Short-term treatment with Uncaria tomentosa aggravates the injury phenotype in mdx mice

INTRODUCTION: Uncaria tomentosa (Willd. ex Roem. & Schult.) DC. (Rubiaceae) or UT is a medicinal plant with antiviral, antimutagenic, anti-inflammatory and antioxidant properties. Duchenne muscular dystrophy (DMD) is a severe muscle wasting disease caused by mutations in the dystrophin gene; this deficiency leads to sarcolemma instability, inflammation, muscle degeneration and fibrosis. OBJECTIVE: Considering the importance of inflammation to dystrophy progression and the anti-inflammatory activity of UT, in the present study we evaluated whether oral administration of UT extract would ameliorate dystrophy in the mdx mice, a DMD model. METHODS: Eight-week-old male mdx mice were submitted to 200 mg/kg body weight daily UT oral administration for 6 weeks. General histopathology was analysed, and muscle tumor necrosis factor α, transforming growth factor-ß, myostatin and osteopontin transcript levels were assessed. The ability of mice to sustain limb tension to oppose their gravitational force was measured. Data were analysed with the unpaired Student's t-test. RESULTS: Morphologically, both untreated and UT-treated animals exhibited internalised nuclei, increased endomysial connective tissue and variations in muscle fibre diameters. Body weight and muscle strength were significantly reduced in the UT-treated animals. Blood creatine kinase was higher in UT-treated compared to untreated animals. In tibialis anterior, myostatin, transcript was more highly expressed in the UT-treated while in the diaphragm muscle, transforming growth factor-ß transcripts were less expressed in the UT-treated. CONCLUSION: While previous studies identified anti-inflammatory, antiproliferative and anticarcinogenic UT effects, the extract indicates worsening of dystrophic muscles phenotype after short-term treatment in mdx mice.

The muscle regeneration marker FOXP3 is associated with muscle injury in Duchenne muscular dystrophy.

BACKGROUND: In Duchenne muscular dystrophy (DMD), the immune system cells (ISC) synthesize molecules to regulate inflammation, a process needed to regenerate muscle. The relationship between those molecules and the muscle injury is unknown. Monocytes belonging to ISC are regulated by omega-3 fatty acids (ω-3 LCPUFAs) in DMD, but whether those fatty acids influence other ISC like T-cells is unknown. OBJECTIVE: We analyzed the expression of the muscle regeneration markers (FOXP3 and AREG) in circulating leukocytes of DMD patients with different lower limb muscle functions and whether ω-3 LCPUFAs regulate the expression of those markers, and the populations of circulating T-cells, their intracellular cytokines, and disease progression (CD69 and CD49d) markers. METHODS: This placebo-controlled, double-blind, randomized study was conducted in DMD boys supplemented with ω-3 LCPUFAs (n = 18) or placebo (sunflower oil, n = 13) for six months. FOXP3 and AREG mRNA expression in leukocytes, immunophenotyping of T-cell populations, CD49d and CD69 markers, and intracellular cytokines in blood samples were analyzed at baseline and months 1, 2, 3, and 6 of supplementation. RESULTS: Patients with assisted ambulation expressed higher (P = 0.015) FOXP3 mRNA levels than ambulatory patients. The FOXP3 mRNA expression correlated (Rho = -0.526, P = 0.03) with the Vignos scale score at month six of supplementation with ω-3 LCPUFAs. CD49d + CD8 + T-cells population was lower (P = 0.037) in the ω -3 LCPUFAs group than placebo at month six of supplementation. CONCLUSION: FOXP3 is highly expressed in circulating leukocytes of DMD patients with the worst muscle function. Omega-3 LCPUFAs might modulate the synthesis of the adhesion marker CD49d + CD8 + T-cells, but their plausible impact on FOXP3 needs more research.

The Early Care (0-3 Years) In Duchenne Muscular Dystrophy Meeting Report.

