expression pattern for the endogenous mouse and exogenous human HSPB1 and show that the ratio of human HSPB1 to the endogenous mouse HspB1 is lower in the sciatic nerve and spinal cord compared to the brain. Conclusion: These results suggest that expressing the transgene at a physiological level using the ROSA26 locus may not be sufficient to model inherited peripheral neuropathies caused by mutation in HSPB1. Show more

Keywords: Peripheral nervous system disease, HSPB1, transgenic mice, ROSA26 locus

DOI: 10.3233/JND-150144

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 183-200, 2016

Authors: Ito, N. | Shimizu, N. | Tanaka, H. | Takeda, S.

Article Type: Research Article

Abstract: Background and Objectives: Cell transplantation is a promising therapy for several muscle diseases, including Duchenne muscular dystrophy. Satellite cells are stem cells in skeletal muscle that provide an important cell source for transplantation therapy. However, culture of satellite cells in vitro causes them to lose their undifferentiated state, associated with reduced transplantation efficiency. It is therefore necessary to develop optimal culture conditions for maintaining the undifferentiated state of satellite cells. Methods: Primary satellite cells were cultured with or without leukemia inhibitory factor (LIF). The expression of undifferentiation and differentiation markers, and the transplantation efficiency were analyzed. …Results: LIF-treated satellite cells showed increased expression of Pax7, and enhanced transplantation efficiency in a mouse model of Duchenne muscular dystrophy. Conclusions: Our study showed that the treatment with LIF effectively maintained the undifferentiated state of satellite cells, and enhanced their transplantation efficiency. These results will contribute to the optimization of culture conditions for cell transplantation therapy. Show more

Keywords: Satellite cell, leukemia inhibitory factor, muscular dystrophy, cell transplantation therapy

DOI: 10.3233/JND-160156

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 201-207, 2016

Authors: Punetha, Jaya | Kesari, Akanchha | Uapinyoying, Prech | Giri, Mamta | Clarke, Nigel F. | Waddell, Leigh B. | North, Kathryn N. | Ghaoui, Roula | O’Grady, Gina L. | Oates, Emily C. | Sandaradura, Sarah A. | Bönnemann, Carsten G. | Donkervoort, Sandra | Plotz, Paul H. | Smith, Edward C. | Tesi-Rocha, Carolina | Bertorini, Tulio E. | Tarnopolsky, Mark A. | Reitter, Bernd | Hausmanowa-Petrusewicz, Irena | Hoffman, Eric P.

Article Type: Research Article

Abstract: Background: Molecular diagnostics in the genetic myopathies often requires testing of the largest and most complex transcript units in the human genome (DMD, TTN , NEB ). Iteratively targeting single genes for sequencing has traditionally entailed high costs and long turnaround times. Exome sequencing has begun to supplant single targeted genes, but there are concerns regarding coverage and needed depth of the very large and complex genes that frequently cause myopathies. Objective: To evaluate efficiency of next-generation sequencing technologies to provide molecular diagnostics for patients with previously undiagnosed myopathies. Methods: We tested a targeted re-sequencing …approach, using a 45 gene emulsion PCR myopathy panel, with subsequent sequencing on the Illumina platform in 94 undiagnosed patients. We compared the targeted re-sequencing approach to exome sequencing for 10 of these patients studied. Results: We detected likely pathogenic mutations in 33 out of 94 patients with a molecular diagnostic rate of approximately 35%. The remaining patients showed variants of unknown significance (35/94 patients) or no mutations detected in the 45 genes tested (26/94 patients). Mutation detection rates for targeted re-sequencing vs. whole exome were similar in both methods; however exome sequencing showed better distribution of reads and fewer exon dropouts. Conclusions: Given that costs of highly parallel re-sequencing and whole exome sequencing are similar, and that exome sequencing now takes considerably less laboratory processing time than targeted re-sequencing, we recommend exome sequencing as the standard approach for molecular diagnostics of myopathies. Show more

