LGMD

Introduction: Limb-girdle muscular dystrophies (LGMD) are a clinical and genetically heterogeneous group of muscle diseases presenting with a phenotypic spectrum. Autosomal dominant LGMDs represent less than 10% of the group and are subdivided, according to the last classification, in five dominant LGMD forms (type D1-D5), in which mutations in various genes have been identified. Dominant LGMD might be due to a defect of DNAJB6 (LGMD D1), transportin-3 (LGMD D2, TNPO3-related), mutations in HNRNPDL gene (LGMD D3), in CAPN3 gene (LGMD D4) and in collagen 6 (LGMD D5, collagen 6-related). Areas covered: We present the pathomechanism of dominant LGMDs and clinical management of respiratory and bulbar complications. We analyze the clinical presentation of cases affected by LGMD D2 in a large Italo-Spanish family. We consider two main phenotypic entities: the childhood form developing the clinical features in the first decade of life and the late-onset phenotype, that present distinct clinical, histopathological and MRI features. The main features in dominant LGMDs, such as LGMD D1, D3, D4, D5 are highlighted, including the differential ones with other dystrophies. Expert opinion: Early diagnosis of LGMD, utilizing Next Generation Sequencing technique (NGS), is crucial for offering an accurate diagnosis. Muscle MRI imaging is emerging both for diagnosis and follow-up of disease progression. Emerging therapeutic strategies are considered for LGMD D2. Q3 �

Introduction: Limb-girdle muscular dystrophies (LGMDs) encompass a clinically heterogeneous group of rare, genetic progressive muscle disorders presenting with weakness and atrophy of predominant pelvic and shoulder muscles. The spectrum of disease severity ranges from severe childhood-onset muscular dystrophy to adult-onset dystrophy. Areas covered: The review presents an update of the clinical phenotypes and diagnostic options for LGMD including both dominant and recessive LGMD and consider their differential clinical and histopathological features. An overview of most common phenotypes and of possible complications is given. The management of the main clinical respiratory, cardiac, and central nervous system complications are covered. The instrumental, muscle imaging, and laboratory exams to assess and reach diagnosis are described. The use of recent genetic techniques such as next generation sequencing (NGS), whole-exome sequencing compared to other techniques (e.g. DNA sequencing, protein analysis) is covered. Currently available drugs or gene therapy and rehabilitation management are focused on. Expert commentary: Many LGMD cases, which for a long time previously remained without a molecular diagnosis, can now be investigated by NGS. Gene mutation analysis is always required to obtain a certain molecular diagnosis, fundamental to select homogeneous group of patients for future pharmaceutical and gene trials.

Objective: To apply next-generation sequencing (NGS) for the investigation of the genetic basis of undiagnosed muscular dystrophies and myopathies in a very large cohort of patients. Methods: We applied an NGS-based platform named MotorPlex to our diagnostic workflow to test muscle disease genes with a high sensitivity and specificity for small DNA variants. We analyzed 504 undiagnosed patients mostly referred as being affected by limb-girdle muscular dystrophy or congenital myopathy. Results: MotorPlex provided a complete molecular diagnosis in 218 cases (43.3%). A further 160 patients (31.7%) showed as yet unproven candidate variants. Pathogenic variants were found in 47 of 93 genes, and in more than 30% of cases, the phenotype was nonconventional, broadening the spectrum of disease presentation in at least 10 genes. Conclusions: Our large DNA study of patients with undiagnosed myopathy is an example of the ongoing revolution in molecular diagnostics, highlighting the advantages in using NGS as a first-tier approach for heterogeneous genetic conditions. Neurology ® 2016;87:1–6 GLOSSARY CM 5 congenital myopathy; LGMD 5 limb-girdle muscular dystrophy; MD 5 muscular dystrophy; NGS 5 next-generation sequencing; NMD 5 neuromuscular disorder.

