Correlations between clinical severity, genotype and muscle pathology in limb girdle muscular dystrophy type 2A

Background

Limb girdle muscular dystrophy type 2A (LGMD2A) is characterised by wide variability in clinical features and rate of progression. Patients with two null mutations usually have a rapid course, but in the remaining cases (two missense mutations or compound heterozygote mutations) prognosis is uncertain.

Methods

We conducted what is to our knowledge the first systematic histopathological, biochemical and molecular investigation of 24 LGMD2A patients, subdivided according to rapid or slow disease progression, to determine if some parameters could correlate with disease progression.

Results

We found that muscle histopathology score and the extent of regenerating and degenerating fibres could be correlated with the rate of disease course when the biochemical and molecular data do not offer sufficient information. Comparison of clinical and muscle histopathological data between LGMD2A and four other types of LGMD (LGMD2B–E) also gave another important and novel result. We found that LGMD2A has significantly lower levels of dystrophic features (ie degenerating and regenerating fibres) and higher levels of chronic changes (ie lobulated fibres) compared with other LGMDs, particularly LGMD2B. These results might explain the observation that atrophic muscle involvement seems to be a clinical feature peculiar to LGMD2A patients.

Conclusions

Distinguishing patterns of muscle histopathological changes in LGMD2A might reflect the effects of a disease‐specific pathogenetic mechanism and provide clues complementary to genetic data.     

New type of X-linked progressive muscular dystrophy involving shoulder girdle and back

A new type of X-linked muscular dystrophy is described in a family in which 7 men had boyhood onset of progressive dystrophy involving muscles of the shoulder and back but not the calves or face. The scapula-back muscles are affected, but the calf muscles are normal. All patients are still able to walk. The oldest patient is now 37 years old. The muscular dystrophy has been specified by electromyography, pathologic tissue microscopic examination, electron microscopic study, and elevated CK. This type of muscular dystrophy has not been reported previously.     

Characterization of lobulated fibers in limb girdle muscular dystrophy type 2A by gene expression profiling

Limb girdle muscular dystrophy type 2A (LGMD2A) is caused by mutations in CAPN3, which encodes an intracellular cysteine protease. To elucidate the fundamental molecular changes that may be responsible for the pathological features of LGMD2A, we employed cDNA microarray analysis. We divided LGMD2A muscles into two groups according to specific pathological features: an early-stage group characterized by the presence of active necrosis and a regeneration process and a later-stage group characterized by the presence of lobulated fibers. After comparing the gene expression profiles of the two groups of LGMD2A muscles with control muscles, we identified 29 genes whose mRNA expression profiles were specifically altered in muscles with lobulated fibers. Interestingly, this group included genes that encode actin filament binding and regulatory proteins, such as gelsolin, PDZ and LIM domain 3 (PDLIM3) and troponin I1. Western blot analysis confirmed the upregulation of these proteins. From these results, we propose that abnormal increased expression of actin filament binding proteins may contribute to the changes of the intra-myofiber structures, observed in lobulated fibers in LGMD2A.     

Pathophysiology of limb girdle muscular dystrophy type 2A: hypothesis and new insights into the IkappaBalpha/NF-kappaB survival pathway in skeletal muscle

Limb girdle muscular dystrophies (LGMDs) are a group of clinically heterogeneous genetic diseases characterized by progressive weakness and atrophy of scapular and pelvic muscles, with either a dominant or recessive autosomic mode of inheritance. The first symptoms of the disorder appear during the first 20 years of life and progresses gradually, and a walking disability develops 10-20 years later. The gene responsible for LGMD2A has been identified and encodes calpain 3, a protease expressed mainly in skeletal muscle. Apoptotic myonuclei were recently detected in muscular biopsy specimens of LGMD2A patients, and apoptosis was found to be correlated with altered subcellular distribution of inhibitory protein kappaBalpha (IkappaBalpha) and nuclear factor kappaB (NF-kappaB), resulting in sarcoplasmic sequestration of NF-kappaB. Calpain 3 dependent IkappaBalpha degradation was reconstituted in vitro, supporting a possible in vivo sequence of events leading from calpain 3 deficiency to IkappaBkappa accumulation, prevention of nuclear translocation of NF-kappaB, and ultimately apoptosis. Therefore calpain 3, present in healthy muscle as sarcoplasmic and nuclear forms, may control IkappaBalpha turnover and indirectly regulate NF-kappaB dependent expression of survival genes. Recent data reported from a new model of LGMD2A in mice and from other muscular disorders strengthen understanding of the molecular links between calpain 3 and the Ikappaalpha/NF-kappaB pathway. Finally, in light of the lack of apoptosis observed in inflammatory myopathies, a unifying model for the control of cell survival in muscle is proposed and discussed     

