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Muscle MRI findings in patients with limb girdle muscular dystrophy with calpain 3 deficiency (LGMD2A) and early contractures
Neuromuscular Disorders 15 (2005) 164–171 www.elsevier.com/locate/nmd
Muscle MRI findings in patients with limb girdle muscular dystrophy with calpain 3 deficiency (LGMD2A) and early contractures Eugenio Mercuria,b,*, Kate Bushbyc, Enzo Riccid, Daniel Birchalle, Marika Paneb, Maria Kinalia, Joanna Allsopf, Vincenzo Nigrog, Amets Sa´enzh, Annachiara Nascimbenii, Luigi Fulizioi, Corrado Angelinii, Francesco Muntonia a
Department of Paediatrics and Neonatal Medicine, Dubowitz Neuromuscular Centre, Imperial College London, Hammersmith Hospital, Du Cane Road, London W12 0NN, UK b Pediatric Neurology Unit, Catholic University Hospital, Rome, Italy c Institute of Human Genetics, International Centre for Life, University of Newcastle, Newcastle upon Tyne, UK d Institute of Neurology, Catholic University Hospital, Rome, Italy e Department of Radiology, University of Newcastle, Newcastle upon Tyne, UK f MRI Robert Steiner Unit, Imperial College London, UK g Telethon Institute of Genetics and Medicine, Naples, Italy h Grupo Neurogene´tica, Unidad Experimental Hospital Donostia, San Sebastia´n, Spain i Department of Neuroscience, University of Padua, Padua, Italy Received 28 July 2004; received in revised form 11 October 2004; accepted 14 October 2004
Abstract Limb girdle muscular dystrophy 2A is a common variant secondary to mutations in the calpain 3 gene. A proportion of patients has early and severe contractures, which can cause diagnostic difficulties with other conditions. We report clinical and muscle magnetic resonance imaging findings in seven limb girdle muscular dystrophy 2A patients (four sporadic and three familial) who had prominent and early contractures. All patients showed a striking involvement of the posterior thigh muscles. The involvement of the other thigh muscles was variable and was related to clinical severity. Young patients with minimal functional motor impairment showed a predominant involvement of the adductors and semimembranosus muscles while patients with restricted ambulation had a more diffuse involvement of the posterolateral muscles of the thigh and of the vastus intermedius with relative sparing of the vastus lateralis, sartorius and gracilis. At calf level all patients showed involvement of the soleus muscle and of the medial head of the gastrocnemius with relative sparing of the lateral head. MRI findings were correlated to those found in two patients with the phenotype of limb girdle muscular dystrophy 2A without early contractures and the pattern observed was quite similar. However, the pattern observed in limb girdle muscular dystrophy 2A is different from that reported in other muscle diseases such as Emery–Dreifuss muscular dystrophy and Bethlem myopathy which have a significant clinical overlap with limb girdle muscular dystrophy 2A once early contractures are present. Our results suggest that muscle MRI may help in recognising patients with limb girdle muscular dystrophy 2A even when the clinical presentation overlaps with other conditions, and may therefore, be used as an additional investigation to target the appropriate biochemical and genetic tests. q 2004 Elsevier B.V. All rights reserved. Keywords: Calpain; MRI; Muscle; Contractures
1. Introduction * Corresponding author. Address: Department of Paediatrics and Neonatal Medicine, Imperial College London, Hammersmith Hospital, Du Cane Road, London W12 0NN, UK. Tel.: C44 20 83833295; fax: C44 20 87408281. E-mail address: [email protected] (E. Mercuri). 0960-8966/$ - see front matter q 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.nmd.2004.10.008
Limb girdle muscular dystrophies are a clinically and geneticallyheterogeneous group of disorders which shareprogressive muscle weakness and degenerative muscle changes. The form of limb girdle muscular dystrophy secondary to
E. Mercuri et al. / Neuromuscular Disorders 15 (2005) 164–171
calpain 3 deficiency (LGMD2A) is one of the most common forms with a reported frequency ranging from 9 to 40% of limb girdle muscular dystrophies. Calpain-3 is a muscle specific intracellular protease. LGMD2A was the first type of muscular dystrophy shown to be caused by an enzymatic defect rather than a defect in a structural muscle protein. From a diagnostic point of view, the definitive diagnosis of LGMD2A relies on the identification of mutations in the calpain 3 gene, located on 15q15-q21. Although a deficiency of calpain 3 expression on western blot analysis can be demonstrated in most cases of LGMD2A, a secondary reduction of calpain 3 has been reported in other forms of recessive limb girdle muscular dystrophy such as LGMD2B or LGMD2J in which the primary defects