Objective: This report summarizes the key discussions from the "Early Care (0-3 years) in Duchenne Muscular Dystrophy" meeting, which aimed to address the challenges and opportunities in the diagnosis and care of Duchenne muscular dystrophy (DMD) and female carriers within the 0-3-year age group. Methods: The meeting brought together experts and healthcare providers who shared insights, discussed advancements in DMD care, and identified research needs. Presentations covered diagnostic challenges, approved therapies, clinical trials, identification of young female carriers, and the importance of clinical care and support for families. Results: The meeting highlighted the importance of timely diagnosis and the lack of evidence-based guidelines for the care of children with DMD aged 0-3 years. Diagnostic challenges were discussed, including delays in receiving a DMD diagnosis and disparities based on ethnicity. The potential benefits and process of newborn screening were addressed.Approved therapeutic interventions, such as corticosteroids and exon-skipping drugs, were explored, with studies indicating the potential benefits of early initiation of corticosteroid therapy and the safety of exon-skipping drugs in DMD. Clinical trials involving infants and young boys were discussed, focusing on drugs like ataluren, vamorolone, and gene therapies.The meeting emphasized the importance of clinical care and support for families, including comprehensive information provision, early intervention services, and individualized support. The identification and care of young female carriers were also addressed. Conclusion: The meeting provided a platform for experts and healthcare providers to discuss and identify key aspects of early care for children with DMD aged 0-3 years. The meeting emphasized the need for early diagnosis, evidence-based guidelines, and comprehensive care and support for affected children and their families. Further research, collaboration, and the development of consensus guidelines are needed to improve early diagnosis, treatment, and outcomes in this population.

A systematic literature review and meta-analysis of the effectiveness of vitamin D supplementation for patients with Duchenne muscular dystrophy.

We conducted a systematic literature review and meta-analysis on the effectiveness of vitamin D supplementation in maintaining or restoring vitamin D levels in Duchenne muscular dystrophy. Due to a lack of randomised controlled trials, cross-sectional and retrospective and prospective cohort studies were taken as the best available evidence. Inclusion criteria included reporting mean serum vitamin D levels in a supplement-taking group. After screening 102 records; 13 were included in a narrative synthesis and eight of these in a meta-analysis. We show that current dosing regimens are preventing severe deficiency but are not effective at maintaining sufficient vitamin D levels within the Duchenne population. Despite high levels of daily vitamin D supplementation (>1000 International Units), at least 20 % of people with Duchenne remain vitamin D deficient. No significant association between dose and serum vitamin D levels was found (r2 = 0.3, p = 0.237). A meta-analysis of mean serum vitamin D levels across eight studies also revealed substantial variability in response to vitamin D supplementation and high heterogeneity (I2 = 99.59 %). These data could impact on an individual's risk and severity of osteoporosis and vertebral fractures.

Determination of equilibria constants of arginine:glycine amidinotransferase (AGAT)-catalyzed reactions using concentrations of circulating amino acids.

Arginine:glycine amidinotransferase (AGAT) catalyzes mainly two reactions that generate 1) L-homoarginine (hArg) from L-arginine and L-lysine (Kharg) and 2) guanidinoacetate (GAA) and L-ornithine from L-arginine and glycine (Kgaa). Previously, we found that pharmacological treatment of Becker muscular dystrophy (BMD) patients with metformin or L-citrulline resulted in antidromic effects on serum hArg and GAA concentrations, seemingly acting as an inhibitor and effector of AGAT activity, respectively. Here, we used data of this study as a model to determine Kharg and Kgaa values by using the concentrations of the participating amino acids measured in serum samples of the BMD patients. The study aimed to prove the general utility of this approach to investigate effects of amino acids and drugs on AGAT-catalyzed reactions in vivo in humans.

Efficacy of N-163 beta-glucan in beneficially improving biomarkers of relevance to muscle function in patients with muscular dystrophies in a pilot clinical study.

