POGLUT1 biallelic mutations cause myopathy with reduced satellite cells, α-dystroglycan hypoglycosylation and a distinctive radiological pattern.

Protein O-glucosyltransferase 1 (POGLUT1) activity is critical for the Notch signaling pathway, being one of the main enzymes responsible for the glycosylation of the extracellular domain of Notch receptors. A biallelic mutation in the POGLUT1 gene has been reported in one family as the cause of an adult-onset limb-girdle muscular dystrophy (LGMD R21; OMIM# 617232). As the result of a collaborative international effort, we have identified the first cohort of 15 patients with LGMD R21, from nine unrelated families coming from different countries, providing a reliable phenotype-genotype and mechanistic insight. Patients carrying novel mutations in POGLUT1 all displayed a clinical picture of limb-girdle muscle weakness. However, the age at onset was broadened from adult to congenital and infantile onset. Moreover, we now report that the unique muscle imaging pattern of "inside-to-outside" fatty degeneration observed in the original cases is indeed a defining feature of POGLUT1 muscular dystrophy. Experiments on muscle biopsies from patients revealed a remarkable and consistent decrease in the level of the NOTCH1 intracellular domain, reduction of the pool of satellite cells (SC), and evidence of α-dystroglycan hypoglycosylation. In vitro biochemical and cell-based assays suggested a pathogenic role of the novel POGLUT1 mutations, leading to reduced enzymatic activity and/or protein stability. The association between the POGLUT1 variants and the muscular phenotype was established by in vivo experiments analyzing the indirect flight muscle development in transgenic Drosophila, showing that the human POGLUT1 mutations reduced its myogenic activity. In line with the well-known role of the Notch pathway in the homeostasis of SC and muscle regeneration, SC-derived myoblasts from patients' muscle samples showed decreased proliferation and facilitated differentiation. Together, these observations suggest that alterations in SC biology caused by reduced Notch1 signaling result in muscular dystrophy in LGMD R21 patients, likely with additional contribution from α-dystroglycan hypoglycosylation. This study settles the muscular clinical phenotype linked to POGLUT1 mutations and establishes the pathogenic mechanism underlying this muscle disorder. The description of a specific imaging pattern of fatty degeneration and muscle pathology with a decrease of α-dystroglycan glycosylation provides excellent tools which will help diagnose and follow up LGMD R21 patients.

MSTO1 mutations cause mtDNA depletion, manifesting as muscular dystrophy with cerebellar involvement.

MSTO1 encodes a cytosolic mitochondrial fusion protein, misato homolog 1 or MSTO1. While the full genotype-phenotype spectrum remains to be explored, pathogenic variants in MSTO1 have recently been reported in a small number of patients presenting with a phenotype of cerebellar ataxia, congenital muscle involvement with histologic findings ranging from myopathic to dystrophic and pigmentary retinopathy. The proposed underlying pathogenic mechanism of MSTO1-related disease is suggestive of impaired mitochondrial fusion secondary to a loss of function of MSTO1. Disorders of mitochondrial fusion and fission have been shown to also lead to mitochondrial DNA (mtDNA) depletion, linking them to the mtDNA depletion syndromes, a clinically and genetically diverse class of mitochondrial diseases characterized by a reduction of cellular mtDNA content. However, the consequences of pathogenic variants in MSTO1 on mtDNA maintenance remain poorly understood. We present extensive phenotypic and genetic data from 12 independent families, including 15 new patients harbouring a broad array of bi-allelic MSTO1 pathogenic variants, and we provide functional characterization from seven MSTO1-related disease patient fibroblasts. Bi-allelic loss-of-function variants in MSTO1 manifest clinically with a remarkably consistent phenotype of childhood-onset muscular dystrophy, corticospinal tract dysfunction and early-onset non-progressive cerebellar atrophy. MSTO1 protein was not detectable in the cultured fibroblasts of all seven patients evaluated, suggesting that pathogenic variants result in a loss of protein expression and/or affect protein stability. Consistent with impaired mitochondrial fusion, mitochondrial networks in fibroblasts were found to be fragmented. Furthermore, all fibroblasts were found to have depletion of mtDNA ranging from 30 to 70% along with alterations to mtDNA nucleoids. Our data corroborate the role of MSTO1 as a mitochondrial fusion protein and highlight a previously unrecognized link to mtDNA regulation. As impaired mitochondrial fusion is a recognized cause of mtDNA depletion syndromes, this novel link to mtDNA depletion in patient fibroblasts suggests that MSTO1-deficiency should also be considered a mtDNA depletion syndrome. Thus, we provide mechanistic insight into the disease pathogenesis associated with MSTO1 mutations and further define the clinical spectrum and the natural history of MSTO1-related disease.