are, respectively, in dysferlin and titin. Furthermore, recent reports suggest that immunoblot for calpain 3 can be normal in some patients carrying missense mutations in the calpain 3 gene [1]. The original paper reporting clinical and genetic findings in Reunion Island patients with LGMD2A described limb girdle distribution of wasting, scapular winging and hip girdle weakness which sparing of the hip abductors [2]. Contractures were often limited to the ankles in the early phases of the diseases and became more diffuse in patients on a wheelchair. Although similar clinical findings have been reported in other patients by several subsequent studies [3,4], variations from this phenotype have also been reported. Fardeau et al. in 1996 described two French metropolitan families with LGMD2A, early onset of contractures, especially of the elbows, and more obvious involvement of the distal muscles [5]. This phenotype, also characterised by early elbow, finger and hip flexor contractures has been reported in a few more cases [4]. In these patients, the diagnosis based on clinical ground only can be difficult as the distribution of muscle wasting, weakness and contractures is similar to Emery–Dreifuss muscular (EDMD) dystrophy and overlaps also with Bethlem myopathy. In the last few years, several studies have reported the value of muscle magnetic resonance imaging (MRI) in identifying patterns of muscle involvement in genetically distinct muscle disorders [6–9]. The aim of this study was to report clinical and MR findings in seven patients with LGMD2A and early contractures. More specifically, we wished to establish whether muscle MRI can identify a specific pattern of muscle involvement in patients with such phenotype and whether MR findings are different from those found in patients with LGMD2A and the ‘non-contracted’ phenotype, and from other conditions such as Bethlem myopathy and EDMD.
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and rigidity of the spine were included in the study. In six out of seven, the diagnosis of LGMD2A was confirmed by genetic analysis showing a mutation in the CAPN3 gene while in the remaining one the diagnosis was suggested not only by the clinical picture but also the complete absence of calpain 3 on Western blot. Mutations in the LMNA gene, responsible for the dominant variant of EDMD were excluded in all. We systematically collected data regarding age and mode of onset, muscle power, contractures, functional abilities, progression of the disease, cardiac and respiratory function. Data on muscle biopsy were also collected. Clinical and immunohistochemical details of three patients have already been reported [4]. Details of histological, histochemical and immunohistochemial procedures have already been described in the same paper [4]. 2.2. Control group Another two patients followed at the Catholic University in Rome, with genetically confirmed diagnosis of LGMD2A and with the phenotype without early contractures were also included in the study as controls for the MRI study. 2.3. Muscle magnetic resonance imaging All the patients were fully co-operative and no sedation or general anaesthesia was required. Muscle MRI was performed on a 1.0-tesla Picker HPQ system, using conventional T1 weighted spin echo. Non-contrast transverse images were obtained in the legs, selecting the axial plane with respect to the long axis of the body. This involved two sequential scans. The slices were 5 mm thick and the gap between slices was 50 mm. TR was 525 msec, TE 14 msec. The scans were examined by two independent observers, blinded to clinical and genetic data, looking for normal and abnormal muscle bulk (atrophy) and for normal and abnormal signal intensity within the different muscles. Sections were generally analysed within the mid-upper section of the thighs and lower legs, for the reason that muscle bulk is greatest at this level and that muscle abnormality could be more clearly visualised as a result. Care was taken to review the remainder of the muscle bulk in order to confirm that the analysed section was representative of the whole muscle. The abnormal signal was classified as mild if only traces of increased signal intensity could be observed in an otherwise well preserved muscle, moderate if less than 50% of the muscle showed increased signal intensity, and severe if at least 50% of it showed increased signal intensity. The muscles examined were:
2. Subjects and methods 2.1. Study group Seven patients (four sporadic and three familial cases) with a diagnosis of LGMD2A, early elbow contractures
anterior thigh: sartorius, gracilis, quadriceps femoris. posterior thigh: semimembranosus, semitendinosus, biceps femoris, adductor muscles. lower leg: gastrocnemius, soleus, tibialis anterior, tibialis posterior, extensor allucis and extensor digitorum longum.
