The Purine Nucleotide Profile in Mouse, Chicken and Human Dystrophic Muscle: An Abnormal Ratio of Inosine plus Adenine Nucleotides to Guanine Nucleotides

1982 ◽  
Vol 62 (1) ◽  
pp. 113-115 ◽  
Author(s):  
R. J. Shuttlewood ◽  
J. R. Griffiths
Keyword(s):  
High Pressure ◽  
Liquid Chromatography ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Adenine Nucleotide ◽  
Adenine Nucleotides ◽  
Purine Nucleotide ◽  
Dystrophic Muscle ◽  
Human Patient ◽  
Abnormal Ratio

1. AMP, ADP, ATP, IMP, GDP, GTP and adenylosuccinate have been measured by high pressure liquid chromatography in three types of animal muscular dystrophy and in a human patient with Duchenne muscular dystrophy. 2. Abnormalities in nucleotide content varied from one dystrophy to another. 3. In each case, however, the ratio [total adenine nucleotide + IMP]/[total guanine nuclotides] was lower in dystrophic muscle, even when severely exercised or ischaemic muscles were used. 4. The practical advantages of this assay for diagnosis of muscular dystrophy are discussed.

scholarly journals Degenerative and regenerative pathways underlying Duchenne muscular dystrophy revealed by single-nucleus RNA sequencing

2020 ◽  
Vol 117 (47) ◽  
pp. 29691-29701 ◽  
Author(s):  
Francesco Chemello ◽  
Zhaoning Wang ◽  
Hui Li ◽  
John R. McAnally ◽  
Ning Liu ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Dystrophin Gene ◽  
Cell Types ◽  
Muscle Pathology ◽  
Dystrophic Muscle ◽  
Nonmuscle Cell ◽  
Prevalent Disease ◽  
Single Nucleus ◽  
Muscle Disorder

Duchenne muscular dystrophy (DMD) is a fatal muscle disorder characterized by cycles of degeneration and regeneration of multinucleated myofibers and pathological activation of a variety of other muscle-associated cell types. The extent to which different nuclei within the shared cytoplasm of a myofiber may display transcriptional diversity and whether individual nuclei within a multinucleated myofiber might respond differentially to DMD pathogenesis is unknown. Similarly, the potential transcriptional diversity among nonmuscle cell types within dystrophic muscle has not been explored. Here, we describe the creation of a mouse model of DMD caused by deletion of exon 51 of the dystrophin gene, which represents a prevalent disease-causing mutation in humans. To understand the transcriptional abnormalities and heterogeneity associated with myofiber nuclei, as well as other mononucleated cell types that contribute to the muscle pathology associated with DMD, we performed single-nucleus transcriptomics of skeletal muscle of mice with dystrophin exon 51 deletion. Our results reveal distinctive and previously unrecognized myonuclear subtypes within dystrophic myofibers and uncover degenerative and regenerative transcriptional pathways underlying DMD pathogenesis. Our findings provide insights into the molecular underpinnings of DMD, controlled by the transcriptional activity of different types of muscle and nonmuscle nuclei.

scholarly journals Mutation-independent Proteomic Signatures of Pathological Progression in Murine Models of Duchenne Muscular Dystrophy

2020 ◽  
Vol 19 (12) ◽  
pp. 2047-2067
Author(s):  
Tirsa L. E. van Westering ◽  
Henrik J. Johansson ◽  
Britt Hanson ◽  
Anna M. L. Coenen-Stass ◽  
Yulia Lomonosova ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Muscle Growth ◽  
Differentially Expressed ◽  
Murine Models ◽  
Dystrophic Muscle ◽  
C2c12 Myoblasts ◽  
Experimental Therapies ◽  
Dystrophin Protein ◽  
Upstream Regulators

