Episodes of exercise-induced dark urine and myalgia in LGMD 2I

Lindberg C, Sixt C, Oldfors A. Episodes of exercise‐induced dark urine and myalgia in LGMD 2I.
Acta Neurol Scand: 2012: 125: 285–287.
© 2011 John Wiley & Sons A/S. Background – Mutations in the fukutin‐related protein gene FKRP (MIM *606596) cause a form of congenital muscular dystrophy (MDC1C) and also limb girdle muscular dystrophy type 2I (LGMD2I). Exercise‐induced myoglobinuria, frequently occurring in metabolic myopathies, has been described in Becker muscular dystrophy and in a few cases of LGMD. Objectives – To describe that episodes with myoglobinuria, often associated with exercise‐induced myalgia, may be common and a presenting symptom in patients with LGMD2I. Methods – Data on episodes of suspected myoglobinuria and myalgia were collected from the patient records on 14 patients with a diagnosis of LGMDI. Results – Five LGMD2I patients reported recurrent episodes of dark urine and myalgia after exercise, and in three of them, this was the only symptom for several years. Conclusions – We conclude that episodes compatible with exercise‐induced myoglobinuria may be frequent in LGMD2I.     

Different localization of dystrophin in developing and adult human skeletal muscle

Duchenne and Becker muscular dystrophy are caused by defects in dystrophin synthesis. Using affinity-purified polyclonal anti-dystrophin antibodies, we have studied immunohistochemically the subcellular localization of dystrophin in embryonic, fetal, and adult human skeletal muscle. In the embryonic stages dystrophin first appears in the sarcoplasm at the peripheral ends of the myotubes, immediately adjacent to the tendons, whereas in fetal stages dystrophin is found throughout the entire myofibers. In agreement with literature data, in adult muscle dystrophin expression was found to be restricted to the sarcolemma. The sarcoplasmic localization in embryonic and fetal tissue and the sarcolemmal localization of dystrophin in mature muscle suggests the accumulation of dystrophin in the cytoplasm prior to its integration into the membrane. These results increase our knowledge of the ontogenesis of dystrophin and may lead to a better understanding of the great diversity in pathological cases of Duchenne and Becker muscular dystrophy.     

Direct dystrophin and reporter gene transfer into dog muscle in vivo

Bacterial β-galactosidase cDNA was injected without lipofectin into 41 sites in dog muscle and expression was seen in 22 of them. The cDNA and lipofectin was injected into 35 similar sites and expression was seen in 21. Expression was seen in a maximum of 2.5% of muscle fibers and 23.21% of nonmuscle cells. A total of 106 muscle sites were injected with the minigene with and without lipofectin. In 4 of the 45 sites injected with the minigene without lipofectin human dystrophin was expressed around the periphery of 0.3% of the fibers. Bacterial β-galactosidase cDNA was injected into the peritoneal cavity of 4 pups, 2 of which also received lipofectin. In all 4, expression was seen in liver, spleen, and mesenteric lymph node. In the 2 pups that received lipofectin, expression was also seen in the diaphragm, intercostal, and abdominal muscles of 1 and in the diagphragm and intercostal muscles of the other. These experiments show that human dystrophin transgene expression can be obtained in dog muscle. However, other methods will be required to increase the degree of expression before gene therapy trials can be undertaken. © 1998 John Wiley & Sons, Inc. Muscle Nerve 21: 159–165, 1998     

Sub-cellular localisation of fukutin related protein in different cell lines and in the muscle of patients with MDC1C and LGMD2I

MDC1C and LGMD2I are two allelic forms of muscular dystrophies caused by mutations in the gene encoding for fukutin related protein (FKRP). FKRP encodes for a putative glycosyltransferase, the precise function of which is unknown. However, the marked reduction of -dystroglycan glycosylation in the muscle of MDC1C and LGMD2I patients suggests a role for FKRP in dystroglycan processing. Using a polyclonal antibody raised against FKRP we now show that endogenous FKRP locates to the Golgi apparatus of neuronal, oligodendroglial, and the cardiac muscle cell line H9c2. In differentiated C2C12 myotubes and in transverse sections of normal skeletal and cardiac muscle, endogenous FKRP surrounded the myonuclei. This localisation was unaffected in the skeletal muscle of patients with MDC1C and LGMD2I carrying various FKRP mutations. These observations imply a specific role for FKRP during striated muscle, neuronal and glial development and suggest that protein mis-localisation is not a common mechanism of disease in FKRP-related dystrophies.     

