Dystrophin In Skeletal Muscle Research Articles

Respiratory performance in Duchenne muscular dystrophy: Clinical manifestations and lessons from animal models.

Duchenne muscular dystrophy (DMD) is a fatal genetic neuromuscular disease. Lack of dystrophin in skeletal muscles leads to intrinsic weakness, injury, subsequent degeneration and fibrosis, decreasing contractile function. Dystropathology eventually presents in all inspiratory and expiratory muscles of breathing, severely curtailing their critical function. In people with DMD, premature death is caused by respiratory or cardiac failure. There is an urgent need to develop therapies that improve quality of life and extend life expectancy in DMD. Surprisingly, there is a dearth of information on respiratory control in animal models of DMD, and respiratory outcome measures are often limited or absent in clinical trials. Characterization of respiratory performance in murine and canine models has revealed extensive remodelling of the diaphragm, the major muscle of inspiration. However, significant compensation by extradiaphragmatic muscles of breathing is evident in early disease, contributing to preservation of peak respiratory system performance. Loss of compensation afforded by accessory muscles in advanced disease is ultimately associated with compromised respiratory performance. A new and potentially more translatable murine model of DMD, the D2.mdx mouse, has recently been developed. Respiratory performance in D2.mdx mice is yet to be characterized fully. However, based on histopathological features, D2.mdx mice might serve as useful preclinical models, facilitating the testing of new therapeutics that rescue respiratory function. This review summarizes the pathophysiological mechanisms associated with DMD both in humans and in animal models, with a focus on breathing. We consider the translational value of each model to human DMD and highlight the urgent need for comprehensive characterization of breathing in representative preclinical models to better inform human trials.

Increase in cathepsin K gene expression in Duchenne muscular dystrophy skeletal muscle.

Dystrophinopathy is caused by alterations in the dystrophin gene. The severe phenotype, Duchenne muscular dystrophy (DMD), is caused by a lack of dystrophin in skeletal muscles, resulting in necrosis and regenerating fibers, inflammatory cells, and muscle fibrosis. Progressive muscle weakness is a characteristic finding of this condition. Here, we encountered a rare case of a 10-year-old patient with asymptomatic dystrophinopathy with no dystrophin expression and investigated the reason for the absence of muscle weakness to obtain therapeutic insights for DMD. Using RNA-seq analysis, gene expression in skeletal muscles was compared among patients with asymptomatic dystrophinopathy, three patients with typical DMD, and two patients without dystrophinopathy who were leading normal daily lives. Cathepsin K (CTSK), myosin heavy chain 3 (MYH3), and nodal modulator 3-like genes exhibited a >8-fold change, whereas crystallin mu gene (CRYM) showed a <1/8-fold change in patients with typical DMD compared with their expression in the patient with asymptomatic dystrophinopathy. Additionally, CTSK and MYH3 expression exhibited a >16-fold change (P < 0.01), whereas CRYM expression showed a <1/16-fold change (P < 0.01) in patients with typical DMD compared with their expression in those without dystrophinopathy. CTSK plays an essential role in skeletal muscle loss, fibrosis, and inflammation in response to muscles injected with cardiotoxin, one of the most common reagents that induce muscle injury. Increased CTSK expression is associated with muscle injury or necrosis in patients with DMD. The lack of muscle weakness in the patient with asymptomatic dystrophinopathy might be attributed to the low CTSK expression in the muscles. To the best of our knowledge, this is the first report to demonstrate that CTSK expression was significantly higher in the skeletal muscles of patients with DMD with a typical phenotype than in those without dystrophinopathy.

Multiomic characterization of disease progression in mice lacking dystrophin.

Duchenne muscular dystrophy (DMD) is caused by genetic mutations leading to lack of dystrophin in skeletal muscle. A better understanding of how objective biomarkers for DMD vary across subjects and over time is needed to model disease progression and response to therapy more effectively, both in pre-clinical and clinical research. We present an in-depth characterization of disease progression in 3 murine models of DMD by multiomic analysis of longitudinal trajectories between 6 and 30 weeks of age. Integration of RNA-seq, mass spectrometry-based metabolomic and lipidomic data obtained in muscle and blood samples by Multi-Omics Factor Analysis (MOFA) led to the identification of 8 latent factors that explained 78.8% of the variance in the multiomic dataset. Latent factors could discriminate dystrophic and healthy mice, as well as different time-points. MOFA enabled to connect the gene expression signature in dystrophic muscles, characterized by pro-fibrotic and energy metabolism alterations, to inflammation and lipid signatures in blood. Our results show that omic observations in blood can be directly related to skeletal muscle pathology in dystrophic muscle.