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3. Results
4. Muscle MRI
3.1. Age of onset
4.1. Thigh muscles
All patients had normal motor milestones and first clinical signs were noted between 2 and 15 years, consisting in toe walking due to Achilles tendon contractures and in one case, in muscle pain and cramps following exercise. 3.2. Maximal functional ability All our patients are still able to walk independently but the four oldest patients have restricted ambulation (details on Table 1).
4.1.1. Study group All seven patients showed a striking involvement of the adductor magnus, associated with variable involvement of the adductor longus and semimembranosus (Fig. 2(a)–(c))in the three youngest children. The oldest patients showed a more severe and diffuse involvement of thigh muscles with prominent involvement of adductors, vastus intermedius and semimembranosus and relative sparing of sartorius and gracilis and partly of the vastus lateralis. (Fig. 2(d) and 3(a,b)).
3.3. Clinical examination None of our patients had facial weakness. All but one showed wasting of the pectoralis and biceps with a striking differential involvement between biceps and triceps. In five cases, the biceps appeared a thin and fibrotic string (Fig. 1). All patients had generalised weakness involving trunk and limbs (proximal more than distal) and the weakest muscle were the hip adductors and extensors which always had subgravity power. Hip abductors were invariably better preserved with antigravity power. All but one also had mild distal weakness in the lower limbs. All patients had TA tightness, which was always the presenting sign but in addition developed marked elbow and hamstrings contractures and spine rigidity which was severe in three cases mostly affecting the lumbar and lower thoracic spine. Five of the seven also had long finger contractures. 3.4. Serum creatine kinase Serum CK was always markedly elevated (15–20 times the normal limits). 3.5. Respiratory function FVC was always relatively well preserved (70–90% of the predicted values) compared to the proximal muscle power and no patients had frequent chest infection or symptoms of nocturnal hypoventilation. 3.6. Cardiac function All patients had serial ECG which were always normal. All seven also had echocardiograms (age 13–37) which were also normal. 3.7. Progression Motor difficulties were progressive after the onset of clinical signs. Although none of our patients has lost independent ambulation they have all experienced increasing difficulties and progressive restriction of walking.
4.1.2. Control group The two patients with the non-contracted phenotype showed a diffuse involvement of the postero-lateral thigh muscles with prominent involvement of adductors, vastus intermedius and semimembranosus and relative sparing of sartorius and gracilis and partly of the vastus lateralis (Fig. 3(c) and (d)). 4.2. Calf muscles 4.2.1. Study group All seven patients showed a relatively normal anterior compartment with relative sparing of the tibialis anterior (7/7) and peroneals muscles (6/7). In the posterior compartment all seven showed a predominant involvement of the medial head of the gastrocnemius, with relative sparing of the lateral head of the gastrocnemius (Fig. 4(a)– (d)), associated with a variable involvement of the soleus. 4.2.2. Control group Both patients with the non-contracted phenotype showed a relatively normal anterior compartment and a predominant involvement of the gastrocnemius, with relative sparing of the lateral head of the gastrocnemius in one of the two and involvement of the soleus.