Background: Muscular dystrophies other than Duchenne muscular dystrophy (DMD) are genetic diseases characterized by increasing muscle weakness, loss of ambulation, and ultimately cardiac and respiratory failure. There are currently no effective therapeutics available. Having demonstrated the efficacy of a N-163 strain of Aureobasidium Pullulans (Neu-REFIX) produced B-1, 3-1,6-Glucan in pre-clinical and clinical studies of Duchenne muscular dystrophy (DMD) earlier, we assessed the effectiveness of this novel Beta glucan in the other muscular dystrophies in the present study. Methods: In this 60-day study, six patients with muscular dystrophies other than DMD consumed one 8g gel of Neu-REFIX beta-glucan along with their usual standard of care treatment regimen, and their biomarkers of relevance to muscle function such as serum calcium (SC), creatine phosphokinase (CPK), and alkaline phosphatase (ALP) levels along with functional improvement criteria, which is, Medical research council (MRC) scale and North Star Ambulatory assessment (NSAA), assessed at baseline and following the intervention. Results: After the intervention, the SC levels significantly decreased from a mean baseline value of 9.28 mg/dL to 8.31 mg/dL (p-value = 0.02). With a p-value of 0.29, the mean CPK value dropped from 2192.33 IU/L to 1567.5 IU/L. Following the intervention, the ALP levels dropped from 200.33 to 75.5 U/L (p-value = 0.15). MRC scale improved in three out of six patients. NSAA remained stable. There were no adverse effects. Conclusion: This study has proven the safety of Neu REFIX beta-glucan food supplement and its efficacy in improving both plasma biomarkers and functional parameters of muscle in a short duration of 2 months. Further validation by evaluation of muscle function for a longer duration is recommended to confirm the efficacy of Neu-REFIX food supplement as a potential adjuvant DMT in muscular dystrophies.

Sulfur amino acid supplementation displays therapeutic potential in a C. elegans model of Duchenne muscular dystrophy.

Mutations in the dystrophin gene cause Duchenne muscular dystrophy (DMD), a common muscle disease that manifests with muscle weakness, wasting, and degeneration. An emerging theme in DMD pathophysiology is an intramuscular deficit in the gasotransmitter hydrogen sulfide (H2S). Here we show that the C. elegans DMD model displays reduced levels of H2S and expression of genes required for sulfur metabolism. These reductions can be offset by increasing bioavailability of sulfur containing amino acids (L-methionine, L-homocysteine, L-cysteine, L-glutathione, and L-taurine), augmenting healthspan primarily via improved calcium regulation, mitochondrial structure and delayed muscle cell death. Additionally, we show distinct differences in preservation mechanisms between sulfur amino acid vs H2S administration, despite similarities in required health-preserving pathways. Our results suggest that the H2S deficit in DMD is likely caused by altered sulfur metabolism and that modulation of this pathway may improve DMD muscle health via multiple evolutionarily conserved mechanisms.

Acute toxicity and regenerative dose finding of an extract of Miconia ferruginata DC. in a mouse model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a severe disease with no cure caused by a genetic abnormality, promoting progressive muscle degeneration. Corticosteroids are used drugs in treatment associated with adverse effects. The extract of Miconia ferruginata (Melastomataceae) (MF) has demonstrated potent antioxidant and anti-inflammatory potential in vitro. This study used a DMD model (mdx) to determine the toxic dose of this plant and found a possible non-toxic dose with therapeutic effects. The mdx groups received an intraperitoneal injection of 0 (control group), 50, 100, 200, 300, and 2000 mg kg-1 of the aqueous leaf extract following a single-dose acute toxicity protocol and were observed for 14 days. The range of toxicity of the extract and LD50 were determined. Histopathological analysis, the quantification of fibrosis, and immunohistochemical analysis of the tissues were performed. The results demonstrated that 2000 mg kg-1 was highly toxic, inducing histopathological changes in the tissues evaluated, with 100% mortality in 48 hours. The other doses caused no behavioral changes or signs of toxicity. The MF extract led reduction in histopathological changes, fibrosis, and inflammation, a reduction in HSP70 and an increase in MCL-1 proteins. Doses of 50-200 mg kg-1 demonstrated regenerative tissue and anti-inflammatory potential.

Exploring the Therapeutic Potential of Ectoine in Duchenne Muscular Dystrophy: Comparison with Taurine, a Supplement with Known Beneficial Effects in the mdx Mouse.