Genetic and clinical findings in a Chinese cohort of patients with collagen VI-related myopathies.

Collagen VI-related myopathy, caused by pathogenic variants in the genes encoding collagen VI, represents a clinical continuum from Ullrich congenital muscular dystrophy (UCMD) to Bethlem myopathy (BM). Clinical data of 60 probands and their family members were collected and muscle biopsies of 26 patients were analyzed. COL6A1, COL6A2 and COL6A3 exons were analyzed by direct sequencing or next generation sequencing (NGS). Sixty patients were characterized by delayed motor milestones, muscle weakness, skin and joint changes with 40 UCMD and 20 BM. Muscle with biopsies revealed dystrophic changes and showed completely deficiency of collagen VI or sarcolemma specific collagen VI deficiency. We identified 62 different pathogenic variants in these 60 patients, with 34 were first reported while 28 were previously known; 72 allelic pathogenic variants in COL6A1 (25/72, 34.7%), COL6A2 (33/72, 45.8%) and COL6A3 (14/72, 19.4%). We also found somatic mosaic variant in the parent of 1 proband by personal genome machine amplicon deep sequencing for mosaicism. Here we provide clinical, histological and genetic evidence of collagen VI-related myopathy in 60 Chinese patients. NGS is a valuable approach for diagnosis and accurate diagnosis provides useful information for genetic counseling of related families.

Expansion of the GLE1-associated arthrogryposis multiplex congenita clinical spectrum.

Mutations in GLE1 cause two recessive subtypes of arthrogryposis multiplex congenita (AMC), a condition characterized by joint contractures at birth, and all previously reported patients died in the perinatal period. GLE1 related AMC has been almost exclusively reported in the Finnish population and is caused by a relatively common pathogenic splicing mutation in that population. Here, we report two non-Finnish brothers with novel compound heterozygous splicing mutations in GLE1, one of whom has survived to 12 years of age. We also demonstrate low levels of residual wild type transcript in fibroblasts from the surviving brother, suggesting that this residual wild-type transcript may contribute to the relatively longer-term survival in this family. We provide a detailed clinical report on the surviving patient, providing the first insight into the natural history of this rare neuromuscular disease. We also suggest that lethal congenital contracture syndrome 1 (LCCS1) and lethal arthrogryposis with anterior horn disease (LAAHD), the two AMC subtypes related to GLE1, do not have sufficient clinical or molecular differentiation to be considered allelic disorders. Rather, GLE1 mutations cause a variable spectrum of AMC severity including a non-lethal variant described herein.

Mutation-specific effects on thin filament length in thin filament myopathy.

OBJECTIVE: Thin filament myopathies are among the most common nondystrophic congenital muscular disorders, and are caused by mutations in genes encoding proteins that are associated with the skeletal muscle thin filament. Mechanisms underlying muscle weakness are poorly understood, but might involve the length of the thin filament, an important determinant of force generation. METHODS: We investigated the sarcomere length-dependence of force, a functional assay that provides insights into the contractile strength of muscle fibers as well as the length of the thin filaments, in muscle fibers from 51 patients with thin filament myopathy caused by mutations in NEB, ACTA1, TPM2, TPM3, TNNT1, KBTBD13, KLHL40, and KLHL41. RESULTS: Lower force generation was observed in muscle fibers from patients of all genotypes. In a subset of patients who harbor mutations in NEB and ACTA1, the lower force was associated with downward shifted force-sarcomere length relations, indicative of shorter thin filaments. Confocal microscopy confirmed shorter thin filaments in muscle fibers of these patients. A conditional Neb knockout mouse model, which recapitulates thin filament myopathy, revealed a compensatory mechanism; the lower force generation that was associated with shorter thin filaments was compensated for by increasing the number of sarcomeres in series. This allowed muscle fibers to operate at a shorter sarcomere length and maintain optimal thin-thick filament overlap. INTERPRETATION: These findings might provide a novel direction for the development of therapeutic strategies for thin filament myopathy patients with shortened thin filament lengths. Ann Neurol 2016;79:959-969.

TPM3 deletions cause a hypercontractile congenital muscle stiffness phenotype.