Dystrophin-associated protein abnormalities in dystrophin-deficient muscle fibers from symptomatic and asymptomatic Duchenne/Becker muscular dystrophy carriers

The absence of dystrophin in muscle fibers is associated with a major reduction in dystrophin-associated proteins (DAPs) and disruption of the linkage between the subsarcolemmal cytoskeleton and the extracellular matrix. We investigated the expression of the DAPs β-dystroglycan, α-sarcoglycan, γ-sarcoglycan and syntrophin as well as utrophin in the muscles of 13 Duchenne muscular dystrophy (DMD) carriers (with variable percentages of dystrophin-deficient fibers and with a range of clinical symptoms), 2 Becker muscular dystrophy (BMD) carriers (expressing a highly truncated protein in some fibers), 2 girls with a DMD-like phenotype, and 11 BMD carriers with almost normal dystrophin expression (reduced or patchy distribution in a few fibers only and rare dystrophin-deficient fibers). DAPs were highly reduced in all fibers lacking dystrophin in the DMD carriers, but were almost normal in the dystrophin-deficient fibers of the 2 BMD carriers with highly truncated dystrophin. In the 11 BMD carriers with nearly normal dystrophin, the few fibers with reduced or patchy dystrophin immunostaining also showed reduced DAP expression in correlation with dystrophin expression. Immunoblot for β-dystroglycan and α-sarcoglycan confirmed the immunohistochemical findings. Utrophin expression was slightly increased in a proportion of fibers in the DMD and BMD carriers with dystrophin mosaicism. We found no correlation between utrophin expression and DAP expression. We conclude that absence or reduction of dystrophin in muscle fibers of DMD and BMD carriers causes a reduction of DAPs in the same fibers, as observed in DMD and BMD patients, while utrophin does not seem to play a role in DAP expression in adult muscle. Received: 11 January 1996 / Revised, accepted: 16 April 1996     

Diagnostic value of muscle MRI in differentiating LGMD2I from other LGMDs

Mutations in the fukutin–related protein (FKRP) have recently been demonstrated to cause limb girdle muscular dystrophy type 2I (LGMD2I), one of the most common forms of the autosomal recessive LGMDs in Europe. We performed a systematic clinical and muscle MRI assessment in 6 LGMD2I patients and compared these findings with those of 14 patients with genetically confirmed diagnosis of other forms of autosomal recessive LGMDs or dystrophinopathies. All LGMD2I patients had a characteristic clinical phenotype with predominant weakness of hip flexion and adduction, knee flexion and ankle dorsiflexion. These findings were also mirrored on MRI of the lower extremities which demonstrated marked signal changes in the adductor muscles, the posterior thigh and posterior calf muscles. This characteristic clinical and MRI phenotype was also seen in LGMD2A. However, in LGMD2A there was a selective involvement of the medial gastrocnemius and soleus muscle in the lower legs which was not seen in LGMD2I. The pattern in LGMD2A and LGMD2I were clearly different from the one seen in alpha–sarcoglycanopathy and dystrophinopathy type Becker which showed marked signal abnormalities in the anterior thigh muscles. Our results indicate that muscular MRI is a powerful tool for differentiating LGMD2I from other forms of autosomal recessive LGMDs and dystrophinopathies.     

