Muscular Dystrophy Stem Cell Treatment

Muscular Dystrophy and Stem Cell Therapy

What is Muscular Dystrophy?

Muscular Dystrophy and Stem Cell Therapy

Muscular Dystrophy and Stem Cell Therapy


Muscular Dystrophy (MD) refers to a group of hereditary muscle diseases that weakens the muscles that move the human body.
Muscular dystrophies are characterized by progressive skeletal muscle weakness, defects in muscle proteins, and the death of muscle cells and tissue.

Nine diseases including Duchenne, Becker, limb girdle, congenital, facioscapulohumeral, myotonic, oculopharyngeal, distal, and Emery-Dreifuss are always classified as muscular dystrophy but there are more than 100 diseases in total with similarities to muscular dystrophy.

Most types of MD are multi-system disorders with manifestations in body systems including the heart, gastrointestinal and nervous systems, endocrine glands, skin, eyes and even brain.

The condition may also lead to mood swings and learning difficulties.

 

Effective myotube formation in human adipose tissue-derived stem cells expressing dystrophin and myosin heavy chain by cellular fusion with mouse C2C12 myoblasts.

Biochem Biophys Res Commun. 2011 Apr 5;

Authors: Eom YW, Lee JE, Yang MS, Jang IK, Kim HE, Lee DH, Kim YJ, Park WJ, Kong JH, Shim KY, Lee JI, Kim HS

Stem cell therapy for muscular dystrophies requires stem cells that are able to participate in the formation of new muscle fibers. However, the differentiation steps that are the most critical for this process are not clear.

We investigated the myogenic phases of human adipose tissue-derived stem cells (hASCs) step by step and the capability of myotube formation according to the differentiation phase by cellular fusion with mouse myoblast C2C12 cells.

In hASCs treated with 5-azacytidine and fibroblast growth factor-2 (FGF-2) for 1day, the early differentiation step to express MyoD and myogenin was induced by FGF-2 treatment for 6days. Dystrophin and myosin heavy chain (MyHC) expression was induced by hASC conditioned medium in the late differentiation step.

Myotubes were observed only in hASCs undergoing the late differentiation step by cellular fusion with C2C12 cells. In contrast, hASCs that were normal or in the early stage were not involved in myotube formation.

Our results indicate that stem cells expressing dystrophin and MyHC are more suitable for myotube formation by co-culture with myoblasts than normal or early differentiated stem cells expressing MyoD and myogenin.

PMID: 21473854 [PubMed - as supplied by publisher]