Keywords: Muscular dystrophy, myopathies, molecular diagnostic testing, high-throughput DNA sequencing, massively-parallel sequencing, Exome, molecular diagnostics, genetic screening

DOI: 10.3233/JND-160151

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 209-225, 2016

Authors: Marey, Isabelle | Ben Yaou, Rabah | Deburgrave, Nathalie | Vasson, Aurélie | Nectoux, Juliette | Leturcq, France | Eymard, Bruno | Laforet, Pascal | Behin, Anthony | Stojkovic, Tanya | Mayer, Michèle | Tiffreau, Vincent | Desguerre, Isabelle | Boyer, François Constant | Nadaj-Pakleza, Aleksandra | Ferrer, Xavier | Wahbi, Karim | Becane, Henri-Marc | Claustres, Mireille | Chelly, Jamel | Cossee, Mireille

Article Type: Research Article

Abstract: Background: Dystrophinopathies are mostly caused by copy number variations, especially deletions, in the dystrophin gene (DMD ). Despite the large size of the gene, deletions do not occur randomly but mainly in two hot spots, the main one involving exons 45 to 55. The underlying mechanisms are complex and implicate two main mechanisms: Non-homologous end joining (NHEJ) and micro-homology mediated replication-dependent recombination (MMRDR). Objective: Our goals were to assess the distribution of intronic breakpoints (BPs) in the genomic sequence of the main hot spot of deletions within DMD gene and to search for specific sequences at …or near to BPs that might promote BP occurrence or be associated with DNA break repair. Methods: Using comparative genomic hybridization microarray, 57 deletions within the intron 44 to 55 region were mapped. Moreover, 21 junction fragments were sequenced to search for specific sequences. Results: Non-randomly distributed BPs were found in introns 44, 47, 48, 49 and 53 and 50% of BPs clustered within genomic regions of less than 700bp. Repeated elements (REs), known to promote gene rearrangement via several mechanisms, were present in the vicinity of 90% of clustered BPs and less frequently (72%) close to scattered BPs, illustrating the important role of such elements in the occurrence of DMD deletions. Palindromic and TTTAAA sequences, which also promote DNA instability, were identified at fragment junctions in 20% and 5% of cases, respectively. Micro-homologies (76%) and insertions or deletions of small sequences were frequently found at BP junctions. Conclusions: Our results illustrate, in a large series of patients, the important role of RE and other genomic features in DNA breaks, and the involvement of different mechanisms in DMD gene deletions: Mainly replication error repair mechanisms, but also NHEJ and potentially aberrant firing of replication origins. A combination of these mechanisms may also be possible. Show more

Keywords: DMD gene, deletion breakpoints, double strand break repair, replication error repair mechanisms

DOI: 10.3233/JND-150134

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 227-245, 2016

Authors: Crowe, Kelly E. | Shao, Guohong | Flanigan, Kevin M. | Martin, Paul T.

Article Type: Research Article

Abstract: Background: Duchenne Muscular Dystrophy (DMD) is a severe, progressive, neuromuscular disorder of childhood. While a number of serum factors have been identified as potential biomarkers of DMD, none, as yet, are proteins within the dystrophin-associated glycoprotein (DAG) complex. Objective: We have developed an immobilized serum ELISA assay to measure the expression of a constitutively cleaved and secreted component of the DAG complex, the N-terminal domain of α dystroglycan (αDG-N), and assayed relative expression in serum from muscular dystrophy patients and normal controls. Methods: ELISAs of immobilized patient or mouse serum and Western blots were used …to assess αDG-N expression. Results: Immobilization of diluted serum on ELISA plates was important for this assay, as methods to measure serum αDG-N in solution were less robust. αDG-N ELISA signals were significantly reduced in DMD serum (27±3% decrease, n  = 9, p  < 0.001) relative to serum from otherwise normal controls (n  = 38), and calculated serum αDG-N concentrations were reduced in DMD relative to normal (p  < 0.01) and Becker Muscular Dystrophy (n  = 11, p  < 0.05) patient serum. By contrast, ELISA signals from patients with Inclusion Body Myositis were not different than normal (4±3% decrease, n  = 8, p  = 0.99). αDG-N serum signals were also significantly reduced in utrophin-deficient mdx mice as compared to mdx and wild type mice. Conclusions: Our results are the first demonstration of a component of the DAG complex as a potential serum biomarker in DMD. Such a serum measure could be further developed as a tool to help reflect overall muscle DAG complex expression or stability. Show more