5. Correlation between clinical severity and MRI changes 5.1. Study group Three of the seven patients were still able to walk independently for relatively long distances. All three showed a selective impairment of adductors muscles and semimembranosus. The other four patients had more restricted ambulation and could only walk short distances with support. All of these patients had a more diffuse involvement of the postero-lateral thigh muscles. Changes in the calf muscles were similar in all seven patients, regardless of severity.
Table 1 Clinical findings Onset
Max functional ability
Age examination (years)
Progress max ability when last seen
Scapular winging
Wasting
Weakness
Contractures
Rigidity of spine
Scoliosis
FVC (%)
Cardiac
CK
1
6 yrs TA contractures, 7 yrs weakness
Walking at 1 year
15
Walking
Yes
YesCC
K
94
ECG: N, Echo: N
X 30
10 yrs TA contractures
Walking at 1 year
13
Walking
Yes
Ankles, hamstrings, fingers, elbows
YesC
K
71
ECG: N, Echo: N
X 15
3
8 yrs: TA contractures, scapular winging 15yrs: pain and cramps, TA and elbow contractures 2 yrs: TA contractures
Walking at 1 year
16
Walking
Yes
Proximal Odistal, adductors subgravity, abductors antigr with res, some distal weakness Proximal Odistal, adductors subgravity, abductors antigr with res, some distal weakness ProximalOdistal, hip add subgravity, abductors antigr with res
Ankles, hamstrings, elbow
2
Pectoralis, biceps, trapezius. medial calves Pectoralis, biceps, hamstrings, medial calves Pectoralis, biceps, deltoid
ITB, ankles, elbow
C
K
94
ECG: N, Echo: N
X 20
Walking at 1 year
38
Walking with support (stick)
Yes
CC
K
80
ECG: N, Echo: N
X 20
14
Yes
Paraspinal, finger, ankle
K
C
80
ECG: N, Echo: N
X 45
6
6 yrs: TA contractures
Walking at 1 year
17
Walking with support, wheelchair part-time since age 13 years Walking with support, climbs stairs with railing
Proximal Odistal, adductors subgravity, abductors antigr with res, some distal weakness Proximal Odistal, adductors subgravity, abductors antigr with res, ankle dorsiflexion weaker than plantar flexion
Ankles, fingers, hamstrings, elbows
Walking at 1 year
Pectoralis, biceps, trapezius. medial calves Pectoralis, biceps, medial calves
Yes
Pectoralis, biceps, medial calves
Paraspinal, neck, ankle, wrist, elbow, finger
K
K
84
ECG: N, Echo: N
7
9 yrs: pain, TA contractures
Walking at 1 year
38
Walking with support, climbs stairs both hands on banister
Yes
Pectoralis, biceps, medial calves
Proximal Odistal, adductors antigr with res, abductors antigr with res, ankle dorsiflexion weaker than plantar flexion Proximal Odistal, adductors subgravity, abductors antigr with res, ankle dorsiflexion weaker than plantar flexion
Ankle
CC
K
90
ECG: N, Echo: N
4
5
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Case
X6
yrs, years; antigr, antigravity; res, resistence; TA, tendon Achilles; N, normal. 167
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Fig. 1. Thin and fibrotic biceps (patient 1).