T Cell Responses to Dystrophin in a Natural History Study of Duchenne Muscular Dystrophy.

Duchenne muscular dystrophy (DMD) is caused by the lack of dystrophin, but many patients have rare revertant fibers that express dystrophin. The skeletal muscle pathology of DMD patients includes immune cell infiltration and inflammatory cascades. There are several strategies to restore dystrophin in skeletal muscles of patients, including exon skipping and gene therapy. There is some evidence that dystrophin restoration leads to a reduction in immune cells, but dystrophin epitopes expressed in revertant fibers or following genome editing, cell therapy, or microdystrophin delivery after adeno-associated viral gene therapy may elicit T cell production in patients. This may affect the efficacy of the therapeutic intervention, and potentially lead to serious adverse events. To confirm and extend previous studies, we performed annual enzyme- linked immunospot interferon-gamma assays on peripheral blood mononuclear cells from 77 pediatric boys with DMD recruited into a natural history study, 69 of whom (89.6%) were treated with corticosteroids. T cell responses to dystrophin were quantified using a total of 368 peptides spanning the entire dystrophin protein, organized into nine peptide pools. Peptide mapping pools were used to further localize the immune response in one positive patient. Six (7.8%) patients had a T cell-mediated immune response to dystrophin at at least one time point. All patients who had a positive result had been treated with corticosteroids, either prednisolone or prednisone. Our results show that ∼8% of DMD individuals in our cohort have a pre-existing T cell-mediated immune response to dystrophin, despite steroid treatment. Although these responses are relatively low level, this information should be considered a useful immunological baseline before undertaking clinical trials and future DMD studies. We further highlight the importance for a robust, reproducible standard operating procedure for collecting, storing, and shipping samples from multiple centers to minimize the number of inconclusive data.

Good news for the mdx mouse community: Improved dystrophin restoration after skipping mouse dystrophin exon 23

Good news for the mdx mouse community: Improved dystrophin restoration after skipping mouse dystrophin exon 23

Dystrophin Restoration after Adeno-Associated Virus U7–Mediated Dmd Exon Skipping Is Modulated by Muscular Exercise in the Severe D2-Mdx Duchenne Muscular Dystrophy Murine Model

Duchenne muscular dystrophy (DMD) is a severe neuromuscular disease caused by Dmd mutations, resulting in the absence of dystrophin in skeletal muscle, with a greater susceptibility to damage during contraction (exercise). In this study, we evaluated whether voluntary exercise impacts a Dmd exon skipping approach and muscle physiology in a severe DMD murine model. We intramuscularly injected an adeno-associated virus (AAV) U7 snRNA that aimed to correct Dmd reading frame in D2-mdx mice, and they voluntary ran in a wheel for 1 month. Our results suggested that voluntary running did not induce muscle fiber regeneration, considering the percentage of centronucleated fibers, Myh3 and Myh4 expression, or maximal force production, and thus possibly not compromise a gene therapy approach. With the AAV-U7 injected just before exercise initiation, we reported that voluntary running did not impact the number of viral genomes and the expression of U7 and Dmd 1 month after injection of AAV-U7, but reduced the amount of dystrophin, from 80% to 65% of the muscle cross-sectional area with dystrophin-expressing fibers. In conclusion, voluntary running did not induce muscle damage and had no drastic detrimental effect on the AAV gene therapy exon skipping approach in a severe murine DMD model. Moreover, our results suggested that exercise should be an additional element to consider in the design and conception of future therapeutic approaches for DMD.

Proteomic profiling of impaired excitation-contraction coupling and abnormal calcium handling in muscular dystrophy.