Duchenne Muscular Dystrophy (DMD) is a debilitating muscle disorder that condemns patients to year-long dependency on glucocorticoids. Chronic glucocorticoid use elicits many unfavourable side-effects without offering satisfying clinical improvement, thus, the search for alternative treatments to alleviate muscle inflammation persists. Taurine, an osmolyte with anti-inflammatory effects, mitigated pathological features in the mdx mouse model for DMD but interfered with murine development. In this study, ectoine is evaluated as an alternative for taurine in vitro in CCL-136 cells and in vivo in the mdx mouse. Pre-treating CCL-136 cells with 0.1 mM taurine and 0.1 mM ectoine prior to exposure with 300 U/mL IFN-γ and 20 ng/mL IL-1ß partially attenuated cell death, whilst 100 mM taurine reduced MHC-I protein levels. In vivo, histopathological features of the tibialis anterior in mdx mice were mitigated by ectoine, but not by taurine. Osmolyte treatment significantly reduced mRNA levels of inflammatory disease biomarkers, respectively, CCL2 and SPP1 in ectoine-treated mdx mice, and CCL2, HSPA1A, TNF-α and IL-1ß in taurine-treated mdx mice. Functional performance was not improved by osmolyte treatment. Furthermore, ectoine-treated mdx mice exhibited reduced body weight. Our results confirmed beneficial effects of taurine in mdx mice and, for the first time, demonstrated similar and differential effects of ectoine.

The Effect of Uridine on the State of Skeletal Muscles and the Functioning of Mitochondria in Duchenne Dystrophy.

Duchenne muscular dystrophy is caused by the loss of functional dystrophin that secondarily causes systemic metabolic impairment in skeletal muscles and cardiomyocytes. The nutraceutical approach is considered as a possible complementary therapy for this pathology. In this work, we have studied the effect of pyrimidine nucleoside uridine (30 mg/kg/day for 28 days, i.p.), which plays an important role in cellular metabolism, on the development of DMD in the skeletal muscles of dystrophin deficient mdx mice, as well as its effect on the mitochondrial dysfunction that accompanies this pathology. We found that chronic uridine administration reduced fibrosis in the skeletal muscles of mdx mice, but it had no effect on the intensity of degeneration/regeneration cycles and inflammation, pseudohypetrophy, and muscle strength of the animals. Analysis of TEM micrographs showed that uridine also had no effect on the impaired mitochondrial ultrastructure of mdx mouse skeletal muscle. The administration of uridine was found to lead to an increase in the expression of the Drp1 and Parkin genes, which may indicate an increase in the intensity of organelle fission and the normalization of mitophagy. Uridine had little effect on OXPHOS dysfunction in mdx mouse mitochondria, and moreover, it was suppressed in the mitochondria of wild type animals. At the same time, uridine restored the transport of potassium ions and reduced the production of reactive oxygen species; however, this had no effect on the impaired calcium retention capacity of mdx mouse mitochondria. The obtained results demonstrate that the used dose of uridine only partially prevents mitochondrial dysfunction in skeletal muscles during Duchenne dystrophy, though it mitigates the development of destructive processes in skeletal muscles.

Early-Onset Diabetes Mellitus in Chromosome 8p11.2 Deletion Syndrome Combined With Becker Muscular Dystrophy - A Case Report.

Background: Chromosome 8p11.2 includes several key genes in development such as the FGFR1, ANK1, KAT6A, and SLC20A2 genes. Deletion of this fragment causes a contiguous gene syndrome. Currently, few cases of interstitial deletion of whole 8p11.2 have been reported. We report a rare case of 8p11.2 deletion syndrome with the unique phenotypes, presenting with early-onset diabetes. Case Description: A 20-year-old man with a 1-year history of diabetes mellitus was admitted to the Endocrinology Clinic. Physical examination revealed the dysmorphic facial features, and broad and foreshortened halluces. Laboratory examination indicated spherocytosis anemia, and hypogonadotropic hypogonadism. Bone mineral density analysis showed decreased bone density in the lumbar vertebrae. Brain CT showed calcification. Whole-exome sequencing revealed a 7.05-Mb deletion in 8p11 containing 43 OMIM genes, and a large in-frame deletion of exons 48-55 in the DMD gene. Metformin was given to the patient after which his blood glucose was well controlled. HCG was injected subcutaneously and was supplemented with calcium and vitamin D, which led to an improvement in the patient's quality of life. Conclusion: We report a rare case of 8p11.2 deletion syndrome with unique phenotypes, and early-onset diabetes. It is challenging for endocrinologists to simultaneously reconcile a combination of these diseases across multiple disciplines. We discussed the influencing factors of early-onset diabetes in this patient and speculated that it was caused by complex interactions of known and unknown genetic backgrounds and environmental factors.

