Degenerative Muscle Disorders Research Articles

Activation of non-myogenic mesenchymal stem cells during the disease progression in dystrophic dystrophin/utrophin knockout mice.

Ectopic calcification as well as fatty and fibrotic tissue accumulation occurs in skeletal muscle during the disease progression of Duchenne muscular dystrophy (DMD), a degenerative muscle disorder caused by mutations in the dystrophin gene. The cellular origin and the environmental cues responsible for this ectopic calcification, fatty and fibrotic infiltration during the disease progression, however, remain unknown. Based on a previously published preplate technique, we isolated two distinct populations of muscle-derived cells from skeletal muscle: (i) a rapidly adhering cell population, which is non-myogenic, Pax7(-) and express the mesenchymal stem cell (MSC) marker platelet-derived growth factor receptor alpha; hence, we termed this population of cells non-myogenic MSCs (nmMSCs); and (ii) a slowly adhering cell population which is Pax7(+) and highly myogenic, termed muscle progenitor cells (MPCs). Previously, we demonstrated that the rapid progression of skeletal muscle histopathologies in dystrophin/utrophin knockout (dys(-/-) utro(-/-) dKO) mice is closely associated with a rapid depletion of the MPC population pool. In the current study, we showed that in contrast to the MPCs, the nmMSCs become activated during the disease progression in dKO mice, displaying increased proliferation and differentiation potentials (adipogenesis, osteogenesis and fibrogenesis). We also found that after co-culturing the dKO-nmMSCs with dKO-MPCs, the myogenic differentiation potential of the dKO-MPCs was reduced. This effect was found to be potentially mediated by the secretion of secreted frizzled-related protein 1 by the dKO-nmMSCs. We therefore posit that the rapid occurrence of fibrosis, ectopic calcification and fat accumulation, in dKO mice, is not only attributable to the rapid depletion of the MPC pool, but is also the consequence of nmMSC activation. Results from this study suggest that approaches to alleviate muscle weakness and wasting in DMD patients should not only target the myogenic MPCs but should also attempt to prevent the activation of the nmMSCs.

TNF-α and IGF1 modify the microRNA signature in skeletal muscle cell differentiation.

BackgroundElevated levels of the inflammatory cytokine TNF-α are common in chronic diseases or inherited or degenerative muscle disorders and can lead to muscle wasting. By contrast, IGF1 has a growth promoting effect on skeletal muscle. The molecular mechanisms mediating the effect of TNF-α and IGF1 on muscle cell differentiation are not completely understood. Muscle cell proliferation and differentiation are regulated by microRNAs (miRNAs) which play a dominant role in this process. This study aims at elucidating how TNF-α or IGF1 regulate microRNA expression to affect myoblast differentiation and myotube formation.ResultsIn this study, we analyzed the impact of TNF-α or IGF1 treatment on miRNA expression in myogenic cells. Results reveal that i) TNF-α and IGF1 regulate miRNA expression during skeletal muscle cell differentiation in vitro, ii) microRNA targets can mediate the negative effect of TNF-α on fusion capacity of skeletal myoblasts by targeting genes associated with axon guidance, MAPK signalling, focal adhesion, and neurotrophin signalling pathway, iii) inhibition of miR-155 in combination with overexpression of miR-503 partially abrogates the inhibitory effect of TNF-α on myotube formation, and iv) MAPK/ERK inhibition might participate in modulating the effect of TNF-α and IGF1 on miRNA abundance.ConclusionsThe inhibitory effects of TNF-α or the growth promoting effects of IGF1 on skeletal muscle differentiation include the deregulation of known muscle-regulatory miRNAs as well as miRNAs which have not yet been associated with skeletal muscle differentiation or response to TNF-α or IGF1. This study indicates that miRNAs are mediators of the inhibitory effect of TNF-α on myoblast differentiation. We show that intervention at the miRNA level can ameliorate the negative effect of TNF-α by promoting myoblast differentiation. Moreover, we cautiously suggest that TNF-α or IGF1 modulate the miRNA biogenesis of some miRNAs via MAPK/ERK signalling. Finally, this study identifies indicative biomarkers of myoblast differentiation and cytokine influence and points to novel RNA targets.Electronic supplementary materialThe online version of this article (doi:10.1186/s12964-015-0083-0) contains supplementary material, which is available to authorized users.