The X-linked inherited neuromuscular disorder Duchenne muscular dystrophy is characterised by primary abnormalities in the membrane cytoskeletal component dystrophin. The almost complete absence of the Dp427-M isoform of dystrophin in skeletal muscles renders contractile fibres more susceptible to progressive degeneration and a leaky sarcolemma membrane. This in turn results in abnormal calcium homeostasis, enhanced proteolysis and impaired excitation-contraction coupling. Biochemical and mass spectrometry-based proteomic studies of both patient biopsy specimens and genetic animal models of dystrophinopathy have demonstrated significant changes in the concentration and/or physiological function of essential calcium-regulatory proteins in dystrophin-lacking voluntary muscles. Abnormalities include dystrophinopathy-associated changes in voltage sensing receptors, calcium release channels, calcium pumps and calcium binding proteins. This review article provides an overview of the importance of the sarcolemmal dystrophin-glycoprotein complex and the wider dystrophin complexome in skeletal muscle and its linkage to depolarisation-induced calcium-release mechanisms and the excitation-contraction-relaxation cycle. Besides chronic inflammation, fat substitution and reactive myofibrosis, a major pathobiochemical hallmark of X-linked muscular dystrophy is represented by the chronic influx of calcium ions through the damaged plasmalemma in conjunction with abnormal intracellular calcium fluxes and buffering. Impaired calcium handling proteins should therefore be included in an improved biomarker signature of Duchenne muscular dystrophy.

Genetic correction strategies for Duchenne muscular dystrophy and their impact on the heart

BackgroundDuchenne muscular dystrophy (DMD) is an X-linked recessive disorder with early childhood onset characterized by profound loss of muscle strength and associated cardiomyopathy. DMD affects is most often caused by deletions involving single or multiple exons that disrupt the open reading frame of the DMD gene. Mutations causing loss or premature truncation of dystrophin result in dystrophin protein deficiency, which renders the plasma membrane of skeletal myofibers and cardiomyocytes weakened. Aim of reviewGenetic correction is in use to treat DMD since several drugs have been already approved which partially restore dystrophin production through the use of antisense oligonucleotides. There are multiple ongoing clinical trials to evaluate the efficacy of treating DMD with micro-dystrophins delivered by adeno-associated viruses. Future approaches entail gene editing to target the single copy of the DMD gene on the X-chromosome. The primary, near-term goal is restoration of skeletal muscle dystrophin, and for some of these treatments, the efficacy in the heart is not fully known. Here, we discuss the anticipated cardiac outcomes of dystrophin-targeted therapies, and how this information informs genomic medicine for cardiomyopathies, especially in childhood. Key scientific concepts of reviewMany genetic treatment strategies are being implemented to treat DMD. Since most preclinical testing has focused on skeletal muscle, there is a gap in knowledge about the expected effects of these approaches on cardiac genetic correction and cardiomyopathy progression in DMD. Additional study is needed.

CLINICAL TRIAL HIGHLIGHTS: O.2 IGNITE DMD Phase I/II ascending dose study of SGT-001 microdystrophin gene therapy for DMD: 1.5-year functional outcomes update

IGNITE DMD is an ongoing, multicenter, first-in-human open-label Phase I/II ascending dose study of the safety and efficacy of a single intravenous infusion of SGT-001, an AAV9 microdystrophin gene therapy, in DMD patients. DMD is caused by mutations that result in the absence of dystrophin in skeletal and cardiac muscle, which leads to muscle fiber deterioration, progressive decline in motor function, and premature death. SGT-001 microdystrophin is a five-repeat dystrophin surrogate that includes the critical actin, dystroglycan, and nNOS binding domains. Skeletal and cardiac muscle-specific expression are driven by the CK8 promotor. Primary outcomes include safety and measurement of microdystrophin expression by Western blot in muscle biopsies at 12 months. Secondary outcomes include evaluations of serum CK, motor function, pulmonary function (PFTs) and patient reported outcome measures (PROMs). Efficacy measures are assessed yearly through 5 years of study, in addition to 1.5-year assessments presented herein. Following the administration of 5E13 vg/kg to 3 subjects the dose was escalated, and all subsequent subjects have received SGT-001 at 2E14 vg/kg (HD). At day 90, microdystrophin expression in the first 3 subjects in the 2E14 vg/kg cohort ranged from approximately 5-17.5% of normal dystrophin by Western blot and 10-70% of positive muscle fibers by immunofluorescence. Importantly, sarcolemmal β-sarcoglycan and nNOS were restored in microdystrophin positive fibers. One-year functional data previously reported showed improvement or stabilization in motor function, PFTs and PROMs compared with expected natural history declines. A 1.5-year update of safety and efficacy of all patients will be discussed. Additionally, an interim update of newly treated patients will be provided. These data continue to suggest the potential for SGT-001 to confer meaningful benefit to patients with DMD and support its continued study as a therapeutic candidate.