Efficacy and Safety of Vamorolone vs Placebo and Prednisone Among Boys With Duchenne Muscular Dystrophy: A Randomized Clinical Trial.

Importance: Corticosteroidal anti-inflammatory drugs are widely prescribed but long-term use shows adverse effects that detract from patient quality of life. Objective: To determine if vamorolone, a structurally unique dissociative steroidal anti-inflammatory drug, is able to retain efficacy while reducing safety concerns with use in Duchenne muscular dystrophy (DMD). Design, Setting, and Participants: Randomized, double-blind, placebo- and prednisone-controlled 24-week clinical trial, conducted from June 29, 2018, to February 24, 2021, with 24 weeks of follow-up. This was a multicenter study (33 referral centers in 11 countries) and included boys 4 to younger than 7 years of age with genetically confirmed DMD not previously treated with corticosteroids. Interventions: The study included 4 groups: placebo; prednisone, 0.75 mg/kg per day; vamorolone, 2 mg/kg per day; and vamorolone, 6 mg/kg per day. Main Outcomes and Measures: Study outcomes monitored (1) efficacy, which included motor outcomes (primary: time to stand from supine velocity in the vamorolone, 6 mg/kg per day, group vs placebo; secondary: time to stand from supine velocity [vamorolone, 2 mg/kg per day], 6-minute walk distance, time to run/walk 10 m [vamorolone, 2 and 6 mg/kg per day]; exploratory: NorthStar Ambulatory Assessment, time to climb 4 stairs) and (2) safety, which included growth, bone biomarkers, and a corticotropin (ACTH)-challenge test. Results: Among the 133 boys with DMD enrolled in the study (mean [SD] age, 5.4 [0.9] years), 121 were randomly assigned to treatment groups, and 114 completed the 24-week treatment period. The trial met the primary end point for change from baseline to week 24 time to stand velocity for vamorolone, 6 mg/kg per day (least-squares mean [SE] velocity, 0.05 [0.01] m/s vs placebo -0.01 [0.01] m/s; 95% CI, 0.02-0.10; P = .002) and the first 4 sequential secondary end points: time to stand velocity, vamorolone, 2 mg/kg per day, vs placebo; 6-minute walk test, vamorolone, 6 mg/kg per day, vs placebo; 6-minute walk test, vamorolone, 2 mg/kg per day, vs placebo; and time to run/walk 10 m velocity, vamorolone, 6 mg/kg per day, vs placebo. Height percentile declined in prednisone-treated (not vamorolone-treated) participants (change from baseline [SD]: prednisone, -1.88 [8.81] percentile vs vamorolone, 6 mg/kg per day, +3.86 [6.16] percentile; P = .02). Bone turnover markers declined with prednisone but not with vamorolone. Boys with DMD at baseline showed low ACTH-stimulated cortisol and high incidence of adrenal insufficiency. All 3 treatment groups led to increased adrenal insufficiency. Conclusions and Relevance: In this pivotal randomized clinical trial, vamorolone was shown to be effective and safe in the treatment of boys with DMD over a 24-week treatment period. Vamorolone may be a safer alternative than prednisone in this disease, in which long-term corticosteroid use is the standard of care. Trial Registration: ClinicalTrials.gov Identifier: NCT03439670.

2-Guanidino-quinazoline promotes the readthrough of nonsense mutations underlying human genetic diseases.

Premature termination codons (PTCs) account for 10 to 20% of genetic diseases in humans. The gene inactivation resulting from PTCs can be counteracted by the use of drugs stimulating PTC readthrough, thereby restoring production of the full-length protein. However, a greater chemical variety of readthrough inducers is required to broaden the medical applications of this therapeutic strategy. In this study, we developed a reporter cell line and performed high-throughput screening (HTS) to identify potential readthrough inducers. After three successive assays, we isolated 2-guanidino-quinazoline (TLN468). We assessed the clinical potential of this drug as a potent readthrough inducer on the 40 PTCs most frequently responsible for Duchenne muscular dystrophy (DMD). We found that TLN468 was more efficient than gentamicin, and acted on a broader range of sequences, without inducing the readthrough of normal stop codons (TC).