The absence of the dystrophin protein in Duchenne muscular dystrophy (DMD) results in myofiber fragility and a plethora of downstream secondary pathologies. Although a variety of experimental therapies are in development, achieving effective treatments for DMD remains exceptionally challenging, not least because the pathological consequences of dystrophin loss are incompletely understood. Here we have performed proteome profiling in tibialis anterior muscles from two murine DMD models (mdx and mdx52) at three ages (8, 16, and 80 weeks of age), all n = 3. High-resolution isoelectric focusing liquid chromatography-tandem MS (HiRIEF-LC–MS/MS) was used to quantify the expression of 4974 proteins across all 27 samples. The two dystrophic models were found to be highly similar, whereas multiple proteins were differentially expressed relative to WT (C57BL/6) controls at each age. Furthermore, 1795 proteins were differentially expressed when samples were pooled across ages and dystrophic strains. These included numerous proteins associated with the extracellular matrix and muscle function that have not been reported previously. Pathway analysis revealed multiple perturbed pathways and predicted upstream regulators, which together are indicative of cross-talk between inflammatory, metabolic, and muscle growth pathways (e.g. TNF, INFγ, NF-κB, SIRT1, AMPK, PGC-1α, PPARs, ILK, and AKT/PI3K). Upregulation of CAV3, MVP and PAK1 protein expression was validated in dystrophic muscle by Western blot. Furthermore, MVP was upregulated during, but not required for, the differentiation of C2C12 myoblasts suggesting that this protein may affect muscle regeneration. This study provides novel insights into mutation-independent proteomic signatures characteristic of the dystrophic phenotype and its progression with aging.

scholarly journals Dependence of mitochondrial and cytosolic adenine nucleotides on oxygen partial pressure in isolated hepatocytes. Application of a new rapid high pressure filtration technique for fractionation

1988 ◽  
Vol 250 (3) ◽  
pp. 641-645 ◽  
Author(s):  
H Hummerich ◽  
H de Groot ◽  
T Noll ◽  
S Soboll
Keyword(s):  
High Pressure ◽  
Oxygen Partial Pressure ◽  
Partial Pressure ◽  
Adenine Nucleotide ◽  
Intact Cell ◽  
Adenine Nucleotides ◽  
Strong Dependence ◽  
Isolated Hepatocytes ◽  
Pressure Filtration ◽  
Filtration Technique

By using a new rapid high pressure filtration technique, mitochondrial and cytosolic ATP and ADP contents were determined in isolated hepatocytes at different oxygen partial pressures. At 670 mmHg, subcellular adenine nucleotide contents and ATP/ADP ratios were comparable with values obtained with the digitonin fractionation technique. However at lower oxygen partial pressure ADP appears to be rephosphorylated during digitonin fractionation whereas with high pressure filtration fractionation rephosphorylation of ADP is avoided due to shorter fractionation times. Cytosolic and mitochondrial ATP/ADP ratios decrease if oxygen partial pressure is lowered. However the absolute values of ATP/ADP ratios depend critically on the incubation conditions. Thus incubation of hepatocytes in an oxystat system, where oxygen partial pressure is maintained constant by infusing oxygen-saturated medium and the hepatocyte suspension is continuously stirred, yields much higher subcellular and overall ATP/ADP ratios than incubation in Erlenmeyer flasks gassed with different gas mixtures and shaken in a water bath. This is ascribed to limited diffusion of oxygen from the medium into the cell if the suspension is not mixed thoroughly by stirring. The strong dependence of subcellular ATP/ADP ratios on incubation conditions indicates that oxygen may be one rate-controlling factor for oxidative phosphorylation in the intact cell.

scholarly journals Function and Genetics of Dystrophin and Dystrophin-Related Proteins in Muscle

2002 ◽  
Vol 82 (2) ◽  
pp. 291-329 ◽  
Author(s):  
Derek J. Blake ◽  
Andrew Weir ◽  
Sarah E. Newey ◽  
Kay E. Davies
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Protein Complex ◽  
Muscle Disease ◽  
Calcium Handling ◽  
Dystrophic Muscle ◽  
Gene Encoding ◽  
Muscle Wasting Disease ◽  
Novel Therapeutic Strategies ◽  
Related Proteins