Electromyographic pattern in Duchenne and Becker muscular dystrophy. Part II. Electromyographic pattern in Becker muscular dystrophy in comparison with Duchenne muscular dystrophy

BACKGROUND: The aim of this study was to compare the electromyographic pattern in Becker muscular dystrophy (BMD) with that found in Duchenne muscular dystrophy (DMD). MATERIAL AND METHOD: Fourteen men with BMD and 51 boys with DMD were investigated. Proximal muscles were examined: m. biceps brachii (BB) and m. rectus femoris (RF). They were divided according to the clinical criteria in two groups: of those with slight changes (group AB) and of those with severe abnormalities (CD). As in the Part I of the paper our own method of Functional-QEMG was applied in the CNEMG examination. RESULTS AND CONCLUSIONS: Spontaneous activity (fibrillations, complex repetitive discharges) was equally frequent in BMD and DMD. Linked potentials were rather frequent in either group. Myopathic features such as MUAPs low amplitude and area, polyphasic shape were seen in either condition, but more marked in DMD than in BMD. Evaluation of IP recordings revealed that IP amplitude (amplitude size) is low in DMD already at the early stage of lesion but normal or only slightly diminished in BMD. It might perhaps suggest different degrees of lesion in type II MUs between the compared types of muscular dystrophy.     

Alpha7beta1 integrin does not alleviate disease in a mouse model of limb girdle muscular dystrophy type 2F

Transgenic expression of the alpha7beta1 integrin in the dystrophic mdx/utr-/- mouse decreases development of muscular dystrophy and enhances longevity. To explore the possibility that elevating alpha7beta1 integrin expression could also ameliorate different forms of muscular dystrophy, we used transgenic technology to enhance integrin expression in mice lacking delta-sarcoglycan (delta sgc), a mouse model for human limb girdle muscular dystrophy type 2F. Unlike alpha7 transgenic mdx/utr-/- mice, enhanced alpha7beta1 integrin expression in the delta sgc-null mouse did not alleviate muscular dystrophy in these animals. Expression of the transgene in the delta sgc-null did not alleviate dystrophic histopathology, nor did it decrease cardiomyopathy or restore exercise tolerance. One hallmark of integrin-mediated alleviation of muscular dystrophy in the mdx/utr-/- background is the restoration of myotendinous junction integrity. As assessed by atomic force microscopy, myotendinous junctions from normal and delta sgc-null mice were indistinguishable, thus suggesting the important influence of myotendinous junction integrity on the severity of muscular dystrophy and providing a possible explanation for the inability of enhanced integrin expression to alleviate dystrophy in the delta sgc-null mouse. These results suggest that distinct mechanisms underlie the development of the diseases that arise from deficiencies in dystrophin and sarcoglycan.     

Electromyographic pattern in Duchenne and Becker muscular dystrophy. Part I: Electromyographic pattern in subsequent stages of muscle lesion in Duchenne muscular dystrophy

BACKGROUND: The study was aimed to evaluate the EMG pattern in myopathy, in the muscles with early, slight abnormalities and in the muscles severely affected, and to analyze the sequence of abnormalities appearing with the progress of the impairment. MATERIAL AND METHOD: Fifty one boys with Duchenne muscular dystrophy were studied. Ninety nine proximal muscles (biceps brachii and rectus femoris) were examined. According to clinical criteria (defective force, atrophy) the muscles were assigned to the group of slight changes (group AB: 50 muscles) and to the group of severe abnormalities (group CD: 49 muscles). An own method of Functional-QEMG was applied in the CNEMG examination. RESULTS AND CONCLUSIONS: The sensitivity of the method applied allowed the detection of myogenic changes even at the subclinical, oligosymptomatic (group AB) stage of impairment. The earliest EMG abnormality detected was an increased percentage of the polyphasic potentials, even if other MUAPs parameters remained normal. However at this stage already a decrease appeared of amplitude, area and MUAPs duration, along with the decrease of IP amplitude and amplitude size. An increase of IP density and a slight increase of number of stable-shaped potentials (simple, but more often polyphasic) was also seen which we believed to reflect the compensatory process. At the further stage, when compensating mechanisms were no longer possible, a dramatic decrease appeared of the IP amplitude, amplitude size and density and, sometimes, also a decrease in the number of stable-shaped potentials. The structural changes such as decrease of amplitude, area and MUAPs duration also progressed. These findings reflect in our study the sequence of EMG abnormalities in the successive stages of myogenic lesion.     