Reduced bone mineral density in adolescents with Duchenne Muscular Dystrophy (DMD) and scoliosis.

Duchenne muscular dystrophy is a progressive disease usually associated with loss of ambulation and progressive scoliosis. Immobilisation and glucocorticoid treatment are predisposing factors for reduced bone mineral density (BMD). Analysis of quantitative computed tomography revealed low BMD in thoracic and lumbar vertebrae in comparison to age- and sex-matched healthy controls. INTRODUCTION: Evaluation of vertebral bone mineral density (BMD) in Duchenne Muscular Dystrophy (DMD) adolescents with untreated advanced scoliosis and comparison with the BMD values of healthy age-matched controls, based on quantitative computer tomography. METHODS: Thirty-seven DMD adolescents (age 15.6 ± 2.5 years) with spinal deformity were evaluated clinically and radiologically prior to definite spinal fusion and compared to 31 male and age-matched healthy individuals (age 15.7 ± 2.3 years). Data related to previous medical treatment, physiotherapy and ambulatory status was also analysed. Scoliotic curves were measured on plain sitting radiographs of the spine. The BMD Z-scores of the thoracic and lumbar vertebrae were calculated with QCTpro® (Mindways Software Inc., USA), based on data sets of preoperative, phantom pre-calibrated spinal computed tomography scans. RESULTS: A statistically significant lower BMD could be found in DMD adolescents, when compared to healthy controls, showing an average value for the lumbar spine of 80.5 ± 30.5 mg/cm3. Z-scores deteriorated from the upper thoracic towards the lower lumbar vertebrae. All but the uppermost thoracic vertebrae had reduced BMD values, with the thoracolumbar and lumbar region demonstrating the lowest BMD. No significant correlation was observed between BMD and the severity of the scoliotic curve, previous glucocorticoid treatment, cardiovascular impairment, vitamin D supplementation, non-invasive ventilation or physiotherapy. CONCLUSION: DMD adolescents with scoliosis have strongly reduced BMD Z-scores, especially in the lumbar spine in comparison to healthy controls. These findings support the implementation of a standardised screening and treatment protocol. Level of evidence/clinical relevance: therapeutic level III.

Effectiveness of Neridronate in the Management of Bone Loss in Patients with Duchenne Muscular Dystrophy: Results from a Pilot Study.

INTRODUCTION: Bone loss is a major issue in patients affected by Duchenne muscular dystrophy (DMD), a rare musculoskeletal disorder, particularly in those treated with glucocorticoids (GCs). We aimed to assess the effectiveness of neridronate in terms of bone mineral density (BMD) changes in this population. METHODS: We retrospectively reviewed the records of patients affected by DMD receiving GCs referred to our outpatient from 2015 to 2020. All patients were treated with an intramuscular (IM) injection of neridronate (25 mg every month). Bone density was measured at the lumbar spine (LS; L1-L4 tract) using dual-energy x-ray absorptiometry (DXA) (GE Lunar), no more than 4 weeks before (T0) and after 1 year from neridronate treatment (T1). RESULTS: Eight boys with DMD were included with a mean age at diagnosis of 4.75 ± 2.81 years. Six of them were non-ambulant and two of them had previous low-trauma fractures (a distal femur fracture and a vertebral compression fracture, respectively). All patients were receiving deflazacort [median duration of therapy 11.5 years (interquartile range 2-25)]. At the DXA evaluation (T0), the mean L1-L4 BMD value was 0.716 ± 0.164 g/cm2. Six patients (75%) showed an L1-L4 Z-score height-adjusted of less than - 2. The mean age of neridronate initiation was 18.87 ± 6.81 years. All patients were supplemented with calcium carbonate and vitamin D at baseline. After 12 months of treatment (T1), the mean L1-L4 BMD value was 0.685 ± 0.190 g/cm2. Seven patients (87.5%) showed an L1-L4 Z-score of less than - 2. Changes in LS BMD and Z-score were not significant between T0 and T1 in our cohort (p = 0.674 and p = 0.208, respectively) as well as among non-ambulant patients with DMD without previous fragility fractures. CONCLUSIONS: In this study, we reported for the first time that neridronate may slow bone loss in GC-treated patients with DMD at 1-year follow-up.