Introduction: Limb girdle muscular dystrophies (LGMDs) are characterized by high molecular heterogeneity, clinical overlap, and a paucity of specific biomarkers. Their molecular definition is fundamental for prognostic and therapeutic purposes. Methods: We created an Italian LGMD registry that included 370 molecularly defined patients. We reviewed detailed retrospective and prospective data and compared each LGMD subtype for differential diagnosis purposes. Results: LGMD types 2A and 2B are the most frequent forms in Italy. The ages at disease onset, clinical progression, and cardiac and respiratory involvement can vary greatly between each LGMD subtype. In a set of extensively studied patients, targeted next-generation sequencing (NGS) identified mutations in 36.5% of cases. Conclusion: Detailed clinical characterization combined with muscle tissue analysis is fundamental to guide differential diagnosis and to address molecular tests. NGS is useful for diagnosing forms without specific biomarkers, although, at least in our study cohort, several LGMD disease mechanisms remain to be identified.

Introduction: Limb girdle muscular dystrophies (LGMDs) are characterized by high molecular heterogeneity, clinical overlap, and a paucity of specific biomarkers. Their molecular definition is fundamental for prognostic and therapeutic purposes. Methods: We created an Italian LGMD registry that included 370 molecularly defined patients. We reviewed detailed retrospective and prospective data and compared each LGMD subtype for differential diagnosis purposes. Results: LGMD types 2A and 2B are the most frequent forms in Italy. The ages at disease onset, clinical progression, and cardiac and respiratory involvement can vary greatly between each LGMD subtype. In a set of extensively studied patients, targeted next-generation sequencing (NGS) identified mutations in 36.5% of cases. Conclusion: Detailed clinical characterization combined with muscle tissue analysis is fundamental to guide differential diagnosis and to address molecular tests. NGS is useful for diagnosing forms without specific biomarkers, although, at least in our study cohort, several LGMD disease mechanisms remain to be identified.

Autosomal recessive limb girdle muscular dystrophies (LGMD type 2) are a clinically and genetically heterogeneous group of disorders, including at least 13 different genetic entities, which are characterized by progressive involvement and wasting of proximal limb girdle muscles. All LGMD2 seem to be present worldwide, though the relative frequency of the different types is variable. The gold standard for LGMD diagnosis is the study of protein involved either by immunoblot or immunohistochemistry on the basis of clinical phenotypes, followed by a mutation study in candidate genes. Pathogenetic mechanisms are various; a different pathophysiology implies different clinical muscle involvement and might require a differential therapeutical approach.

Background: Dystroglycanopathy (α-DG) is a relatively common, clinically and genetically heterogeneous category of congenital forms of muscular dystrophy (CMD) and limb-girdle muscular dystrophy (LGMD) associated with hypoglycosylated α-dystroglycan. To date, mutations in at least 19 genes have been associated with α-DG. One of them, GMPPB, encoding the guanosine-diphosphate-mannose (GDP-mannose) pyrophosphorylase B protein, has recently been associated with a wide clinical spectrum ranging from severe Walker-Warburg syndrome to pseudo-metabolic myopathy and even congenital myasthenic syndromes. We re-sequenced the full set of known disease genes in 73 Italian patients with evidence of either reduced or nearly absent α-dystroglycan to assess genotype-phenotype correlations in this cohort. We used innovative bioinformatic tools to calculate the effects of all described GMPPB mutations on protein function and attempted to correlate them with phenotypic expressions. Results: We identified 13 additional cases from 12 families and defined seven novel mutations. Patients displayed variable phenotypes including less typical pictures, ranging from asymptomatic hyperCKemia, to arthrogryposis and congenital clubfoot at birth, and also showed neurodevelopmental comorbidities, such as seizures and ataxic gait, as well as autism-spectrum disorder, which is seldom described in clinical reports of dystroglycanopathies. We also demonstrated that few mutations recur in the Italian GMPPB-mutated population and that alterations of protein stability are the main effects of GMPPB missense variants.

Limb-girdle muscular dystrophies (LGMD) are clinically and genetically heterogeneous conditions, presenting with a wide clinical spectrum, leading to progressive proximal weakness caused by loss of muscle fibers. MiR-206 is a member of myomiRNAs, a group of miRNAs with important function in skeletal muscle. Our aim is to determine the value of miR-206 in detecting muscle disease evolution in patients affected by LGMD. We describe clinical features, disease history and progression of eleven patients affected by various types of LGMD: transportinopathy, sarcoglycanopathy and calpainopathy. We analyzed the patients' mutations and we studied the circulating miR-206 in serum by qRT-PCR; muscle MRI was done with a 1.5 Tesla apparatus. The severe evolution of disease type is associated with the expression levels of miR-206, which was significantly elevated in our LGMD patient cohort in comparison with a control group. In particular, we observed an over-expression of miR-206 that was 50...