OBJECTIVE: Mutations in TPM3, encoding Tpm3.12, cause a clinically and histopathologically diverse group of myopathies characterized by muscle weakness. We report two patients with novel de novo Tpm3.12 single glutamic acid deletions at positions ΔE218 and ΔE224, resulting in a significant hypercontractile phenotype with congenital muscle stiffness, rather than weakness, and respiratory failure in one patient. METHODS: The effect of the Tpm3.12 deletions on the contractile properties in dissected patient myofibers was measured. We used quantitative in vitro motility assay to measure Ca(2+) sensitivity of thin filaments reconstituted with recombinant Tpm3.12 ΔE218 and ΔE224. RESULTS: Contractility studies on permeabilized myofibers demonstrated reduced maximal active tension from both patients with increased Ca(2+) sensitivity and altered cross-bridge cycling kinetics in ΔE224 fibers. In vitro motility studies showed a two-fold increase in Ca(2+) sensitivity of the fraction of filaments motile and the filament sliding velocity concentrations for both mutations. INTERPRETATION: These data indicate that Tpm3.12 deletions ΔE218 and ΔE224 result in increased Ca(2+) sensitivity of the troponin-tropomyosin complex, resulting in abnormally active interaction of the actin and myosin complex. Both mutations are located in the charged motifs of the actin-binding residues of tropomyosin 3, thus disrupting the electrostatic interactions that facilitate accurate tropomyosin binding with actin necessary to prevent the on-state. The mutations destabilize the off-state and result in excessively sensitized excitation-contraction coupling of the contractile apparatus. This work expands the phenotypic spectrum of TPM3-related disease and provides insights into the pathophysiological mechanisms of the actin-tropomyosin complex.

The neuromuscular differential diagnosis of joint hypermobility.

Joint hypermobility is the defining feature of various inherited connective tissue disorders such as Marfan syndrome and various types of Ehlers-Danlos syndrome and these will generally be the first conditions to be considered by geneticists and pediatricians in the differential diagnosis of a patient presenting with such findings. However, several congenital and adult-onset inherited myopathies also present with joint hypermobility in the context of often only mild-to-moderate muscle weakness and should, therefore, be included in the differential diagnosis of joint hypermobility. In fact, on the molecular level disorders within both groups represent different ends of the same spectrum of inherited extracellular matrix (ECM) disorders. In this review we will summarize the measures of joint hypermobility, illustrate molecular mechanisms these groups of disorders have in common, and subsequently discuss the clinical features of: 1) the most common connective tissue disorders with myopathic or other neuromuscular features: Ehlers-Danlos syndrome, Marfan syndrome and Loeys-Dietz syndrome; 2) myopathy and connective tissue overlap disorders (muscle extracellular matrix (ECM) disorders), including collagen VI related dystrophies and FKBP14 related kyphoscoliotic type of Ehlers-Danlos syndrome; and 3) various (congenital) myopathies with prominent joint hypermobility including RYR1- and SEPN1-related myopathy. The aim of this review is to assist clinical geneticists and other clinicians with recognition of these disorders.

Beta-sarcoglycan (A3b) mutations cause autosomal recessive muscular dystrophy with loss of the sarcoglycan complex.

The dystrophin associated proteins (DAPs) are good candidates for harboring primary mutations in the genetically heterogeneous autosomal recessive muscular dystrophies (ARMD). The transmembrane components of the DAPs can be separated into the dystroglycan and the sarcoglycan complexes. Here we report the isolation of cDNAs encoding the 43 kD sarcoglycan protein beta-sarcoglycan (A3b) and the localization of the human gene to chromosome 4q12. We describe a young girl with ARMD with truncating mutations on both alleles. Immunostaining of her muscle biopsy shows specific loss of the components of the sarcoglycan complex (beta-sarcoglycan, alpha-sarcoglycan (adhalin), and 35 kD sarcoglycan). Thus secondary destabilization of the sarcoglycan complex may be an important pathophysiological event in ARMD.

Zebrafish models of collagen VI-related myopathies.