Frequent low penetrance mutations in the Lamin A/C gene,causing Emery Dreifuss muscular dystrophy

Emery Dreifuss muscular dystrophy is a genetically heterogeneous disorder characterized by the clinical triad of early onset contractures, progressive muscular wasting and weakness with humeroperoneal distribution and cardiac conduction defects. Mutations in the Lamin A/C (LMNA) gene are responsible for the autosomal dominant and the autosomal recessive forms. Familiar and sporadic patients carrying mutations in the LMNA gene show high variability in the clinical symptomatology and age of onset. In this report, we describe four families harboring missense mutations in the LMNA gene and we show that the effect of mutations ranges from silent to fully penetrant. We suggest that incomplete penetrance of dominant mutations in the LMNA gene is a common feature and we emphasize the significance of mutational analysis in relatives of sporadic cases of laminopathies, as asymptomatic carriers face high risk of sudden cardiac death.     

How does dystrophin deficiency lead to muscle degeneration? — Evidence from the MDX mouse

The mdx mouse has a defect in the same gene as boys with Duchenne muscular dystrophy, which results in the absence of the protein product, dystrophin. A large number of recent studies have used the mdx mouse model to examine the potential role of dystrophy in normal muscle and the mechanisms by which dystrophin-deficiency leads to myopathy. This review discusses critically the results of these studies and their relevance to understanding the mechanisms by which dystrophin-deficiency leads to muscle necrosis.     

Binding of Ricinus communis I lectin to the muscle cell plasma membrane in diseased muscle

Using an electron histochemical technique, we have observed the binding of a D-galactose-specific lectin to the muscle cell plasma membrane in muscle biopsies taken from patients with various neuromuscular disorders. In spinal muscular atrophy, the only neurogenic disease studied, the plasma membrane stained as in normal muscle. However, in the myopathies Becker and limb-girdle muscular dystrophy and in the polymyositis there was a reduction in both the occurrence and the intensity of staining of the plasma membrane. Reduced lectin binding by the plasma membrane probably reflects secondary changes in the composition of glycoproteins and/or glycolipids in the membrane and seems to be common to all these myopathies to varying degrees.     

Dysregulation and cellular mislocalization of specific miRNAs in myotonic dystrophy type 1

Myotonic Dystrophy Type-1 (DM1) is caused by the expansion of a CTG repeat with a peculiar pattern of multisystemic involvement affecting skeletal muscles, the heart, the eye, the central nervous system and the endocrine system. Since microRNA expression is disrupted in several myopathies, the expression of 24 candidate microRNAs was analyzed in skeletal muscle biopsies of 15 DM1 patients. Controls were constituted by biopsies without overt pathological features derived from 14 subjects with suspected neuromuscular disorder of undetermined nature. We found that miR-1 and miR-335 were up-regulated, whereas miR-29b and c, and miR-33 were down-regulated in DM1 biopsies compared to controls. We also found that the cellular distribution of muscle specific miR-1, miR-133b and miR-206 was severely altered in DM1 skeletal muscles. MicroRNA dysregulation was likely functionally relevant, since it impacted on the expression of the predicted miR-1, and miR-29 targets. The observed miRNA dysregulations and myslocalizations may contribute to DM1 pathogenetic mechanisms.     

Regeneration-blocked mdx muscle: in vivo model for testing treatments

We have refined the mdx mouse as a clinical model for Duchenne dystrophy. Our power estimates, primary measures, regular sacrifice intervals, and quality checks constitute a high-speed, low-cost system for preclinically testing therapies designed to slow muscle destruction in Duchenne dystrophy. Irradiated (18 Gy) and contralateral shielded anterior tibial muscles were studied in 21-day-old mdx and normal mice at the time of irradiation and 4, 8, 12, 16, and 20 weeks thereafter. Regeneration-blocked mdx (irradiated) muscle expressed muscular dystrophy as progressive wasting after a brief (4 week) period of growth. Regeneration-blocked normal muscle showed stunted growth but neither progressive wasting nor microscopic pathological changes. © 1997 John Wiley & Sons, Inc. Muscle Nerve 20: 1016–1023, 1997     

Skeletal muscle pathology in autosomal dominant Emery-Dreifuss muscular dystrophy with lamin A/C mutations