The X-linked muscle-wasting disease Duchenne muscular dystrophy is caused by mutations in the gene encoding dystrophin. There is currently no effective treatment for the disease; however, the complex molecular pathology of this disorder is now being unravelled. Dystrophin is located at the muscle sarcolemma in a membrane-spanning protein complex that connects the cytoskeleton to the basal lamina. Mutations in many components of the dystrophin protein complex cause other forms of autosomally inherited muscular dystrophy, indicating the importance of this complex in normal muscle function. Although the precise function of dystrophin is unknown, the lack of protein causes membrane destabilization and the activation of multiple pathophysiological processes, many of which converge on alterations in intracellular calcium handling. Dystrophin is also the prototype of a family of dystrophin-related proteins, many of which are found in muscle. This family includes utrophin and α-dystrobrevin, which are involved in the maintenance of the neuromuscular junction architecture and in muscle homeostasis. New insights into the pathophysiology of dystrophic muscle, the identification of compensating proteins, and the discovery of new binding partners are paving the way for novel therapeutic strategies to treat this fatal muscle disease. This review discusses the role of the dystrophin complex and protein family in muscle and describes the physiological processes that are affected in Duchenne muscular dystrophy.

scholarly journals Emerging strategies for cell and gene therapy of the muscular dystrophies

2009 ◽  
Vol 11 ◽  
Author(s):  
Lindsey A. Muir ◽  
Jeffrey S. Chamberlain
Keyword(s):  
Gene Therapy ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Myotonic Dystrophy ◽  
Trinucleotide Repeat ◽  
Chromosome 1 ◽  
Muscular Dystrophies ◽  
Dystrophic Muscle ◽  
Kinase Gene ◽  
Wide Range

The muscular dystrophies are a heterogeneous group of over 40 disorders that are characterised by muscle weakness and wasting. The most common are Duchenne muscular dystrophy and Becker muscular dystrophy, which result from mutations within the gene encoding dystrophin; myotonic dystrophy type 1, which results from an expanded trinucleotide repeat in the myotonic dystrophy protein kinase gene; and facioscapulohumeral dystrophy, which is associated with contractions in the subtelomeric region of human chromosome 1. Currently the only treatments involve clinical management of symptoms, although several promising experimental strategies are emerging. These include gene therapy using adeno-associated viral, lentiviral and adenoviral vectors and nonviral vectors, such as plasmid DNA. Exon-skipping and cell-based therapies have also shown promise in the effective treatment and regeneration of dystrophic muscle. The availability of numerous animal models for Duchenne muscular dystrophy has enabled extensive testing of a wide range of therapeutic approaches for this type of disorder. Consequently, we focus here on the therapeutic developments for Duchenne muscular dystrophy as a model of the types of approaches being considered for various types of dystrophy. We discuss the advantages and limitations of each therapeutic strategy, as well as prospects and recent successes in the context of future clinical applications.

scholarly journals Adipose Tissue-Derived Stem Cell Secreted IGF-1 Protects Myoblasts from the Negative Effect of Myostatin

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Sebastian Gehmert ◽  
Carina Wenzel ◽  
Markus Loibl ◽  
Gero Brockhoff ◽  
Michaela Huber ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Stem Cell ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Protective Effect ◽  
Conditioned Medium ◽  
Therapeutic Option ◽  
Dystrophic Muscle ◽  
Rnai Technology ◽  
Myod Expression