G.P.177: Constipation in Duchenne muscular dystrophy: Common, underdiagnosed and undertreated

Duchenne muscular dystrophy (DMD) is an X-linked progressive degenerative muscle disorder. Constipation as a co-morbidity in DMD has not been subject to a systematic study. To determine the prevalence of constipation, identify its predictive factors and evaluate different screening methods in a cohort of DMD patients. Design/Methods: This was a prospective cross-sectional study that evaluated a cohort of DMD patients for constipation using the pediatric Rome-III questionnaires (QPGS-RIII) and the Bristol stool chart. Additional clinical data collected included functional assessments and medications. Fecal load was assessed on routine spine X-rays. Logistic regression with backward elimination as a variable selection procedure was used to determine possible predictors of constipation. Among 98 male DMD patients (average age 11.3±3.8years), the overall prevalence of constipation fulfilling Rome-III criteria was 44.9%, of which 45% received symptomatic treatment and 11% were asymptomatic. The age distribution of patients with constipation (<i>n</i>=44) was similar to that without (10.8±2.6 vs 11.1±3years, <i>p</i>=0.62). Prevalence of constipation did not correlate with functional status (Northstar ambulatory assessment scale, <i>p</i>=0.39), treatment with calcium supplements (<i>p</i>=0.92) or with stool load on abdominal X-ray (<i>p</i>=0.82). The regression model failed to identify any predictive variables for constipation. The Bristol stool chart, abnormal abdominal exam (fullness, distension, tenderness, palpable masses or abnormal bowel sounds) and fecal load on X-ray were inferior to the Rome-III criteria in diagnosing constipation. (1) Constipation is a common co-morbidity in DMD, irrespective of age or the stage of disease. (2) Of the tests evaluated, the Rome III criteria may be the most appropriate screening tool for constipation in DMD patients. (3) Constipation in this cohort was underdiagnosed and either not treated or treated inadequately.

“Cute Girl in Wheelchair—Why?”: Cripping YouTube

“Cute Girl in Wheelchair—Why?”Cripping YouTube Rachel Reinke (bio) and Anastasia Todd (bio) Hi I’m Kaley! I am trans and proud! I go to Salisbury University to study Social Work, I’m Hispanic, and I have a muscle degenerative disorder called Duchenne Muscular Dystrophy. In November of 2013 I opened up about being a trans girl! Coming out has been a tough emotional roller coaster and I will do my best to overcome those difficulties. I will give it all I’ve got! I made this channel for others that are going through what I’m going through and show them they are not alone. The show is about me trying to become who I am. We will get to see my progress to become Kaley! I hope you can join me in my awesome journey! -Kaley Kutie’s YouTube “About” Page Channels like that of Kaley Vides—better known by her channel title, “Kaley Kutie”—are prime examples of young people using YouTube to (re)create the meanings of commonsense notions—of youth, of gender, and of disability—out of their everyday lived experience. Kaley’s channel includes both informative and “silly” videos, ranging from 6 to 36 minutes long, forming a collective archive that often directly addresses her transgender and disabled subjectivities. In addition to documenting the “journey” of Kaley’s “everyday life,” some of her most-watched videos include question and answer sessions about being transgender and about her particular disability, and among the most popular, “Tip &Tricks: Makeup Tips for Physical Disabled Women.”1 Putting these lived experiences at the forefront of a Disability Studies curriculum can make the classroom an interactive space for action, reaction, and for deeper affective understanding of disabled subjectivities. Kaley, who identifies as transgender and disabled, provides an interesting example of how YouTube vloggers can affectively disorient space online in order to create a physical presence of those who are excluded from normative understandings of “being.” As Kate O’Riordan and David Phillips have noted, the Internet allows us to investigate “the cultural and political economies of places, and the reflexive construction of places and identities” (4). Under close analysis, Kaley Kutie’s YouTube channel allows students to do the work that O’Riordan and Phillips describe. Kaley Kutie’s YouTube channel, and others like it, can be understood as “forms of cultural pedagogy that confront, challenge and redefine knowledge and practice, and mediate altered sensibilities” (Hickey-Moody and Crowley 400).2 [End Page 168] Facilitating such interactions is crucial in classrooms where disability or transgender subject positions often figure as diversity commodities rather than complex identities—if they appear at all. Incorporating YouTube content from Kaley Kutie and other disabled and trans* vloggers, and allowing meaningful interaction to emerge from students’ affective responses to that content, shifts disabled content so that it is intrinsically valued, rather than marginalized as “an instrumental tool for diagnosis and therapy” (Adkins et al. qtd. in Alper 3). Understanding this content from an intersectional perspective also allows that intrinsic value provokes students to see that multiple subject positions affect Kaley’s experiences of the world—and her desire to share them with us. Cripping the Classroom Teaching Disability Studies in the neoliberal academy means recognizing that, as Merri Lisa Johnson and Robert McRuer argue, disability identity has become “part of capitalism’s array of target markets,” where certain forms of knowledge about disability are valued more than others (128). To crip is to uncover and explore the nebulous infrastructure of oppressive systems. Cripping the classroom enables those who teach Disability Studies to participate in a collective troubling, rupturing, and questioning of oppressive knowledges and structures. Pedagogically, this requires an intersectional approach to knowledge creation within the classroom, as well as paying attention to disabled people whose claims to self-representation and whose positions as knowledge-creators have been marginalized. Cripping the classroom questions what kinds of knowledge are valued. In Anastasia’s experience teaching Disability Studies, affect and affective ways of knowing have been a productive tool for cripping the classroom and for “question[ing] what we think we know about disability” (Johnson and McRuer 130). Attention to affect “creates space for embodied...