DMD – ANIMAL MODELS: EP.77 Subcellular organization of skeletal muscle dystrophin is restored by tricyclo-DNA mediated exon skipping in the DmdEGFP-mdx mouse model

DMD – ANIMAL MODELS: EP.77 Subcellular organization of skeletal muscle dystrophin is restored by tricyclo-DNA mediated exon skipping in the DmdEGFP-mdx mouse model

Characterization of Alzheimer's Disease‐like Neuropathology in Duchenne Muscular Dystrophy Using the D2 mdx Mouse Model

Duchenne Muscular Dystrophy (DMD) is a progressive muscle wasting disorder which affects approximately 1 in 3500 male births and is caused by a mutation in the dystrophin gene. Although most research focuses on the role of dystrophin in skeletal and cardiac muscle, dystrophin is also expressed in the brain, specifically in memory-associated brain regions like the hippocampus and cortex. Furthermore, patients with DMD and mdx mice (rodent model of DMD) exhibit significant memory and learning impairments, implicating dystrophin loss in neurodegeneration. A recent study found patients with DMD had higher serum amyloid-β42, with this marker being heavily associated with cognitive impairments and Alzheimer's disease pathology. The purpose of this study was to characterize the neuropathology associated with DMD, focusing on markers of amyloidogenesis and synaptic function, in a preclinical mdx mouse model. Male mdx and age matched (8-9 weeks old) wild-type (DBA-2J) mice were purchased from Jackson laboratories (n = 12 per group). Novel object recognition testing (NORT) was performed to determine cognitive function. Prefrontal cortex and hippocampus brain regions were extracted for analysis. NORT testing demonstrated that mdx mice had reduced exploration time of the novel object (WT 35.2 sec vs mdx 18.7 sec; p = 0.02) and a lower exploration index (-25%, p < 0.05) compared to WT mice. Western blot analysis revealed higher soluble APP β (+40%) and total APP (+20%) in the prefrontal cortex of mdx mice compared to WT, and lower soluble APP α (-30%) fragment in the hippocampus (p <0.05). Furthermore, PSD95 content was nearly 2-fold higher in the prefrontal cortex of mdx mice compared to WT (p <0.05). This study provides novel information about the cellular pathways associated with memory impairment with DMD. Specifically, we show a shift towards amyloidogenesis in the prefrontal cortex and hippocampus brain regions of mdx mice, suggesting the possibility of similar pathogenesis to Alzheimer's disease. These changes are accompanied by reduced performance in the NORT test. Importantly, our results demonstrate that these changes occur at a relatively young age in D2 mdx mice, similar to what is observed in patients with DMD.

LATE BREAKING NEWS ORAL PRESENTATION: LBO 3 Expression of apparent full-length dystrophin in skeletal muscle in a first-in-human gene therapy trial using the scAAV9.U7-ACCA vector

Exon duplications that cause Duchenne muscular dystrophy (DMD) are promising candidates for exon skipping therapies because skipping a single exon copy should result in wild-type transcript and full-length dystrophin expression. We developed a therapeutic exon skipping viral vector (scAAV9.U7-ACCA) comprising four copies of a modified U7snRNA containing antisense sequences targeting the splice donor (2 copies) and splice acceptor (2 copies) of the DMD exon 2, the most commonly duplicated exon. Following dose finding studies in Dup2 mice and after demonstrating lack of toxicity in non-human primates, we initiated a first-in-human clinical trial (NCT04240314).

Decreased smooth muscle function, peristaltic activity, and gastrointestinal transit in dystrophic (mdx) mice.