6. Discussion The seven cases with LGMD2A presented in this paper have calf hypertrophy, rigidity of the spine and early contractures affecting Achilles tendons, elbow and long finger flexors. The distribution of muscle weakness and contractures in these patients is in keeping with what
previously described by Fardeau et al. in 1996 as a variant of the phenotype originally described in patients with LGMD2A. A similar ‘contracted’ phenotype has been previously reported in a few other patients with LGMD2A [4] but it is not well recognised in the literature. As a significant number of our patients presented with these features, we want to draw attention to this phenotype, which has a significant clinical overlap with other conditions, in particular with Emery–Dreifuss muscular dystrophy (EDMD) and the severe end of the Bethlem myopathy spectrum. Using muscle MRI we have been able to identify a distinct pattern of muscle involvement. The severity of the involvement was variable and was related to the clinical severity of motor impairment. The three younger patients who were able to walk independently for relatively long distances had a selective impairment of the adductor magnus and of the semimembranosus. In the four oldest patients, who all had restricted ambulation, there was a similar involvement of the adductor magnus, but this was associated with a more diffuse involvement of the posterior thigh muscles and vasti intermedius and a with selective sparing of sartorius and gracilis muscles and vastus lateralis. Although none of the previous studies has systematically evaluated muscle imaging in patients with LGMD2A,
Fig. 2. Transverse T1 weighted images through thigh muscles in four patients with contracted phenotype (cases 1–4). Note the predominant involvement of the adductors muscles and the early signs of involvement of the semimembranosus in the three young patients (cases 1–3, panels a,b,c) and the more diffuse involvement in the oldest patient (case 4, panel d) with relative sparing of the sartorius.
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Fig. 3. Transverse T1 weighted images through thigh muscles in two patients with contracted phenotype (cases 5, 6, panel a and b) and in two patients with the non-contracted phenotype (panel c and d). Note the predominant involvement of the posterior muscles and of the vastus intermedius and medialis with relative sparing of the sartorius, gracilis and partly of the vastus lateralis and of the rectus femoris in all four patients regardless of their clinical phenotype.
the picture of muscle MRI [10] and CT [2,5] previously reported show similar findings at thigh level. The pattern of muscle involvement in the calves consisted in a selective involvement of the medial head of the gastrocnemius muscle compared to the lateral head, which was always relatively spared. The involvement of the gastrocnemius was associated with variable involvement of the soleus muscle. It is of interest that a similar pattern of muscle involvement was also found in the control group of patients with LGMD2A but with the non-contracted phenotype. Although we had only two patients with the non-contracted phenotype who had a scan, when patients with the same age and level of function were compared, the pattern observed was very similar with prominent involvement of the posterior thigh muscles and vastus intermedius and with relative sparing of vastus lateralis, sartorius and gracilis. These findings, therefore, suggest that, despite the differences in muscle wasting and in contractures, both forms of LGMD2A have a similar pattern of muscle involvement on muscle MR, which appears to vary in relation to the severity of clinical functional impairment. This is in keeping with our studies in a number of genetically distinct forms of muscular dystrophies including EDMD, a variety of congenital myopathies and in Bethlem myopathy and neurogenic disorder, in which we
were able to recognise a disease specific pattern of selective muscle involvement. The pattern of involvement on muscle MRI observed in LGMD2A was, in contrast, different from those previously reported in other forms of muscle disorders with overlapping clinical phenotype, such as Emery–Dreifuss muscular dystrophy. Although the differential involvement of the medial and lateral head of the gastrocnemius observed in the calf muscles in patients with LGMD2A was similar to that reported in patients with the dominant form of EDMD [6], the pattern of muscle involvement at thigh level was different. While patients with LGMD2A had a more striking involvement of the adductor muscles and relative sparing of the vastus lateralis, the vasti muscles were selectively affected in the dominant form of EDMD. The pattern of selective muscle involvement observed in patients with LGMD2A was also different from that observed in the X-linked form of Emery–Dreifuss muscular dystrophy, characterised by a more prominent involvement of the soleus and relative sparing of the gastrocnemii muscles [6]. This information can be important at the time of directing mutation analysis as the two forms have a significant clinical overlap. In both forms the age of onset is variable but the most common presentation is TA tightness at school age. Other common features are scapuloperoneal wasting and scapular winging, rigidity of the spine, elbow
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Fig. 4. Transverse T1 weighted images through calf muscles in four patients with contracted phenotype (cases 1–4). Note the differential involvement between medial and lateral head of the gastrocnemius and the variable involvement of the soleus. Perneus muscles were only mildly affected in case 1 (panel a).