Collagen VI is an integral part of the skeletal muscle extracellular matrix, providing mechanical stability and facilitating matrix-dependent cell signaling. Mutations in collagen VI result in either Ullrich congenital muscular dystrophy (UCMD) or Bethlem myopathy (BM), with UCMD being clinically more severe. Recent studies demonstrating increased apoptosis and abnormal mitochondrial function in Col6a1 knockout mice and in human myoblasts have provided the first mechanistic insights into the pathophysiology of these diseases. However, how loss of collagen VI causes mitochondrial dysfunction remains to be understood. Progress is hindered in part by the lack of an adequate animal model for UCMD, as knockout mice have a mild motor phenotype. To further the understanding of these disorders, we have generated zebrafish models of the collagen VI myopathies. Morpholinos designed to exon 9 of col6a1 produced a severe muscle disease reminiscent of UCMD, while ones to exon 13 produced a milder phenotype similar to BM. UCMD-like zebrafish have increased cell death and abnormal mitochondria, which can be attenuated by treatment with the proton pump modifier cyclosporin A (CsA). CsA improved the motor deficits in UCMD-like zebrafish, but failed to reverse the sarcolemmal membrane damage. In all, we have successfully generated the first vertebrate model matching the clinical severity of UCMD and demonstrated that CsA provides phenotypic improvement, thus corroborating data from knockout mice supporting the use of mitochondrial permeability transition pore modifiers as therapeutics in patients, and providing proof of principle for the utility of the zebrafish as a powerful preclinical model.

Myopathy in a 20-year-old female patient with D4ST-1 deficient Ehlers-Danlos syndrome due to a homozygous CHST14 mutation.

We here report on a 20-year-old female patient with EDS due to a homozygous CHST14 single nucleotide deletion resulting in D4ST-1 deficiency, accompanied by muscle hypoplasia and muscle weakness. Findings of muscle ultrasound, electromyography, and muscle biopsy pointed to a myopathy, similarly as in other EDS types. This myopathy probably contributes to the gross motor developmental delay in this type of EDS.

Familial reducing body myopathy with cytoplasmic bodies and rigid spine revisited: identification of a second LIM domain mutation in FHL1.

OBJECTIVE: Reducing body myopathy (RBM) is a rare progressive disorder of muscle characterized by intracytoplasmic inclusions, which stain strongly with menadione-NBT (nitroblue tetrazolium). We recently identified the four and a half LIM domain gene FHL1 located on chromosome Xq26 as the causative gene for RBM. So far eight familial cases and 21 sporadic patients with RBM have been reported in the literature. METHODS: We ascertained a total of 8 members of a German family initially reported by Goebel et al. as a mixed myopathy with rigid spine myopathy and reducing as well as cytoplasmic bodies. Clinical findings in the original and additional family members have been reviewed. Mutation detection was performed by direct sequencing of FHL1 exons. RESULTS: We identified a novel mutation (p.C150R) in the second LIM domain of FHL1 in six family members (1 male, 5 females). The male index patient was the most affected member presenting with rigid spine, followed by rapidly progressive muscle weakness. He died from the consequences of respiratory insufficiency at the age of 29.5 years. His sister, mother, grandmother, aunt and female cousin all carried the mutation in the heterozygous state. The sister is clinically unaffected; their mother had myopathic changes in her muscle biopsy, while the grandmother showed first signs of weakness at 50 years of age. The 54-year-old aunt and her daughter are clinically asymptomatic. CONCLUSION: We report a novel LIM2 domain mutation in FHL1 in a previously reported family with RBM with cytoplasmic bodies and spinal rigidity. While the male index patient was significantly affected, female carriers show varying manifestations and may be asymptomatic, likely reflecting varying degrees of X-inactivation. RBM continues to be associated with mutations in the LIM2 domain of FHL1. We also confirm our earlier observation that mutations at the N-terminal end of the LIM2 domain seem to be milder compared to mutations seen at the C-terminal part of the domain which cause severe disease even in female carriers.

Molecular organization of sarcoglycan complex in mouse myotubes in culture.

The sarcoglycans are a complex of four transmembrane proteins (alpha, beta, gamma, and delta) which are primarily expressed in skeletal muscle and are closely associated with dystrophin and the dystroglycans in the muscle membrane. Mutations in the sarcoglycans are responsible for four autosomal recessive forms of muscular dystrophy. The function and the organization of the sarcoglycan complex are unknown. We have used coimmunoprecipitation and in vivo cross-linking techniques to analyze the sarcoglycan complex in cultured mouse myotubes. We demonstrate that the interaction between beta- and delta-sarcoglycan is resistant to high concentrations of SDS and alpha-sarcoglycan is less tightly associated with other members of the complex. Cross-linking experiments show that beta-, gamma-, and delta-sarcoglycan are in close proximity to one another and that delta-sarcoglycan can be cross-linked to the dystroglycan complex. In addition, three of the sarcoglycans (beta, gamma, and delta) are shown to form intramolecular disulfide bonds. These studies further our knowledge of the structure of the sarcoglycan complex. Our proposed model of their interactions helps to explain some of the emerging data on the consequences of mutations in the individual sarcoglycans, their effect on the complex, and potentially the clinical course of muscular dystrophies.