Duchenne muscular dystrophy (DMD) is characterized by the lack of dystrophin in skeletal, cardiac, and smooth muscle. Slow colonic transit and constipation are common in DMD patients and animal models of DMD. However, the cause of this hypocontractility and the expression of contractile proteins in smooth muscle are unknown. The aim of the study was to investigate the expression of contractile proteins in the colonic smooth muscle and the function of the colon in control and mdx mice. Muscle contraction was measured in muscle strips and isolated muscle cells. Peristaltic activity was measured in ex vivo preparations by spatiotemporal mapping, and gastrointestinal (GI) transit in vivo was measured by the distribution of fluorescent marker along the intestine and colon. mRNA expression of contractile proteins smoothelin, caldesmon, calponin, and tropomyosin was measured by qRT-PCR. Expression of mRNA for contractile proteins was decreased in colonic smooth muscle of mdx mice compared with control. Contraction in response to acetylcholine and KCl was decreased in colonic muscle strips and in isolated muscle cells of mdx mice. Distension of ex vivo colons with Krebs buffer induced peristalsis in both control and mdx mice; however, significantly fewer full peristaltic waves were recorded in the colons of mdx mice. GI transit was also inhibited in mdx mice. The data indicate that the lack of dystrophin causes decrease in colonic smooth muscle contractility, peristalsis, and GI transit and provides the basis for analysis of mechanisms involved in smooth muscle dysfunction in DMD.

Protective Effects of Rocuronium Bromide on Ischemia-Reperfusion Injury in Skeletal Muscle Induced by Tourniquet in Patients Undergoing Elective Unilateral Total Knee Arthroplasty: A Prospective, Double Blind, Randomized, Controlled Study.

PurposeTo investigate the effects of different doses of rocuronium on ischemia-reperfusion injury in skeletal muscle induced by tourniquet in patients undergoing elective unilateral total knee arthroplasty.Patients and MethodsA total of 90 patients undergoing elective unilateral knee arthroplasty under general anesthesia combined with femoral nerve block were randomly divided into 3 groups: normal saline group (group S), rocuronium 0.6 mg/kg group (group L), and rocuronium 1.2 mg/kg group (group H). The primary outcome was the expression of dystrophin in skeletal muscle at 60 min after ischemia. Secondary outcomes included the concentration of malondialdehyde (MDA) and neuronal nitric oxide synthase (nNOS) in blood at 5 min and 30 min after reperfusion. In addition, thigh girth at 24 h and 48 h after operation, the leaving bed time, the incidence of tourniquet-related hypertension and short-term (3 days after operation) complications (nausea and vomiting, swelling, blister, wound infection) and long-term (3 months after operation) complications (joint instability, stiffness, nerve paralysis, pain) were recorded.Main ResultsThe expression of dystrophin in the rocuronium group was higher than that in group S after ischemia (P <0.05). The concentration of MDA in the rocuronium 1.2 mg/kg group was lower at 30 min after reperfusion (P < 0.05). There was no significant difference in nNOS among groups at each time point (P > 0.05). The change of thigh girth was the smallest in the rocuronium 1.2 mg/kg group after operation (P<0.05). The leaving bed time was significantly earlier after operation in the rocuronium group than that in group S (P <0.05).ConclusionRocuronium can protect skeletal muscle from ischemia-reperfusion injury induced by tourniquet. The mechanism may be related to the fact that rocuronium can reduce the loss of dystrophin in skeletal muscle and have the effects of anti-oxidation and anti-stress.Trial RegistrationThe study was registered at http://www.chictr.org.cn (ChiCTR1800019221, registered on 2018–10-31).

Severe cardiac involvement with preserved truncated dystrophin expression in Becker muscular dystrophy by +1G>A DMD splice-site mutation: a case report

Becker muscular dystrophy (BMD) is caused by specific mutations in the DMD gene that causes progressive muscle weakness and primarily affects skeletal and cardiac muscle. Although cardiac involvement is a significant cause of mortality in BMD, the genetic–phenotype correlation for skeletal and cardiac muscles has not been elucidated. Here, we described a 39-year-old man with BMD, who presented with subtle skeletal muscle weakness in the right leg in his 20s and underwent left ventricular restoration for severe dilated cardiomyopathy at the age of 29. He had difficulty climbing stairs after the age of 35. Neither duplication nor deletion of exons was detected by multiplex ligation-dependent probe amplification. A hemizygous c.264 + 1G>A mutation in intron 4 of the DMD was identified by next-generation sequencing. Furthermore, exon 4 skipping of the DMD was confirmed in both skeletal and cardiac muscles evaluated by reverse transcriptase PCR. Endomyocardial and skeletal muscle biopsies revealed dystrophic pathology characterized by muscle fiber atrophy and hypertrophy with a mild degree of interstitial fibrosis. Interestingly, dystrophin immunohistochemistry demonstrated patchy and faint staining of the skeletal muscle membranes but almost normal staining of the cardiac muscle membranes. Western blot analysis revealed a decreased amount of truncated dystrophin in skeletal muscle but surprisingly almost normal amount in cardiac muscle. This case indicates that BMD patients may have severe cardiac dysfunction despite preserved cardiac truncated dystrophin expression.