Cardiac dysrhythmias,cardiomyopathy and muscular dystrophy in patients with Emery-Dreifuss muscular dystrophy and limb-girdle muscular dystrophy type 1B

Emery-Dreifuss muscular dystrophy (EDMD) and limb-girdle muscular dystrophy type 1B (LGMD1B) are characterized by cardiac dysrhythmias, late-onset cardiomyopathy, slowly progressive skeletal myopathy and contractures of the neck, elbows and ankles. The causative mutation is either in the emerin gene (X-linked recessive EDMD) or lamin A/C gene (autosomal dominant EDMD2 or LGMD1B). We report three cases of EDMD, EDMD2 and LGMD1B. A 14-yr-old boy showed limitation of cervical flexion and contractures of both elbows and ankles. Sinus arrest with junctional escape beats was noted. He was diagnosed as X-linked recessive EDMD (MIM 310300). A 28-yr-old female showed severe wasting and weakness of humeroperoneal muscles. Marked limitation of cervical flexion and contractures of both elbows and ankles were noted. Varying degrees of AV block were noted. She was diagnosed as autosomal dominant EDMD2 (MIM 181350). A 41-yr-old female had contractures of both ankles and limb-girdle type muscular dystrophy. ECG revealed atrial tachycardia with high grade AV block. She was diagnosed as autosomal dominant LGMD1B (MIM 159001). Cardiac dysrhythmias in EDMD and LGMD1B include AV block, bradycardia, atrial tachycardia, atrial fibrillation, and atrial standstill, causing sudden death necessitating pacemaker implantation. Cardiologists should know about these unusual genetic diseases with conduction defects, especially in young adults.     

Identical mutation in patients with limb girdle muscular dystrophy type 2B or Miyoshi myopathy suggests a role for modifier gene(s)

Limb girdle muscular dystrophy type 2B (LGMD2B) and Miyoshi myopathy (MM), a distal muscular dystrophy, are both caused by mutations in the recently cloned gene dysferlin, gene symbol DYSF. Two large pedigrees have been described which have both types of patient in the same families. Moreover, in both pedigrees LGMD2B and MM patients are homozygous for haplotypes of the critical region. This suggested that the same mutation in the same gene would lead to both LGMD2B or MM in these families and that additional factors were needed to explain the development of the different clinical phenotypes. In the present paper we show that in one of these families Pro791 of dysferlin is changed to an Arg residue. Both the LGMD2B and MM patients in this kindred are homozygous for this mutation, as are four additional patients from two previously unpublished families. Haplotype analyses suggest a common origin of the mutation in all the patients. On western blots of muscle, LGMD2B and MM patients show a similar abundance in dysferlin staining of 15 and 11%, respectively. Normal tissue sections show that dysferlin localizes to the sarcolemma while tissue sections from MM and LGMD patients show minimal staining which is indistinguishable between the two types. These findings emphasize the role for the dysferlin gene as being responsible for both LGMD2B and MM, but that the distinction between these two clinical phenotypes requires the identification of additional factor(s), such as modifier gene(s).     

Hutterite brothers both affected with two forms of limb girdle muscular dystrophy: LGMD2H and LGMD2I

Limb girdle muscular dystrophy (LGMD) is very common in the Hutterite population of the North American Prairies. We have recently reported the homozygous c.1459G>A mutation in TRIM32 associated with LGMD2H. We have also identified Hutterite patients with LGMD2I, homozygous for the common c.826C>A mutation in FKRP. To date, all Hutterites with LGMD have been shown to be homozygous for either the TRIM32 or FKRP mutation. We now report a Hutterite family in which both parents and five sons were all found to be homozygous for the TRIM32 mutation. The father had slowly progressive proximal muscle weakness, whereas three sons and their mother, all currently asymptomatic, had normal physical examinations. The remaining two sons (7 and 10 years old), presented with mild decrease in stamina, had normal neuromuscular examinations and were found to be homozygous for the FKRP mutation in addition to the TRIM32 mutation. These two boys do not differ in age at or mode of presentation, physical findings, or serum CK levels compared to age-matched individuals affected with LGMD2I alone. This suggests that the effects of these two mutations are not acting synergistically at this time. It remains to be seen whether there will be signs of interaction between these two mutations as the patients get older.     