Effects of moderate aerobic exercise, low-level laser therapy, or their combination on muscles pathology, oxidative stress and irisin levels in the mdx mouse model of Duchenne muscular dystrophy.

This study aimed to investigate how the combined use of low-level laser therapy (LLLT) and exercise, to reduce the possible side effects and/or increase the benefits of exercise, would affect oxidative stress, utrophin, irisin peptide, and skeletal, diaphragmatic, and cardiac muscle pathologies. In our study, 20 mdx mice were divided into four groups. Groups; sedentary and placebo LLLT (SC), sedentary and LLLT (SL), 30-min swimming exercise (Ex), and 30-min swimming exercise and LLLT (ExL). After 8 weeks of swimming exercise, muscle tests, biochemically; oxidative stress index (OSI), utrophin and irisin levels were measured. Skeletal, diaphragmatic and cardiac muscle histopathological scores, skeletal and cardiac muscle myocyte diameters were determined under the light and electron microscope. While only irisin levels were increased in group SL compared to SC, it was determined that OSI, heart muscle histopathological scores decreased and irisin levels increased in both exercise groups (p < 0.05). In addition, in the ExL group, an increase in rotarod and utrophin levels, and a decrease in muscle and diaphragm muscle histopathological scores were observed (p < 0.05). It was determined that the application of swimming exercise in the mdx mouse model increased the irisin level in the skeletal muscle, while reducing the OSI, degeneration in the heart muscle, inflammation and cardiopathy. When LLLT was applied in addition to exercise, muscle strength, skeletal muscle utrophin levels increased, and skeletal and diaphragmatic muscle degeneration and inflammation decreased. In addition, it was determined that only LLLT application increased the level of skeletal muscle irisin.

Martial Arts Training for Boys With Duchenne Muscular Dystrophy.

PURPOSE: The primary aim of this pilot study was to investigate the safety and feasibility of a 3-month martial arts-based training (MAT) program for patients with Duchenne muscular dystrophy (DMD). The secondary aim was to examine changes in physical and psychosocial abilities after participating in the MAT program. METHODS: Twelve patients with DMD (10 ambulant and 2 nonambulant) were included. The MAT program was evaluated on feasibility and safety. Changes in physical abilities were measured using the Motor Function Measure, Performance of Upper Limb scale, and the North Star Ambulatory Assessment. Changes in psychosocial abilities were measured using the Strength and Difficulties Questionnaire, Personal Adjustment and Role Skills for DMD, and the Self-Perception Profile for Children/Adolescents. RESULTS: Two participants did not complete the MAT program. Attendance rate for the 10 remaining participants was 91%. Eleven falls were reported during the training, but these falls did not result in injuries. Therefore, the MAT program was found feasible and safe. After completing the MAT program, most participants showed an improvement of their psychosocial abilities, and their physical abilities did not show deterioration. CONCLUSION: The MAT program is feasible and safe for boys with DMD.

Iron overload and impaired iron handling contribute to the dystrophic pathology in models of Duchenne muscular dystrophy.