Related Articles Creation of a Novel Humanized Dystrophic Mouse Model of Duchenne Muscular Dystrophy and Application of a CRISPR/Cas9 Gene Editing Therapy. J Neuromuscul Dis. 2017;4(2):139-145 Authors: Young CS, Mokhonova E, Quinonez M, Pyle AD, Spencer MJ Abstract Duchenne muscular dystrophy is caused by mutations in DMD which disrupt the reading frame. Therapeutic strategies that restore DMD's reading frame, such as exon skipping and CRISPR/Cas9, need to be tested in the context of the human DMD sequence in vivo. We have developed a novel dystrophic mouse model by using CRISPR/Cas9 to delete exon 45 in the human DMD gene in hDMD mice, which places DMD out-of-frame. We have utilized this model to demonstrate that our clinically-relevant CRISPR/Cas9 platform, which targets deletion of human DMD exons 45-55, can be directly applied in vivo to restore dystrophin. PMID: 28505980 [PubMed - in process]
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Related Articles The nuclear pore protein Nup153 associates with chromatin and regulates cardiac gene expression in dystrophic mdx hearts. Cardiovasc Res. 2016 Nov 01;112(2):555-567 Authors: Nanni S, Re A, Ripoli C, Gowran A, Nigro P, D'Amario D, Amodeo A, Crea F, Grassi C, Pontecorvi A, Farsetti A, Colussi C Abstract Aims: Beyond the control of nuclear-cytoplasmic trafficking nucleoporins regulate gene expression and are involved in cardiac diseases. Notably, a number of cardiovascular disorders have been linked to alterations in epigenetic mechanisms. Here we aimed to determine the contribution of Nup153 to the epigenetic alterations occurring in cardiomyopathy of dystrophin-deficient mdx mice (C57BL/10ScSn-Dmd mdx /J). Methods and results: Nup153 was lysine-acetylated and its expression was significantly increased at protein level in mdx hearts compared with controls. Accordingly, lysine acetyl transferase (KAT) activity associated with Nup153 was higher in mdx hearts paralleling increased binding with the lysine acetylases P300/CBP-associated factor (PCAF) and p300. Interestingly, Nup153 silencing in mdx organotypic heart tissue slices caused a reduction in PCAF- and p300-specific activities. Remarkably, the level of nitric oxide (NO), which is reduced in mdx mice, was important for KAT-dependent regulation of Nup153. In fact, treatment of mdx heart tissue with an NO donor or the KAT inhibitor anacardic acid normalized Nup153 protein expression. Nup153 was recruited to chromatin and regulated the transcription of genes involved in cardiac remodelling, including the actin-binding protein nexilin. Accordingly, nexilin protein expression was abrogated by Nup153 silencing in mdx organotypic cultures. Electrophysiological and molecular experiments revealed that Nup153 overexpression in normal cardiomyocytes increases Ca v 1.2 calcium channel expression and function. Alterations in Nup153 protein expression and intracellular localization were also found in dystrophic cardiomyocytes derived from patient-specific induced pluripotent stem cells. Importantly, Nup153 up-regulation and increased acetylation were also found in the heart of Duchenne muscular dystrophy patients. Conclusions: Our data indicate that Nup153 is an epigenetic regulator which, upon altered NO signalling, mediates the activation of genes potentially associated with early dystrophic cardiac remodelling. PMID: 28513807 [PubMed - indexed for MEDLINE]
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Related Articles Therapeutic developments: Masters of medicine. Nature. 2017 05 17;545(7654):S4-S9 Authors: Bourzac K, Bender E, Dolgin E, Mullard A, Savage N, Gruber K PMID: 28514412 [PubMed - indexed for MEDLINE]
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Related Articles Repression of phosphatidylinositol transfer protein α ameliorates the pathology of Duchenne muscular dystrophy. Proc Natl Acad Sci U S A. 2017 06 06;114(23):6080-6085 Authors: Vieira NM, Spinazzola JM, Alexander MS, Moreira YB, Kawahara G, Gibbs DE, Mead LC, Verjovski-Almeida S, Zatz M, Kunkel LM Abstract Duchenne muscular dystrophy (DMD) is a progressive muscle wasting disease caused by X-linked inherited mutations in the DYSTROPHIN (DMD) gene. Absence of dystrophin protein from the sarcolemma causes severe muscle degeneration, fibrosis, and inflammation, ultimately leading to cardiorespiratory failure and premature death. Although there are several promising strategies under investigation to restore dystrophin protein expression, there is currently no cure for DMD, and identification of genetic modifiers as potential targets represents an alternative therapeutic strategy. In a Brazilian golden retriever muscular dystrophy (GRMD) dog colony, two related dogs demonstrated strikingly mild dystrophic phenotypes compared with those typically observed in severely affected GRMD dogs despite lacking dystrophin. Microarray analysis of these "escaper" dogs revealed reduced expression of phosphatidylinositol transfer protein-α (PITPNA) in escaper versus severely affected GRMD dogs. Based on these findings, we decided to pursue investigation of modulation of PITPNA expression on dystrophic pathology in GRMD dogs, dystrophin-deficient sapje zebrafish, and human DMD myogenic cells. In GRMD dogs, decreased expression of Pitpna was associated with increased phosphorylated Akt (pAkt) expression and decreased PTEN levels. PITPNA knockdown by injection of morpholino oligonucleotides in sapje zebrafish also increased pAkt, rescued the abnormal muscle phenotype, and improved long-term sapje mutant survival. In DMD myotubes, PITPNA knockdown by lentiviral shRNA increased pAkt and increased myoblast fusion index. Overall, our findings suggest PIPTNA as a disease modifier that accords benefits to the abnormal signaling, morphology, and function of dystrophic skeletal muscle, and may be a target for DMD and related neuromuscular diseases. PMID: 28533404 [PubMed - indexed for MEDLINE]
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Related Articles [Skeletal muscle stem cell.] Clin Calcium. 2017;27(6):789-794 Authors: Yuasa S Abstract Adult skeletal muscle has its own stem cell population known as satellite cells. After muscle injury, quiescent satellite cells are activated and then proliferate and differentiate into mature skeletal muscle to ensure that muscle function is recovered. In our screen for myocyte differentiation-promoting factors, we noted markedly elevated expression of granulocyte-colony stimulating factor receptor(G-CSFR, encoded by csf3r)in the skeletal muscle developing area. Furthermore, G-CSFR was transiently expressed in regenerating myocytes of adult injured skeletal muscle, and extrinsic G-CSF supported short-term and long-term muscle regeneration in mouse model of skeletal muscle injury. PMID: 28536315 [PubMed - indexed for MEDLINE]
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Related Articles Subtly Modulating Glycogen Synthase Kinase 3 β: Allosteric Inhibitor Development and Their Potential for the Treatment of Chronic Diseases. J Med Chem. 2017 06 22;60(12):4983-5001 Authors: Palomo V, Perez DI, Roca C, Anderson C, Rodríguez-Muela N, Perez C, Morales-Garcia JA, Reyes JA, Campillo NE, Perez-Castillo AM, Rubin LL, Timchenko L, Gil C, Martinez A Abstract Glycogen synthase kinase 3 β (GSK-3β) is a central target in several unmet diseases. To increase the specificity of GSK-3β inhibitors in chronic treatments, we developed small molecules allowing subtle modulation of GSK-3β activity. Design synthesis, structure-activity relationships, and binding mode of quinoline-3-carbohydrazide derivatives as allosteric modulators of GSK-3β are presented here. Furthermore, we show how allosteric binders may overcome the β-catenin side effects associated with strong GSK-3β inhibition. The therapeutic potential of some of these modulators has been tested in human samples from patients with congenital myotonic dystrophy type 1 (CDM1) and spinal muscular atrophy (SMA) patients. We found that compound 53 improves delayed myogenesis in CDM1 myoblasts, while compounds 1 and 53 have neuroprotective properties in SMA-derived cells. These findings suggest that the allosteric modulators of GSK-3β may be used for future development of drugs for DM1, SMA, and other chronic diseases where GSK-3β inhibition exhibits therapeutic effects. PMID: 28548834 [PubMed - indexed for MEDLINE]
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Related Articles RhoA/ROCK inhibition improves the beneficial effects of glucocorticoid treatment in dystrophic muscle: implications for stem cell depletion. Hum Mol Genet. 2017 08 01;26(15):2813-2824 Authors: Mu X, Tang Y, Takayama K, Chen W, Lu A, Wang B, Weiss K, Huard J Abstract Glucocorticoid treatment represents a standard palliative treatment for Duchenne muscular dystrophy (DMD) patients, but various adverse effects have limited this treatment. In an effort to understand the mechanism(s) by which glucocorticoids impart their effects on the dystrophic muscle, and potentially reduce the adverse effects, we have studied the effect of prednisolone treatment in dystrophin/utrophin double knockout (dKO) mice, which exhibit a severe dystrophic phenotype due to rapid muscle stem cell depletion. Our results indicate that muscle stem cell depletion in dKO muscle is related to upregulation of mTOR, and that prednisolone treatment reduces the expression of mTOR and other pro-inflammatory mediators, consequently slowing down muscle stem cell depletion. However, prednisolone treatment was unable to improve the myogenesis of stem cells and reduce fibrosis in dKO muscle. We then studied whether glucocorticoid treatment can be improved by co-administration of an inhibitor of RhoA/ROCK signaling, which can be activated by glucocorticoids and was found in our previous work to be over-activated in dystrophic muscle. Our results indicate that the combination of RhoA/ROCK inhibition and glucocorticoid treatment in dystrophic muscle have a synergistic effect in alleviating the dystrophic phenotype. Taken together, our study not only shed light on the mechanism by which glucocorticoid imparts its beneficial effect on dystrophic muscle, but also revealed the synergistic effect of RhoA/ROCK inhibition and glucocorticoid treatment, which could lead to the development of more efficient therapeutic approaches for treating DMD patients. PMID: 28549178 [PubMed - indexed for MEDLINE]
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Related Articles In silico discovery of substituted pyrido[2,3-d]pyrimidines and pentamidine-like compounds with biological activity in myotonic dystrophy models. PLoS One. 2017;12(6):e0178931 Authors: González ÀL, Konieczny P, Llamusi B, Delgado-Pinar E, Borrell JI, Teixidó J, García-España E, Pérez-Alonso M, Estrada-Tejedor R, Artero R Abstract Myotonic dystrophy type 1 (DM1) is a rare multisystemic disorder associated with an expansion of CUG repeats in mutant DMPK (dystrophia myotonica protein kinase) transcripts; the main effect of these expansions is the induction of pre-mRNA splicing defects by sequestering muscleblind-like family proteins (e.g. MBNL1). Disruption of the CUG repeats and the MBNL1 protein complex has been established as the best therapeutic approach for DM1, hence two main strategies have been proposed: targeted degradation of mutant DMPK transcripts and the development of CUG-binding molecules that prevent MBNL1 sequestration. Herein, suitable CUG-binding small molecules were selected using in silico approaches such as scaffold analysis, similarity searching, and druggability analysis. We used polarization assays to confirm the CUG repeat binding in vitro for a number of candidate compounds, and went on to evaluate the biological activity of the two with the strongest affinity for CUG repeats (which we refer to as compounds 1-2 and 2-5) in DM1 mutant cells and Drosophila DM1 models with an impaired locomotion phenotype. In particular, 1-2 and 2-5 enhanced the levels of free MBNL1 in patient-derived myoblasts in vitro and greatly improved DM1 fly locomotion in climbing assays. This work provides new computational approaches for rational large-scale virtual screens of molecules that selectively recognize CUG structures. Moreover, it contributes valuable knowledge regarding two compounds with desirable biological activity in DM1 models. PMID: 28582438 [PubMed - indexed for MEDLINE]
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Related Articles SPP1 genotype and glucocorticoid treatment modify osteopontin expression in Duchenne muscular dystrophy cells. Hum Mol Genet. 2017 09 01;26(17):3342-3351 Authors: Vianello S, Pantic B, Fusto A, Bello L, Galletta E, Borgia D, Gavassini BF, Semplicini C, Sorarù G, Vitiello L, Pegoraro E Abstract Glucocorticoids are beneficial in Duchenne muscular dystrophy (DMD). Osteopontin (OPN), the protein product of SPP1, plays a role in DMD pathology modulating muscle inflammation and regeneration. A polymorphism in the SPP1 promoter (rs28357094) has been recognized as a genetic modifier of DMD, and there is evidence suggesting that it modifies response to glucocorticoid treatment. The effect of the glucocorticoid deflazacort on SPP1 mRNA and protein expression was investigated in DMD primary human myoblasts and differentiated myotubes with defined rs28357094 genotype (TT versus TG). Both healthy and DMD myoblasts/myotubes abundantly express OPN. In immunoblot, OPN was detected as a doublet of 55 and 50 kDa bands, with a shift towards the lighter isoform in the transition from myoblasts to myotubes and to mature muscle. A significant increase in OPN expression was observed in DMD myotubes carrying the TG compared to the TT genotype at rs28357094. Deflazacort treatment led to a significant increase of OPN only in myotubes carrying the TG genotype, leading to OPN overexpression. Our study shows a strong effect of the rs28357094 G allele in increasing OPN expression in the presence of deflazacort, and adds to the evidence that rs28357094 polymorphism may predict response to glucocorticoids in DMD. PMID: 28595270 [PubMed - indexed for MEDLINE]
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Related Articles CXCL12 and osteopontin from bone marrow-derived mesenchymal stromal cells improve muscle regeneration. Sci Rep. 2017 06 12;7(1):3305 Authors: Maeda Y, Yonemochi Y, Nakajyo Y, Hidaka H, Ikeda T, Ando Y Abstract Muscle satellite cells are essential for muscle regeneration. However, efficient regeneration does not occur without muscle-resident mesenchymal progenitor cells. We show here that bone marrow-derived mesenchymal stromal cells (Bm-MSCs) also facilitate muscle regeneration in Duchenne muscular dystrophy (DMD) model mice. Bm-MSCs transplanted into peritoneal cavities of DMD model mice with severe muscle degeneration strongly suppressed dystrophic pathology and improved death-related symptoms, which resulted in dramatic lifespan extension. Isolated single myofibers from Bm-MSC-transplanted mice manifested considerably less myofiber splitting compared with myofibers from non-transplanted mice, which indicated that transplantation significantly ameliorated abnormal regeneration. With regard to the number of satellite cells, several cells remained on myofibers from Bm-MSC-transplanted model mice, but satellite cells rarely occurred on myofibers from non-transplanted mice. Also, CXCL12 was crucial for muscle regeneration. CXCL12 facilitated muscle regeneration and paired box protein-7 (PAX7) expression after cardiotoxin-related muscle injury in vivo. The majority of primary muscle satellite cells sorted by integrin-α7 and CD34 expressed CXCR4, a receptor specific for CXCL12. CXCL12 strongly suppressed p-STAT3 expression in these sorted cells in vitro. CXCL12 may therefore influence muscle regeneration through STAT3 signaling in satellite cells. Targeting these proteins in or on muscle satellite cells may improve many degenerative muscle diseases. PMID: 28607396 [PubMed - indexed for MEDLINE]
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Related Articles Cellular Reprogramming, Genome Editing, and Alternative CRISPR Cas9 Technologies for Precise Gene Therapy of Duchenne Muscular Dystrophy. Stem Cells Int. 2017;2017:8765154 Authors: Gee P, Xu H, Hotta A Abstract In the past decade, the development of two innovative technologies, namely, induced pluripotent stem cells (iPSCs) and the CRISPR Cas9 system, has enabled researchers to model diseases derived from patient cells and precisely edit DNA sequences of interest, respectively. In particular, Duchenne muscular dystrophy (DMD) has been an exemplary monogenic disease model for combining these technologies to demonstrate that genome editing can correct genetic mutations in DMD patient-derived iPSCs. DMD is an X-linked genetic disorder caused by mutations that disrupt the open reading frame of the dystrophin gene, which plays a critical role in stabilizing muscle cells during contraction and relaxation. The CRISPR Cas9 system has been shown to be capable of targeting the dystrophin gene and rescuing its expression in in vitro patient-derived iPSCs and in vivo DMD mouse models. In this review, we highlight recent advances made using the CRISPR Cas9 system to correct genetic mutations and discuss how emerging CRISPR technologies and iPSCs in a combined platform can play a role in bringing a therapy for DMD closer to the clinic. PMID: 28607562 [PubMed]
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Related Articles Genome Editing and Muscle Stem Cells as a Therapeutic Tool for Muscular Dystrophies. Curr Stem Cell Rep. 2017;3(2):137-148 Authors: Pini V, Morgan JE, Muntoni F, O'Neill HC Abstract PURPOSE OF REVIEW: Muscular dystrophies are a group of severe degenerative disorders characterized by muscle fiber degeneration and death. Therapies designed to restore muscle homeostasis and to replace dying fibers are being experimented, but none of those in clinical trials are suitable to permanently address individual gene mutation. The purpose of this review is to discuss genome editing tools such as CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated), which enable direct sequence alteration and could potentially be adopted to correct the genetic defect leading to muscle impairment. RECENT FINDINGS: Recent findings show that advances in gene therapy, when combined with traditional viral vector-based approaches, are bringing the field of regenerative medicine closer to precision-based medicine. SUMMARY: The use of such programmable nucleases is proving beneficial for the creation of more accurate in vitro and in vivo disease models. Several gene and cell-therapy studies have been performed on satellite cells, the primary skeletal muscle stem cells involved in muscle regeneration. However, these have mainly been based on artificial replacement or augmentation of the missing protein. Satellite cells are a particularly appealing target to address these innovative technologies for the treatment of muscular dystrophies. PMID: 28616376 [PubMed]
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Related Articles Effects of omega-3 on matrix metalloproteinase-9, myoblast transplantation and satellite cell activation in dystrophin-deficient muscle fibers. Cell Tissue Res. 2017 09;369(3):591-602 Authors: de Carvalho SC, Hindi SM, Kumar A, Marques MJ Abstract In Duchenne muscular dystrophy (DMD), lack of dystrophin leads to progressive muscle degeneration, with DMD patients suffering from cardiorespiratory failure. Cell therapy is an alternative to life-long corticoid therapy. Satellite cells, the stem cells of skeletal muscles, do not completely compensate for the muscle damage in dystrophic muscles. Elevated levels of proinflammatory and profibrotic factors, such as metalloproteinase 9 (MMP-9), impair muscle regeneration, leading to extensive fibrosis and poor results with myoblast transplantation therapies. Omega-3 is an anti-inflammatory drug that protects against muscle degeneration in the mdx mouse model of DMD. In the present study, we test our hypothesis that omega-3 affects MMP-9 and thereby benefits muscle regeneration and myoblast transplantation in the mdx mouse. We observe that omega-3 reduces MMP-9 gene expression and improves myoblast engraftment, satellite cell activation, and muscle regeneration by mechanisms involving, at least in part, the regulation of macrophages, as shown here with the fluorescence-activated cell sorting technique. The present study demonstrates the benefits of omega-3 on satellite cell survival and muscle regeneration, further supporting its use in clinical trials and cell therapies in DMD. PMID: 28623422 [PubMed - indexed for MEDLINE]
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Related Articles Making muscle: skeletal myogenesis in vivo and in vitro. Development. 2017 06 15;144(12):2104-2122 Authors: Chal J, Pourquié O Abstract Skeletal muscle is the largest tissue in the body and loss of its function or its regenerative properties results in debilitating musculoskeletal disorders. Understanding the mechanisms that drive skeletal muscle formation will not only help to unravel the molecular basis of skeletal muscle diseases, but also provide a roadmap for recapitulating skeletal myogenesis in vitro from pluripotent stem cells (PSCs). PSCs have become an important tool for probing developmental questions, while differentiated cell types allow the development of novel therapeutic strategies. In this Review, we provide a comprehensive overview of skeletal myogenesis from the earliest premyogenic progenitor stage to terminally differentiated myofibers, and discuss how this knowledge has been applied to differentiate PSCs into muscle fibers and their progenitors in vitro. PMID: 28634270 [PubMed - indexed for MEDLINE]
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Related Articles An Examination of the Role of Transcriptional and Posttranscriptional Regulation in Rhabdomyosarcoma. Stem Cells Int. 2017;2017:2480375 Authors: Hron AJ, Asakura A Abstract Rhabdomyosarcoma (RMS) is an aggressive family of soft tissue tumors that most commonly manifests in children. RMS variants express several skeletal muscle markers, suggesting myogenic stem or progenitor cell origin of RMS. In this review, the roles of both recently identified and well-established microRNAs in RMS are discussed and summarized in a succinct, tabulated format. Additionally, the subtypes of RMS are reviewed along with the involvement of basic helix-loop-helix (bHLH) proteins, Pax proteins, and microRNAs in normal and pathologic myogenesis. Finally, the current and potential future treatment options for RMS are outlined. PMID: 28638414 [PubMed]
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Related Articles Multi-harmonic Imaging in the Second Near-Infrared Window of Nanoparticle-Labeled Stem Cells as a Monitoring Tool in Tissue Depth. ACS Nano. 2017 07 25;11(7):6672-6681 Authors: Dubreil L, Leroux I, Ledevin M, Schleder C, Lagalice L, Lovo C, Fleurisson R, Passemard S, Kilin V, Gerber-Lemaire S, Colle MA, Bonacina L, Rouger K Abstract In order to assess the therapeutic potential of cell-based strategies, it is of paramount importance to elaborate and validate tools for monitoring the behavior of injected cells in terms of tissue dissemination and engraftment properties. Here, we apply bismuth ferrite harmonic nanoparticles (BFO HNPs) to in vitro expanded human skeletal muscle-derived stem cells (hMuStem cells), an attractive therapeutic avenue for patients suffering from Duchenne muscular dystrophy (DMD). We demonstrate the possibility of stem cell labeling with HNPs. We also show that the simultaneous acquisition of second- and third-harmonic generation (SHG and THG) from BFO HNPs helps separate their response from tissue background, with a net increase in imaging selectivity, which could be particularly important in pathologic context that is defined by a highly remodelling tissue. We demonstrate the possibility of identifying <100 nm HNPs in depth of muscle tissue at more than 1 mm from the surface, taking full advantage of the extended imaging penetration depth allowed by multiphoton microscopy in the second near-infrared window (NIR-II). Based on this successful assessment, we monitor over 14 days any modification on proliferation and morphology features of hMuStem cells upon exposure to PEG-coated BFO HNPs at different concentrations, revealing their high biocompatibility. Successively, we succeed in detecting individual HNP-labeled hMuStem cells in skeletal muscle tissue after their intramuscular injection. PMID: 28644009 [PubMed - in process]