Myostatin, a TGF-βfamily member, is associated with inhibition of muscle growth and differentiation and might interact with the IGF-1 signaling pathway. Since IGF-1 is secreted at a bioactive level by adipose tissue-derived mesenchymal stem cells (ASCs), these cells (ASCs) provide a therapeutic option for Duchenne Muscular Dystrophy (DMD). But the protective effect of stem cell secreted IGF-1 on myoblast under high level of myostatin remains unclear. In the present study murine myoblasts were exposed to myostatin under presence of ASCs conditioned medium and investigated for proliferation and apoptosis. The protective effect of IGF-1 was further examined by using IGF-1 neutralizing and receptor antibodies as well as gene silencing RNAi technology. MyoD expression was detected to identify impact of IGF-1 on myoblasts differentiation when exposed to myostatin. IGF-1 was accountable for 43.6% of the antiapoptotic impact and 48.8% for the proliferative effect of ASCs conditioned medium. Furthermore, IGF-1 restored mRNA and protein MyoD expression of myoblasts under risk. Beside fusion and transdifferentiation the beneficial effect of ASCs is mediated by paracrine secreted cytokines, particularly IGF-1. The present study underlines the potential of ASCs as a therapeutic option for Duchenne muscular dystrophy and other dystrophic muscle diseases.

scholarly journals BETs inhibition attenuates oxidative stress and preserves muscle integrity in Duchenne muscular dystrophy

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Marco Segatto ◽  
Roberta Szokoll ◽  
Raffaella Fittipaldi ◽  
Cinzia Bottino ◽  
Lorenzo Nevi ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Skeletal Muscle ◽  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Mdx Mouse ◽  
Early Event ◽  
Dystrophic Muscle ◽  
Protein Levels ◽  
Bet Inhibitors ◽  
Ros Metabolism

AbstractDuchenne muscular dystrophy (DMD) affects 1 in 3500 live male births. To date, there is no effective cure for DMD, and the identification of novel molecular targets involved in disease progression is important to design more effective treatments and therapies to alleviate DMD symptoms. Here, we show that protein levels of the Bromodomain and extra-terminal domain (BET) protein BRD4 are significantly increased in the muscle of the mouse model of DMD, the mdx mouse, and that pharmacological inhibition of the BET proteins has a beneficial outcome, tempering oxidative stress and muscle damage. Alterations in reactive oxygen species (ROS) metabolism are an early event in DMD onset and they are tightly linked to inflammation, fibrosis, and necrosis in skeletal muscle. By restoring ROS metabolism, BET inhibition ameliorates these hallmarks of the dystrophic muscle, translating to a beneficial effect on muscle function. BRD4 direct association to chromatin regulatory regions of the NADPH oxidase subunits increases in the mdx muscle and JQ1 administration reduces BRD4 and BRD2 recruitment at these regions. JQ1 treatment reduces NADPH subunit transcript levels in mdx muscles, isolated myofibers and DMD immortalized myoblasts. Our data highlight novel functions of the BET proteins in dystrophic skeletal muscle and suggest that BET inhibitors may ameliorate the pathophysiology of DMD.

scholarly journals A human in vitro model of Duchenne muscular dystrophy muscle formation and contractility

2016 ◽  
Vol 215 (1) ◽  
pp. 47-56 ◽  
Author(s):  
Alexander P. Nesmith ◽  
Matthew A. Wagner ◽  
Francesco S. Pasqualini ◽  
Blakely B. O’Connor ◽  
Mark J. Pincus ◽  
...  
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Dystrophic Muscle ◽  
Muscle Stem Cells ◽  
Functional Behavior ◽  
Muscle Tissues ◽  
Muscle Formation ◽  
Structural Aberrations ◽  
Vitro Model

Tongue weakness, like all weakness in Duchenne muscular dystrophy (DMD), occurs as a result of contraction-induced muscle damage and deficient muscular repair. Although membrane fragility is known to potentiate injury in DMD, whether muscle stem cells are implicated in deficient muscular repair remains unclear. We hypothesized that DMD myoblasts are less sensitive to cues in the extracellular matrix designed to potentiate structure–function relationships of healthy muscle. To test this hypothesis, we drew inspiration from the tongue and engineered contractile human muscle tissues on thin films. On this platform, DMD myoblasts formed fewer and smaller myotubes and exhibited impaired polarization of the cell nucleus and contractile cytoskeleton when compared with healthy cells. These structural aberrations were reflected in their functional behavior, as engineered tongues from DMD myoblasts failed to achieve the same contractile strength as healthy tongue structures. These data suggest that dystrophic muscle may fail to organize with respect to extracellular cues necessary to potentiate adaptive growth and remodeling.