Muscle‐derived cells (MDCs) responsible for myogenesis differ from MDCs involved in adipogenesis in dystrophin/utrophin−/− mice

Ectopic fat formation/accumulation and infiltration in skeletal muscle is closely associated with several disorders including Duchenne muscular dystrophy (DMD), a degenerative muscle disorder. However, neither the origin of ectopic adipocytes, nor the mechanism of their formation in disease, is known. Based on a previously published preplate technique, we isolated two types of muscle derived cells; rapidly adhering cells (RACs) and slowly adhering cells (SACs) from skeletal muscle of C57BL (wt) and utrophin/dystrophin double knockout (dys−/−utro−/−, dKO) mice, which emulates the phenotype seen in DMD patients. Previously, we have shown that SACs are myogenic progenitor cells and are mainly involved in muscle fiber regeneration or degeneration. Here, we identify RACs as PDGFRá + multipotent, mesenchymal progenitor cells and observe that dKO‐RACs display increased adipogenic, osteogenic, and chondrogenic potentials compared to wt‐RACs. We also show that, compared to other muscle‐derived cell populations from dKO mice, RACs display significantly increased proliferation and adipogenic potentials both in vitro and in vivo. This study suggests that RACs may be the major contributor to ectopic fat cell formation within skeletal muscle in pathological conditions such as DMD. Therefore, targeting RACs to block adipogenesis in skeletal muscle may open new opportunities to treat muscle diseases.Grant Funding Source: Department of Defense (DOD) – Army

Yoga and Neurobehavioral Sciences: Indian Perspective

The issue carries a series of research reports by the team from Advanced Center of Yoga in collaboration with Morarji Desai National Institute of Yoga. I wish to congratulate the team for making this possible. I also thank the editors, Dr. M. S. Keshavan, Dr. T. S. S. Rao and Dr. Naren for their effort in bringing out this supplement. National Institute of Mental Health and Neuro Sciences (NIMHANS) has identified yoga as an important area for service as well as research and the word yoga itself figures in the logo of NIMHANS. This is researched in both clinical and preclinical departments over the four decades or longer. The advanced center was set up with a memorandum of understanding and collaboration with the “Morarji Desai National Institute of Yoga” in 2007. This led to opening of a regular yoga service for the patients. Every day nearly 40 patients and some of their caregivers receive yoga intervention in this center. Yoga is also given as an add-on intervention in the children's ward. The team has trained the nurses in other wards too to provide yoga-based physical activity. In the past 5 years, the team conducted several seminars and symposia to popularize yoga in relation to mental health and neurosciences. The populations included doctors, teachers, special groups like the elderly and caregivers. The group also released a booklet on yoga in depression. Interdisciplinary research was undertaken as part of this collaboration in important areas. Students were guided in these researches to complete their PhD and MD degrees successfully. Some of these research papers are being published as a compilation in this special issue of the Indian Journal of Psychiatry. Yoga approaches used in these studies include postures (Asanas), breathing techniques (Pranayamas) and chanting (OM). These procedures were chosen from traditional texts and the modules were validated by experts in the field of Yoga. Such validated modules were used in these studies. The studies demonstrate the efficacy of these modules in select conditions, but with some limitations. That many of the measurements are objective suggests that there was no bias in these results. There is also evidence that the yoga intervention produced predictable biological changes. These biological changes too were relevant to the therapeutic effects. Yoga also was found beneficial in certain divergent groups, caregivers and children and acutely ill in-patients. Persons with difficult-to-treat epilepsy attending out-patient care too expressed a need for yoga as an add-on intervention. This is only a modest attempt to project evidences in favor of yoga. I am also aware that some studies are not published here as they are yet incomplete, have not shown such positive results in the preliminary analyses. More careful documentation/verification of yoga's adverse effects, if any, deserves to be brought to the notice of clinicians. The team should also extend its work to include other clinical populations in whom the allopathic interventions have limitations. Examples include dementias, degenerative muscle disorders, acquired brain damage-related deficits, childhood/developmental disorders and substance use disorders. There is a need to develop strategies that help to overcome barriers in reaching yoga to the patients living in the community. Demonstrating the effectiveness of such strategies will go a long way in introducing comprehensive/integrated approaches for the care of chronically ill. There could also be potential to include these in public health programs. Given the presence at NIMHANS of advanced and cutting edge research facilities in basic science, the team must explore the research with reference to the basic mechanisms of yoga as a composite module as well as its independent components. Demonstration of such biological evidence in correcting the underlying disease by yoga helps to motivate modern practitioners to recommend yoga as a treatment in certain conditions. The quantum of evidence in favor of yoga could well prompt in the days to come inclusion of a yoga specialist in the mental health Team. I wish the yoga research team all success in meeting these objectives in the near future. I also wish all the readers a productive reading and I hope they will be the individuals who will speak for yoga in the modern medicine clinics.