Limb-girdle muscular dystrophies (LGMD) are genetically and clinically heterogeneous conditions. We investigated a large family with autosomal dominant transmission pattern, previously classified as LGMD1F and mapped to chromosome 7q32. Affected members are characterized by muscle weakness affecting earlier the pelvic girdle and the ileopsoas muscles. We sequenced the whole exome of four family members and identified a shared heterozygous frame-shift variant in the Transportin 3 (TNPO3) gene, encoding a member of the importin-b super-family. The TNPO3 gene is mapped within the LGMD1F critical interval and its 923-amino acid human gene product is also expressed in skeletal muscle. In addition, we identified an isolated case of LGMD with a new missense mutation in the same gene. We localized the mutant TNPO3 around the nucleus, but not inside. The involvement of gene related to the nuclear transport suggests a novel disease mechanism leading to muscular dystrophy.

LGMD D2 is a disease caused by TNPO3 mutation. We describe the expression of TNPO3 and selected proteins, likely modified by TNPO3 mutation, in muscle biopsies of affected patients. We also aim to find other genes involved in pathways correlated to TNPO3. Our morphological study on LGMD D2 muscle described the expression of TNPO3 and SRSF1, a splicing factor transported by TNPO3. Moreover, we investigated some sarcomeric and nuclear proteins, likely altered by TNPO3 mutation. Through an in silico approach we tried to identify genes involved in pathways that include, besides TNPO3 and SRSF1, p62 and Murf-1, altered in LGMD D2. In patients' muscles TNPO3 appeared weaker and randomly organized, with sporadic cytoplasmic aggregates positive for TNPO3; both SRSF1 and sarcomeric alpha actinin showed a different expression, while there were no alterations in the expression of the nuclear proteins. The in silic o study lead to identify five genes, all coding for proteins responsible for muscle contraction. Our data suggest a possible interference in the morphology and function of myofibrillar network by mutated TNPO3 ; these findings are supported by the in silico identification of genes involved in muscle contraction that could help to explain the pathogenic mechanisms of LGMD D2.

We read with interest the recent clinical research article by Abraham et al 1 entitled "European Federation of Neurological Societies cutoff values significantly reduce creatine kinase sensitivity for diagnosing neu-romuscular disorders." We should stress that the European Federation of Neurological Societies (EFNS) guidelines refer to the investigation of patients who have either paucisymptomatic or asymptomatic hyper-CKemia. 2 The purpose of the guideline was to rationalize investigation of individuals who are otherwise well but have a raised creatine kinase (CK). The guidelines are not meant to be used to diagnose neuromuscular disorders. The EFNS guidelines clearly state that nonneuromuscular and nonmyopathic conditions, which include three of the four categories in the article by Abraham et al 1 (motor neuron disease, polyneuropathy and chronic inflammatory demyelinating polyneuropathy), must be considered before investigating paucisymptomatic or asymptomatic hyper-CKemia. Patients with any one of the above three categories are unlikely to be paucisymptomatic or asymptomatic; indeed, how did their patients present? Diagnosis in these three categories can be achieved by clinical examination and electrophysiological studies, while the role of CK is less important. Even in the authors myopathy category, how many of their patients were paucisymptomatic or asymptomatic, and how many had weakness at presentation? The authors used CK levels from several different laboratories, did these have the same normal range? The EFNS guidelines suggest that, for nonblack women and nonblack men, hyperCKemia, for the purposes of investigation, should be defined as >1.5 × upper limit of normal (ULN) because, according to the study by Brewster et al, 3 shifting the threshold of investigation to 1.5 × ULN approximately halves the number of patients to be investigated. 2,3 Levels of 1.5 × ULN correspond to EFNS values of 325 IU/L and 504 IU/L, which were quoted by Abra-ham et al. 1 The question arises whether it is appropriate to have this threshold, and this remains to be settled by prospective studies. In the meantime, what is the evidence so far? The Tromso study, 4 which is probably the best epidemiological study on CK in a non-black well population and included 12 828 participants, defined the ULN as ≥210 IU/L for women, ≥400 IU/L for men <50 years, and ≥ 280 IU/L for men ≥50 years. The 1.5 × ULN of these figures are 315 IU/L for women and 420-600 IU/L for men, which are similar to the values proposed by the last EFNS guidelines. In a follow-up of the Tromso study, 120 individuals with persistent hyperCKemia (1 × ULN) were investigated clinically and by electro-myography, and four (3%) individuals were diagnosed with a myopa-thy. 5 There were three women with CK values of 293 IU/L, 391 IU/ L, and 766 IU/L and one 59-year-old man with 528 IU/L. Had the Tromso study 4 investigators adopted the 1.5 × ULN cutoff, they would have missed one case out of 120 paucisymptomatic or asymptomatic individuals with hyperCKemia due to a myopathy. The authors concluded that their findings supported the EFNS guidelines on the diagnostic approach to paucisymptomatic or asymptomatic hyperCKemia. 5