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Related Articles Dental stem cells: recent progresses in tissue engineering and regenerative medicine. Ann Med. 2017 Dec;49(8):644-651 Authors: Botelho J, Cavacas MA, Machado V, Mendes JJ Abstract Since the disclosure of adult mesenchymal stem cells (MSCs), there have been an intense investigation on the characteristics of these cells and their potentialities. Dental stem cells (DSCs) are MSC-like populations with self-renewal capacity and multidifferentiation potential. Currently, there are five main DSCs, dental pulp stem cells (DPSCs), stem cells from exfoliated deciduous teeth (SHED), stem cells from apical papilla (SCAP), periodontal ligament stem cells (PDLSCs) and dental follicle precursor cells (DFPCs). These cells are extremely accessible, prevail during all life and own an amazing multipotency. In the past decade, DPSCs and SHED have been thoroughly studied in regenerative medicine and tissue engineering as autologous stem cells therapies and have shown amazing therapeutic abilities in oro-facial, neurologic, corneal, cardiovascular, hepatic, diabetic, renal, muscular dystrophy and auto-immune conditions, in both animal and human models, and most recently some of them in human clinical trials. In this review, we focus the characteristics, the multiple roles of DSCs and its potential translation to clinical settings. These new insights of the apparently regenerative aptitude of these DSCs seems quite promising to investigate these cells abilities in a wide variety of pathologies. Key messages Dental stem cells (DSCs) have a remarkable self-renewal capacity and multidifferentiation potential; DSCs are extremely accessible and prevail during all life; DSCs, as stem cells therapies, have shown amazing therapeutic abilities in oro-facial, neurologic, corneal, cardiovascular, hepatic, diabetic, renal, muscular dystrophy and autoimmune conditions; DSCs are becoming extremely relevant in tissue engineering and regenerative medicine. PMID: 28649865 [PubMed - indexed for MEDLINE]
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Related Articles PAX7 Targets, CD54, Integrin α9β1, and SDC2, Allow Isolation of Human ESC/iPSC-Derived Myogenic Progenitors. Cell Rep. 2017 06 27;19(13):2867-2877 Authors: Magli A, Incitti T, Kiley J, Swanson SA, Darabi R, Rinaldi F, Selvaraj S, Yamamoto A, Tolar J, Yuan C, Stewart R, Thomson JA, Perlingeiro RCR Abstract Pluripotent stem (PS)-cell-derived cell types hold promise for treating degenerative diseases. However, PS cell differentiation is intrinsically heterogeneous; therefore, clinical translation requires the development of practical methods for isolating progenitors from unwanted and potentially teratogenic cells. Muscle-regenerating progenitors can be derived through transient PAX7 expression. To better understand the biology, and to discover potential markers for these cells, here we investigate PAX7 genomic targets and transcriptional changes in human cells undergoing PAX7-mediated myogenic commitment. We identify CD54, integrin α9β1, and Syndecan2 (SDC2) as surface markers on PAX7-induced myogenic progenitors. We show that these markers allow for the isolation of myogenic progenitors using both fluorescent- and CGMP-compatible magnetic-based sorting technologies and that CD54+α9β1+SDC2+ cells contribute to long-term muscle regeneration in vivo. These findings represent a critical step toward enabling the translation of PS-cell-based therapies for muscle diseases. PMID: 28658631 [PubMed - indexed for MEDLINE]
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Related Articles Autologous intramuscular transplantation of engineered satellite cells induces exosome-mediated systemic expression of Fukutin-related protein and rescues disease phenotype in a murine model of limb-girdle muscular dystrophy type 2I. Hum Mol Genet. 2017 10 01;26(19):3682-3698 Authors: Frattini P, Villa C, De Santis F, Meregalli M, Belicchi M, Erratico S, Bella P, Raimondi MT, Lu Q, Torrente Y Abstract α-Dystroglycanopathies are a group of muscular dystrophies characterized by α-DG hypoglycosylation and reduced extracellular ligand-binding affinity. Among other genes involved in the α-DG glycosylation process, fukutin related protein (FKRP) gene mutations generate a wide range of pathologies from mild limb girdle muscular dystrophy 2I (LGMD2I), severe congenital muscular dystrophy 1C (MDC1C), to Walker-Warburg Syndrome and Muscle-Eye-Brain disease. FKRP gene encodes for a glycosyltransferase that in vivo transfers a ribitol phosphate group from a CDP -ribitol present in muscles to α-DG, while in vitro it can be secreted as monomer of 60kDa. Consistently, new evidences reported glycosyltransferases in the blood, freely circulating or wrapped within vesicles. Although the physiological function of blood stream glycosyltransferases remains unclear, they are likely released from blood borne or distant cells. Thus, we hypothesized that freely or wrapped FKRP might circulate as an extracellular glycosyltransferase, able to exert a "glycan remodelling" process, even at distal compartments. Interestingly, we firstly demonstrated a successful transduction of MDC1C blood-derived CD133+ cells and FKRP L276IKI mouse derived satellite cells by a lentiviral vector expressing the wild-type of human FKRP gene. Moreover, we showed that LV-FKRP cells were driven to release exosomes carrying FKRP. Similarly, we observed the presence of FKRP positive exosomes in the plasma of FKRP L276IKI mice intramuscularly injected with engineered satellite cells. The distribution of FKRP protein boosted by exosomes determined its restoration within muscle tissues, an overall recovery of α-DG glycosylation and improved muscle strength, suggesting a systemic supply of FKRP protein acting as glycosyltransferase. PMID: 28666318 [PubMed - indexed for MEDLINE]
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Related Articles Can Human Embryonic Stem Cell-Derived Stromal Cells Serve a Starting Material for Myoblasts? Stem Cells Int. 2017;2017:7541734 Authors: Ando Y, Saito M, Machida M, Yoshida-Noro C, Akutsu H, Takahashi M, Toyoda M, Umezawa A Abstract A large number of myocytes are necessary to treat intractable muscular disorders such as Duchenne muscular dystrophy with cell-based therapies. However, starting materials for cellular therapy products such as myoblasts, marrow stromal cells, menstrual blood-derived cells, and placenta-derived cells have a limited lifespan and cease to proliferate in vitro. From the viewpoints of manufacturing and quality control, cells with a long lifespan are more suitable as a starting material. In this study, we generated stromal cells for future myoblast therapy from a working cell bank of human embryonic stem cells (ESCs). The ESC-derived CD105+ cells with extensive in vitro proliferation capability exhibited myogenesis and genetic stability in vitro. These results imply that ESC-derived CD105+ cells are another cell source for myoblasts in cell-based therapy for patients with genetic muscular disorders. Since ESCs are immortal, mesenchymal stromal cells generated from ESCs can be manufactured at a large scale in one lot for pharmaceutical purposes. PMID: 28706537 [PubMed]
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Related Articles Skeletal muscle generated from induced pluripotent stem cells - induction and application. World J Stem Cells. 2017 Jun 26;9(6):89-97 Authors: Miyagoe-Suzuki Y, Takeda S Abstract Human induced pluripotent stem cells (hiPS cells or hiPSCs) can be derived from cells of patients with severe muscle disease. If skeletal muscle induced from patient-iPSCs shows disease-specific phenotypes, it can be useful for studying the disease pathogenesis and for drug development. On the other hand, human iPSCs from healthy donors or hereditary muscle disease-iPSCs whose genomes are edited to express normal protein are expected to be a cell source for cell therapy. Several protocols for the derivation of skeletal muscle from human iPSCs have been reported to allow the development of efficient treatments for devastating muscle diseases. In 2017, the focus of research is shifting to another stage: (1) the establishment of mature myofibers that are suitable for study of the pathogenesis of muscle disease; (2) setting up a high-throughput drug screening system; and (3) the preparation of highly regenerative, non-oncogenic cells in large quantities for cell transplantation, etc. PMID: 28717411 [PubMed]
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Related Articles Osteopontin is linked with AKT, FoxO1, and myostatin in skeletal muscle cells. Muscle Nerve. 2017 Dec;56(6):1119-1127 Authors: Nghiem PP, Kornegay JN, Uaesoontrachoon K, Bello L, Yin Y, Kesari A, Mittal P, Schatzberg SJ, Many GM, Lee NH, Hoffman EP Abstract INTRODUCTION: Osteopontin (OPN) polymorphisms are associated with muscle size and modify disease progression in Duchenne muscular dystrophy (DMD). We hypothesized that OPN may share a molecular network with myostatin (MSTN). METHODS: Studies were conducted in the golden retriever (GRMD) and mdx mouse models of DMD. Follow-up in-vitro studies were employed in myogenic cells and the mdx mouse treated with recombinant mouse (rm) or human (Hu) OPN protein. RESULTS: OPN was increased and MSTN was decreased and levels correlated inversely in GRMD hypertrophied muscle. RM-OPN treatment led to induced AKT1 and FoxO1 phosphorylation, microRNA-486 modulation, and decreased MSTN. An AKT1 inhibitor blocked these effects, whereas an RGD-mutant OPN protein and an RGDS blocking peptide showed similar effects to the AKT inhibitor. RMOPN induced myotube hypertrophy and minimal Feret diameter in mdx muscle. DISCUSSION: OPN may interact with AKT1/MSTN/FoxO1 to modify normal and dystrophic muscle. Muscle Nerve 56: 1119-1127, 2017. PMID: 28745831 [PubMed - indexed for MEDLINE]
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Related Articles Human dental pulp pluripotent-like stem cells promote wound healing and muscle regeneration. Stem Cell Res Ther. 2017 07 27;8(1):175 Authors: Martínez-Sarrà E, Montori S, Gil-Recio C, Núñez-Toldrà R, Costamagna D, Rotini A, Atari M, Luttun A, Sampaolesi M Abstract BACKGROUND: Dental pulp represents an easily accessible autologous source of adult stem cells. A subset of these cells, named dental pulp pluripotent-like stem cells (DPPSC), shows high plasticity and can undergo multiple population doublings, making DPPSC an appealing tool for tissue repair or maintenance. METHODS: DPPSC were harvested from the dental pulp of third molars extracted from young patients. Growth factors released by DPPSC were analysed using antibody arrays. Cells were cultured in specific differentiation media and their endothelial, smooth and skeletal muscle differentiation potential was evaluated. The therapeutic potential of DPPSC was tested in a wound healing mouse model and in two genetic mouse models of muscular dystrophy (Scid/mdx and Sgcb-null Rag2-null γc-null). RESULTS: DPPSC secreted several growth factors involved in angiogenesis and extracellular matrix deposition and improved vascularisation in all three murine models. Moreover, DPPSC stimulated re-epithelialisation and ameliorated collagen deposition and organisation in healing wounds. In dystrophic mice, DPPSC engrafted in the skeletal muscle of both dystrophic murine models and showed integration in muscular fibres and vessels. In addition, DPPSC treatment resulted in reduced fibrosis and collagen content, larger cross-sectional area of type II fast-glycolytic fibres and infiltration of higher numbers of proangiogenic CD206+ macrophages. CONCLUSIONS: Overall, DPPSC represent a potential source of stem cells to enhance the wound healing process and slow down dystrophic muscle degeneration. PMID: 28750661 [PubMed - indexed for MEDLINE]
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Related Articles Direct reprogramming of fibroblasts into skeletal muscle progenitor cells by transcription factors enriched in undifferentiated subpopulation of satellite cells. Sci Rep. 2017 Aug 14;7(1):8097 Authors: Ito N, Kii I, Shimizu N, Tanaka H, Takeda S Abstract Satellite cells comprise a functionally heterogeneous population of stem cells in skeletal muscle. Separation of an undifferentiated subpopulation and elucidation of its molecular background are necessary to identify the reprogramming factors to induce skeletal muscle progenitor cells. In this study, we found that intracellular esterase activity distinguishes a subpopulation of cultured satellite cells with high stemness using esterase-sensitive cell staining reagent, calcein-AM. Gene expression analysis of this subpopulation revealed that defined combinations of transcription factors (Pax3, Mef2b, and Pitx1 or Pax7, Mef2b, and Pitx1 in embryonic fibroblasts, and Pax7, Mef2b and MyoD in adult fibroblasts) reprogrammed fibroblasts into skeletal muscle progenitor cells. These reprogrammed cells formed Dystrophin-positive mature muscle fibers when transplanted into a mouse model of Duchenne muscular dystrophy. These results highlight the new marker for heterogenous population of cultured satellite cells, potential therapeutic approaches and cell sources for degenerative muscle diseases. PMID: 28808339 [PubMed - in process]
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Related Articles Follistatin N terminus differentially regulates muscle size and fat in vivo. Exp Mol Med. 2017 09 15;49(9):e377 Authors: Zheng H, Qiao C, Tang R, Li J, Bulaklak K, Huang Z, Zhao C, Dai Y, Li J, Xiao X Abstract Delivery of follistatin (FST) represents a promising strategy for both muscular dystrophies and diabetes, as FST is a robust antagonist of myostatin and activin, which are critical regulators of skeletal muscle and adipose tissues. FST is a multi-domain protein, and deciphering the function of different domains will facilitate novel designs for FST-based therapy. Our study aims to investigate the role of the N-terminal domain (ND) of FST in regulating muscle and fat mass in vivo. Different FST constructs were created and packaged into the adeno-associated viral vector (AAV). Overexpression of wild-type FST in normal mice greatly increased muscle mass while decreasing fat accumulation, whereas overexpression of an N terminus mutant or N terminus-deleted FST had no effect on muscle mass but moderately decreased fat mass. In contrast, FST-I-I containing the complete N terminus and double domain I without domain II and III had no effect on fat but increased skeletal muscle mass. The effects of different constructs on differentiated C2C12 myotubes were consistent with the in vivo finding. We hypothesized that ND was critical for myostatin blockade, mediating the increase in muscle mass, and was less pivotal for activin binding, which accounts for the decrease in the fat tissue. An in vitro TGF-beta1-responsive reporter assay revealed that FST-I-I and N terminus-mutated or -deleted FST showed differential responses to blockade of activin and myostatin. Our study provided direct in vivo evidence for a role of the ND of FST, shedding light on future potential molecular designs for FST-based gene therapy. PMID: 28912572 [PubMed - indexed for MEDLINE]
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Related Articles ECM-Related Myopathies and Muscular Dystrophies: Pros and Cons of Protein Therapies. Compr Physiol. 2017 Sep 12;7(4):1519-1536 Authors: Van Ry PM, Fontelonga TM, Barraza-Flores P, Sarathy A, Nunes AM, Burkin DJ Abstract Extracellular matrix (ECM) myopathies and muscular dystrophies are a group of genetic diseases caused by mutations in genes encoding proteins that provide critical links between muscle cells and the extracellular matrix. These include structural proteins of the ECM, muscle cell receptors, enzymes, and intracellular proteins. Loss of adhesion within the myomatrix results in progressive muscle weakness. For many ECM muscular dystrophies, symptoms can occur any time after birth and often result in reduced life expectancy. There are no cures for the ECM-related muscular dystrophies and treatment options are limited to palliative care. Several therapeutic approaches have been explored to treat muscular dystrophies including gene therapy, gene editing, exon skipping, embryonic, and adult stem cell therapy, targeting genetic modifiers, modulating inflammatory responses, or preventing muscle degeneration. Recently, protein therapies that replace components of the defective myomatrix or enhance muscle and/or extracellular matrix integrity and function have been explored. Preclinical studies for many of these biologics have been promising in animal models of these muscle diseases. This review aims to summarize the ECM muscular dystrophies for which protein therapies are being developed and discuss the exciting potential and possible limitations of this approach for treating this family of devastating genetic muscle diseases. © 2017 American Physiological Society. Compr Physiol 7:1519-1536, 2017. PMID: 28915335 [PubMed - indexed for MEDLINE]
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Related Articles Myogenic Progenitor Cells Exhibit Type I Interferon-Driven Proangiogenic Properties and Molecular Signature During Juvenile Dermatomyositis. Arthritis Rheumatol. 2018 01;70(1):134-145 Authors: Gitiaux C, Latroche C, Weiss-Gayet M, Rodero MP, Duffy D, Bader-Meunier B, Glorion C, Nusbaum P, Bodemer C, Mouchiroud G, Chelly J, Germain S, Desguerre I, Chazaud B Abstract OBJECTIVE: Juvenile dermatomyositis (JDM) is an inflammatory pediatric myopathy characterized by focal capillary loss in muscle, followed by progressive recovery upon adequate treatment with immunomodulating drugs, although some patients remain refractory to treatment. While the underlying mechanism of capillary depletion remains uncertain, recent studies have identified an up-regulation of type I interferon (IFN) expression specific to JDM. Given that myogenic precursor cells (MPCs) exert proangiogenic activity during normal skeletal muscle regeneration, we hypothesized that they may also modulate vascular remodeling/angiogenesis during JDM. The aim of this study was to investigate that hypothesis. METHODS: Human cell cocultures were used to analyze angiogenic properties in patients with JDM, patients with Duchenne's muscular dystrophy (DMD) (control patients for vascular remodeling), and healthy control subjects. Transcriptome analysis was used to examine muscle-derived MPCs. Histologic analysis of type I IFN in muscle biopsy samples was also performed. RESULTS: Using human cell cocultures, we showed highly angiogenic properties of MPCs from JDM patients in association with the expression of an angiogenic molecular signature. Transcriptome analysis of MPCs freshly isolated from muscle samples revealed type I IFN as the master regulator of the most up-regulated genes in JDM-derived MPCs. Functionally, treatment of normal MPCs with type I IFN recapitulated the molecular pattern and the proangiogenic functions of JDM-derived MPCs. In vivo histologic investigation showed that MPCs synthesized type I IFN and major proangiogenic molecules in JDM muscle. Moreover, MPCs derived from JDM muscles that were characterized by strong vasculopathy produced higher levels of type I IFN, confirming MPCs as a cellular source of type I IFN during JDM, and this correlated with the severity of the disease. CONCLUSION: These results demonstrate a new type I IFN pathway in JDM that activates the production of angiogenic effectors by MPCs, triggering their proangiogenic function to promote vessel recovery and muscle reconstruction. PMID: 28941175 [PubMed - indexed for MEDLINE]
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Related Articles Myogenic Satellite Cells: Biological Milieu and Possible Clinical Applications. Pak J Biol Sci. 2017;20(1):1-11 Authors: S Said R, G Mustafa A, A Asfour H, I Shaqoura E Abstract Adult skeletal muscle is a post-mitotic terminally differentiated tissue that possesses an immense potential for regeneration after injury. This regeneration can be achieved by adult stem cells named satellite cells that inhabit the muscular tissue. These cells were first identified in 1961 and were described as being wedged between the plasma membrane of the muscle fiber and the surrounding basement membrane. Since their discovery, many researchers investigated their embryological origin and the exact role they play in muscle regeneration and repair. Under normal conditions, satellite cells are retained in a quiescent state and when required, these cells are activated to proliferate and differentiate to repair pre-existing muscle fibers or to a lesser extent fuse with each other to form new myofibers. During skeletal muscle regeneration, satellite cell actions are regulated through a cascade of complex signaling pathways that are influenced by multiple extrinsic factors within the satellite cell micro-environment. Here, the basic concepts were studied about satellite cells, their development, function, distribution and the different cellular and molecular mechanisms that regulate these cells. The recent findings about some of their clinical applications and potential therapeutic use were also discussed. PMID: 29023009 [PubMed - indexed for MEDLINE]
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Related Articles Myoblasts and macrophages are required for therapeutic morpholino antisense oligonucleotide delivery to dystrophic muscle. Nat Commun. 2017 10 16;8(1):941 Authors: Novak JS, Hogarth MW, Boehler JF, Nearing M, Vila MC, Heredia R, Fiorillo AA, Zhang A, Hathout Y, Hoffman EP, Jaiswal JK, Nagaraju K, Cirak S, Partridge TA Abstract Exon skipping is a promising therapeutic strategy for Duchenne muscular dystrophy (DMD), employing morpholino antisense oligonucleotides (PMO-AO) to exclude disruptive exons from the mutant DMD transcript and elicit production of truncated dystrophin protein. Clinical trials for PMO show variable and sporadic dystrophin rescue. Here, we show that robust PMO uptake and efficient production of dystrophin following PMO administration coincide with areas of myofiber regeneration and inflammation. PMO localization is sustained in inflammatory foci where it enters macrophages, actively differentiating myoblasts and newly forming myotubes. We conclude that efficient PMO delivery into muscle requires two concomitant events: first, accumulation and retention of PMO within inflammatory foci associated with dystrophic lesions, and second, fusion of PMO-loaded myoblasts into repairing myofibers. Identification of these factors accounts for the variability in clinical trials and suggests strategies to improve this therapeutic approach to DMD.Exon skipping is a strategy for the treatment of Duchenne muscular dystrophy, but has variable efficacy. Here, the authors show that dystrophin restoration occurs preferentially in areas of myofiber regeneration, where antisense oligonucleotides are stored in macrophages and delivered to myoblasts and newly formed myotubes. PMID: 29038471 [PubMed - indexed for MEDLINE]