BACKGROUND: Oxidative stress is implicated in the pathophysiology of Duchenne muscular dystrophy (DMD, caused by mutations in the dystrophin gene), which is the most common and severe of the muscular dystrophies. To our knowledge, the distribution of iron, an important modulator of oxidative stress, has not been assessed in DMD. We tested the hypotheses that iron accumulation occurs in mouse models of DMD and that modulation of iron through the diet or chelation could modify disease severity. METHODS: We assessed iron distribution and total elemental iron using LA-ICP-MS on skeletal muscle cross-sections of 8-week-old Bl10 control mice and dystrophic mdx mice (with moderate dystrophy) and dystrophin/utrophin-null mice (dko, with severe dystrophy). In addition, mdx mice (4 weeks) were treated with either an iron chelator (deferiprone 150 mg/kg/day) or iron-enriched feed (containing 1% added iron as carbonyl iron). Immunoblotting was used to determine the abundance of iron- and mitochondria-related proteins. (Immuno)histochemical and mRNA assessments of fibrosis and inflammation were also performed. RESULTS: We observed a significant increase in total elemental iron in hindlimb muscles of dko mice (+50%, P < 0.05) and in the diaphragm of mdx mice (+80%, P < 0.05), with both tissues exhibiting severe pathology. Iron dyshomeostasis was further evidenced by an increase in the storage protein ferritin (dko: +39%, P < 0.05) and ferroportin compared with Bl10 control mice (mdx: +152% and dko: +175%, P < 0.05). Despite having features of iron overload, dystrophic muscles had lower protein expression of ALAS-1, the rate-limiting enzyme for haem synthesis (dko -44%, P < 0.05), and the haem-containing protein myoglobin (dko -54%, P < 0.05). Deferiprone treatment tended to decrease muscle iron levels in mdx mice (-30%, P < 0.1), which was associated with lower oxidative stress and fibrosis, but suppressed haem-containing proteins and mitochondrial content. Increasing iron via dietary intervention elevated total muscle iron (+25%, P < 0.05) but did not aggravate the pathology. CONCLUSIONS: Muscles from dystrophic mice have increased iron levels and dysregulated iron-related proteins that are associated with dystrophic pathology. Muscle iron levels were manipulated by iron chelation and iron enriched feed. Iron chelation reduced fibrosis and reactive oxygen species (ROS) but also suppressed haem-containing proteins and mitochondrial activity. Conversely, iron supplementation increased ferritin and haem-containing proteins but did not alter ROS, fibrosis, or mitochondrial activity. Further studies are required to investigate the contribution of impaired ferritin breakdown in the dysregulation of iron homeostasis in DMD.

Paeonia lactiflora extract improves the muscle function of mdx mice, an animal model of Duchenne muscular dystrophy, via downregulating the high mobility group box 1 protein.

ETHNOPHARMACOLOGICAL RELEVANCE: Paeonia lactiflora Pall. is an ethnopharmacological medicine with a long history of human use for treating various inflammatory diseases in many Asian countries. AIM OF THE STUDY: Duchenne muscular dystrophy (DMD) is an X-linked degenerative muscle disease affecting 1 in 3500 males and is characterized by severe muscle inflammation and a progressive decline in muscle function. This study aimed to elucidate the effects of an ethanol extract of the root of Paeonia lactiflora Pall. (PL) on the muscle function in the muscular dystrophy X-linked (mdx) mouse, the most commonly used animal model of DMD. MATERIALS AND METHODS: Male mdx mice and wild-type controls aged 5 weeks were orally treated with PL for 4 weeks. The corticosteroid prednisolone was used as a comparator drug. Muscle strength and motor coordination were assessed via the grip-strength and rotarod tests, respectively. Muscle damage was evaluated via histological examination and assessment of plasma creatine-kinase activity. Proteomic analyses were conducted to identify the muscle proteins whose levels were significantly affected by PL (ProteomeXchange identifier: PXD028886). Muscle and plasma levels of these proteins, and their corresponding mRNAs were measured using western blotting and ELISA, and quantitative reverse transcription-polymerase chain reaction, respectively. RESULTS: The muscle strength and motor coordination of mdx mice were significantly increased by the oral treatment of PL. PL significantly reduced the histological muscle damage and plasma creatine-kinase activity. Proteomic analyses of the muscle showed that PL significantly downregulated the high mobility group box 1 (HMGB1) protein and Toll-like receptor (TLR) 4, thus suppressing the HMGB1-TLR4-NF-κB signaling, in the muscle of mdx mice. Consequently, the muscle levels of proinflammatory cytokines/chemokines, which play crucial roles in inflammation, were downregulated. CONCLUSION: PL improves the muscle function and reduces the muscle damage in mdx mice via suppressing the HMGB1-TLR4-NF-κB signaling and downregulating proinflammatory cytokines/chemokines.