Mutations in the dystrophin-associated protein gamma-sarcoglycan in chromosome 13 muscular dystrophy.

Severe childhood autosomal recessive muscular dystrophy (SCARMD) is a progressive muscle-wasting disorder common in North Africa that segregates with microsatellite markers at chromosome 13q12. Here, it is shown that a mutation in the gene encoding the 35-kilodalton dystrophin-associated glycoprotein, gamma-sarcoglycan, is likely to be the primary genetic defect in this disorder. The human gamma-sarcoglycan gene was mapped to chromosome 13q12, and deletions that alter its reading frame were identified in three families and one of four sporadic cases of SCARMD. These mutations not only affect gamma-sarcoglycan but also disrupt the integrity of the entire sarcoglycan complex.

[Documentation of outpatient treatment of Paget's disease in Westphalia-Lippe. Descriptive analysis of statutory health insurance data for the year 2005]. / Dokumentation der ambulanten Morbus-Paget-Behandlung am Beispiel der Region Westfalen-Lippe: Deskriptive Analyse der Abrechnungsdaten gesetzlich Krankenversicherter für das Jahr 2005.

GOAL: Inpatient health care data are often used as a source of information for health reporting in Germany, despite the fact that a lot of diseases are predominantly treated in the outpatient sector. This study provides a first overview of the outpatient care situation in relation to Paget's disease. METHOD: Outpatient care data from the Association of Statutory Health Insurance Physicians constituted the database for a descriptive analysis, capturing the state of medical care for the rare rheumatic illness Paget's disease (Osteodystrophia deformans) in the region of East Westphalia-Lippe in 2005. RESULTS: While the health report of North Rhine-Westphalia documents a total of 56 cases of M. Paget discharged from hospital for the year 2003, 166 patients suffering from Paget's disease consulted an practice-based physician in 2005 in the district of Detmold alone. The latter figure corresponds to 8.0 treated patients per 100.000 inhabitants. The treatment rates of men and women are comparable. The probability of treatment increases with advancing age. On average, patients with Paget's disease are 65.6 years old (SD=15.4 years). Almost 90% of the diagnoses of Paget's disease are classified as M88.9 according to ICD 10; more exact localisations are provided only for relatively few cases. Nearly a quarter of the cases (24.1%) are treated by general practitioners and internists. Anaesthetists treated 17.5% of the patients and orthopaedists 16.9%. In addition, ophthalmologists treat a considerable proportion of cases (12.0%). CONCLUSION: Consistent with the rareness of Paget's disease, the treatment data are as low as expected. However, the results show that routinely collected health care data allow insights into morbidity structures within the outpatient sector. It follows that for statutory health insurants (approximately 90% of the population) there should be an extension of health reporting to diseases that are mainly treated in outpatient settings.

Two-minute versus 6-minute walk distances during 6-minute walk test in neuromuscular disease: Is the 2-minute walk test an effective alternative to a 6-minute walk test?

Functional tests such as Motor Function Measure-32 (MFM-32), supine to stand, ascend/descend stairs permit the assessment of task-specific motor function in neuromuscular disease (NMD). The 6-min walk test (6MWT), though functional, is primarily used to assess endurance and disease progression in children with neuromuscular disorders. Barriers to 6MWT administration, in this population, can include reduced attention span due to age and inability to tolerate test length due to weakness. We propose task-specific functional deficits are related to endurance. Additionally, the 2-min walk test (2MWT) could effectively replace the 6MWT in this population. Seventy-seven participants, ages 5-18, with a variety of neuromuscular disorders performed the 6MWT, timed functional tests (TFT), and the MFM-32. Correlation and paired t-test analyses were used to compare the distance walked in the first 2 min (2MWD) to the distance walked in the entire 6 min (6MWD) and to the functional outcome measures above. The 2MWD strongly correlated with 6MWD and the other outcome measures. Paired t-test analysis also showed that the 2MWD did not differ from the distance walked in the last 2 min of the 6MWT. Although equivalence testing could not reject the claim that this difference exceeded the upper practical limit of 9.5 m, it only showed a modest overestimation of the 4-6MWD compared with the 2MWD. Together, our results support the ability of the 2MWD to predict the 6MWD, specifically in the pediatric neuromuscular disease population.

Exon skipping mutations in collagen VI are common and are predictive for severity and inheritance.