Sarco (Endo) Plasmic Reticulum Calcium Atpases (SERCA) Isoforms in the Normal and Diseased Cardiac, Vascular and Skeletal Muscle

Deregulated or enhanced calcium ion (Ca2+) influx across an unstable sarcolemma has been proposed to directly affect cardiac hypertrophic remodelling, vascular proliferative diseases and degenerative muscle disorders. Aberrant intracellular handling is partly due to a defect in Sarcoplasmic Reticulum (SR) function. Decreased Ca2+ uptake in cardiac, vascular and skeletal myocytes is associated with a decrease in the expression and activity of the fast sarco/endoplasmic reticulum Ca2+ ATPase (SERCA2a or SERCA1a isoforms). SERCA2a gene transfer was successfully used in heart failure; this approach holds further therapeutic promises in vascular proliferative diseases and dystrophin-deficient muscular diseases. The growing family of human SERCA isoforms comprises at least 14 mRNA and proteins with different functional characteristics and cell-specific expression. This review focuses on the biological role and therapeutic potential of different isoforms of SERCA in the physiology and pathology of cardiac, vascular and skeletal muscle cells.

Subcellular Localization of Dystrophin Isoforms in Cardiomyocytes and Phenotypic Analysis of Dystrophin-deficient Mice Reveal Cardiac Myopathy is Predominantly Caused by a Deficiency in Full-length Dystrophin

Duchenne muscular dystrophy (DMD) is an X-linked recessive progressive muscle degenerative disorder that causes dilated cardiomyopathy in the second decade of life in affected males. Dystrophin, the gene responsible for DMD, encodes full-length dystrophin and various short dystrophin isoforms. In the mouse heart, full-length dystrophin Dp427 and a short dystrophin isoform, Dp71, are expressed. In this study, we intended to clarify the functions of these dystrophin isoforms in DMD-related cardiomyopathy. We used two strains of mice: mdx mice, in which Dp427 was absent but Dp71 was present, and DMD-null mice, in which both were absent. By immunohistochemical staining and density-gradient centrifugation, we found that Dp427 was located in the cardiac sarcolemma and also at the T-tubules, whereas Dp71 was specifically located at the T-tubules. In order to determine whether T tubule-associated Dp71 was involved in DMD-related cardiac disruption, we compared the cardiac phenotypes between DMD-null mice and mdx mice. Both DMD-null mice and mdx mice exhibited severe necrosis, which was followed by fibrosis in cardiac muscle. However, we could not detect a significant difference in myocardial fibrosis between mdx mice and DMD-null mice. Based on the present results, we have shown that cardiac myopathy is caused predominantly by a deficiency of full-length dystrophin Dp427.