Muscle fatigability and atrophy are frequent clinical signs in limb girdle muscular dystrophy (LGMD), but their pathogenetic mechanisms are still poorly understood. We review a series of different factors that may be connected in causing fatigue and atrophy, particularly considering the role of neuronal nitric oxide synthase (nNOS) and additional factors such as gender in different forms of LGMD (both recessive and dominant) underlying different pathogenetic mechanisms. In sarcoglycanopathies, the sarcolemmal nNOS reactivity varied from absent to reduced, depending on the residual level of sar-coglycan complex: in cases with complete sarcoglycan complex deficiency (mostly in beta-sarcoglycanopathy), the sarcolemmal nNOS reaction was absent and it was always associated with early severe clinical phenotype and cardiomyopathy. Calpainopathy, dysferlinopathy, and caveolinopathy present gradual onset of fatigability and had normal sarcolemmal nNOS reactivity. Notably, as compared with caveolinopathy and sar-coglycanopathies, calpainopathy and dysferlinopathy showed a higher degree of muscle fiber atrophy. Males with calpainopathy and dysferlinopathy showed significantly higher fiber atrophy than control males, whereas female patients have similar values than female controls, suggesting a gender difference in muscle fiber atrophy with a relative protection in females. In female patients, the smaller initial muscle fiber size associated to endocrine factors and less physical effort might attenuate gender-specific muscle loss and atrophy.

Limb-girdle muscular dystrophies (LGMD) are clinically and genetically heterogeneous conditions, presenting with a wide clinical spectrum, leading to progressive proximal weakness caused by loss of muscle fibers. MiR-206 is a member of myomiRNAs, a group of miRNAs with important function in skeletal muscle. Our aim is to determine the value of miR-206 in detecting muscle disease evolution in patients affected by LGMD. We describe clinical features, disease history and progression of eleven patients affected by various types of LGMD: transportinopathy, sarcoglycanopathy and calpainopathy. We analyzed the patients' mutations and we studied the circulating miR-206 in serum by qRT-PCR; muscle MRI was done with a 1.5 Tesla apparatus. The severe evolution of disease type is associated with the expression levels of miR-206, which was significantly elevated in our LGMD patient cohort in comparison with a control group. In particular, we observed an over-expression of miR-206 that was 50-80 folds elevated in two patients with a severe and early disease course in the transportinopathy and calpainopathy sub-types. The functional impairment was observed clinically and muscle loss and atrophy documented by muscle MRI. This study provides the first evidence that miR-206 is associated with phenotypic expression and it could be used as a prognostic indicator of LGMD disease progression.

Introduction: By reviewing the literature from the last twenty years we present an accurate assessment of the state of the art in the pathogenesis and clinical presentations of sarcoglycanopathies, as well as the progress in diagnosis and treatment. Areas covered: Sarcoglycanopathies usually have a childhood onset but they can occur in adults with a limb girdle phenotype. Four main genes are expressed in the sarcoglycan complex. Cases with beta-or delta-sarcoglycan primary deficiency often present severe cardiac and respiratory complications. A defect of nitric oxide synthase might contribute to the pathogenesis of cardiac involvement and fatigue. Neuroimaging shows that muscle involvement affects mainly proximal muscle groups, followed by fibro-fatty replacement. Expert opinion: In cases of children with high creatine kinase levels and weakness or adults with limb girdle weakness, immunolabelling of muscle biopsy with anti-sarcoglycan antibodies may suggest the diagnosis, which should be confirmed by mutation analysis in the sarcoglycan genes. New genetic technologies, such as next generation sequencing, might be useful to obtain a molecular diagnosis, which is necessary for genetic counselling.