Social stress is lethal in the mdx model of Duchenne muscular dystrophy

BackgroundDuchenne muscular dystrophy (DMD) is caused by the loss of dystrophin. Severe and ultimately lethal, DMD progresses relatively slowly in that patients become wheelchair bound only around age twelve with a survival expectancy reaching the third decade of life. MethodsThe mildly-affected mdx mouse model of DMD, and transgenic DysΔMTB-mdx and Fiona-mdx mice expressing dystrophin or utrophin, respectively, were exposed to either mild (scruffing) or severe (subordination stress) stress paradigms and profiled for their behavioral and physiological responses. A subgroup of mdx mice exposed to subordination stress were pretreated with the beta-blocker metoprolol. FindingsSubordination stress caused lethality in ∼30% of mdx mice within 24 h and ∼70% lethality within 48 h, which was not rescued by metoprolol. Lethality was associated with heart damage, waddling gait and hypo-locomotion, as well as marked up-regulation of the hypothalamus-pituitary-adrenocortical axis. A novel cardiovascular phenotype emerged in mdx mice, in that scruffing caused a transient drop in arterial pressure, while subordination stress caused severe and sustained hypotension with concurrent tachycardia. Transgenic expression of dystrophin or utrophin in skeletal muscle protected mdx mice from scruffing and social stress-induced responses including mortality. InterpretationWe have identified a robust new stress phenotype in the otherwise mildly affected mdx mouse that suggests relatively benign handling may impact the outcome of behavioural experiments, but which should also expedite the knowledge-based therapy development for DMD. FundingGreg Marzolf Jr. Foundation, Summer's Wish Fund, NIAMS, Muscular Dystrophy Association, University of Minnesota and John and Cheri Gunvalson Trust.

A case report: X-linked dystrophin gene mutation causing severe isolated dilated cardiomyopathy

Background X-linked dilated cardiomyopathy (XLDCM) is a rare but rapidly progressive cardiomyopathy caused by dystrophin gene mutation. Mutations are more often associated with Duchenne and Becker Muscular Dystrophy, which are characterized by skeletal muscle weakness or limb girdle dystrophy. However, patients with isolated XLDCM have normal skeletal muscle but complete dystrophin loss in cardiac muscle resulting in isolated myocardial involvement without overt signs of skeletal myopathy.Case summary A previously well 16-year-old boy developed sudden onset dense left-sided weakness and facial droop. Computed tomography (CT) angiography and CT brain showed an occluded right internal carotid artery extending to the right middle cerebral artery. He underwent successful endovascular clot retrieval but developed frank pulmonary oedema and cardiogenic shock requiring inotropic support and intubation. Transthoracic echocardiography demonstrated severe left ventricular (LV) cardiomyopathy and an apical thrombus. Subsequent cardiac magnetic resonance (CMR) imaging confirmed the LV parameters and diffuse late gadolinium enhancement. Despite absence of skeletal manifestations, subsequent genetic testing revealed an X-linked dystrophin gene mutation [c.31+G>T (IVS1G>T)]. He was commenced on empirical heart failure therapy and underwent successful cardiac transplantation.Discussion X-linked dilated cardiomyopathy is a rare, rapidly progressing cardiomyopathy. Patients show normal skeletal muscle dystrophin but absent expression in cardiac muscle, resulting fibrosis, and atrophy. About 20% of affected young males have significantly reduced survival and thus the diagnosis must be considered in cases of idiopathic cardiomyopathy with CMR and genetic testing key to the diagnosis. Whilst evidence exists for empirical heart failure medications, cardiac transplantation remains the definitive treatment.