Development of Multiexon Skipping Antisense Oligonucleotide Therapy for Duchenne Muscular Dystrophy

Duchenne muscular dystrophy (DMD) is an incurable, X-linked progressive muscle degenerative disorder that results from the absence of dystrophin protein and leads to premature death in affected individuals due to respiratory and/or cardiac failure typically by age of 30. Very recently the exciting prospect of an effective oligonucleotide therapy has emerged which restores dystrophin protein expression to affected tissues in DMD patients with highly promising data from a series of clinical trials. This therapeutic approach is highly mutation specific and thus is personalised. Therefore DMD has emerged as a model genetic disorder for understanding and overcoming of the challenges of developing personalised genetic medicines. One of the greatest weaknesses of the current oligonucleotide approach is that it is a mutation-specific therapy. To address this limitation, we have recently demonstrated that exons 45–55 skipping therapy has the potential to treat clusters of mutations that cause DMD, which could significantly reduce the number of compounds that would need to be developed in order to successfully treat all DMD patients. Here we discuss and review the latest preclinical work in this area as well as a variety of accompanying issues, including efficacy and potential toxicity of antisense oligonucleotides, prior to human clinical trials.

Loss of cIAP1 attenuates soleus muscle pathology and improves diaphragm function in mdx mice

The cellular inhibitor of apoptosis 1 (cIAP1) protein is an essential regulator of canonical and noncanonical nuclear factor κB (NF-κB) signaling pathways. NF-κB signaling is known to play important roles in myogenesis and degenerative muscle disorders such as Duchenne muscular dystrophy (DMD), but the involvement of cIAP1 in muscle disease has not been studied directly. Here, we asked whether the loss of cIAP1 would influence the pathology of skeletal muscle in the mdx mouse model of DMD. Double-mutant cIAP1(-/-);mdx mice exhibited reduced muscle damage and decreased fiber centronucleation in the soleus, compared with single-mutant cIAP1(+/+);mdx mice. This improvement in pathology was associated with a reduction in muscle infiltration by macrophages and diminished expression of inflammatory cytokines such as IL-6 and tumor necrosis factor-α. Furthermore, the cIAP1(-/-);mdx mice exhibited reduced serum creatine kinase, and improved exercise endurance associated with improved exercise resilience by the diaphragm. Mechanistically, the loss of cIAP1 was sufficient to drive constitutive activation of the noncanonical NF-κB pathway, which led to increased myoblast fusion in vitro and in vivo. Collectively, these results show that the loss of cIAP1 protects skeletal muscle from the degenerative pathology resulting from systemic loss of dystrophin.

Dystrophin and utrophin “double knockout” dystrophic mice exhibit a spectrum of degenerative musculoskeletal abnormalities

Duchenne muscular dystrophy (DMD) is a degenerative muscle disorder characterized by the lack of dystrophin expression at the sarcolemma of muscle fibers. In addition, DMD patients acquire osteopenia, fragility fractures, and scoliosis indicating that a deficiency in skeletal homeostasis coexists but little is known about the effects of DMD on bone and other connective tissues within the musculoskeletal system. Recent evidence has emerged implicating adult stem cell dysfunction in DMD myopathogenesis. Given the common mesenchymal origin of muscle and bone, we sought to investigate bone and other musculoskeletal tissues in a DMD mouse model. Here, we report that dystrophin-utrophin double knockout (dko) mice exhibit a spectrum of degenerative changes, outside skeletal muscle, in bone, articular cartilage, and intervertebral discs, in addition to reduced lifespan, muscle degeneration, spinal deformity, and cardiomyopathy previously reported. We also report these mice to have a reduced capacity for bone healing and exhibit spontaneous heterotopic ossification in the hind limb muscles. Therefore, we propose the dko mouse as a model for premature musculoskeletal aging and posit that a similar phenomenon may occur in patients with DMD.

Dystrophin-compromised sarcoglycan-δ knockout diaphragm requires full wild type embryonic stem cell reconstitution for correction

Limb-girdle muscular dystrophy-2F (LGMD-2F) is an incurable degenerative muscle disorder caused by a mutation in the sarcoglycan-δ (SGδ)-encoding gene (SGCD in humans). The lack of SGδ results in the complete disruption of the sarcoglycan complex (SGC) in the skeletal and cardiac muscle within the larger dystrophin-glycoprotein complex (DGC). The long-term consequences of SG ablation on other members of the DGC are currently unknown. We produced mosaic mice through the injection of wild-type (WT) embryonic stem cells (ESCs) into SGδ-knockout (KO) blastocysts. ESC-derived SGδ was supplied to the sarcolemma of 18-month-old chimeric muscle, which resulted in the restoration of the SGC. Despite SGC rescue, and contrary to previous observations obtained with WT/mdx chimeras (a mouse rescue paradigm for Duchenne muscular dystrophy), low levels of ESC incorporation were insufficient to produce histological corrections in SGδ-KO skeletal muscle or heart. The inefficient process of ESC rescue was more evident in the SGδ-KO diaphragm, which had reduced levels of dystrophin and no compensatory utrophin, and needed almost full WT ESC reconstitution for histological improvement. The results suggest that the SGδ-KO mouse model of LGMD is not amenable to ESC treatment.