Biochemical characterization of muscle tissue of limb girdle muscular dystrophy: an 1H and 13C NMR study

The metabolic differences between the muscle biopsies of patients with limb girdle muscular dystrophy (LGMD) and normal controls were characterized using high-resolution 1H and 13C NMR spectroscopy. In all, 44 metabolites were unambiguously assigned in the perchloric acid extracts of skeletal muscle tissue, using 2D double quantum filtered (DQF COSY), total correlation (TOCSY), and 1H/13C heteronuclear multiple quantum coherence (HMQC) spectroscopy. The concentrations of glycolytic substrate, glucose (p=0.03), gluconeogenic amino acids, glutamine (p=0.02) and alanine (p=0.009) together with glycolytic product, lactate (p=0.04), were found to be significantly lowered in LGMD patients as compared with controls. The reduction in the concentration of glucose may be attributed to the decrease in the concentration of gluconeogenic amino acids in the degenerated muscle. Reduction in the rate of anaerobic glycolysis and lowered substrate concentration appear to be the possible reasons for the decrease in the concentration of lactate. A significant reduction in the concentration of choline in LGMD patients was also observed compared with controls. Lower concentration of choline may be the result of decreased rate of membrane turnover in LGMD patients. The data presented here provide an insight into the potentials of in-vitro NMR spectroscopy in the study of muscle metabolism.     

A novel biallelic variant in the Popeye domain-containing protein 1 (POPDC1) underlies limb girdle muscle dystrophy type 25

POPDC1 also known as BVES, is a highly conserved transmembrane protein, important for striated muscle function and homeostasis. Pathogenic variants in the POPDC1 gene are associated with limb-girdle muscular dystrophy type 25 (LGMDR25). In the present study, we performed trio-whole exome sequencing (WES) followed by Sanger sequencing on a single family having LGMD clinical features. Protein modeling of all POPDC1 missense variants (POPDC1^Pro134Leu, POPDC1^Ile193Ser, and POPDC1^Ser201Phe) associated with LGMDR25 were performed using Molecular Dynamics (MD) simulation. We identified a homozygous missense variant (c.401C>T; p.Pro134Leu) in the POPDC1 gene. Altered 3D structure, disruptive fluctuation, less compactness, and instability were observed in all the three variants of POPDC1 protein models. In comparison, POPDC1^Ser201Phe protein dynamics were more unstable than other variants. Functional study of newly identified variant would add key answers to underlying mechanisms of the disease.     

LIMS2 mutations are associated with a novel muscular dystrophy,severe cardiomyopathy and triangular tongues

Limb girdle muscular dystrophy (LGMD) is a heterogeneous group of genetic disorders leading to progressive muscle degeneration and often associated with cardiac complications. We present two adult siblings with childhood‐onset of weakness progressing to a severe quadriparesis with the additional features of triangular tongues and biventricular cardiac dysfunction. Whole exome sequencing identified compound heterozygous missense mutations that are predicted to be pathogenic in LIMS2. Biopsy of skeletal muscle demonstrated disrupted immunostaining of LIMS2. This is the first report of mutations in LIMS2 and resulting disruption of the integrin linked kinase (ILK)–LIMS–parvin complex associated with LGMD.     

A rare form of limb girdle muscular dystrophy (type 2E) seen in an Iranian family detected by autozygosity mapping

Sarcoglycanopathies (SGPs) constitute a subgroup of autosomal recessive limb girdle muscular dystrophies (LGMDs) which are caused by mutations in sarcoglycan (SGs) genes. SG proteins form a core complex consisting of α, β, γ and δ sarcoglycans which are encoded by SGCA, SGCB, SGCG and SGCD genes, respectively. Genetic defect, in any of these SG proteins, results in instability of the whole complex. This effect can be helpful in interpreting muscle biopsy results. Autozygosity mapping is a gene mapping approach which can be applied in large consanguineous families for tracking the defective gene in most autosomal recessive disorders. In the present study, we used autozygosity mapping, to find the gene responsible for muscular dystrophy. Proband was a 10-year-old boy referred to our center for ruling out DMD (Duchenne muscular dystrophy). According to the pedigree and clinical reports, we assessed him for SGPs. Haplotyping, using the four short tandem repeat (STR) markers for each of the SG genes, showed that the phenotype may segregate with SGCB gene; and observing two crossing overs which occurred within the gene suggested that the mutation might be in the first two exons of SGCB gene. Mutation analysis showed a 26?bp duplication (10?bp before the initiation codon till 13?bp after the ATG start codon). This will cause a frameshift in protein synthesis.     

Mutations in the fukutin-related protein gene (FKRP) identify limb girdle muscular dystrophy 2I as a milder allelic variant of congenital muscular dystrophy MDC1C.