The pathogenesis of Duchenne muscular dystrophy

A new concept of pathogenesis Duchenne Muscular Dystrophy (DMD) based on authors experience and discussion of results of other studies. Author estimate DMD as a cerebro-muscular pathology depend on absent dystrophin (D) in skeletal muscles, heart, brain. Author suppose that all family D work as one functional system, which play the important role in organization of walking. Destroy of this system is cause of the disease.

The root cause of Duchenne muscular dystrophy is the lack of dystrophin in smooth muscle of blood vessels rather than in skeletal muscle per se

Background:The dystrophin protein is part of the dystrophin associated protein complex (DAPC) linking the intracellular actin cytoskeleton to the extracellular matrix. Mutations in the dystrophin gene cause Duchenne and Becker muscular dystrophy (D/BMD). Neuronal nitric oxide synthase associates with dystrophin in the DAPC to generate the vasodilator nitric oxide (NO). Systemic dystrophin deficiency, such as in D/BMD, results in muscle ischemia, injury and fatigue during exercise as dystrophin is lacking, affecting NO production and hence vasodilation. The role of neuregulin 1 (NRG) signaling through the epidermal growth factor family of receptors ERBB2 and ERBB4 in skeletal muscle has been controversial, but it was shown to phosphorylate α-dystrobrevin 1 (α-DB1), a component of the DAPC. The aim of this investigation was to determine whether NRG signaling had a functional role in muscular dystrophy.Methods:Primary myoblasts (muscle cells) were isolated from conditional knock-out mice containing lox P flanked ERBB2 and ERBB4 receptors, immortalized and exposed to Cre recombinase to obtainErbb2/4double knock-out (dKO) myoblasts where NRG signaling would be eliminated. Myotubes, thein vitroequivalent of muscle fibers, formed by fusion of the lox P flankedErbb2/4myoblasts as well as theErbb2/4dKO myoblasts were then used to identify changes in dystrophin expression.Results:Elimination of NRG signaling resulted in the absence of dystrophin demonstrating that it is essential for dystrophin expression. However, unlike the DMD mouse model mdx, with systemic dystrophin deficiency, lack of dystrophin in skeletal muscles ofErbb2/4dKO mice did not result in muscular dystrophy. In these mice, ERBB2/4, and thus dystrophin, is still expressed in the smooth muscle of blood vessels allowing normal blood flow through vasodilation during exercise.Conclusions:Dystrophin deficiency in smooth muscle of blood vessels, rather than in skeletal muscle, is the main cause of disease progression in DMD.

Dystrophinopathy-associated dysfunction of Krebs cycle metabolism.

Duchenne muscular dystrophy is a deadly muscle-wasting disorder caused by loss of dystrophin protein. Studies suggest that metabolic alterations are important to disease pathogenesis. Because muscle accounts for ~40% of body mass, we hypothesized that dystrophy-mediated metabolic changes would be measurable in biofluids and that a metabolomic analysis of urine would provide insight into the metabolic status of dystrophic muscle. Using the mdx mouse model, we performed a large-scale metabolomic screen at 1 and 3months. While 10% of metabolites were altered at age 1month, 40% were changed at 3months. Principal component analysis distinguished wild-type from mdx animals, with the greatest separation at 3months. A critical distinguishing pathway was Krebs cycle metabolite depletion in mdx urine. Five of seven detected Krebs cycle metabolites were depleted in mdx urine, with succinate being the most robustly affected metabolite. Using selected reaction monitoring mass spectrometry, we demonstrated that muscle-specific dystrophin expression corrects mdx succinate depletion. When subjected to downhill treadmill running, wild-type and mdx mice expressing recombinant dystrophin in skeletal muscle displayed significant increases in urinary succinate levels. However, mdx succinate levels were unchanged, suggesting urinary succinate depletion may reflect an inability to upregulate the Krebs cycle following exercise. Finally, we show that supplementing the Krebs cycle in an ex vivo fatigue/recovery assay significantly impacts mdx muscle performance but has no effect on wild-type muscle. Our results suggest that global metabolic impairment is associated with mdx disease progression and that Krebs cycle deficiencies are a downstream consequence of dystrophin loss.