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Related Articles Impeding Transcription of Expanded Microsatellite Repeats by Deactivated Cas9. Mol Cell. 2017 Nov 02;68(3):479-490.e5 Authors: Pinto BS, Saxena T, Oliveira R, Méndez-Gómez HR, Cleary JD, Denes LT, McConnell O, Arboleda J, Xia G, Swanson MS, Wang ET Abstract Transcription of expanded microsatellite repeats is associated with multiple human diseases, including myotonic dystrophy, Fuchs endothelial corneal dystrophy, and C9orf72-ALS/FTD. Reducing production of RNA and proteins arising from these expanded loci holds therapeutic benefit. Here, we tested the hypothesis that deactivated Cas9 enzyme impedes transcription across expanded microsatellites. We observed a repeat length-, PAM-, and strand-dependent reduction of repeat-containing RNAs upon targeting dCas9 directly to repeat sequences; targeting the non-template strand was more effective. Aberrant splicing patterns were rescued in DM1 cells, and production of RAN peptides characteristic of DM1, DM2, and C9orf72-ALS/FTD cells was drastically decreased. Systemic delivery of dCas9/gRNA by adeno-associated virus led to reductions in pathological RNA foci, rescue of chloride channel 1 protein expression, and decreased myotonia. These observations suggest that transcription of microsatellite repeat-containing RNAs is more sensitive to perturbation than transcription of other RNAs, indicating potentially viable strategies for therapeutic intervention. PMID: 29056323 [PubMed - indexed for MEDLINE]
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Related Articles Silencing Nfix rescues muscular dystrophy by delaying muscle regeneration. Nat Commun. 2017 10 20;8(1):1055 Authors: Rossi G, Bonfanti C, Antonini S, Bastoni M, Monteverde S, Innocenzi A, Saclier M, Taglietti V, Messina G Abstract Muscular dystrophies are severe disorders due to mutations in structural genes, and are characterized by skeletal muscle wasting, compromised patient mobility, and respiratory functions. Although previous works suggested enhancing regeneration and muscle mass as therapeutic strategies, these led to no long-term benefits in humans. Mice lacking the transcription factor Nfix have delayed regeneration and a shift toward an oxidative fiber type. Here, we show that ablating or silencing the transcription factor Nfix ameliorates pathology in several forms of muscular dystrophy. Silencing Nfix in postnatal dystrophic mice, when the first signs of the disease already occurred, rescues the pathology and, conversely, Nfix overexpression in dystrophic muscles increases regeneration and markedly exacerbates the pathology. We therefore offer a proof of principle for a novel therapeutic approach for muscular dystrophies based on delaying muscle regeneration. PMID: 29057908 [PubMed - indexed for MEDLINE]
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Related Articles Treatment with the anti-IL-6 receptor antibody attenuates muscular dystrophy via promoting skeletal muscle regeneration in dystrophin-/utrophin-deficient mice. Skelet Muscle. 2017 10 27;7(1):23 Authors: Wada E, Tanihata J, Iwamura A, Takeda S, Hayashi YK, Matsuda R Abstract BACKGROUND: Chronic increases in the levels of the inflammatory cytokine interleukin-6 (IL-6) in serum and skeletal muscle are thought to contribute to the progression of muscular dystrophy. Dystrophin/utrophin double-knockout (dKO) mice develop a more severe and progressive muscular dystrophy than the mdx mice, the most common murine model of Duchenne muscular dystrophy (DMD). In particular, dKO mice have smaller body sizes and muscle diameters, and develop progressive kyphosis and fibrosis in skeletal and cardiac muscles. As mdx mice and DMD patients, we found that IL-6 levels in the skeletal muscle were significantly increased in dKO mice. Thus, in this study, we aimed to analyze the effects of IL-6 receptor (IL-6R) blockade on the muscle pathology of dKO mice. METHODS: Male dKO mice were administered an initial injection (200 mg/kg intraperitoneally (i.p.)) of either the anti-IL-6R antibody MR16-1 or an isotype-matched control rat IgG at the age of 14 days, and were then given weekly injections (25 mg/kg i.p.) until 90 days of age. RESULTS: Treatment of dKO mice with the MR16-1 antibody successfully inhibited the IL-6 pathway in the skeletal muscle and resulted in a significant reduction in the expression levels of phosphorylated signal transducer and activator of transcription 3 in the skeletal muscle. Pathologically, a significant increase in the area of embryonic myosin heavy chain-positive myofibers and muscle diameter, and reduced fibrosis in the quadriceps muscle were observed. These results demonstrated the therapeutic effects of IL-6R blockade on promoting muscle regeneration. Consistently, serum creatine kinase levels were decreased. Despite these improvements observed in the limb muscles, degeneration of the diaphragm and cardiac muscles was not ameliorated by the treatment of mice with the MR16-1 antibody. CONCLUSION: As no adverse effects of treatment with the MR16-1 antibody were observed, our results indicate that the anti-IL-6R antibody is a potential therapy for muscular dystrophy particularly for promoting skeletal muscle regeneration. PMID: 29078808 [PubMed - indexed for MEDLINE]
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Related Articles Preclinical and clinical advances in transposon-based gene therapy. Biosci Rep. 2017 Dec 22;37(6): Authors: Tipanee J, Chai YC, VandenDriessche T, Chuah MK Abstract Transposons derived from Sleeping Beauty (SB), piggyBac (PB), or Tol2 typically require cotransfection of transposon DNA with a transposase either as an expression plasmid or mRNA. Consequently, this results in genomic integration of the potentially therapeutic gene into chromosomes of the desired target cells, and thus conferring stable expression. Non-viral transfection methods are typically preferred to deliver the transposon components into the target cells. However, these methods do not match the efficacy typically attained with viral vectors and are sometimes associated with cellular toxicity evoked by the DNA itself. In recent years, the overall transposition efficacy has gradually increased by codon optimization of the transposase, generation of hyperactive transposases, and/or introduction of specific mutations in the transposon terminal repeats. Their versatility enabled the stable genetic engineering in many different primary cell types, including stem/progenitor cells and differentiated cell types. This prompted numerous preclinical proof-of-concept studies in disease models that demonstrated the potential of DNA transposons for ex vivo and in vivo gene therapy. One of the merits of transposon systems relates to their ability to deliver relatively large therapeutic transgenes that cannot readily be accommodated in viral vectors such as full-length dystrophin cDNA. These emerging insights paved the way toward the first transposon-based phase I/II clinical trials to treat hematologic cancer and other diseases. Though encouraging results were obtained, controlled pivotal clinical trials are needed to corroborate the efficacy and safety of transposon-based therapies. PMID: 29089466 [PubMed - indexed for MEDLINE]
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Related Articles ACVR2B/Fc counteracts chemotherapy-induced loss of muscle and bone mass. Sci Rep. 2017 Oct 31;7(1):14470 Authors: Barreto R, Kitase Y, Matsumoto T, Pin F, Colston KC, Couch KE, O'Connell TM, Couch ME, Bonewald LF, Bonetto A Abstract Chemotherapy promotes the development of cachexia, a debilitating condition characterized by muscle and fat loss. ACVR2B/Fc, an inhibitor of the Activin Receptor 2B signaling, has been shown to preserve muscle mass and prolong survival in tumor hosts, and to increase bone mass in models of osteogenesis imperfecta and muscular dystrophy. We compared the effects of ACVR2B/Fc on muscle and bone mass in mice exposed to Folfiri. In addition to impairing muscle mass and function, Folfiri had severe negative effects on bone, as shown by reduced trabecular bone volume fraction (BV/TV), thickness (Tb.Th), number (Tb.N), connectivity density (Conn.Dn), and by increased separation (Tb.Sp) in trabecular bone of the femur and vertebra. ACVR2B/Fc prevented the loss of muscle mass and strength, and the loss of trabecular bone in femurs and vertebrae following Folfiri administration. Neither Folfiri nor ACVR2B/Fc had effects on femoral cortical bone, as shown by unchanged cortical bone volume fraction (Ct.BV/TV), thickness (Ct.Th) and porosity. Our results suggest that Folfiri is responsible for concomitant muscle and bone degeneration, and that ACVR2B/Fc prevents these derangements. Future studies are required to determine if the same protective effects are observed in combination with other anticancer regimens or in the presence of cancer. PMID: 29089584 [PubMed - in process]
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Related Articles Skeletal Muscle Cells Generated from Pluripotent Stem Cells. Stem Cells Int. 2017;2017:7824614 Authors: Miyagoe-Suzuki Y, Asakura A, Suzuki M PMID: 29098011 [PubMed]
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Related Articles A human iPS cell myogenic differentiation system permitting high-throughput drug screening. Stem Cell Res. 2017 12;25:98-106 Authors: Uchimura T, Otomo J, Sato M, Sakurai H Abstract Muscular dystrophy is a disease characterized by progressive muscle weakness and degeneration. There are currently no available treatments for most muscular diseases, such as muscular dystrophy. Moreover, current therapeutics are focused on improving the quality of life of patients by relieving the symptoms or stress caused by the disease. Although the causative genes for many muscular diseases have been identified, the mechanisms underlying their pathogenesis remain unclear. Patient-derived induced pluripotent stem cells (iPSCs) have become a powerful tool for understanding the pathogenesis of intractable diseases, as well as for phenotype screening, which can serve as the basis for developing new drugs. However, it is necessary to develop an efficient and reproducible myogenic differentiation system. Previously, we reported a tetracycline-inducible MyoD overexpression model of myogenic differentiation using human iPSCs (hiPSCs). However, this model has certain disadvantages that limit its use in various applications, such as a drug screening. In this study, we developed an efficient and reproducible myogenic differentiation system by further modifying our previous protocol. The new protocol achieves efficient differentiation of feeder-free hiPSCs to myogenic cells via small-scale culture in six-well microplates to large-scale culture in 384-well microplates for high-throughput applications. PMID: 29125995 [PubMed - indexed for MEDLINE]
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Related Articles Killing two birds with one stone: The multifunctional roles of mesenchymal stem cells in the treatment of neurodegenerative and muscle diseases. Histol Histopathol. 2018 Jul;33(7):629-638 Authors: Lee NK, Na DL, Chang JW Abstract Neurodegenerative and muscle diseases bear both complex and multifactorial pathologies. An efficacious and robust therapeutic option to treat these diseases is yet to be elucidated. At such a time, mesenchymal stem cells have drawn significant attention due to their immunomodulatory and regenerative properties. Accumulating evidence has proposed the capability of MSCs to serve multiple roles in a broad spectrum of diseases by secretion of trophic or paracrine factors. In the present review, we will look into the recent literature and discuss the therapeutic functions of MSCs and their potential to treat various neurodegenerative (Alzheimer's, Parkinson's, and Huntington's disease) and muscle (Duchenne muscular dystrophy, myopathy, and multiple sclerosis) diseases. PMID: 29188600 [PubMed - indexed for MEDLINE]
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Related Articles Glucocorticoids Improve Myogenic Differentiation In Vitro by Suppressing the Synthesis of Versican, a Transitional Matrix Protein Overexpressed in Dystrophic Skeletal Muscles. Int J Mol Sci. 2017 Dec 06;18(12): Authors: McRae N, Forgan L, McNeill B, Addinsall A, McCulloch D, Van der Poel C, Stupka N Abstract In Duchenne muscular dystrophy (DMD), a dysregulated extracellular matrix (ECM) directly exacerbates pathology. Glucocorticoids are beneficial therapeutics in DMD, and have pleiotropic effects on the composition and processing of ECM proteins in other biological contexts. The synthesis and remodelling of a transitional versican-rich matrix is necessary for myogenesis; whether glucocorticoids modulate this transitional matrix is not known. Here, versican expression and processing were examined in hindlimb and diaphragm muscles from mdx dystrophin-deficient mice and C57BL/10 wild type mice. V0/V1 versican (Vcan) mRNA transcripts and protein levels were upregulated in dystrophic compared to wild type muscles, especially in the more severely affected mdx diaphragm. Processed versican (versikine) was detected in wild type and dystrophic muscles, and immunoreactivity was highly associated with newly regenerated myofibres. Glucocorticoids enhanced C2C12 myoblast fusion by modulating the expression of genes regulating transitional matrix synthesis and processing. Specifically, Tgfβ1, Vcan and hyaluronan synthase-2 (Has2) mRNA transcripts were decreased by 50% and Adamts1 mRNA transcripts were increased three-fold by glucocorticoid treatment. The addition of exogenous versican impaired myoblast fusion, whilst glucocorticoids alleviated this inhibition in fusion. In dystrophic mdx muscles, versican upregulation correlated with pathology. We propose that versican is a novel and relevant target gene in DMD, given its suppression by glucocorticoids and that in excess it impairs myoblast fusion, a process key for muscle regeneration. PMID: 29211034 [PubMed - indexed for MEDLINE]
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Related Articles In Vivo Target Gene Activation via CRISPR/Cas9-Mediated Trans-epigenetic Modulation. Cell. 2017 Dec 14;171(7):1495-1507.e15 Authors: Liao HK, Hatanaka F, Araoka T, Reddy P, Wu MZ, Sui Y, Yamauchi T, Sakurai M, O'Keefe DD, Núñez-Delicado E, Guillen P, Campistol JM, Wu CJ, Lu LF, Esteban CR, Izpisua Belmonte JC Abstract Current genome-editing systems generally rely on inducing DNA double-strand breaks (DSBs). This may limit their utility in clinical therapies, as unwanted mutations caused by DSBs can have deleterious effects. CRISPR/Cas9 system has recently been repurposed to enable target gene activation, allowing regulation of endogenous gene expression without creating DSBs. However, in vivo implementation of this gain-of-function system has proven difficult. Here, we report a robust system for in vivo activation of endogenous target genes through trans-epigenetic remodeling. The system relies on recruitment of Cas9 and transcriptional activation complexes to target loci by modified single guide RNAs. As proof-of-concept, we used this technology to treat mouse models of diabetes, muscular dystrophy, and acute kidney disease. Results demonstrate that CRISPR/Cas9-mediated target gene activation can be achieved in vivo, leading to measurable phenotypes and amelioration of disease symptoms. This establishes new avenues for developing targeted epigenetic therapies against human diseases. VIDEO ABSTRACT. PMID: 29224783 [PubMed - indexed for MEDLINE]
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Related Articles Reversible immortalisation enables genetic correction of human muscle progenitors and engineering of next-generation human artificial chromosomes for Duchenne muscular dystrophy. EMBO Mol Med. 2018 02;10(2):254-275 Authors: Benedetti S, Uno N, Hoshiya H, Ragazzi M, Ferrari G, Kazuki Y, Moyle LA, Tonlorenzi R, Lombardo A, Chaouch S, Mouly V, Moore M, Popplewell L, Kazuki K, Katoh M, Naldini L, Dickson G, Messina G, Oshimura M, Cossu G, Tedesco FS Abstract Transferring large or multiple genes into primary human stem/progenitor cells is challenged by restrictions in vector capacity, and this hurdle limits the success of gene therapy. A paradigm is Duchenne muscular dystrophy (DMD), an incurable disorder caused by mutations in the largest human gene: dystrophin. The combination of large-capacity vectors, such as human artificial chromosomes (HACs), with stem/progenitor cells may overcome this limitation. We previously reported amelioration of the dystrophic phenotype in mice transplanted with murine muscle progenitors containing a HAC with the entire dystrophin locus (DYS-HAC). However, translation of this strategy to human muscle progenitors requires extension of their proliferative potential to withstand clonal cell expansion after HAC transfer. Here, we show that reversible cell immortalisation mediated by lentivirally delivered excisable hTERT and Bmi1 transgenes extended cell proliferation, enabling transfer of a novel DYS-HAC into DMD satellite cell-derived myoblasts and perivascular cell-derived mesoangioblasts. Genetically corrected cells maintained a stable karyotype, did not undergo tumorigenic transformation and retained their migration ability. Cells remained myogenic in vitro (spontaneously or upon MyoD induction) and engrafted murine skeletal muscle upon transplantation. Finally, we combined the aforementioned functions into a next-generation HAC capable of delivering reversible immortalisation, complete genetic correction, additional dystrophin expression, inducible differentiation and controllable cell death. This work establishes a novel platform for complex gene transfer into clinically relevant human muscle progenitors for DMD gene therapy. PMID: 29242210 [PubMed - in process]
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Related Articles Immunoglobulin therapy ameliorates the phenotype and increases lifespan in the severely affected dystrophin-utrophin double knockout mice. Eur J Hum Genet. 2017 12;25(12):1388-1396 Authors: Nunes BG, Loures FV, Bueno HMS, Cangussu EB, Goulart E, Coatti GC, Caldini EG, Condino-Neto A, Zatz M Abstract Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder, caused by mutations in the dystrophin gene, affecting 1:3500-5000 boys worldwide. The lack of dystrophin induces degeneration of muscle cells and elicits an immune response characterized by an intensive secretion of pro-inflammatory cytokines. Immunoglobulins modulate the inflammatory response through several mechanisms and have been widely used as an adjuvant therapy for autoimmune diseases. Here we evaluated the effect of immunoglobulin G (IG) injected intraperitoneally in a severely affected double knockout (dko) mouse model for Duchenne muscular dystrophy. The IG dko treated mice were compared regarding activity rates, survival and histopathology with a control untreated group. Additionally, dendritic cells and naïve lymphocytes from these two groups and WT mice were obtained to study in vitro the role of the immune system associated to DMD pathophysiology. We show that IG therapy significantly enhances activity rate and lifespan of dko mice. It diminishes muscle tissue inflammation by decreasing the expression of costimulatory molecules MHC, CD86 and CD40 and reducing Th1-related cytokines IFN-γ, IL-1β and TNF-α release. IG therapy dampens the effector immune responses supporting the hypothesis according to which the immune response accelerates DMD progression. As IG therapy is already approved by FDA for treating autoimmune disorders, with less side-effects than currently used glucocorticoids, our results may open a new therapeutic option aiming to improve life quality and lifespan of DMD patients. PMID: 29255177 [PubMed - indexed for MEDLINE]
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Related Articles PAX7 target genes are globally repressed in facioscapulohumeral muscular dystrophy skeletal muscle. Nat Commun. 2017 12 18;8(1):2152 Authors: Banerji CRS, Panamarova M, Hebaishi H, White RB, Relaix F, Severini S, Zammit PS Abstract Facioscapulohumeral muscular dystrophy (FSHD) is a prevalent, incurable myopathy, linked to hypomethylation of D4Z4 repeats on chromosome 4q causing expression of the DUX4 transcription factor. However, DUX4 is difficult to detect in FSHD muscle biopsies and it is debatable how robust changes in DUX4 target gene expression are as an FSHD biomarker. PAX7 is a master regulator of myogenesis that rescues DUX4-mediated apoptosis. Here, we show that suppression of PAX7 target genes is a hallmark of FSHD, and that it is as major a signature of FSHD muscle as DUX4 target gene expression. This is shown using meta-analysis of over six FSHD muscle biopsy gene expression studies, and validated by RNA-sequencing on FSHD patient-derived myoblasts. DUX4 also inhibits PAX7 from activating its transcriptional target genes and vice versa. Furthermore, PAX7 target gene repression can explain oxidative stress sensitivity and epigenetic changes in FSHD. Thus, PAX7 target gene repression is a hallmark of FSHD that should be considered in the investigation of FSHD pathology and therapy. PMID: 29255294 [PubMed - indexed for MEDLINE]
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Related Articles Aberrant Caspase Activation in Laminin-α2-Deficient Human Myogenic Cells is Mediated by p53 and Sirtuin Activity. J Neuromuscul Dis. 2018;5(1):59-73 Authors: Yoon S, Beermann ML, Yu B, Shao D, Bachschmid M, Miller JB Abstract BACKGROUND: Mutations in the LAMA2 gene encoding laminin-α2 cause congenital muscular dystrophy Type 1A (MDC1A), a severe recessive disease with no effective treatment. Previous studies have shown that aberrant activation of caspases and cell death through a pathway regulated by BAX and KU70 is a significant contributor to pathogenesis in laminin-α2-deficiency. OBJECTIVES: To identify mechanisms of pathogenesis in MDC1A. METHODS: We used immunocytochemical and molecular studies of human myogenic cells and mouse muscles-comparing laminin-α2-deficient vs. healthy controls-to identify mechanisms that regulate pathological activation of caspase in laminin-α2-deficiency. RESULTS: In cultures of myogenic cells from MDC1A donors, p53 accumulated in a subset of nuclei and aberrant caspase activation was inhibited by the p53 inhibitor pifithrin-alpha. Also, the p53 target BBC3 (PUMA) was upregulated in both MDC1A myogenic cells and Lama2-/- mouse muscles. In addition, studies with sirtuin inhibitors and SIRT1 overexpression showed that caspase activation in MDC1A myotubes was inversely related to sirtuin deacetylase activity. Caspase activation in laminin-α2-deficiency was, however, not associated with increased phosphorylation of p38 MAPK. CONCLUSIONS: Aberrant caspase activation in MDC1A cells was mediated both by sirtuin deacetylase activity and by p53. Interventions that inhibit aberrant caspase activation by targeting sirtuin or p53 function could potentially be useful in ameliorating MDC1A. PMID: 29278895 [PubMed - indexed for MEDLINE]