The limb girdle and congenital muscular dystrophies (LGMD and CMD) are characterized by skeletal muscle weakness and dystrophic muscle changes. The onset of symptoms in CMD is within the first few months of life, whereas in LGMD they can occur in late childhood, adolescence or adult life. We have recently demonstrated that the fukutin-related protein gene (FKRP) is mutated in a severe form of CMD (MDC1C), characterized by the inability to walk, leg muscle hypertrophy and a secondary deficiency of laminin alpha2 and alpha-dystroglycan. Both MDC1C and LGMD2I map to an identical region on chromosome 19q13.3. To investigate whether these are allelic disorders, we undertook mutation analysis of FKRP in 25 potential LGMD2I families, including some with a severe and early onset phenotype. Mutations were identified in individuals from 17 families. A variable reduction of alpha-dystroglycan expression was observed in the skeletal muscle biopsy of all individuals studied. In addition, several cases showed a deficiency of laminin alpha2 either by immunocytochemistry or western blotting. Unexpectedly, affected individuals from 15 families had an identical C826A (Leu276Ileu) mutation, including five that were homozygous for this change. Linkage analysis identified at least two possible haplotypes in linkage disequilibrium with this mutation. Patients with the C826A change had the clinically less severe LGMD2I phenotype, suggesting that this is a less disruptive FKRP mutation than those found in MDC1C. The spectrum of LGMD2I phenotypes ranged from infants with an early presentation and a Duchenne-like disease course including cardiomyopathy, to milder phenotypes compatible with a favourable long-term outcome.     

Herbal medicine significantly improved muscle function in a patient with type 1 facioscapulohumeral muscular dystrophy: A case report

Background and aimFacioscapulohumeral muscular dystrophy (FSHD) is a common muscular disorder. At present, treatments for FSHD have limited effects on the muscle function of patients. A famous Chinese medicine formula, Buzhong Yiqi (BZYQ), has shown promising effects on several muscular diseases, but evidence regarding its effect on FSHD is lacking. This study aimed to examine the effect of BZYQ on FSHD.Case presentationA 15-year-old girl suffered from progressive muscle weakness, with a genetically confirmed diagnosis of FSHD. Except for routine FSHD management, the patient received BZQY every day. The muscle strength of the patient remarkably increased after discharge.ConclusionsThis study was novel in reporting a significant improvement in muscle function in a patient with FSHD treated with an integrated approach of BZYQ and routine management. Therefore, BZYQ might be a potential treatment for FSHD, requiring further investigations.     

Clinical and pathological characteristics of four Korean patients with limb-girdle muscular dystrophy type 2B

Limb-girdle muscular dystrophy type 2B (LGMD2B), a subtype of autosomal recessive limb-girdle muscular dystrophy (ARLGMD), is characterized by a relatively late onset and slow progressive course. LGMD2B is known to be caused by the loss of the dysferlin protein at sarcolemma in muscle fibers. In this study, the clinical and pathological characteristics of Korean LGMD2B patients were investigated. Seventeen patients with ARLGMD underwent muscle biopsy and the histochemical examination was performed. For the immunocytochemistry, a set of antibodies against dystrophin, alpha, beta, gamma, delta-sarcoglycans, dysferlin, caveolin-3, and beta-dystroglycan was used. Four patients (24%) showed selective loss of immunoreactivity against dysferlin at the sarcolemma on the muscle specimens. Therefore, they were classified into the LGMD2B category. The age at the onset of disease ranged from 9 yr to 33 yr, and none of the patients was wheelchair bound at the neurological examination. The serum creatine kinase (CK) was high in all the patients (4010-5310 IU/L). The pathologic examination showed mild to moderate dystrophic features. These are the first Korean LGMD2B cases with a dysferlin deficiency confirmed by immunocytochemistry. The clinical, pathological, and immunocytochemical findings of the patients with LGMD2B in this study were in accordance with those of other previous reports.     

Analysis of motor control and kinesthetic perception in patients with progressive muscular dystrophy

The purpose of this study was to examine motor control and kinesthetic perception of upper extremity in patients with Duchenne progressive muscular dystrophy. Nine normal subjects and nine subjects with muscular dystrophy performed a pursuit tracking task with step wave target by means of isometric contraction, and simultaneously estimated magnitude of muscular tension during tracking behavior. The results were as follows: (1) the muscular electrical activity measured from the EMGs was directly proportional to the muscular tension for both the normal and the muscular dystrophy groups, (2) the speed of step response for the muscular dystrophy group was slower than that for the normal group, and (3) the exponent of power function for the muscular dystrophy group tended to be smaller than that for the normal group. These results were discussed in terms of the stage of disability in progressive muscular dystrophy.