Mutations in the genes encoding collagen VI (COL6A1, COL6A2, and COL6A3) cause Bethlem myopathy (BM) and Ullrich congenital muscular dystrophy (UCMD), two related conditions of differing severity. BM is a relatively mild dominantly inherited disorder characterized by proximal weakness and distal joint contractures. UCMD was originally regarded as an exclusively autosomal recessive condition causing severe muscle weakness with proximal joint contractures and distal hyperlaxity. We and others have subsequently modified this model when we described UCMD patients with heterozygous in-frame deletions acting in a dominant-negative way. Here we report 10 unrelated patients with a UCMD clinical phenotype and de novo dominant negative heterozygous splice mutations in COL6A1, COL6A2, and COL6A3 and contrast our findings with four UCMD patients with recessively acting splice mutations and two BM patients with heterozygous splice mutations. We find that the location of the skipped exon relative to the molecular structure of the collagen chain strongly correlates with the clinical phenotype. Analysis by immunohistochemical staining of muscle biopsies and dermal fibroblast cultures, as well as immunoprecipitation to study protein biosynthesis and assembly, suggests different mechanisms each for exon skipping mutations underlying dominant UCMD, dominant BM, and recessive UCMD. We provide further evidence that de novo dominant mutations in severe UCMD occur relatively frequently in all three collagen VI chains and offer biochemical insight into genotype-phenotype correlations within the collagen VI-related disorders by showing that severity of the phenotype depends on the ability of mutant chains to be incorporated in the multimeric structure of collagen VI.

Clinical and molecular overlap between myopathies and inherited connective tissue diseases.

This review presents an overview of myopathies and inherited connective tissue disorders that are caused by defects in or deficiencies of molecules within the extracellular matrix (ECM). We will cover the myopathies caused by defects in transmembrane protein complexes (dystroglycan, sarcoglycan, and integrins), laminin, and collagens (collagens VI, XIII, and XV). Clinical characteristics of several of these myopathies imply skin and joint features. We subsequently describe the inherited connective tissue disorders that are characterized by mild to moderate muscle involvement in addition to the dermal, vascular, or articular symptoms. These disorders are caused by defects of matrix-embedded ECM molecules that are also present within muscle (collagens I, III, V, IX, lysylhydroxylase, tenascin, fibrillin, fibulin, elastin, and perlecan). By focussing on the structure and function of these ECM molecules, we aim to point out the clinical and molecular overlap between the groups of disorders. We argue that clinicians and researchers dealing with myopathies and inherited connective tissue disorders should be aware of this overlap. Only a multi-disciplinary approach will allow full recognition of the wide variety of symptoms present in the spectrum of ECM defects, which has important implications for scientific research, diagnosis, and for the treatment of these disorders.

Phenotypic variability in siblings with calpainopathy (LGMD2A).

Calpainopathy is an autosomal-recessive limb girdle muscular dystrophy (LGMD2A) characterized by selective atrophy and weakness of proximal limb girdle muscles. The clinical phenotype of the disease is highly variable inter-familial, but little is known about intra-familial variability. This study reports the phenotypic variability in eight sibling pairs with genetically proven LGMD2A. Although siblings with identical mutations were often similarly affected, in some families the age of onset and the clinical course varied considerably.

A comparative analysis of collagen VI production in muscle, skin and fibroblasts from 14 Ullrich congenital muscular dystrophy patients with dominant and recessive COL6A mutations.

Ullrich congenital muscular dystrophy (UCMD) is caused by recessive and dominant mutations in COL6A genes. We have analysed collagen VI expression in 14 UCMD patients. Sequencing of COL6A genes had identified homozygous and heterozygous mutations in 12 cases. Analysis of collagen VI in fibroblast cultures derived from eight of these patients showed reduced extracellular deposition in all cases and intracellular collagen VI staining in seven cases. This was observed even in cases that showed normal collagen VI labelling in skin biopsies. Collagen VI immunolabelling was reduced in all the available muscle biopsies. When comparisons were possible no correlation was seen between the extent of the reduction in the muscle and fibroblast cultures, the mode of inheritance or the severity of the clinical phenotype. Mutations affecting glycine substitutions in the conserved triple helical domain were common and all resulted in reduced collagen VI. This study expands the spectrum of collagen VI defects and shows that analysis of skin fibroblasts may be a useful technique for the detection of collagen VI abnormalities. In contrast, immunohistochemical analysis of skin biopsies may not always reveal an underlying collagen VI defect.