We present our observations on the skeletal muscle pathology of nine cases from seven families of autosomal dominant Emery-Dreifuss muscular dystrophy (ADEDMD) with identified mutations in the lamin A/C gene, aged 2-35 years at the time of biopsy. The severity of pathological change was moderate and the most common features were variation in fibre size (hypertrophy and atrophy), an increase in internal nuclei and smaller diameter fibres with high oxidative enzyme activity. Only one case showed necrosis, which was present in two separate samples taken from the quadriceps and tibialis anterior, at different ages. Immunocytochemistry detected an age-related reduction of laminin beta1 on the muscle fibres in adolescent and adult cases. Antibodies to lamins A and A/C, and emerin did not reveal any detectable differences from controls. Electron microscopy of two out of three cases showed an abnormal distribution of heterochromatin in many fibre nuclei. Our results show that dystrophic changes in skeletal muscle are not a major feature of ADEDMD, and that nuclear abnormalities may be detected with electron microscopy. Immunodetection of reduced laminin beta1 may be a useful secondary marker in adults with this disorder, as immunocytochemistry of lamins is not yet of diagnostic use.     

Fukuyama-type congenital muscular dystrophy: a case report in the Japanese population living in Brazil

INTRODUCTION: We present herein clinical, histological and magnetic resonance imaging (MRI) findings in a patient with Fukuyama-type congenital muscular dystrophy (FCMD). He is the first case report in the Japanese population living in Brazil. CASE REPORT: The child presented with neonatal hypotonia, delayed motor abilities and speech, seizures, cerebral and cerebellar gyrus abnormalities with signal intensity change in the white matter by MRI, high serum level of creatinephosphokinase (CK), and dystrophic skeletal muscle with normal merosin, alpha-sarcoglycan and dystrophin expression. The fukutin gene study showed one founder 3-kb retrotransposal insertion in the 3'-non-coding region, and in the other allele no mutation was detected after screening all exons and flanking introns by sequencing. DISCUSSION: This case report emphasizes the importance to consider FCMD in Japanese people living in other countries.     

Electrical impedance myography for the in vivo and ex vivo assessment of muscular dystrophy (mdx) mouse muscle

Introduction: Sensitive, non‐invasive techniques are needed that can provide biomarkers of disease status and the effects of therapy in muscular dystrophy. Methods: We evaluated electrical impedance myography (EIM) to serve in this role by studying 2‐month‐old and 18‐month‐old mdx and wild‐type (WT) animals (10 animals in each of 4 groups). Results: Marked differences were observed in EIM values between mdx and WT animals; the differences were more pronounced between the older age groups (e.g., reactance of 92.6 ±4.3 Ω for mdx animals vs. 130 ± 4.1 Ω for WT animals, P < 0.001). In addition, in vivo EIM parameters correlated significantly with the extent of connective tissue deposition in the mdx animals. Conclusions: EIM has the potential to serve as a valuable non‐invasive method for evaluating muscular dystrophy. It can be a useful biomarker to assist with therapeutic testing in both pre‐clinical and clinical studies. Muscle Nerve 49 : 829–835, 2014     

DMD、BMD患者骨骼肌、皮肤立毛肌肌营养不良蛋白同时欠损

目的研究、对比肌营养不良蛋白(dystrophin)在杜兴型肌营养不良(Duchenne muscular dystrophy,DMD)和贝克型肌营养不良(Becker muscular dystrophy,BMD)患者活检骨骼肌、皮肤立毛肌中的表达。方法用肌营养不良蛋白三个不同区域的单克隆抗体(Dystrophin-N、-C、-R)对11例DMD患者,5例BMD患者和3例其他神经肌病患者同时行活检骨骼肌、皮肤免疫组织化学染色分析。结果与对照例相比,11例DMD患者抗Dystro-phin-N、-C、-R单克隆抗体免疫组织化学染色显示:骨骼肌肌纤维膜Dystrophin-N、-C、-R呈完全欠损;皮肤立毛肌Dystrophin-N、-R完全欠损,Dystrophin-C轻微表达。5例BMD患者抗Dystrophin-N、-C、-R单克隆抗体免疫组织化学染色显示:肌营养不良蛋白在骨骼肌肌纤维膜和皮肤均呈不完全欠损。结论DMD和BMD患者肌营养不良蛋白在骨骼肌肌纤维膜、皮肤立毛肌呈完全/不完全欠损,与骨骼肌活检相同,皮肤活检也是分子病理学诊断DMD、BMD简便、易行、可靠的方法。     