scholarly journals Mechanisms and Clinical Applications of Glucocorticoid Steroids in Muscular Dystrophy

2021 ◽  
Vol 8 (1) ◽  
pp. 39-52
Author(s):  
Mattia Quattrocelli ◽  
Aaron S. Zelikovich ◽  
Isabella M. Salamone ◽  
Julie A. Fischer ◽  
Elizabeth M. McNally
Keyword(s):  
Duchenne Muscular Dystrophy ◽  
Muscular Dystrophy ◽  
Bone Health ◽  
Chronic Conditions ◽  
Clinical Applications ◽  
Dystrophic Muscle ◽  
Immunomodulatory Agents ◽  
Growth Impairment ◽  
Dosing Strategies ◽  
Glucocorticoid Action

Glucocorticoid steroids are widely used as immunomodulatory agents in acute and chronic conditions. Glucocorticoid steroids such as prednisone and deflazacort are recommended for treating Duchenne Muscular Dystrophy where their use prolongs ambulation and life expectancy. Despite this benefit, glucocorticoid use in Duchenne Muscular Dystrophy is also associated with significant adverse consequences including adrenal suppression, growth impairment, poor bone health and metabolic syndrome. For other forms of muscular dystrophy like the limb girdle dystrophies, glucocorticoids are not typically used. Here we review the experimental evidence supporting multiple mechanisms of glucocorticoid action in dystrophic muscle including their role in dampening inflammation and myofiber injury. We also discuss alternative dosing strategies as well as novel steroid agents that are in development and testing, with the goal to reduce adverse consequences of prolonged glucocorticoid exposure while maximizing beneficial outcomes.

scholarly journals Measurement of Nucleotide Pools in Platelets and Red Blood Cells Using High Pressure Liquid Chromatography

1977 ◽  
Author(s):  
L. D’Souza ◽  
H. I. Glueck
Keyword(s):  
High Pressure ◽  
Platelet Aggregation ◽  
Adenine Nucleotide ◽  
Platelet Rich Plasma ◽  
Adenine Nucleotides ◽  
Single Step ◽  
Small Sample ◽  
Normal Donor ◽  
Chromatographic Technique ◽  
Nucleotide Pools

Release of adenine nucleotides during platelet aggregation is well established. The released adenosine diphosphate (ADP) is thought to cause secondary platelet aggregation. A high pressure liquid chromatographic technique has been used to measure accurately the various nucleotide pools in intact platelets, as well as their levels during various stages of aggregation. Accurate and highly reproducible measurements of 10-12 moles of all nucleotides are possible using the bonded microparticulate, Partisil −10 SAX column, a strong anion exchanger (Whatman Inc., Clifton, N. J.) and an electronic integrator. The method is a modification of an earlier method of P. Brown (1973), and G. Rao et al (1975). Using this technique we are able to obtain within 75 minutes a single step gradient separation of cyclic nucleotides (cAMP, cCMP, cGMP) as well as mono-, di-, and tri-phosphonucleosides. Sample sizes of 0.5 ml of platelet rich plasma are adequate for at least 3 estimations. Normal donor platelets show the following adenine nucleotide levels;* AMP 0.13 (0.14); ADP 1.64 (0.81); and ATP 4.90 (0.42) nanomoles per 1011 platelets. The method has certain advantages over the enzymatic or luciferase assays, since all the nucleotides can be simultaneously measured in a single step from a very small sample and there is no cross reactivity with any of the nucleotides. Similar studies of plasma and red blood cell nucleotides are possible with this technique.*Mean (standard deviation)

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