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Related Articles "Known Unknowns": Current Questions in Muscle Satellite Cell Biology. Curr Top Dev Biol. 2018;126:205-233 Authors: Cornelison D Abstract Our understanding of satellite cells, now known to be the obligate stem cells of skeletal muscle, has increased dramatically in recent years due to the introduction of new molecular, genetic, and technical resources. In addition to their role in acute repair of damaged muscle, satellite cells are of interest in the fields of aging, exercise, neuromuscular disease, and stem cell therapy, and all of these applications have driven a dramatic increase in our understanding of the activity and potential of satellite cells. However, many fundamental questions of satellite cell biology remain to be answered, including their emergence as a specific lineage, the degree and significance of heterogeneity within the satellite cell population, the roles of their interactions with other resident and infiltrating cell types during homeostasis and regeneration, and the relative roles of intrinsic vs extrinsic factors that may contribute to satellite cell dysfunction in the context of aging or disease. This review will address the current state of these open questions in satellite cell biology. PMID: 29304999 [PubMed - in process]
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Related Articles Creation of Dystrophin Expressing Chimeric Cells of Myoblast Origin as a Novel Stem Cell Based Therapy for Duchenne Muscular Dystrophy. Stem Cell Rev. 2018 04;14(2):189-199 Authors: Siemionow M, Cwykiel J, Heydemann A, Garcia-Martinez J, Siemionow K, Szilagyi E Abstract Over the past decade different stem cell (SC) based approaches were tested to treat Duchenne Muscular Dystrophy (DMD), a lethal X-linked disorder caused by mutations in dystrophin gene. Despite research efforts, there is no curative therapy for DMD. Allogeneic SC therapies aim to restore dystrophin in the affected muscles; however, they are challenged by rejection and limited engraftment. Thus, there is a need to develop new more efficacious SC therapies. Chimeric Cells (CC), created via ex vivo fusion of donor and recipient cells, represent a promising therapeutic option for tissue regeneration and Vascularized Composite Allotransplantation (VCA) due to tolerogenic properties that eliminate the need for lifelong immunosuppression. This proof of concept study tested feasibility of myoblast fusion for Dystrophin Expressing. Chimeric Cell (DEC) therapy through in vitro characterization and in vivo assessment of engraftment, survival, and efficacy in the mdx mouse model of DMD. Murine DEC were created via ex vivo fusion of normal (snj) and dystrophin-deficient (mdx) myoblasts using polyethylene glycol. Efficacy of myoblast fusion was confirmed by flow cytometry and dystrophin immunostaining, while proliferative and myogenic differentiation capacity of DEC were assessed in vitro. Therapeutic effect after DEC transplant (0.5 × 106) into the gastrocnemius muscle (GM) of mdx mice was assessed by muscle functional tests. At 30 days post-transplant dystrophin expression in GM of injected mdx mice increased to 37.27 ± 12.1% and correlated with improvement of muscle strength and function. Our study confirmed feasibility and efficacy of DEC therapy and represents a novel SC based approach for treatment of muscular dystrophies. PMID: 29305755 [PubMed - indexed for MEDLINE]
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Related Articles Effective regeneration of dystrophic muscle using autologous iPSC-derived progenitors with CRISPR-Cas9 mediated precise correction. Med Hypotheses. 2018 Jan;110:97-100 Authors: Hagan M, Ashraf M, Kim IM, Weintraub NL, Tang Y Abstract Duchenne muscular dystrophy (DMD) is a lethal muscle wasting disease caused by a lack of dystrophin, which eventually leads to apoptosis of muscle cells and impaired muscle contractility. Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein 9 (CRISPR/Cas9) gene editing of induced pluripotent stem cells (IPSC) offers the potential to correct the DMD gene defect and create healthy IPSC for autologous cell transplantation without causing immune activation. However, IPSC carry a risk of tumor formation, which can potentially be mitigated by differentiation of IPSC into myogenic progenitor cells (MPC). We hypothesize that precise genetic editing in IPSC using CRISPR-Cas9 technology, coupled with MPC differentiation and autologous transplantation, can lead to safe and effective muscle repair. With future research, our hypothesis may provide an optimal autologous stem cell-based approach to treat the dystrophic pathology and improve the quality of life for patients with DMD. PMID: 29317080 [PubMed - indexed for MEDLINE]
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Related Articles Overexpression of the double homeodomain protein DUX4c interferes with myofibrillogenesis and induces clustering of myonuclei. Skelet Muscle. 2018 01 12;8(1):2 Authors: Vanderplanck C, Tassin A, Ansseau E, Charron S, Wauters A, Lancelot C, Vancutsem K, Laoudj-Chenivesse D, Belayew A, Coppée F Abstract BACKGROUND: Facioscapulohumeral muscular dystrophy (FSHD) is associated with DNA hypomethylation at the 4q35 D4Z4 repeat array. Both the causal gene DUX4 and its homolog DUX4c are induced. DUX4c is immunodetected in every myonucleus of proliferative cells, while DUX4 is present in only 1/1000 of myonuclei where it initiates a gene deregulation cascade. FSHD primary myoblasts differentiate into either atrophic or disorganized myotubes. DUX4 expression induces atrophic myotubes and associated FSHD markers. Although DUX4 silencing normalizes the FSHD atrophic myotube phenotype, this is not the case for the disorganized phenotype. DUX4c overexpression increases the proliferation rate of human TE671 rhabdomyosarcoma cells and inhibits their differentiation, suggesting a normal role during muscle differentiation. METHODS: By gain- and loss-of-function experiments in primary human muscle cells, we studied the DUX4c impact on proliferation, differentiation, myotube morphology, and FSHD markers. RESULTS: In primary myoblasts, DUX4c overexpression increased the staining intensity of KI67 (a proliferation marker) in adjacent cells and delayed differentiation. In differentiating cells, DUX4c overexpression led to the expression of some FSHD markers including β-catenin and to the formation of disorganized myotubes presenting large clusters of nuclei and cytoskeletal defects. These were more severe when DUX4c was expressed before the cytoskeleton reorganized and myofibrils assembled. In addition, endogenous DUX4c was detected at a higher level in FSHD myotubes presenting abnormal clusters of nuclei and cytoskeletal disorganization. We found that the disorganized FSHD myotube phenotype could be rescued by silencing of DUX4c, not DUX4. CONCLUSION: Excess DUX4c could disturb cytoskeletal organization and nuclear distribution in FSHD myotubes. We suggest that DUX4c up-regulation could contribute to DUX4 toxicity in the muscle fibers by favoring the clustering of myonuclei and therefore facilitating DUX4 diffusion among them. Defining DUX4c functions in the healthy skeletal muscle should help to design new targeted FSHD therapy by DUX4 or DUX4c inhibition without suppressing DUX4c normal function. PMID: 29329560 [PubMed - indexed for MEDLINE]
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Related Articles To roll the eyes and snap a bite - function, development and evolution of craniofacial muscles. Semin Cell Dev Biol. 2018 Jan 10;: Authors: Schubert FR, Singh AJ, Afoyalan O, Kioussi C, Dietrich S Abstract Craniofacial muscles, muscles that move the eyes, control facial expression and allow food uptake and speech, have long been regarded as a variation on the general body muscle scheme. However, evidence has accumulated that the function of head muscles, their developmental anatomy and the underlying regulatory cascades are distinct. This article reviews the key aspects of craniofacial muscle and muscle stem cell formation and discusses how this differs from the trunk programme of myogenesis; we show novel RNAseq data to support this notion. We also trace the origin of head muscle in the chordate ancestors of vertebrates and discuss links with smooth-type muscle in the primitive chordate pharynx. We look out as to how the special properties of head muscle precursor and stem cells, in particular their competence to contribute to the heart, could be exploited in regenerative medicine. PMID: 29331210 [PubMed - as supplied by publisher]
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Related Articles Efficient differentiation of human pluripotent stem cells into skeletal muscle cells by combining RNA-based MYOD1-expression and POU5F1-silencing. Sci Rep. 2018 01 19;8(1):1189 Authors: Akiyama T, Sato S, Chikazawa-Nohtomi N, Soma A, Kimura H, Wakabayashi S, Ko SBH, Ko MSH Abstract Direct generation of skeletal muscle cells from human pluripotent stem cells (hPSCs) would be beneficial for drug testing, drug discovery, and disease modelling in vitro. Here we show a rapid and robust method to induce myogenic differentiation of hPSCs by introducing mRNA encoding MYOD1 together with siRNA-mediated knockdown of POU5F1 (also known as OCT4 or OCT3/4). This integration-free approach generates functional skeletal myotubes with sarcomere-like structure and a fusion capacity in several days. The POU5F1 silencing facilitates MYOD1 recruitment to the target promoters, which results in the significant activation of myogenic genes in hPSCs. Furthermore, deep sequencing transcriptome analyses demonstrated that POU5F1-knockdown upregulates the genes associated with IGF- and FGF-signaling and extracellular matrix that may also support myogenic differentiation. This rapid and direct differentiation method may have potential applications in regenerative medicine and disease therapeutics for muscle disorders such as muscular dystrophy. PMID: 29352121 [PubMed - indexed for MEDLINE]
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Related Articles Temporal requirement of dystroglycan glycosylation during brain development and rescue of severe cortical dysplasia via gene delivery in the fetal stage. Hum Mol Genet. 2018 04 01;27(7):1174-1185 Authors: Sudo A, Kanagawa M, Kondo M, Ito C, Kobayashi K, Endo M, Minami Y, Aiba A, Toda T Abstract Congenital muscular dystrophies (CMDs) are characterized by progressive weakness and degeneration of skeletal muscle. In several forms of CMD, abnormal glycosylation of α-dystroglycan (α-DG) results in conditions collectively known as dystroglycanopathies, which are associated with central nervous system involvement. We recently demonstrated that fukutin, the gene responsible for Fukuyama congenital muscular dystrophy, encodes the ribitol-phosphate transferase essential for dystroglycan function. Brain pathology in patients with dystroglycanopathy typically includes cobblestone lissencephaly, mental retardation, and refractory epilepsy; however, some patients exhibit average intelligence, with few or almost no structural defects. Currently, there is no effective treatment for dystroglycanopathy, and the mechanisms underlying the generation of this broad clinical spectrum remain unknown. Here, we analysed four distinct mouse models of dystroglycanopathy: two brain-selective fukutin conditional knockout strains (neuronal stem cell-selective Nestin-fukutin-cKO and forebrain-selective Emx1-fukutin-cKO), a FukutinHp strain with the founder retrotransposal insertion in the fukutin gene, and a spontaneous Large-mutant Largemyd strain. These models exhibit variations in the severity of brain pathology, replicating the clinical heterogeneity of dystroglycanopathy. Immunofluorescence analysis of the developing cortex suggested that residual glycosylation of α-DG at embryonic day 13.5 (E13.5), when cortical dysplasia is not yet apparent, may contribute to subsequent phenotypic heterogeneity. Surprisingly, delivery of fukutin or Large into the brains of mice at E12.5 prevented severe brain malformation in Emx1-fukutin-cKO and Largemyd/myd mice, respectively. These findings indicate that spatiotemporal persistence of functionally glycosylated α-DG may be crucial for brain development and modulation of glycosylation during the fetal stage could be a potential therapeutic strategy for dystroglycanopathy. PMID: 29360985 [PubMed - in process]
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Related Articles The roles of muscle stem cells in muscle injury, atrophy and hypertrophy. J Biochem. 2018 May 01;163(5):353-358 Authors: Fukada SI Abstract Skeletal muscle is composed of multinuclear cells called myofibers. Muscular dystrophy (a genetic muscle disorder) induces instability in the cell membrane of myofibers and eventually causes myofibre damage. Non-genetic muscle disorders, including sarcopenia, diabetes, bedridden immobility and cancer cachexia, lead to atrophy of myofibres. In contrast, resistance training induces myofibre hypertrophy. Thus, myofibres exhibit a plasticity that is strongly affected by both intrinsic and extrinsic factors. There is no doubt that muscle stem cells (MuSCs, also known as muscle satellite cells) are indispensable for muscle repair/regeneration, but their contributions to atrophy and hypertrophy are still controversial. The present review focuses on the relevance of MuSCs to (i) muscle diseases and (ii) hypertrophy. Further, this review addresses fundamental questions about MuSCs to clarify the onset or progression of these diseases and which might lead to development of a MuSC-based therapy. PMID: 29394360 [PubMed - indexed for MEDLINE]
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Related Articles Inhibition of Methyltransferase Setd7 Allows the In Vitro Expansion of Myogenic Stem Cells with Improved Therapeutic Potential. Cell Stem Cell. 2018 Feb 01;22(2):177-190.e7 Authors: Judson RN, Quarta M, Oudhoff MJ, Soliman H, Yi L, Chang CK, Loi G, Vander Werff R, Cait A, Hamer M, Blonigan J, Paine P, Doan LTN, Groppa E, He W, Su L, Zhang RH, Xu P, Eisner C, Low M, Barta I, Lewis CB, Zaph C, Karimi MM, Rando TA, Rossi FM Abstract The development of cell therapy for repairing damaged or diseased skeletal muscle has been hindered by the inability to significantly expand immature, transplantable myogenic stem cells (MuSCs) in culture. To overcome this limitation, a deeper understanding of the mechanisms regulating the transition between activated, proliferating MuSCs and differentiation-primed, poorly engrafting progenitors is needed. Here, we show that methyltransferase Setd7 facilitates such transition by regulating the nuclear accumulation of β-catenin in proliferating MuSCs. Genetic or pharmacological inhibition of Setd7 promotes in vitro expansion of MuSCs and increases the yield of primary myogenic cell cultures. Upon transplantation, both mouse and human MuSCs expanded with a Setd7 small-molecule inhibitor are better able to repopulate the satellite cell niche, and treated mouse MuSCs show enhanced therapeutic potential in preclinical models of muscular dystrophy. Thus, Setd7 inhibition may help bypass a key obstacle in the translation of cell therapy for muscle disease. PMID: 29395054 [PubMed - in process]
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Related Articles Generation of GZKHQi001-A and GZWWTi001-A, two induced pluripotent stem cell lines derived from peripheral blood mononuclear cells of Duchenne muscular dystrophy patients. Stem Cell Res. 2018 04;28:25-28 Authors: Yuhuan X, Yingjun X, Yanting X, Yuchang C, Bing S, Shaoying L, Haoxian L, Yexing X, Shuming O, Zeyu X, Xiaofang S Abstract Duchenne muscular dystrophy (DMD) is an X-linked disease caused by mutations in the DMD gene, which spans ~2.4Mb of genomic sequence at locus Xp21. This mutation results in the loss of the protein dystrophin. DMD patients die in their second or third decade due to either respiratory failure or cardiomyopathy, as the absence of dystrophin leads to myofiber membrane fragility and necrosis, eventually resulting in muscle atrophy and contractures. Currently, there is no effective treatment for DMD, therefore induced pluripotent stem cells from DMD patients would be a powerful tool for studying disease mechanisms. PMID: 29414414 [PubMed - indexed for MEDLINE]
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Related Articles Re-education begins at home: an overview of the discovery of in vivo-active small molecule modulators of endogenous stem cells. Expert Opin Drug Discov. 2018 04;13(4):307-326 Authors: Um J, Lee JH, Jung DW, Williams DR Abstract INTRODUCTION: Degenerative diseases, such as Alzheimer's disease, heart disease and arthritis cause great suffering and are major socioeconomic burdens. An attractive treatment approach is stem cell transplantation to regenerate damaged or destroyed tissues. However, this can be problematic. For example, donor cells may not functionally integrate into the host tissue. An alternative methodology is to deliver bioactive agents, such as small molecules, directly into the diseased tissue to enhance the regenerative potential of endogenous stem cells. Areas covered: In this review, the authors discuss the necessity of developing these small molecules to treat degenerative diseases and survey progress in their application as therapeutics. They describe both the successes and caveats of developing small molecules that target endogenous stem cells to induce tissue regeneration. This article is based on literature searches which encompass databases for biomedical research and clinical trials. These small molecules are also categorized per their target disease and mechanism of action. Expert opinion: The development of small molecules targeting endogenous stem cells is a high-profile research area. Some compounds have made the successful transition to the clinic. Novel approaches, such as modulating the stem cell niche or targeted delivery to disease sites, should increase the likelihood of future successes in this field. PMID: 29421943 [PubMed - indexed for MEDLINE]
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Related Articles Age Is Relative-Impact of Donor Age on Induced Pluripotent Stem Cell-Derived Cell Functionality. Front Cardiovasc Med. 2018;5:4 Authors: Strässler ET, Aalto-Setälä K, Kiamehr M, Landmesser U, Kränkel N Abstract Induced pluripotent stem cells (iPSCs) avoid many of the restrictions that hamper the application of human embryonic stem cells: limited availability of source material due to legal restrictions in some countries, immunogenic rejection and ethical concerns. Also, the donor's clinical phenotype is often known when working with iPSCs. Therefore, iPSCs seem ideal to tackle the two biggest tasks of regenerative medicine: degenerative diseases with genetic cause (e.g., Duchenne's muscular dystrophy) and organ replacement in age-related diseases (e.g., end-stage heart or renal failure), especially in combination with recently developed gene-editing tools. In the setting of autologous transplantation in elderly patients, donor age becomes a potentially relevant factor that needs to be assessed. Here, we review and critically discuss available data pertinent to the questions: How does donor age influence the reprogramming process and iPSC functionality? Would it even be possible to reprogram senescent somatic cells? How does donor age affect iPSC differentiation into specialised cells and their functionality? We also identify research needs, which might help resolve current unknowns. Until recently, most hallmarks of ageing were attributed to an accumulation of DNA damage over time, and it was thus expected that DNA damage from a somatic cell would accumulate in iPSCs and the cells derived from them. In line with this, a decreased lifespan of cloned organisms compared with the donor was also observed in early cloning experiments. Therefore, it was questioned for a time whether iPSC derived from an old individual's somatic cells would suffer from early senescence and, thus, may not be a viable option either for disease modelling nor future clinical applications. Instead, typical signs of cellular ageing are reverted in the process of iPSC reprogramming, and iPSCs from older donors do not show diminished differentiation potential nor do iPSC-derived cells from older donors suffer early senescence or show functional impairments when compared with those from younger donors. Thus, the data would suggest that donor age does not limit iPSC application for modelling genetic diseases nor regenerative therapies. However, open questions remain, e.g., regarding the potential tumourigenicity of iPSC-derived cells and the impact of epigenetic pattern retention. PMID: 29423397 [PubMed]
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Related Articles Exosome-Mediated Benefits of Cell Therapy in Mouse and Human Models of Duchenne Muscular Dystrophy. Stem Cell Reports. 2018 Mar 13;10(3):942-955 Authors: Aminzadeh MA, Rogers RG, Fournier M, Tobin RE, Guan X, Childers MK, Andres AM, Taylor DJ, Ibrahim A, Ding X, Torrente A, Goldhaber JM, Lewis M, Gottlieb RA, Victor RA, Marbán E Abstract Genetic deficiency of dystrophin leads to disability and premature death in Duchenne muscular dystrophy (DMD), affecting the heart as well as skeletal muscle. Here, we report that clinical-stage cardiac progenitor cells, known as cardiosphere-derived cells (CDCs), improve cardiac and skeletal myopathy in the mdx mouse model of DMD. Injection of CDCs into the hearts of mdx mice augments cardiac function, ambulatory capacity, and survival. Exosomes secreted by human CDCs reproduce the benefits of CDCs in mdx mice and in human induced pluripotent stem cell-derived Duchenne cardiomyocytes. Surprisingly, CDCs and their exosomes also transiently restored partial expression of full-length dystrophin in mdx mice. The findings further motivate the testing of CDCs in Duchenne patients, while identifying exosomes as next-generation therapeutic candidates. PMID: 29478899 [PubMed - in process]
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Related Articles Humanizing the mdx mouse model of DMD: the long and the short of it. NPJ Regen Med. 2018;3:4 Authors: Yucel N, Chang AC, Day JW, Rosenthal N, Blau HM Abstract Duchenne muscular dystrophy (DMD) is a common fatal heritable myopathy, with cardiorespiratory failure occurring by the third decade of life. There is no specific treatment for DMD cardiomyopathy, in large part due to a lack of understanding of the mechanisms underlying the cardiac failure. Mdx mice, which have the same dystrophin mutation as human patients, are of limited use, as they do not develop early dilated cardiomyopathy as seen in patients. Here we summarize the usefulness of the various commonly used DMD mouse models, highlight a model with shortened telomeres like humans, and identify directions that warrant further investigation. PMID: 29479480 [PubMed]
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Related Articles Autologous Cell Therapy Approach for Duchenne Muscular Dystrophy using PiggyBac Transposons and Mesoangioblasts. Mol Ther. 2018 Apr 04;26(4):1093-1108 Authors: Iyer PS, Mavoungou LO, Ronzoni F, Zemla J, Schmid-Siegert E, Antonini S, Neff LA, Dorchies OM, Jaconi M, Lekka M, Messina G, Mermod N Abstract Duchenne muscular dystrophy (DMD) is a lethal muscle-wasting disease currently without cure. We investigated the use of the PiggyBac transposon for full-length dystrophin expression in murine mesoangioblast (MABs) progenitor cells. DMD murine MABs were transfected with transposable expression vectors for full-length dystrophin and transplanted intramuscularly or intra-arterially into mdx/SCID mice. Intra-arterial delivery indicated that the MABs could migrate to regenerating muscles to mediate dystrophin expression. Intramuscular transplantation yielded dystrophin expression in 11%-44% of myofibers in murine muscles, which remained stable for the assessed period of 5 months. The satellite cells isolated from transplanted muscles comprised a fraction of MAB-derived cells, indicating that the transfected MABs may colonize the satellite stem cell niche. Transposon integration site mapping by whole-genome sequencing indicated that 70% of the integrations were intergenic, while none was observed in an exon. Muscle resistance assessment by atomic force microscopy indicated that 80% of fibers showed elasticity properties restored to those of wild-type muscles. As measured in vivo, transplanted muscles became more resistant to fatigue. This study thus provides a proof-of-principle that PiggyBac transposon vectors may mediate full-length dystrophin expression as well as functional amelioration of the dystrophic muscles within a potential autologous cell-based therapeutic approach of DMD. PMID: 29503200 [PubMed - in process]