Extreme variability of skeletal and cardiac muscle involvement in patients with mutations in exon 11 of the lamin A/C gene

Mutations of the LMNA gene, encoding the nuclear envelope proteins lamins A and C, give rise to Emery-Dreifuss muscular dystrophy and to limb-girdle muscular dystrophy 1B (EDMD and LGMD1B). With one exception, all the reported EDMD and LGMD1B mutations are confined to the first 10 exons of the gene. We report four separate cases, with mutations in the same codon of LMNA exon 11, characterized by remarkable variability of clinical findings, in addition to features not previously reported. One patient had congenital weakness and died in early childhood. In two other patients, severe cardiac problems arose early and, in one of these, cardiac signs preceded by many years the onset of skeletal muscle weakness. The fourth case had a mild and late-onset LGMD1B phenotype. Our cases further expand the clinical spectrum associated with mutations in the LMNA gene and provide new evidence of the role played by the C-terminal domain of lamin A.     

Global brain ischemia and reperfusion: Golgi apparatus ultrastructure in neurons selectively vulnerable to death

The neocortex and the hippocampus were examined for lipid peroxidation products and ultrastructural alterations by fluorescence and electron microscopy, respectively, in rats subjected to 10 min of cardiac arrest or 10 min cardiac arrest and either 90 or 360 min reperfusion. Lipid peroxidation products were observed after 90 min reperfusion in the perikarya and proximal dendrites of neocortical pyramidal neurons and in the hippocampal hilar cells and CA₁, region; the fluorescene was most intense at the base of the apical dendrite, the region of the Golgi apparatus. After 90 min of reperfusion, the CA₁, showed considerable stretches of rough endoplasmic reticulum devoid of ribosomes and the Golgi cisternae were shorter and widely dilated. The neocortex showed similar endoplasmic reticulum changes, but no significant alterations to the Golgi were noted. In addition there were areas where strings of ribosomes appear to be detaching from the endoplasmic reticulum. After 360 min reperfusion in both the neocortex and the hippocampus, the damage appeared more severe. The Golgi was fragmented into vacuoles, membranous whorls had appeared, and dense aggregates of smooth vesicles were seen coalescing with each other and the vacuoles. These observations suggest that early Golgi involvement is a more important marker of lethal injury than ribosome release from the endoplasmic reticulum. The areas of disturbed Golgi ultrastructure correspond to those areas that show evidence of lipid peroxidation and imply that lipid peroxidation may be causally related to the disturbance in Golgi ultrastructure.     

Deflazacort but not prednisone improves both muscle repair and fiber growth in diaphragm and limb muscle in vivo in the mdx dystrophic mouse

The effects of the glucocorticoids deflazacort and prednisone on mdx mouse dystrophy and muscle regeneration were evaluated in a 4.5-week double-blind study to test whether they would decrease dystrophy by anti-inflammatory effects [in intact diaphragm and left tibialis anterior (TA) muscle] and increase new muscle formation (after crush injury). In the left TA, fiber diameter was greater after deflazacort and prednisone compared to placebo. However, only deflazacort increased the centronucleation index of accumulated damage and repair, and myotube growth over the long term. In crush-injured TA, the fusion of proliferative muscle precursors to myotubes (by autoradiography) was increased only after deflazacort. Diaphragm muscle was much less inflamed, and fiber diameter was greater after deflazacort. Results suggest that glucocorticoids decreased the severe phenotype of dystrophy in the mdx diaphragm. Moreover, deflazacort uniquely promoted myogenic repair over short and longer terms, in addition to stimulating fiber growth. These first clues to the targets of deflazacort action on muscle repair have important positive implications for treating Duchenne dystrophy. © 1996 John Wiley & Sons, Inc.