Keywords: Dystroglycan, muscular dystrophy, biomarker, dystrophin, utrophin

DOI: 10.3233/JND-150127

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 247-260, 2016

Authors: Ramos, Edwardo | Conde, José G. | Berrios, Rafael Arias | Pardo, Sherly | Gómez, Omar | Mas Rodríguez, Manuel F.

Article Type: Research Article

Abstract: Background: Duchenne and Becker Muscular Dystrophy (DMD and BMD, respectively), are common forms of inherited muscle disease. Information regarding the epidemiology of these conditions, including genotype, is still sparse. Objective: To establish the prevalence and genetic profile of DMD and BMD in Puerto Rico. Methods: We collected data from medical records in all Muscular Dystrophy Association (MDA) clinics in Puerto Rico in order to estimate the prevalence of DMD and BMD and to describe the genotypic profile of these patients. Patients selected for data analysis matched “definite”, “probable” and “possible” case definitions as established by …MD STARnet. Results: A total of 141 patients matched the inclusion criteria, with 64.5% and 35.5% being categorized into DMD and BMD, respectively. DMD and BMD prevalence in Puerto Rico was estimated at 5.18 and 2.84 per 100,000 males, respectively. Deletion was the most common form of mutation (66.7%) in the dystrophin gene, with exons in segment 45 to 47 being the most frequently affected. Conclusions: This is the first report of the prevalence and genetic profile characteristics of DMD and BMD in Puerto Rico. Prevalence of DMD was similar to that reported worldwide, while prevalence of BMD was higher. Genetic profile was consistent with that reported in the literature. Show more

Keywords: Duchenne muscular dystrophy, Becker muscular dystrophy, dystrophinopathy, muscular dystrophy, Puerto Rico

DOI: 10.3233/JND-160147

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 261-266, 2016

Authors: Cruz, Simao | Figueroa-Bonaparte, Sebastian | Gallardo, Eduard | de Becdelièvre, Alix | Gartioux, Corine | Allamand, Valérie | Piñol, Patricia | Garcia, Maria Angels Rodriguez | Jiménez-Mallebriera, Cecilia | Llauger, Jaume | González-Rodríguez, Lidia | Cortes-Vicente, Elena | Illa, Isabel | Díaz-Manera, Jordi

Article Type: Research Article

Abstract: The classical phenotypes of collagen VI-associated myopathies are well described. Little is known, however, about the progression of patients at the mildest end of the clinical spectrum. In this report, we describe the clinical findings and the results of MRI, muscle biopsy, collagen VI expression in cultured skin fibroblasts and genetic tests of a series of patients with Bethlem myopathy. Our series highlights the existence of mild presentations of this disorder that progresses only slightly and can easily be overlooked. Analysis of the genetic studies suggests that missense mutations can be associated to a milder clinical presentation. Muscle MRI is …extremely useful as it shows a pathognomonic pattern in most patients, especially those with some degree of muscle weakness. Show more

Keywords: Bethlem disease, collagen VI deficiency, muscle dystrophy, muscle MRI, Ulrich disease

DOI: 10.3233/JND-150135

Citation: Journal of Neuromuscular Diseases, vol. 3, no. 2, pp. 267-274, 2016