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Related Articles Muscle Stem/Progenitor Cells and Mesenchymal Stem Cells of Bone Marrow Origin for Skeletal Muscle Regeneration in Muscular Dystrophies. Arch Immunol Ther Exp (Warsz). 2018 Oct;66(5):341-354 Authors: Klimczak A, Kozlowska U, Kurpisz M Abstract Muscular dystrophies represent a group of diseases which may develop in several forms, and severity of the disease is usually associated with gene mutations. In skeletal muscle regeneration and in muscular dystrophies, both innate and adaptive immune responses are involved. The regenerative potential of mesenchymal stem/stromal cells (MSCs) of bone marrow origin was confirmed by the ability to differentiate into diverse tissues and by their immunomodulatory and anti-inflammatory properties by secretion of a variety of growth factors and anti-inflammatory cytokines. Skeletal muscle comprises different types of stem/progenitor cells such as satellite cells and non-satellite stem cells including MSCs, interstitial stem cells positive for stress mediator PW1 expression and negative for PAX7 called PICs (PW1+/PAX7- interstitial cells), fibro/adipogenic progenitors/mesenchymal stem cells, muscle side population cells and muscle resident pericytes, and all of them actively participate in the muscle regeneration process. In this review, we present biological properties of MSCs of bone marrow origin and a heterogeneous population of muscle-resident stem/progenitor cells, their interaction with the inflammatory environment of dystrophic muscle and potential implications for cellular therapies for muscle regeneration. Subsequently, we propose-based on current research results, conclusions, and our own experience-hypothetical mechanisms for modulation of the complete muscle regeneration process to treat muscular dystrophies. PMID: 29536116 [PubMed - indexed for MEDLINE]
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Related Articles Dystrophin Expressing Chimeric (DEC) Human Cells Provide a Potential Therapy for Duchenne Muscular Dystrophy. Stem Cell Rev. 2018 06;14(3):370-384 Authors: Siemionow M, Cwykiel J, Heydemann A, Garcia J, Marchese E, Siemionow K, Szilagyi E Abstract Duchenne Muscular Dystrophy (DMD) is a progressive and lethal disease caused by mutations of the dystrophin gene. Currently no cure exists. Stem cell therapies targeting DMD are challenged by limited engraftment and rejection despite the use of immunosuppression. There is an urgent need to introduce new stem cell-based therapies that exhibit low allogenic profiles and improved cell engraftment. In this proof-of-concept study, we develop and test a new human stem cell-based approach to increase engraftment, limit rejection, and restore dystrophin expression in the mdx/scid mouse model of DMD. We introduce two Dystrophin Expressing Chimeric (DEC) cell lines created by ex vivo fusion of human myoblasts (MB) derived from two normal donors (MBN1/MBN2), and normal and DMD donors (MBN/MBDMD). The efficacy of fusion was confirmed by flow cytometry and confocal microscopy based on donor cell fluorescent labeling (PKH26/PKH67). In vitro, DEC displayed phenotype and genotype of donor parent cells, expressed dystrophin, and maintained proliferation and myogenic differentiation. In vivo, local delivery of both DEC lines (0.5 × 106) restored dystrophin expression (17.27%±8.05-MBN1/MBN2 and 23.79%±3.82-MBN/MBDMD) which correlated with significant improvement of muscle force, contraction and tolerance to fatigue at 90 days after DEC transplant to the gastrocnemius muscles (GM) of dystrophin-deficient mdx/scid mice. This study establishes DEC as a potential therapy for DMD and other types of muscular dystrophies. PMID: 29546607 [PubMed - indexed for MEDLINE]
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Related Articles Immune-Mediated Necrotizing Myopathy. Curr Rheumatol Rep. 2018 03 26;20(4):21 Authors: Pinal-Fernandez I, Casal-Dominguez M, Mammen AL Abstract PURPOSE OF REVIEW: Immune-mediated necrotizing myopathy (IMNM) is a type of autoimmune myopathy characterized by relatively severe proximal weakness, myofiber necrosis with minimal inflammatory cell infiltrate on muscle biopsy, and infrequent extra-muscular involvement. Here, we will review the characteristics of patients with IMNM. RECENT FINDINGS: Anti-signal recognition particle (SRP) and anti-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) autoantibodies are closely associated with IMNM and define unique subtypes of patients. Importantly, the new European Neuromuscular Centre criteria recognize anti-SRP myopathy, anti-HMGCR myopathy, and autoantibody-negative IMNM as three distinct subtypes of IMNM. Anti-SRP myopathy patients have more severe muscle involvement, have more common extra-muscular features, and may respond best to immunosuppressive regimens that include rituximab. In contrast, anti-HMGCR myopathy is often associated with statin exposure and intravenous immunoglobulin treatment may be an effective treatment, even as monotherapy. Both anti-SRP and anti-HMGCR myopathy tend to be most severe in younger patients. Furthermore, children with these forms of IMNM may present with dystrophy-like features which are potentially reversible with immunosuppressant treatment. IMNM patients with either autoantibody may experience fatty replacement of muscle soon after disease onset, suggesting that intense and early immunosuppressant therapy may provide the best chance to avoid long-term disability. IMNM is composed of anti-SRP myopathy, anti-HMGCR myopathy, and autoantibody-negative IMNM. Both anti-SRP and anti-HMGCR myopathy can cause severe weakness, especially in younger patients. Anti-SRP myopathy patients tend to have the most severe weakness and most prevalent extra-muscular features. Autoantibody-negative IMNM remains poorly described. PMID: 29582188 [PubMed - in process]
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Related Articles Three-Dimensional Human iPSC-Derived Artificial Skeletal Muscles Model Muscular Dystrophies and Enable Multilineage Tissue Engineering. Cell Rep. 2018 Apr 17;23(3):899-908 Authors: Maffioletti SM, Sarcar S, Henderson ABH, Mannhardt I, Pinton L, Moyle LA, Steele-Stallard H, Cappellari O, Wells KE, Ferrari G, Mitchell JS, Tyzack GE, Kotiadis VN, Khedr M, Ragazzi M, Wang W, Duchen MR, Patani R, Zammit PS, Wells DJ, Eschenhagen T, Tedesco FS Abstract Generating human skeletal muscle models is instrumental for investigating muscle pathology and therapy. Here, we report the generation of three-dimensional (3D) artificial skeletal muscle tissue from human pluripotent stem cells, including induced pluripotent stem cells (iPSCs) from patients with Duchenne, limb-girdle, and congenital muscular dystrophies. 3D skeletal myogenic differentiation of pluripotent cells was induced within hydrogels under tension to provide myofiber alignment. Artificial muscles recapitulated characteristics of human skeletal muscle tissue and could be implanted into immunodeficient mice. Pathological cellular hallmarks of incurable forms of severe muscular dystrophy could be modeled with high fidelity using this 3D platform. Finally, we show generation of fully human iPSC-derived, complex, multilineage muscle models containing key isogenic cellular constituents of skeletal muscle, including vascular endothelial cells, pericytes, and motor neurons. These results lay the foundation for a human skeletal muscle organoid-like platform for disease modeling, regenerative medicine, and therapy development. PMID: 29669293 [PubMed - in process]
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Related Articles Premyogenic progenitors derived from human pluripotent stem cells expand in floating culture and differentiate into transplantable myogenic progenitors. Sci Rep. 2018 Apr 26;8(1):6555 Authors: Sakai-Takemura F, Narita A, Masuda S, Wakamatsu T, Watanabe N, Nishiyama T, Nogami K, Blanc M, Takeda S, Miyagoe-Suzuki Y Abstract Human induced pluripotent stem cells (hiPSCs) are a potential source for cell therapy of Duchenne muscular dystrophy. To reliably obtain skeletal muscle progenitors from hiPSCs, we treated hiPS cells with a Wnt activator, CHIR-99021 and a BMP receptor inhibitor, LDN-193189, and then induced skeletal muscle cells using a previously reported sphere-based culture. This protocol greatly improved sphere formation efficiency and stably induced the differentiation of myogenic cells from hiPS cells generated from both healthy donors and a patient with congenital myasthenic syndrome. hiPSC-derived myogenic progenitors were enriched in the CD57(-) CD108(-) CD271(+) ERBB3(+) cell fraction, and their differentiation was greatly promoted by TGF-β inhibitors. TGF-β inhibitors down-regulated the NFIX transcription factor, and NFIX short hairpin RNA (shRNA) improved the differentiation of iPS cell-derived myogenic progenitors. These results suggest that NFIX inhibited differentiation of myogenic progenitors. hiPSC-derived myogenic cells differentiated into myofibers in muscles of NSG-mdx 4Cv mice after direct transplantation. Our results indicate that our new muscle induction protocol is useful for cell therapy of muscular dystrophies. PMID: 29700358 [PubMed - in process]
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Related Articles New Hope for Treatment of Duchene Dystrophy by Employing Dystrophin Expressing Chimeric Cells - Studies Published in Stem Cell Reviews and Reports. Stem Cell Rev. 2018 06;14(3):295-296 Authors: Ratajczak MZ PMID: 29705868 [PubMed - indexed for MEDLINE]
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Related Articles Human serum and platelet lysate are appropriate xeno-free alternatives for clinical-grade production of human MuStem cell batches. Stem Cell Res Ther. 2018 May 02;9(1):128 Authors: Saury C, Lardenois A, Schleder C, Leroux I, Lieubeau B, David L, Charrier M, Guével L, Viau S, Delorme B, Rouger K Abstract BACKGROUND: Canine MuStem cells have demonstrated regenerative efficacy in a dog model of muscular dystrophy, and the recent characterization of human counterparts (hMuStem) has highlighted the therapeutic potential of this muscle-derived stem cell population. To date, these cells have only been generated in research-grade conditions. However, evaluation of the clinical efficacy of any such therapy will require the production of hMuStem cells in compliance with good manufacturing practices (GMPs). Because the current use of fetal bovine serum (FBS) to isolate and expand hMuStem cells raises several ethical, safety, and supply concerns, we assessed the use of two alternative xeno-free blood derivatives: human serum (HS) and a human platelet lysate (hPL). METHODS: hMuStem cells were isolated and expanded in vitro in either HS-supplemented or hPL-supplemented media and the proliferation rate, clonogenicity, myogenic commitment potential, and oligopotency compared with that observed in FBS-supplemented medium. Flow cytometry and high-throughput 3'-digital gene expression RNA sequencing were used to characterize the phenotype and global gene expression pattern of hMuStem cells cultured with HS or hPL. RESULTS: HS-supplemented and hPL-supplemented media both supported the isolation and long-term proliferation of hMuStem cells. Compared with FBS-based medium, both supplements enhanced clonogenicity and allowed for a reduction in growth factor supplementation. Neither supplement altered the cell lineage pattern of hMuStem cells. In vitro differentiation assays revealed a decrease in myogenic commitment and in the fusion ability of hMuStem cells when cultured with hPL. In return, this reduction of myogenic potential in hPL-supplemented cultures was rapidly reversed by substitution of hPL with HS or fibrinogen-depleted hPL. Moreover, culture of hMuStem cells in hPL hydrogel and fibrinogen-depleted hPL demonstrated that myogenic differentiation potential is maintained in heparin-free hPL derivatives. CONCLUSIONS: Our findings indicate that HS and hPL are efficient and viable alternatives to FBS for the preparation of hMuStem cell batches in compliance with GMPs. PMID: 29720259 [PubMed - in process]
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Related Articles Efficient exon skipping of SGCG mutations mediated by phosphorodiamidate morpholino oligomers. JCI Insight. 2018 May 03;3(9): Authors: Wyatt EJ, Demonbreun AR, Kim EY, Puckelwartz MJ, Vo AH, Dellefave-Castillo LM, Gao QQ, Vainzof M, Pavanello RCM, Zatz M, McNally EM Abstract Exon skipping uses chemically modified antisense oligonucleotides to modulate RNA splicing. Therapeutically, exon skipping can bypass mutations and restore reading frame disruption by generating internally truncated, functional proteins to rescue the loss of native gene expression. Limb-girdle muscular dystrophy type 2C is caused by autosomal recessive mutations in the SGCG gene, which encodes the dystrophin-associated protein γ-sarcoglycan. The most common SGCG mutations disrupt the transcript reading frame abrogating γ-sarcoglycan protein expression. In order to treat most SGCG gene mutations, it is necessary to skip 4 exons in order to restore the SGCG transcript reading frame, creating an internally truncated protein referred to as Mini-Gamma. Using direct reprogramming of human cells with MyoD, myogenic cells were tested with 2 antisense oligonucleotide chemistries, 2'-O-methyl phosphorothioate oligonucleotides and vivo-phosphorodiamidate morpholino oligomers, to induce exon skipping. Treatment with vivo-phosphorodiamidate morpholino oligomers demonstrated efficient skipping of the targeted exons and corrected the mutant reading frame, resulting in the expression of a functional Mini-Gamma protein. Antisense-induced exon skipping of SGCG occurred in normal cells and those with multiple distinct SGCG mutations, including the most common 521ΔT mutation. These findings demonstrate a multiexon-skipping strategy applicable to the majority of limb-girdle muscular dystrophy 2C patients. PMID: 29720576 [PubMed - as supplied by publisher]
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Related Articles ANKK1 is found in myogenic precursors and muscle fibers subtypes with glycolytic metabolism. PLoS One. 2018;13(5):e0197254 Authors: Rubio-Solsona E, Martí S, Vílchez JJ, Palau F, Hoenicka J Abstract Ankyrin repeat and kinase domain containing 1 (ANKK1) gene has been widely related to neuropsychiatry disorders. The localization of ANKK1 in neural progenitors and its correlation with the cell cycle has suggested its participation in development. However, ANKK1 functions still need to be identified. Here, we have further characterized the ANKK1 localization in vivo and in vitro, by using immunolabeling, quantitative real-time PCR and Western blot in the myogenic lineage. Histologic investigations in mice and humans revealed that ANKK1 is expressed in precursors of embryonic and adult muscles. In mice embryos, ANKK1 was found in migrating myotubes where it shows a polarized cytoplasmic distribution, while proliferative myoblasts and satellite cells show different isoforms in their nuclei and cytoplasm. In vitro studies of ANKK1 protein isoforms along the myogenic progression showed the decline of nuclear ANKK1-kinase until its total exclusion in myotubes. In adult mice, ANKK1 was expressed exclusively in the Fast-Twitch muscles fibers subtype. The induction of glycolytic metabolism in C2C12 cells with high glucose concentration or treatment with berberine caused a significant increase in the ANKK1 mRNA. Similarly, C2C12 cells under hypoxic conditions caused the increase of nuclear ANKK1. These results altogether show a relationship between ANKK1 gene regulation and the metabolism of muscles during development and in adulthood. Finally, we found ANKK1 expression in regenerative fibers of muscles from dystrophic patients. Future studies in ANKK1 biology and the pathological response of muscles will reveal whether this protein is a novel muscle disease biomarker. PMID: 29758057 [PubMed - indexed for MEDLINE]
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Related Articles Aggregate mesenchymal stem cell delivery ameliorates the regenerative niche for muscle repair. J Tissue Eng Regen Med. 2018 Aug;12(8):1867-1876 Authors: Ruehle MA, Stevens HY, Beedle AM, Guldberg RE, Call JA Abstract Duchenne muscular dystrophy is a severe muscle wasting disease due to the absence of the dystrophin protein from the muscle cell membrane, which renders the muscle susceptible to continuous damage. In Duchenne muscular dystrophy patients, muscle weakness, together with cycles of degeneration/regeneration and replacement with noncontractile tissue, limit mobility and lifespan. Because the loss of dystrophin results in loss of polarity and a reduction in the number of self-renewing satellite cells, it is postulated that these patients could achieve an improved quality of life if delivered cells could restore satellite cell function. In this study, we used both an established myotoxic injury model in wild-type (WT) mice and mdx mice alone (spontaneous muscle damage). Single (SC) and aggregated (AGG) mesenchymal stem cells (MSCs) were injected into the gastrocnemius muscles 4 hr after injury (WT mice). The recovery of peak isometric torque was longitudinally assessed over 5 weeks, with earlier takedowns for histological assessment of healing (fibre cross-section area and central nucleation) and MSC retention. AGG-treated WT mice had significantly greater torque recovery at Day 14 than SC or saline-treated mice and a greater CSA at Day 10, compared with SC/saline. AGG-treated mdx mice had a greater peak isometric torque compared with SC/saline. In vitro immunomodulatory factor secretion of AGG-MSCs was higher than SC-MSCs for all tested growth factors with the largest difference observed in hepatocyte growth factor. Future studies are necessary to pair immunomodulatory factor secretion with functional attributes, to better predict the potential therapeutic value of MSC treatment modalities. PMID: 29774991 [PubMed - in process]
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Related Articles CD133+ cells derived from skeletal muscles of Duchenne muscular dystrophy patients have a compromised myogenic and muscle regenerative capability. Stem Cell Res. 2018 Jul;30:43-52 Authors: Meng J, Muntoni F, Morgan J Abstract Cell-mediated gene therapy is a possible means to treat muscular dystrophies like Duchenne muscular dystrophy. Autologous patient stem cells can be genetically-corrected and transplanted back into the patient, without causing immunorejection problems. Regenerated muscle fibres derived from these cells will express the missing dystrophin protein, thus improving muscle function. CD133+ cells derived from normal human skeletal muscle contribute to regenerated muscle fibres and form muscle stem cells after their intra-muscular transplantation into an immunodeficient mouse model. But it is not known whether CD133+ cells derived from DMD patient muscles have compromised muscle regenerative function. To test this, we compared CD133+ cells derived from DMD and normal human muscles. DMD CD133+ cells had a reduced capacity to undergo myogenic differentiation in vitro compared with CD133+ cells derived from normal muscle. In contrast to CD133+ cells derived from normal human muscle, those derived from DMD muscle formed no satellite cells and gave rise to significantly fewer muscle fibres of donor origin, after their intra-muscular transplantation into an immunodeficient, non-dystrophic, mouse muscle. DMD CD133+ cells gave rise to more clones of smaller size and more clones that were less myogenic than did CD133+ cells derived from normal muscle. The heterogeneity of the progeny of CD133+ cells, combined with the reduced proliferation and myogenicity of DMD compared to normal CD133+ cells, may explain the reduced regenerative capacity of DMD CD133+ cells. PMID: 29783100 [PubMed - in process]
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Related Articles Placenta-derived mesenchymal stromal cells and their exosomes exert therapeutic effects in Duchenne muscular dystrophy. Biomaterials. 2018 Aug;174:67-78 Authors: Bier A, Berenstein P, Kronfeld N, Morgoulis D, Ziv-Av A, Goldstein H, Kazimirsky G, Cazacu S, Meir R, Popovtzer R, Dori A, Brodie C Abstract Duchenne muscular dystrophy (DMD) is a degenerative lethal, X-linked disease of skeletal and cardiac muscles caused by mutations in the dystrophin gene. Cell therapy using different cell types, including mesenchymal stromal cells (MSCs), has been considered as a potential approach for the treatment of DMD. MSCs can be obtained from autologous sources such as bone marrow and adipose tissues or from allogeneic placenta and umbilical cord. The safety and therapeutic impact of these cells has been demonstrated in pre-clinical and clinical studies and their functions are attributed to paracrine effects that are mediated by secreted cytokines and extracellular vesicles. Here, we studied the therapeutic effects of placenta-derived MSCs (PL-MSCs) and their secreted exosomes using mouse and human myoblasts from healthy controls, Duchenne patients and mdx mice. Treatment of myoblasts with conditioned medium or exosomes secreted by PL-MSCs increased the differentiation of these cells and decreased the expression of fibrogenic genes in DMD patient myoblasts. In addition, these treatments also increased the expression of utrophin in these cells. Using a quantitative miR-29c reporter, we demonstrated that the PL-MSC effects were partly mediated by the transfer of exosomal miR-29c. Intramuscular transplantation of PL-MSCs in mdx mice resulted in decreased creatine kinase levels. PL-MSCs significantly decreased the expression of TGF-β and the level of fibrosis in the diaphragm and cardiac muscles, inhibited inflammation and increased utrophin expression. In vivo imaging analyses using MSCs labeled with gold nanoparticles or fluorescent dyes demonstrated localization of the cells in the muscle tissues up to 3 weeks post treatment. Altogether, these results demonstrate that PL-MSCs and their secreted exosomes have important clinical applications in cell therapy of DMD partly via the targeted delivery of exosomal miR-29c. PMID: 29783118 [PubMed - in process]
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Related Articles Monogenic diseases in India. Mutat Res. 2018 Apr - Jun;776:23-31 Authors: Venugopal A, Chandran M, Eruppakotte N, Kizhakkillach S, Breezevilla SC, Vellingiri B Abstract Studies on monogenic diseases are considered valuable because they give insights and expand our knowledge on gene function and regulation. Despite all the current advancement in science and technology, a deep understanding and knowledge as to why only those particular genes are affected in a disease is still vague. We also lack profound illumination as to why only certain mutations are seen in a disease. Though useful from a research perspective, a majority of these diseases are lethal resulting in death of the affected individual. Unfortunately, in the fast - growing land of India, the incidence of monogenic diseases is very high with few counter-measures in place. This article encompasses a list of all monogenic diseases ever to be reported in India with special focus on five diseases which has been stated to have the highest incidence in India. Here, we discuss about the limited research carried out in India on these high incidence monogenic diseases, the other diseases related to those genes, the range of treatments available for these diseases in India in contrast to its availability around the world and the need to develop treatment strategies to reduce the mortality and morbidity due to these rare but daunting diseases. PMID: 29807575 [PubMed - indexed for MEDLINE]