Myogenic Reprogramming of Bone Marrow Derived Cells in a W41Dmdmdx Deficient Mouse Model

Lack of expression of dystrophin leads to degeneration of muscle fibers and infiltration of connective and adipose tissue. Cell transplantation therapy has been proposed as a treatment for intractable muscle degenerative disorders. Several reports have demonstrated the ability of bone-marrow derived cells (BMDC) to contribute to non-haematopoietic tissues including epithelium, heart, liver, skeletal muscle and brain following transplantation by means of fusion and reprogramming. A key issue is the extent to which fusion and reprogramming can occur in vivo, particularly under conditions of myogenic deterioration.To investigate the therapeutic potential of bone marrow transplantation in monogenetic myopathy, green fluorescent protein-positive (GFP+) bone marrow cells were transplanted into non-irradiated c-kit receptor – deficient (W41) mdx mice. This model allows BMDC reconstitution in the absence of irradiation induced myeloablation. We provide the first report of BMDC fusion in a W41/Dmdmdx deficient mouse model.In the absence of irradiation induced injury, few GFP+ cardiomyocytes and muscle fibres were detected 24 weeks post BMT. It was expected that the frequency of fusion in the hearts of W41Dmdmdx mice would be similar to frequencies observed in infarcted mice [1].Although, it is clear from this study that individual cardiomyocytes with monogenetic deficiencies can be rescued by fusion, it is as clear that in the absence of irradiation, the formation of stable and reprogrammed fusion hybrids occurs, with the current techniques, at very low levels in non-irradiated recipients.

Long‐Term Repeated Cricopharyngeal Dilatation for OPMD

Objective: Oculopharyngeal muscular dystrophy (OPMD) is a rare, autosomal dominant, progressive degenerative muscle disorder featuring dysphagia with limited therapeutic options. Cricopharyngeal myotomy improves symptoms, yet dysphagia frequently recurs, failing to prevent aspiration and enteral feeding. The objective is to evaluate safety and efficacy of repeated endoscopic dilatation for OPMD over a 15-year period. Method: All patients at our unit with genetically confirmed OPMD were included. Cricopharyngeal dilatation was performed with a wire-guided 18 mm Savary-Gilliard bougie. Repeat dilatation was offered when symptoms recurred. Symptom severity prior to initial dilatation and at follow-up was evaluated using the Sydney Swallow Questionnaire (SSQ). Results: Nine patients (7 female, 2 male) were included. Median total treatment period was 13 years (range, 3-15 years), median number of dilatations per patient was 7.2 (1-16), and median interval between treatments was 15 months (range, 4.5-45 months). All patients recorded sustained symptom improvement. Mean SSQ score was 1108 out of 1700 (SD ± 272.9) prior to first dilatation and 298 out of 1700 (SD ± 189.1) at last follow-up, representing a 73% decrease (95% CI 52-94) in degree of dysphagia symptoms (Paired t test, P = .0001). All mean scores for individual questions also showed significant improvement ( P &lt; .05). No adverse events were reported with all patients maintaining oral feeding at last follow-up. Conclusion: Repeated cricopharyngeal dilatation is a safe, effective, well-tolerated, and long-lasting treatment for dysphagia in OPMD.