Limb-girdle muscular dystrophy 2I (LGMD2I) is a neuromuscular disorder with a heterogeneous phenotype. It is caused by mutations in the Fukutin Related Protein (FKRP) gene, which is ubiquitously expressed in human tissues. FKRP functions in CNS are largely unknown. To investigate possible cognitive impairment in LGMD2I and to describe brain MRI features. Ten LGMD2I patients (four males and six females, mean age 44 years, age range 19–69 years) were assessed with an extensive neuropsychological battery, psychopathological tests and neuromuscular specific quality-of-life questionnaire. Adults were compared with ten matched healthy controls. All patients underwent complete neurological examination, and nine underwent brain MRI scanning. Patients showed a fairly specific cognitive profile with mild impairment in executive functions and visuo-spatial planning without substantial impairment in global and logic IQ. MRI findings were heterogeneous: four patients showed non-specific white matter abnormalities; two patients showed moderate ventriculomegaly; three patients showed mild enlargement of subarachnoid spaces, without a specific pattern. Cerebellar atrophy was marked in one patient. Abnormal glycosylation of α-dystroglycan in LGMD2I may interfere with brain development and cognitive performances involving the frontal and posterior parietal regions, but does not result in specific brain MRI abnormalities.

Article abstract—Background: Mutations in the genes encoding for calpain-3 and dysferlin are responsible for limb– girdle muscular dystrophy (LGMD) type 2A and 2B, the most common forms of autosomal recessive LGMD. Objective: To identify calpain-3 or dysferlin deficiency in a large cohort of patients with as yet unclassified LGMD and myopathy through candidate protein analysis. Methods: The authors' muscle biopsy database search identified 407 candidate muscle biopsies with normal dystrophin and sarcoglycan. Calpain-3 and dysferlin were studied by Western blotting and immuno

Objectives: The aim of this study was to investigate the pathologic changes, evaluated by MRI, which involve the muscles of patients affected by calpainopathy (LGMD2A) and dys-ferlinopathy (LGMD2B), and evaluate their correlation with muscle strength and muscle performance. Methods: 18 patients affected by LGMD2B and 10 patients affected by LGMD2A have been evaluated clinically and by MRI imaging. We also tested, in our patients, the muscle strength of 11 upper and lower limbs muscle groups using MRC scale. Their muscular performance has been evaluated using the GSGCA scale (Gait, Stairs, Gowers, Chair, Arms functional tests). We have studied the skeletal muscles of upper and lower limbs by using MRI imaging (T1, T2, STIR sequences). The fibro-fatty replacement has been evaluated on T1 sequences by using a specific score. Myoedema has been evaluated on STIR sequences by using a new oedema score. Results: We observed an inverse linear correlation between muscle strength (MRC scale) and functional muscular performance (GSGCA scale) in patients with LGMD2A and 2B. We, also, found a significant inverse linear correlation between the degree of fibro-fatty changes, on MRI, and the strength of the same muscles tested with MRC scale. A direct linear correlation was found between the degree of fibro-fatty changes, on MRI, and the functional muscular performance, (GSGCA scale). The presence of myoedema was detected in upper and lower limbs of LGMD2B and was described for the first time, also, in LGMD2A patients. These findings have been evaluated on STIR sequences and, in both muscle disorders , a peculiar pattern of myoedema distribution was found: the anterior compartment of lower limb was more involved than the posterior compartment in both dystrophies, the leg more than the thigh for LGMD2B, in equivalent extent in thigh and leg for LGMD2A. Conclusions: MRI imaging is useful to study LGMD, because the quantification of fibro-fatty replacement and myoedema, associated with the clinical examination resulted of prognostic value in disease progression.