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Related Articles Vascular Delivery of Allogeneic MuStem Cells in Dystrophic Dogs Requires Only Short-Term Immunosuppression to Avoid Host Immunity and Generate Clinical/Tissue Benefits. Cell Transplant. 2018 Jul;27(7):1096-1110 Authors: Lorant J, Larcher T, Jaulin N, Hedan B, Lardenois A, Leroux I, Dubreil L, Ledevin M, Goubin H, Moullec S, Deschamps JY, Thorin C, André C, Adjali O, Rouger K Abstract Growing demonstrations of regenerative potential for some stem cells led recently to promising therapeutic proposals for neuromuscular diseases. We have shown that allogeneic MuStem cell transplantation into Golden Retriever muscular dystrophy (GRMD) dogs under continuous immunosuppression (IS) leads to persistent clinical stabilization and muscle repair. However, long-term IS in medical practice is associated with adverse effects raising safety concerns. Here, we investigate whether the IS removal or its restriction to the transplantation period could be considered. Dogs aged 4-5 months old received vascular infusions of allogeneic MuStem cells without IS (GRMDMU/no-IS) or under transient IS (GRMDMU/tr-IS). At 5 months post-infusion, persisting clinical status improvement of the GRMDMU/tr-IS dogs was observed while GRMDMU/no-IS dogs exhibited no benefit. Histologically, only 9-month-old GRMDMU/tr-IS dogs showed an increased muscle regenerative activity. A mixed cell reaction with the host peripheral blood mononucleated cells (PBMCs) and corresponding donor cells revealed undetectable to weak lymphocyte proliferation in GRMDMU/tr-IS dogs compared with a significant proliferation in GRMDMU/no-IS dogs. Importantly, any dog group showed neither cellular nor humoral anti-dystrophin responses. Our results show that transient IS is necessary and sufficient to sustain allogeneic MuStem cell transplantation benefits and prevent host immunity. These findings provide useful critical insight to designing therapeutic strategies. PMID: 29871519 [PubMed - in process]
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Related Articles Abnormalities in Skeletal Muscle Myogenesis, Growth, and Regeneration in Myotonic Dystrophy. Front Neurol. 2018;9:368 Authors: André LM, Ausems CRM, Wansink DG, Wieringa B Abstract Myotonic dystrophy type 1 (DM1) and 2 (DM2) are autosomal dominant degenerative neuromuscular disorders characterized by progressive skeletal muscle weakness, atrophy, and myotonia with progeroid features. Although both DM1 and DM2 are characterized by skeletal muscle dysfunction and also share other clinical features, the diseases differ in the muscle groups that are affected. In DM1, distal muscles are mainly affected, whereas in DM2 problems are mostly found in proximal muscles. In addition, manifestation in DM1 is generally more severe, with possible congenital or childhood-onset of disease and prominent CNS involvement. DM1 and DM2 are caused by expansion of (CTG•CAG)n and (CCTG•CAGG)n repeats in the 3' non-coding region of DMPK and in intron 1 of CNBP, respectively, and in overlapping antisense genes. This critical review will focus on the pleiotropic problems that occur during development, growth, regeneration, and aging of skeletal muscle in patients who inherited these expansions. The current best-accepted idea is that most muscle symptoms can be explained by pathomechanistic effects of repeat expansion on RNA-mediated pathways. However, aberrations in DNA replication and transcription of the DM loci or in protein translation and proteome homeostasis could also affect the control of proliferation and differentiation of muscle progenitor cells or the maintenance and physiological integrity of muscle fibers during a patient's lifetime. Here, we will discuss these molecular and cellular processes and summarize current knowledge about the role of embryonic and adult muscle-resident stem cells in growth, homeostasis, regeneration, and premature aging of healthy and diseased muscle tissue. Of particular interest is that also progenitor cells from extramuscular sources, such as pericytes and mesoangioblasts, can participate in myogenic differentiation. We will examine the potential of all these types of cells in the application of regenerative medicine for muscular dystrophies and evaluate new possibilities for their use in future therapy of DM. PMID: 29892259 [PubMed]
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Related Articles Galectin-3 and N-acetylglucosamine promote myogenesis and improve skeletal muscle function in the mdx model of Duchenne muscular dystrophy. FASEB J. 2018 Jun 12;:fj201701151RRR Authors: Rancourt A, Dufresne SS, St-Pierre G, Lévesque JC, Nakamura H, Kikuchi Y, Satoh MS, Frenette J, Sato S Abstract The muscle membrane, sarcolemma, must be firmly attached to the basal lamina. The failure of proper attachment results in muscle injury, which is the underlying cause of Duchenne muscular dystrophy (DMD), in which mutations in the dystrophin gene disrupts the firm adhesion. In patients with DMD, even moderate contraction causes damage, leading to progressive muscle degeneration. The damaged muscles are repaired through myogenesis. Consequently, myogenesis is highly active in patients with DMD, and the repeated activation of myogenesis leads to the exhaustion of the myogenic stem cells. Therefore, approaches to reducing the risk of the exhaustion are to develop a treatment that strengthens the interaction between the sarcolemma and the basal lamina and increases the efficiency of the myogenesis. Galectin-3 is an oligosaccharide-binding protein and is known to be involved in cell-cell interactions and cell-matrix interactions. Galectin-3 is expressed in myoblasts and skeletal muscle, although its function in muscle remains elusive. In this study, we found evidence that galectin-3 and the monosaccharide N-acetylglucosamine, which increases the synthesis of binding partners (oligosaccharides) of galectin-3, promote myogenesis in vitro. Moreover, in the mdx mouse model of DMD, treatment with N-acetylglucosamine increased muscle-force production. The results suggest that treatment with N-acetylglucosamine might mitigate the burden of DMD.-Rancourt, A., Dufresne, S. S., St-Pierre, G., Lévesque, J.-C., Nakamura, H., Kikuchi, Y., Satoh, M. S., Frenette, J., Sato, S. Galectin-3 and N-acetylglucosamine promote myogenesis and improve skeletal muscle function in the mdx model of Duchenne muscular dystrophy. PMID: 29894670 [PubMed - as supplied by publisher]
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Related Articles Recapitulating muscle disease phenotypes with myotonic dystrophy 1 induced pluripotent stem cells: a tool for disease modeling and drug discovery. Dis Model Mech. 2018 07 18;11(7): Authors: Mondragon-Gonzalez R, Perlingeiro RCR Abstract Myotonic dystrophy 1 (DM1) is a multisystem disorder primarily affecting the central nervous system, heart and skeletal muscle. It is caused by an expansion of the CTG trinucleotide repeats in the 3' untranslated region of the DMPK gene. Although patient myoblasts have been used for studying the disease in vitro, the invasiveness as well as the low accessibility to muscle biopsies motivate the development of alternative reliable myogenic models. Here, we established two DM1 induced pluripotent stem (iPS) cell lines from patient-derived fibroblasts and, using the PAX7 conditional expression system, differentiated these into myogenic progenitors and, subsequently, terminally differentiated myotubes. Both DM1 myogenic progenitors and myotubes were found to express the intranuclear RNA foci exhibiting sequestration of MBNL1. Moreover, we found the DM1-related mis-splicing, namely BIN1 exon 11 in DM1 myotubes. We used this model to test a specific therapy, antisense oligonucleotide treatment, and found that this efficiently abolished RNA foci and rescued BIN1 mis-splicing in DM1 iPS cell-derived myotubes. Together, our results demonstrate that myotubes derived from DM1 iPS cells recapitulate the critical molecular features of DM1 and are sensitive to antisense oligonucleotide treatment, confirming that these cells can be used for in vitro disease modeling and candidate drug testing or screening.This article has an associated First Person interview with the first author of the paper. PMID: 29898953 [PubMed - indexed for MEDLINE]
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Related Articles miR-23b and miR-218 silencing increase Muscleblind-like expression and alleviate myotonic dystrophy phenotypes in mammalian models. Nat Commun. 2018 06 26;9(1):2482 Authors: Cerro-Herreros E, Sabater-Arcis M, Fernandez-Costa JM, Moreno N, Perez-Alonso M, Llamusi B, Artero R Abstract Functional depletion of the alternative splicing factors Muscleblind-like (MBNL 1 and 2) is at the basis of the neuromuscular disease myotonic dystrophy type 1 (DM1). We previously showed the efficacy of miRNA downregulation in Drosophila DM1 model. Here, we screen for miRNAs that regulate MBNL1 and MBNL2 in HeLa cells. We thus identify miR-23b and miR-218, and confirm that they downregulate MBNL proteins in this cell line. Antagonists of miR-23b and miR-218 miRNAs enhance MBNL protein levels and rescue pathogenic missplicing events in DM1 myoblasts. Systemic delivery of these "antagomiRs" similarly boost MBNL expression and improve DM1-like phenotypes, including splicing alterations, histopathology, and myotonia in the HSALR DM1 model mice. These mammalian data provide evidence for therapeutic blocking of the miRNAs that control Muscleblind-like protein expression in myotonic dystrophy. PMID: 29946070 [PubMed - indexed for MEDLINE]
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Related Articles Human Adipose-Derived CD146+ Stem Cells Increase Life Span of a Muscular Dystrophy Mouse Model More Efficiently than Mesenchymal Stromal Cells. DNA Cell Biol. 2018 Sep;37(9):798-804 Authors: Gomes JP, Coatti GC, Valadares MC, Assoni AF, Pelatti MV, Secco M, Zatz M Abstract Duchenne muscular dystrophy is the most common and severe form of progressive muscular dystrophy. Previous results showed an increased survival in double knockout mice (dko) when treated with adipose-derived CD146+ cells. In this study, we analyzed the effect of CD146+ cells compared to mesenchymal stem/stromal cells (MSCs) derived from the same human adipose sample when injected in the dko mouse model without immunosuppression. Both CD146+ cells and MSCs increased the survival of treated mice when compared to vehicle-injected mice, with a more prominent effect of CD146+ cells than MSCs. Both CD146+ cells and MSCs suppressed peripheral blood mononuclear cell proliferation, indicating immunomodulatory properties. Co-culture experiments showed that MSCs have a more inflammatory profile expression, and angiogenesis assay showed that CD146+ cells can improve blood vessel formation. CD146+ cells can extend survival of muscular dystrophy mice more efficiently than MSCs, possibly due to immunomodulatory and angiogenic properties. Further investigations focusing on exogenous CD146+ cell role in vivo will improve cell therapy understanding and effectiveness. PMID: 30059260 [PubMed - indexed for MEDLINE]
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Related Articles Cardiovascular Precision Medicine in the Genomics Era. JACC Basic Transl Sci. 2018 Apr;3(2):313-326 Authors: Dainis AM, Ashley EA Abstract Precision medicine strives to delineate disease using multiple data sources-from genomics to digital health metrics-in order to be more precise and accurate in our diagnoses, definitions, and treatments of disease subtypes. By defining disease at a deeper level, we can treat patients based on an understanding of the molecular underpinnings of their presentations, rather than grouping patients into broad categories with one-size-fits-all treatments. In this review, the authors examine how precision medicine, specifically that surrounding genetic testing and genetic therapeutics, has begun to make strides in both common and rare cardiovascular diseases in the clinic and the laboratory, and how these advances are beginning to enable us to more effectively define risk, diagnose disease, and deliver therapeutics for each individual patient. PMID: 30062216 [PubMed]
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Related Articles Agent-based model illustrates the role of the microenvironment in regeneration in healthy and mdx skeletal muscle. J Appl Physiol (1985). 2018 Aug 02;: Authors: Virgilio KM, Martin KS, Peirce SM, Blemker SS Abstract Duchenne muscular dystrophy (DMD) is a progressive muscle wasting disease with no effective treatment. Multiple mechanisms are thought to contribute to muscle wasting, including increased susceptibility to contraction-induced damage, chronic inflammation, fibrosis, altered satellite stem cell (SSC) dynamics, and impaired regenerative capacity. The goals of this project were to: (i) develop an agent-based model of skeletal muscle that predicts the dynamic regenerative response of muscle cells, fibroblasts, SSCs, and inflammatory cells as a result of contraction-induced injury, (ii) calibrate and validate the model parameters based on comparison with published experimental measurements, and (iii) use the model to investigate how changing isolated and combined factors known to be associated with DMD (e.g. altered fibroblast or SSC behaviors) influence muscle regeneration. Our predictions revealed that the percent of injured muscle that recovered 28 days post injury was dependent on the peak SSC counts following injury. In simulations with near-full CSA recovery (healthy, 4 week mdx, 3 month mdx), the SSC counts correlated with the extent of initial injury; however, in simulations with impaired regeneration (9 month mdx), the peak SSC counts were suppressed relative to initial injury. The differences in SSC counts between these groups were emergent predictions dependent on altered microenvironment factors known to be associated with DMD. Multiple cell types influenced the peak number of SSCs, but no individual parameter predicted the differences in SSC counts. This finding suggests that interventions to target the microenvironment rather than SSCs directly, could be an effective method for improving regeneration in impaired muscle. PMID: 30070607 [PubMed - as supplied by publisher]
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Related Articles Muscle Stem Cell Immunostaining. Curr Protoc Mouse Biol. 2018 Sep;8(3):e47 Authors: Wang S, Zhang B, Addicks GC, Zhang H, J Menzies K, Zhang H Abstract Muscle stem cells (MuSCs) are essential for maintaining muscle homeostasis by providing progenitor cells for muscle regeneration after injury and in muscular diseases. MuSC properties dynamically change, reflecting physiology or pathological status. For instance, MuSCs are activated after muscle injury, but become exhausted in late stages of Duchenne Muscular Dystrophy (DMD) disease and senescent during aging. Therefore, characterization of MuSCs, including proliferation, activation, senescence, and apoptosis, etc., is very important in applying MuSC knowledge to regenerative medicine, such as in the treatment of DMD and to improve muscle function in aging. Here, we describe a detailed method for characterizing MuSCs in situ using immunostaining techniques in the mouse. This method can also be easily adapted to analyze other skeletal muscle properties. © 2018 by John Wiley & Sons, Inc. PMID: 30106515 [PubMed - indexed for MEDLINE]
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Related Articles Tbx1 regulates inherited metabolic and myogenic abilities of progenitor cells derived from slow- and fast-type muscle. Cell Death Differ. 2018 Aug 28;: Authors: Motohashi N, Uezumi A, Asakura A, Ikemoto-Uezumi M, Mori S, Mizunoe Y, Takashima R, Miyagoe-Suzuki Y, Takeda S, Shigemoto K Abstract Skeletal muscle is divided into slow- and fast-type muscles, which possess distinct contractile and metabolic properties. Myogenic progenitors associated with each muscle fiber type are known to intrinsically commit to specific muscle fiber lineage during embryonic development. However, it is still unclear whether the functionality of postnatal adult myogenic cells is attributable to the muscle fiber in which they reside, and whether the characteristics of myogenic cells derived from slow- and fast-type fibers can be distinguished at the genetic level. In this study, we isolated adult satellite cells from slow- and fast-type muscle individually and observed that satellite cells from each type of muscle generated myotubes expressing myosin heavy chain isoforms similar to their original muscle, and showed different metabolic features. Notably, we discovered that slow muscle-derived cells had low potential to differentiate but high potential to self-renew compared with fast muscle-derived cells. Additionally, cell transplantation experiments of slow muscle-derived cells into fast-type muscle revealed that slow muscle-derived cells could better contribute to myofiber formation and satellite cell constitution than fast muscle-derived cells, suggesting that the recipient muscle fiber type may not affect the predetermined abilities of myogenic cells. Gene expression analyses identified T-box transcriptional factor Tbx1 as a highly expressed gene in fast muscle-derived myoblasts. Gain- and loss-of-function experiments revealed that Tbx1 modulated muscle fiber types and oxidative metabolism in myotubes, and that Tbx1 stimulated myoblast differentiation, but did not regulate myogenic cell self-renewal. Our data suggest that metabolic and myogenic properties of myogenic progenitor cells vary depending on the type of muscle from which they originate, and that Tbx1 expression partially explains the functional differences of myogenic cells derived from fast-type and slow-type muscles. PMID: 30154444 [PubMed - as supplied by publisher]
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Related Articles Engineered DNA plasmid reduces immunity to dystrophin while improving muscle force in a model of gene therapy of Duchenne dystrophy. Proc Natl Acad Sci U S A. 2018 09 25;115(39):E9182-E9191 Authors: Ho PP, Lahey LJ, Mourkioti F, Kraft PE, Filareto A, Brandt M, Magnusson KEG, Finn EE, Chamberlain JS, Robinson WH, Blau HM, Steinman L Abstract In gene therapy for Duchenne muscular dystrophy there are two potential immunological obstacles. An individual with Duchenne muscular dystrophy has a genetic mutation in dystrophin, and therefore the wild-type protein is "foreign," and thus potentially immunogenic. The adeno-associated virus serotype-6 (AAV6) vector for delivery of dystrophin is a viral-derived vector with its own inherent immunogenicity. We have developed a technology where an engineered plasmid DNA is delivered to reduce autoimmunity. We have taken this approach into humans, tolerizing to myelin proteins in multiple sclerosis and to proinsulin in type 1 diabetes. Here, we extend this technology to a model of gene therapy to reduce the immunogenicity of the AAV vector and of the wild-type protein product that is missing in the genetic disease. Following gene therapy with systemic administration of recombinant AAV6-microdystrophin to mdx/mTRG2 mice, we demonstrated the development of antibodies targeting dystrophin and AAV6 capsid in control mice. Treatment with the engineered DNA construct encoding microdystrophin markedly reduced antibody responses to dystrophin and to AAV6. Muscle force in the treated mice was also improved compared with control mice. These data highlight the potential benefits of administration of an engineered DNA plasmid encoding the delivered protein to overcome critical barriers in gene therapy to achieve optimal functional gene expression. PMID: 30181272 [PubMed - indexed for MEDLINE]
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Related Articles Protein disulfide isomerase as a prosurvival factor in cell therapy for muscular and vascular diseases. Stem Cell Res Ther. 2018 Sep 26;9(1):250 Authors: Di Rocco G, Baldari S, Gentile A, Capogrossi M, Toietta G Abstract BACKGROUND: Cell therapy for degenerative diseases aims at rescuing tissue damage by delivery of precursor cells. Thus far, this strategy has been mostly unsuccessful due to massive loss of donor cells shortly after transplantation. Several strategies have been applied to increase transplanted cell survival but only with limited success. The endoplasmic reticulum (ER) is an organelle involved in protein folding, calcium homeostasis, and lipid biosynthesis. Protein disulfide isomerase (PDI) is a molecular chaperone induced and activated by ER stress. PDI is induced by hypoxia in neuronal, cardiac, and endothelial cells, supporting increased cell survival to hypoxic stress and protection from apoptosis in response to ischemia. METHODS: We achieved ex vivo PDI gene transfer into luciferase-expressing myoblasts and endothelial cells. We assessed cell engraftment upon intramuscular transplantation into a mouse model of Duchenne muscular dystrophy (mdx mouse) and into a mouse model of ischemic disease. RESULTS: We observed that loss of full-length dystrophin expression in mdx mice muscle leads to an increase of PDI expression, possibly in response to augmented ER protein folding load. Moreover, we determined that overexpression of PDI confers a survival advantage for muscle cells in vitro and in vivo to human myoblasts injected into murine dystrophic muscle and to endothelial cells administered upon hindlimb ischemia damage, improving the therapeutic outcome of the cell therapy treatment. CONCLUSIONS: Collectively, these results suggest that overexpression of PDI may protect transplanted cells from hypoxia and other possibly occurring ER stresses, and consequently enhance their regenerative properties. PMID: 30257707 [PubMed - in process]
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Related Articles Supplementation with a selective amino acid formula ameliorates muscular dystrophy in mdx mice. Sci Rep. 2018 Oct 02;8(1):14659 Authors: Banfi S, D'Antona G, Ruocco C, Meregalli M, Belicchi M, Bella P, Erratico S, Donato E, Rossi F, Bifari F, Lonati C, Campaner S, Nisoli E, Torrente Y Abstract Duchenne muscular dystrophy (DMD) is one of the most common and severe forms of muscular dystrophy. Oxidative myofibre content, muscle vasculature architecture and exercise tolerance are impaired in DMD. Several studies have demonstrated that nutrient supplements ameliorate dystrophic features, thereby enhancing muscle performance. Here, we report that dietary supplementation with a specific branched-chain amino acid-enriched mixture (BCAAem) increased the abundance of oxidative muscle fibres associated with increased muscle endurance in dystrophic mdx mice. Amelioration of the fatigue index in BCAAem-treated mdx mice was caused by a cascade of events in the muscle tissue, which were promoted by endothelial nitric oxide synthase (eNOS) activation and vascular endothelial growth factor (VEGF) expression. VEGF induction led to recruitment of bone marrow (BM)-derived endothelial progenitors (EPs), which increased the capillary density of dystrophic skeletal muscle. Functionally, BCAAem mitigated the dystrophic phenotype of mdx mice without inducing dystrophin protein expression or replacing the dystrophin-associated glycoprotein (DAG) complex in the membrane, which is typically lost in DMD. BCAAem supplementation could be an effective adjuvant strategy in DMD treatment. PMID: 30279586 [PubMed - in process]