Cricopharyngeal Dilatation for the Long-term Treatment of Dysphagia in Oculopharyngeal Muscular Dystrophy

Oculopharyngeal muscular dystrophy (OPMD) is a rare autosomal dominant, progressive degenerative muscle disorder featuring dysphagia with limited therapeutic options. The aim of this study was to evaluate the safety and efficacy of repeated endoscopic dilatation for OPMD over a 15-year period. All patients seen at our Regional Swallowing Clinic with OPMD confirmed by genetic analysis were included. Cricopharyngeal dilatation was performed as an outpatient procedure using a wire-guided 18-mm (54 Fr) Savary-Gilliard bougie with the patient under sedation. Patients were offered repeat endoscopic dilatation when symptoms recurred. Symptom severity prior to initial dilatation and at follow-up was evaluated using the Sydney Swallow Questionnaire (SSQ). Nine patients (7 female, 2 male) were included for analysis. Median total treatment period was 13years (range=3-15), median number of dilatations per patient was 7.2 (range=1-16), and median interval between treatments was 15months (range=4.5-45). All patients recorded sustained symptom improvement. Mean SSQ score (out of 1,700) was 1,108.11 (SD±272.85) prior to first dilatation and 297.78 (SD±189.14) at last follow-up, representing a 73% decrease (95% CI=52-94) in degree of dysphagia symptoms (paired t-test, P=0.0001). All mean scores for individual questions also showed significant improvement (P<0.05). No adverse events were reported with all patients maintaining oral feeding at last follow-up. Repeated cricopharyngeal dilatation is a safe, effective, well-tolerated, and long-lasting treatment for dysphagia in OPMD.

Differential Requirement for Utrophin in the Induced Pluripotent Stem Cell Correction of Muscle versus Fat in Muscular Dystrophy Mice

Duchenne muscular dystrophy (DMD) is an incurable degenerative muscle disorder. We injected WT mouse induced pluripotent stem cells (iPSCs) into mdx and mdx∶utrophin mutant blastocysts, which are predisposed to develop DMD with an increasing degree of severity (mdx <<< mdx∶utrophin). In mdx chimeras, iPSC-dystrophin was supplied to the muscle sarcolemma to effect corrections at morphological and functional levels. Dystrobrevin was observed in dystrophin-positive and, at a lesser extent, utrophin-positive areas. In the mdx∶utrophin mutant chimeras, although iPSC-dystrophin was also supplied to the muscle sarcolemma, mice still displayed poor skeletal muscle histopathology, and negligible levels of dystrobrevin in dystrophin- and utrophin-negative areas. Not only dystrophin-expressing tissues are affected by iPSCs. Mdx and mdx∶utrophin mice have reduced fat/body weight ratio, but iPSC injection normalized this parameter in both mdx and mdx∶utrophin chimeras, despite the fact that utrophin was compromised in the mdx∶utrophin chimeric fat. The results suggest that the presence of utrophin is required for the iPSC-corrections in skeletal muscle. Furthermore, the results highlight a potential (utrophin-independent) non-cell autonomous role for iPSC-dystrophin in the corrections of non-muscle tissue like fat, which is intimately related to the muscle.

Mitigation of muscular dystrophy in mice by SERCA overexpression in skeletal muscle

Muscular dystrophies (MDs) comprise a group of degenerative muscle disorders characterized by progressive muscle wasting and often premature death. The primary defect common to most MDs involves disruption of the dystrophin-glycoprotein complex (DGC). This leads to sarcolemmal instability and Ca(2+) influx, inducing cellular necrosis. Here we have shown that the dystrophic phenotype observed in δ-sarcoglycan–null (Sgcd(–/–)) mice and dystrophin mutant mdx mice is dramatically improved by skeletal muscle–specific overexpression of sarcoplasmic reticulum Ca(2+) ATPase 1 (SERCA1). Rates of myofiber central nucleation, tissue fibrosis, and serum creatine kinase levels were dramatically reduced in Sgcd(–/–) and mdx mice with the SERCA1 transgene, which also rescued the loss of exercise capacity in Sgcd(–/–) mice. Adeno-associated virus–SERCA2a (AAV-SERCA2a) gene therapy in the gastrocnemius muscle of Sgcd(–/–) mice mitigated dystrophic disease. SERCA1 overexpression reversed a defect in sarcoplasmic reticulum Ca(2+) reuptake that characterizes dystrophic myofibers and reduced total cytosolic Ca(2+). Further, SERCA1 overexpression almost completely rescued the dystrophic phenotype in a mouse model of MD driven solely by Ca(2+) influx. Mitochondria isolated from the muscle of SERCA1-Sgcd(–/–) mice were no longer swollen and calpain activation was reduced, suggesting protection from Ca(2+)-driven necrosis. Our results suggest a novel therapeutic approach using SERCA1 to abrogate the altered intracellular Ca(2+) levels that underlie most forms of MD.