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Related Articles Metabolomic Analyses Reveal Extensive Progenitor Cell Deficiencies in a Mouse Model of Duchenne Muscular Dystrophy. Metabolites. 2018 Oct 03;8(4): Authors: Joseph J, Cho DS, Doles JD Abstract Duchenne muscular dystrophy (DMD) is a musculoskeletal disorder that causes severe morbidity and reduced lifespan. Individuals with DMD have an X-linked mutation that impairs their ability to produce functional dystrophin protein in muscle. No cure exists for this disease and the few therapies that are available do not dramatically delay disease progression. Thus, there is a need to better understand the mechanisms underlying DMD which may ultimately lead to improved treatment options. The muscular dystrophy (MDX) mouse model is frequently used to explore DMD disease traits. Though some studies of metabolism in dystrophic mice exist, few have characterized metabolic profiles of supporting cells in the diseased environment. Using nontargeted metabolomics we characterized metabolic alterations in muscle satellite cells (SCs) and serum of MDX mice. Additionally, live-cell imaging revealed MDX-derived adipose progenitor cell (APC) defects. Finally, metabolomic studies revealed a striking elevation of acylcarnitines in MDX APCs, which we show can inhibit APC proliferation. Together, these studies highlight widespread metabolic alterations in multiple progenitor cell types and serum from MDX mice and implicate dystrophy-associated metabolite imbalances in APCs as a potential contributor to adipose tissue disequilibrium in DMD. PMID: 30282911 [PubMed]
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Related Articles Dusp6 is a genetic modifier of growth through enhanced ERK activity. Hum Mol Genet. 2019 Jan 15;28(2):279-289 Authors: Vo AH, Swaggart KA, Woo A, Gao QQ, Demonbreun AR, Fallon KS, Quattrocelli M, Hadhazy M, Page PGT, Chen Z, Eskin A, Squire K, Nelson SF, McNally EM Abstract Like other single-gene disorders, muscular dystrophy displays a range of phenotypic heterogeneity even with the same primary mutation. Identifying genetic modifiers capable of altering the course of muscular dystrophy is one approach to deciphering gene-gene interactions that can be exploited for therapy development. To this end, we used an intercross strategy in mice to map modifiers of muscular dystrophy. We interrogated genes of interest in an interval on mouse chromosome 10 associated with body mass in muscular dystrophy as skeletal muscle contributes significantly to total body mass. Using whole-genome sequencing of the two parental mouse strains combined with deep RNA sequencing, we identified the Met62Ile substitution in the dual-specificity phosphatase 6 (Dusp6) gene from the DBA/2 J (D2) mouse strain. DUSP6 is a broadly expressed dual-specificity phosphatase protein, which binds and dephosphorylates extracellular-signal-regulated kinase (ERK), leading to decreased ERK activity. We found that the Met62Ile substitution reduced the interaction between DUSP6 and ERK resulting in increased ERK phosphorylation and ERK activity. In dystrophic muscle, DUSP6 Met62Ile is strongly upregulated to counteract its reduced activity. We found that myoblasts from the D2 background were insensitive to a specific small molecule inhibitor of DUSP6, while myoblasts expressing the canonical DUSP6 displayed enhanced proliferation after exposure to DUSP6 inhibition. These data identify DUSP6 as an important regulator of ERK activity in the setting of muscle growth and muscular dystrophy. PMID: 30289454 [PubMed - in process]
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Related Articles Muscle Satellite Cell Cross-Talk with a Vascular Niche Maintains Quiescence via VEGF and Notch Signaling. Cell Stem Cell. 2018 Oct 04;23(4):530-543.e9 Authors: Verma M, Asakura Y, Murakonda BSR, Pengo T, Latroche C, Chazaud B, McLoon LK, Asakura A Abstract Skeletal muscle is a complex tissue containing tissue resident muscle stem cells (satellite cells) (MuSCs) important for postnatal muscle growth and regeneration. Quantitative analysis of the biological function of MuSCs and the molecular pathways responsible for a potential juxtavascular niche for MuSCs is currently lacking. We utilized fluorescent reporter mice and muscle tissue clearing to investigate the proximity of MuSCs to capillaries in 3 dimensions. We show that MuSCs express abundant VEGFA, which recruits endothelial cells (ECs) in vitro, whereas blocking VEGFA using both a vascular endothelial growth factor (VEGF) inhibitor and MuSC-specific VEGFA gene deletion reduces the proximity of MuSCs to capillaries. Importantly, this proximity to the blood vessels was associated with MuSC self-renewal in which the EC-derived Notch ligand Dll4 induces quiescence in MuSCs. We hypothesize that MuSCs recruit capillary ECs via VEGFA, and in return, ECs maintain MuSC quiescence though Dll4. PMID: 30290177 [PubMed - in process]
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Related Articles Orienting Muscle Stem Cells for Regeneration in Homeostasis, Aging, and Disease. Cell Stem Cell. 2018 Nov 01;23(5):653-664 Authors: Feige P, Brun CE, Ritso M, Rudnicki MA Abstract Muscle stem cells, or satellite cells, are required for skeletal muscle maintenance, growth, and repair. Following satellite cell activation, several factors drive asymmetric cell division to generate a stem cell and a proliferative progenitor that forms new muscle. The balance between symmetric self-renewal and asymmetric division significantly impacts the efficiency of regeneration. In this Review, we discuss the relationship of satellite cell heterogeneity and the establishment of polarity to asymmetric division, as well as how these processes are impacted in homeostasis, aging, and disease. We also highlight therapeutic opportunities for targeting satellite cell polarity and self-renewal to stimulate muscle regeneration. PMID: 30388423 [PubMed - in process]
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Related Articles Modeling Skeletal Muscle Laminopathies Using Human Induced Pluripotent Stem Cells Carrying Pathogenic LMNA Mutations. Front Physiol. 2018;9:1332 Authors: Steele-Stallard HB, Pinton L, Sarcar S, Ozdemir T, Maffioletti SM, Zammit PS, Tedesco FS Abstract Laminopathies are a clinically heterogeneous group of disorders caused by mutations in LMNA. The main proteins encoded by LMNA are Lamin A and C, which together with Lamin B1 and B2, form the nuclear lamina: a mesh-like structure located underneath the inner nuclear membrane. Laminopathies show striking tissue specificity, with subtypes affecting striated muscle, peripheral nerve, and adipose tissue, while others cause multisystem disease with accelerated aging. Although several pathogenic mechanisms have been proposed, the exact pathophysiology of laminopathies remains unclear, compounded by the rarity of these disorders and lack of easily accessible cell types to study. To overcome this limitation, we used induced pluripotent stem cells (iPSCs) from patients with skeletal muscle laminopathies such as LMNA-related congenital muscular dystrophy and limb-girdle muscular dystrophy 1B, to model disease phenotypes in vitro. iPSCs can be derived from readily accessible cell types, have unlimited proliferation potential and can be differentiated into cell types that would otherwise be difficult and invasive to obtain. iPSC lines from three skeletal muscle laminopathy patients were differentiated into inducible myogenic cells and myotubes. Disease-associated phenotypes were observed in these cells, including abnormal nuclear shape and mislocalization of nuclear lamina proteins. Nuclear abnormalities were less pronounced in monolayer cultures of terminally differentiated skeletal myotubes than in proliferating myogenic cells. Notably, skeletal myogenic differentiation of LMNA-mutant iPSCs in artificial muscle constructs improved detection of myonuclear abnormalities compared to conventional monolayer cultures across multiple pathogenic genotypes, providing a high-fidelity modeling platform for skeletal muscle laminopathies. Our results lay the foundation for future iPSC-based therapy development and screening platforms for skeletal muscle laminopathies. PMID: 30405424 [PubMed]
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Related Articles Fibrosis Rescue Improves Cardiac Function in Dystrophin-Deficient Mice and Duchenne Patient-Specific Cardiomyocytes by Immunoproteasome Modulation. Am J Pathol. 2018 Nov 16;: Authors: Farini A, Gowran A, Bella P, Sitzia C, Scopece A, Castiglioni E, Rovina D, Nigro P, Villa C, Fortunato F, Comi GP, Milano G, Pompilio G, Torrente Y Abstract Patients affected by Duchenne muscular dystrophy (DMD) develop a progressive dilated cardiomyopathy characterized by inflammatory cell infiltration, necrosis, and cardiac fibrosis. Standard treatments consider the use of β-blockers and angiotensin-converting enzyme inhibitors that are symptomatic and unspecific toward DMD disease. Medications that target DMD cardiac fibrosis are in the early stages of development. We found immunoproteasome dysregulation in affected hearts of mdx mice (murine animal model of DMD) and cardiomyocytes derived from induced pluripotent stem cells of patients with DMD. Interestingly, immunoproteasome inhibition ameliorated cardiomyopathy in mdx mice and reduced the development of cardiac fibrosis. Establishing the immunoproteasome inhibition-dependent cardioprotective role suggests the possibility of modulating the immunoproteasome as new and clinically relevant treatment to rescue dilated cardiomyopathy in patients with DMD. PMID: 30448404 [PubMed - as supplied by publisher]
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Related Articles The Danger Signal Extracellular ATP Is Involved in the Immunomediated Damage of α-Sarcoglycan-Deficient Muscular Dystrophy. Am J Pathol. 2018 Nov 16;: Authors: Gazzerro E, Baratto S, Assereto S, Baldassari S, Panicucci C, Raffaghello L, Scudieri P, De Battista D, Fiorillo C, Volpi S, Chaabane L, Malnati M, Messina G, Bruzzone S, Traggiai E, Grassi F, Minetti C, Bruno C Abstract In muscular dystrophies, muscle membrane fragility results in a tissue-specific increase of danger-associated molecular pattern molecules (DAMPs) and infiltration of inflammatory cells. The DAMP extracellular ATP (eATP) released by dying myofibers steadily activates muscle and immune purinergic receptors exerting dual negative effects: a direct damage linked to altered intracellular calcium homeostasis in muscle cells and an indirect toxicity through the triggering of the immune response and inhibition of regulatory T cells. Accordingly, pharmacologic and genetic inhibition of eATP signaling improves the phenotype in models of chronic inflammatory diseases. In α-sarcoglycanopathy, eATP effects may be further amplified because α-sarcoglycan extracellular domain binds eATP and displays an ecto-ATPase activity, thus controlling eATP concentration at the cell surface and attenuating the magnitude and/or the duration of eATP-induced signals. Herein, we show that in vivo blockade of the eATP/P2X purinergic pathway by a broad-spectrum P2XR-antagonist delayed the progression of the dystrophic phenotype in α-sarcoglycan-null mice. eATP blockade dampened the muscular inflammatory response and enhanced the recruitment of forkhead box protein P3-positive immunosuppressive regulatory CD4+ T cells. The improvement of the inflammatory features was associated with increased strength, reduced necrosis, and limited expression of profibrotic factors, suggesting that pharmacologic purinergic antagonism, altering the innate and adaptive immune component in muscle infiltrates, might provide a therapeutic approach to slow disease progression in α-sarcoglycanopathy. PMID: 30448410 [PubMed - as supplied by publisher]
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Related Articles Applications of CRISPR/Cas9 for the Treatment of Duchenne Muscular Dystrophy. J Pers Med. 2018 Nov 24;8(4): Authors: Lim KRQ, Yoon C, Yokota T Abstract 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. PMID: 30477208 [PubMed]
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Related Articles The Trithorax protein Ash1L promotes myoblast fusion by activating Cdon expression. Nat Commun. 2018 11 28;9(1):5026 Authors: Castiglioni I, Caccia R, Garcia-Manteiga JM, Ferri G, Caretti G, Molineris I, Nishioka K, Gabellini D Abstract Myoblast fusion (MF) is required for muscle growth and repair, and its alteration contributes to muscle diseases. The mechanisms governing this process are incompletely understood, and no epigenetic regulator has been previously described. Ash1L is an epigenetic activator belonging to the Trithorax group of proteins and is involved in FSHD muscular dystrophy, autism and cancer. Its physiological role in skeletal muscle is unknown. Here we report that Ash1L expression is positively correlated with MF and reduced in Duchenne muscular dystrophy. In vivo, ex vivo and in vitro experiments support a selective and evolutionary conserved requirement for Ash1L in MF. RNA- and ChIP-sequencing indicate that Ash1L is required to counteract Polycomb repressive activity to allow activation of selected myogenesis genes, in particular the key MF gene Cdon. Our results promote Ash1L as an important epigenetic regulator of MF and suggest that its activity could be targeted to improve cell therapy for muscle diseases. PMID: 30487570 [PubMed - indexed for MEDLINE]
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Related Articles Efficacy of stem cell therapy in ambulatory and nonambulatory children with Duchenne muscular dystrophy - Phase I-II. Degener Neurol Neuromuscul Dis. 2018;8:63-77 Authors: Dai A, Baspinar O, Yeşilyurt A, Sun E, Aydemir Çİ, Öztel ON, Capkan DU, Pinarli F, Agar A, Karaöz E Abstract Purpose: Duchenne muscular dystrophy (DMD) is an X-linked recessive pediatric disorder that ultimately leads to progressive muscle degeneration. It has been known that cell-based therapies were used to promote muscle regeneration. The main purpose of this study was to investigate the effects of allogeneic Wharton jelly-derived mesenchymal stem cells therapy in Duchenne muscular dystrophy. Patients and methods: Four ambulatory and five nonambulatory male patients were assessed as having acceptance criteria. Gene expression and immunohistochemical analysis were performed for dystrophin gene expression. The fluorescent in situ hybridization method was used for detection of chimerism and donor-recipient compatibility. Complement dependent lymphocytotoxic crossmatch test and detection of panel reactive antigen were performed. All patients were treated with 2 × 106 cells/kg dose of allogeneic Wharton jelly-derived mesenchymal stem cells via intra-arterial and intramuscular administration. Stability was maintained in patient follow-up tests, which are respiratory capacity tests, cardiac measurements, and muscle strength tests. Results: The vastus intermedius muscle was observed in one patient with MRI. Chimerism was detected by fluorescent in situ hybridization and mean gene expression was increased to 3.3-fold. An increase in muscle strength measurements and pulmonary function tests was detected. Additionally, we observed two of nine patients with positive panel reactive antigen result. Conclusion: All our procedures are well tolerated, and we have not seen any application-related complications so far. Our main purpose of this study was to investigate the effects of allogeneic mesenchymal stem cell therapy and determine its suitability and safety as a form of treatment in this untreatable disorder. PMID: 30498389 [PubMed]
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Related Articles Human iPSC Models to Study Orphan Diseases: Muscular Dystrophies. Curr Stem Cell Rep. 2018;4(4):299-309 Authors: Xia G, Terada N, Ashizawa T Abstract Purpose of Review: Muscular dystrophies (MDs) are a spectrum of muscle disorders, which are caused by a number of gene mutations. The studies of MDs are limited due to lack of appropriate models, except for Duchenne muscular dystrophy (DMD), myotonic dystrophy type 1 (DM1), facioscapulohumeral muscular dystrophy (FSHD), and certain type of limb-girdle muscular dystrophy (LGMD). Human induced pluripotent stem cell (iPSC) technologies are emerging to offer a useful model for mechanistic studies, drug discovery, and cell-based therapy to supplement in vivo animal models. This review will focus on current applications of iPSC as disease models of MDs for studies of pathogenic mechanisms and therapeutic development. Recent Findings: Many and more human disease-specific iPSCs have been or being established, which carry the natural mutation of MDs with human genomic background. These iPSCs can be differentiated into specific cell types affected in a particular MDs such as skeletal muscle progenitor cells, skeletal muscle fibers, and cardiomyocytes. Human iPSCs are particularly useful for studies of the pathogenicity at the early stage or developmental phase of MDs. High-throughput screening using disease-specific human iPSCs has become a powerful technology in drug discovery. While MD iPSCs have been generated for cell-based replacement therapy, recent advances in genome editing technologies enabled correction of genetic mutations in these cells in culture, raising hope for in vivo genome therapy, which offers a fundamental cure for these daunting inherited MDs. Summary: Human disease-specific iPSC models for MDs are emerging as an additional tool to current disease models for elucidating disease mechanisms and developing therapeutic intervention. PMID: 30524939 [PubMed]
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Related Articles Induced Pluripotent Stem Cells for Duchenne Muscular Dystrophy Modeling and Therapy. Cells. 2018 Dec 07;7(12): Authors: Danisovic L, Culenova M, Csobonyeiova M Abstract Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder, caused by mutation of the DMD gene which encodes the protein dystrophin. This dystrophin defect leads to the progressive degeneration of skeletal and cardiac muscles. Currently, there is no effective therapy for this disorder. However, the technology of cell reprogramming, with subsequent controlled differentiation to skeletal muscle cells or cardiomyocytes, may provide a unique tool for the study, modeling, and treatment of Duchenne muscular dystrophy. In the present review, we describe current methods of induced pluripotent stem cell generation and discuss their implications for the study, modeling, and development of cell-based therapies for Duchenne muscular dystrophy. PMID: 30544588 [PubMed]
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Related Articles The multi-functional roles of menstrual blood-derived stem cells in regenerative medicine. Stem Cell Res Ther. 2019 Jan 03;10(1):1 Authors: Chen L, Qu J, Xiang C Abstract Menstrual blood-derived stem cells (MenSCs) are a novel source of mesenchymal stem cells (MSCs). MenSCs are attracting more and more attention since their discovery in 2007. MenSCs also have no moral dilemma and show some unique features of known adult-derived stem cells, which provide an alternative source for the research and application in regenerative medicine. Currently, people are increasingly interested in their clinical potential due to their high proliferation, remarkable versatility, and periodic acquisition in a non-invasive manner with no other sources of MSCs that are comparable in adult tissue. In this review, the plasticity of pluripotent biological characteristics, immunophenotype and function, differentiative potential, and immunomodulatory properties are assessed. Furthermore, we also summarize their therapeutic effects and functional characteristics in various diseases, including liver disease, diabetes, stroke, Duchenne muscular dystrophy, ovarian-related disease, myocardial infarction, Asherman syndrome, Alzheimer's disease, acute lung injury, cutaneous wound, endometriosis, and neurodegenerative diseases. Subsequently, the clinical potential of MenSCs is investigated. There is a need for a deeper understanding of its immunomodulatory and diagnostic properties with safety concern on a variety of environmental conditions (such as epidemiological backgrounds, age, hormonal status, and pre-contraceptive). In summary, MenSC has a great potential for reducing mortality and improving the quality of life of severe patients. As a kind of adult stem cells, MenSCs have multiple properties in treating a variety of diseases in regenerative medicine for future clinical applications. PMID: 30606242 [PubMed - in process]
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Related Articles iPSCs as a Platform for Disease Modeling, Drug Screening, and Personalized Therapy in Muscular Dystrophies. Cells. 2019 Jan 03;8(1): Authors: Ortiz-Vitali JL, Darabi R Abstract Induced pluripotent stem cells (iPSCs) are the foundation of modern stem cell-based regenerative medicine, especially in the case of degenerative disorders, such as muscular dystrophies (MDs). Since their introduction in 2006, many studies have used iPSCs for disease modeling and identification of involved mechanisms, drug screening, as well as gene correction studies. In the case of muscular dystrophies, these studies commenced in 2008 and continue to address important issues, such as defining the main pathologic mechanisms in different types of MDs, drug screening to improve skeletal/cardiac muscle cell survival and to slow down disease progression, and evaluation of the efficiency of different gene correction approaches, such as exon skipping, Transcription activator-like effector nucleases (TALENs), Zinc finger nucleases (ZFNs) and RNA-guided endonuclease Cas9 (CRISPR/Cas9). In the current short review, we have summarized chronological progress of these studies and their key findings along with a perspective on the future road to successful iPSC-based cell therapy for MDs and the potential hurdles in this field. PMID: 30609814 [PubMed]
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Related Articles Restoring Dystrophin Expression in Duchenne Muscular Dystrophy: Current Status of Therapeutic Approaches. J Pers Med. 2019 Jan 07;9(1): Authors: Shimizu-Motohashi Y, Komaki H, Motohashi N, Takeda S, Yokota T, Aoki Y Abstract Duchenne muscular dystrophy (DMD), a rare genetic disorder characterized by progressive muscle weakness, is caused by the absence or a decreased amount of the muscle cytoskeletal protein dystrophin. Currently, several therapeutic approaches to cure DMD are being investigated, which can be categorized into two groups: therapies that aim to restore dystrophin expression, and those that aim to compensate for the lack of dystrophin. Therapies that restore dystrophin expression include read-through therapy, exon skipping, vector-mediated gene therapy, and cell therapy. Of these approaches, the most advanced are the read-through and exon skipping therapies. In 2014, ataluren, a drug that can promote ribosomal read-through of mRNA containing a premature stop codon, was conditionally approved in Europe. In 2016, eteplirsen, a morpholino-based chemical capable of skipping exon 51 in premature mRNA, received conditional approval in the USA. Clinical trials on vector-mediated gene therapy carrying micro- and mini- dystrophin are underway. More innovative therapeutic approaches include CRISPR/Cas9-based genome editing and stem cell-based cell therapies. Here we review the current status of therapeutic approaches for DMD, focusing on therapeutic approaches that can restore dystrophin. PMID: 30621068 [PubMed]
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