Cricopharyngeal dilatation for the long-term treatment of dysphagia in oculopharyngeal muscular dystrophy

Oculopharyngeal muscular dystrophy (OPMD) is a rare autosomal dominant, progressive degenerative muscle disorder featuring dysphagia with limited therapeutic options. The aim of this study was to evaluate the safety and efficacy of repeated endoscopic dilatation for OPMD over a 15-year period. All patients seen at our Regional Swallowing Clinic with OPMD confirmed by genetic analysis were included. Cricopharyngeal dilatation was performed as an outpatient procedure using a wire-guided 18-mm (54 Fr) Savary-Gilliard bougie with the patient under sedation. Patients were offered repeat endoscopic dilatation when symptoms recurred. Symptom severity prior to initial dilatation and at follow-up was evaluated using the Sydney Swallow Questionnaire (SSQ). Nine patients (7 female, 2 male) were included for analysis. Median total treatment period was 13 years (range = 3–15), median number of dilatations per patient was 7.2 (range = 1–16), and median interval between treatments was 15 months (range = 4.5–45). All patients recorded sustained symptom improvement. Mean SSQ score (out of 1,700) was 1,108.11 (SD ± 272.85) prior to first dilatation and 297.78 (SD ± 189.14) at last follow-up, representing a 73% decrease (95% CI = 52–94) in degree of dysphagia symptoms (paired t-test, P = 0.0001). All mean scores for individual questions also showed significant improvement (P < 0.05). No adverse events were reported with all patients maintaining oral feeding at last follow-up. Repeated cricopharyngeal dilatation is a safe, effective, well-tolerated, and long-lasting treatment for dysphagia in OPMD.

Genetic Ablation of TWEAK Augments Regeneration and Post-Injury Growth of Skeletal Muscle in Mice

Impairment in the regeneration process is a critical determinant for skeletal muscle wasting in chronic diseases and degenerative muscle disorders. Inflammatory cytokines are known to cause significant muscle wasting, however, their role in myofiber regeneration is less clear. In this study we have investigated the role of tumor necrosis factor-like weak inducer of apoptosis (TWEAK) in skeletal muscle regeneration in vivo. Our results show that expression levels of TWEAK and its receptor Fn14 are significantly increased in skeletal muscles of mice after injury. Genetic deletion of TWEAK increased the fiber cross-sectional area and levels of embryonic isoform of myosin heavy chain in regenerating tibial anterior muscle. Conversely, muscle-specific transgenic overexpression of TWEAK reduced the fiber cross-sectional area and levels of the embryonic myosin heavy chain in regenerating muscle. TWEAK induced the expression of several inflammatory molecules and increased interstitial fibrosis in regenerating muscle. Genetic ablation of TWEAK suppressed, whereas overexpression of TWEAK increased, the activation of nuclear factor-kappa B without affecting the activation of Akt or p38 kinase in regenerating myofibers. Primary myoblasts from TWEAK-null mice showed enhanced differentiation in vitro, whereas myoblasts from TWEAK-Tg mice showed reduced differentiation compared with wild-type mice. Collectively, our study suggests that TWEAK negatively regulates muscle regeneration and that TWEAK is a potential therapeutic target to enhance skeletal muscle regeneration in vivo.

Functional Rescue of Dystrophin-deficient mdx Mice by a Chimeric Peptide-PMO

Splice modulation using antisense oligonucleotides (AOs) has been shown to yield targeted exon exclusion to restore the open reading frame and generate truncated but partially functional dystrophin protein. This has been successfully demonstrated in dystrophin-deficient mdx mice and in Duchenne muscular dystrophy (DMD) patients. However, DMD is a systemic disease; successful therapeutic exploitation of this approach will therefore depend on effective systemic delivery of AOs to all affected tissues. We have previously shown the potential of a muscle-specific/arginine-rich chimeric peptide-phosphorodiamidate morpholino (PMO) conjugate, but its long-term activity, optimized dosing regimen, capacity for functional correction and safety profile remain to be established. Here, we report the results of this chimeric peptide-PMO conjugate in the mdx mouse using low doses (3 and 6 mg/kg) administered via a 6 biweekly systemic intravenous injection protocol. We show 100% dystrophin-positive fibers and near complete correction of the dystrophin transcript defect in all peripheral muscle groups, with restoration of 50% dystrophin protein over 12 weeks, leading to correction of the DMD pathological phenotype and restoration of muscle function in the absence of detectable toxicity or immune response. Chimeric muscle-specific/cell-penetrating peptides therefore represent highly promising agents for systemic delivery of splice-correcting PMO oligomers for DMD therapy.