contractures and, in some cases, a striking wasting of biceps muscles in the arms with relatively preserved triceps muscles. Both the dominant form of EDMD and LGMD2A show some wasting in the medial aspects of the calves but there are some differences in the distribution of muscle wasting in the thighs. While patients with LGMD2A have a striking wasting of the medial and posterior aspects of the thighs with a relative hypertrophy of the quadriceps, patients with the dominant form of EDMD generally have more diffuse wasting [11–13]. The absent cardiac involvement in LGMD2A is a major difference compared to what found in EDMD. However, in EDMD cardiac involvement is found invariably only in patients above the age of 30 years and as all our patients came to our observation before the age of 20, a normal cardiac assessment cannot exclude the diagnosis. Both forms also have dystrophic changes on muscle biopsy and markedly increased CK, although in the dominant form of EDMD CK levels are more variable and overall tend to be lower than in our cases with LGMD2A [12,13]. The phenotype with early contractures also has some clinical overlap with Bethlem myopathy [12,14], especially if patients are in the first decade or in their teens. CK levels, however, are generally lower in Bethlem myopathy and the changes on muscle biopsy are generally non-dystrophic.
However, serum CK of several thousands have been reported in patients with proven BM and this is also our experience. Muscle MRI can help in the differential diagnosis as patients with Bethlem myopathy show a typical pattern of muscle involvement with a concentric atrophy of the rectus and vastus lateralis [7]. Our results suggest that muscle MRI may help in the differential diagnosis between patients with LGMD2A and ADED and Bethlem myopathy. These results are most helpful when patients are young and less affected and patterns of selective involvement are better observed. This is typically the time when the various diagnostic investigations are requested, and the identification of a specific MRI pattern might help to target the appropriate gene. The finding of a selective pattern of skeletal muscle involvement in LGMD2A despite their unusual phenotype will also help to persevere in the search of mutations in the calpain 3 gene in cases with apparently normal protein expression on western blot. We also predict that it will help the study of cases with partial reduction of calpain 3, in whom the deficiency could be either primary or represent a secondary defect due to mutations in other genes. In this study, we used a short protocol, suitable for pediatric age [6–9] with standard T1transverse sequences.
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This short protocol provides reliable information on the presence and the degree of abnormal signal within individual muscles in the legs but has some limitations related to the short duration of the exam. Further information could be achieved by imaging the upper limbs and the paravertebral muscles. A more quantitative analysis may also provide further information on the degree of atrophy and of abnormal signal which are difficult to obtain with our short protocol.
Acknowledgements This study was supported by the MDC centre grant, United Kingdom, Banca Telethon GTF02009, ’EurobioEurobiobank Network (EC Fifth Framework Programme (QLRI-CT-2002-02769) and by the Italian Ministry of Health. The authors thank Caroline Sewry, Louise Anderson and Lucy Feng for their help with muscle biopsies.
References [1] Fanin M, Nascimbeni AC, Fulizio L, et al. Loss of calpain-3 autocatalytic activity in LGMD2A patients with normal protein expression. Am J Pathol 2003;163:1929–36. [2] Fardeau M, Hillaire D, Mignard C, et al. Juvenile limb-girdle muscular dystrophy. Clinical, histopatological and genetic data from a small community living in the Reunion Island. Brain 1996;119: 295–308. [3] Dincer P, Leturcq F, Richard I, et al. A biochemical, genetic, and clinical survey of autosomal recessive limb girdle muscular dystrophies in Turkey. Ann Neurol 1997;42:222–9.
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[4] Pollit C, Anderson LVB, Pogue P, et al. The phenotype of calpainopathy: diagnosis based on a multidisciplinary approach. Neuromuscul Disord 2001;11:287–96. [5] Fardeau M, Eymard B, Mignard C, et al. Chromosome 15-linked limb-girdle muscular dystrophy: clinical phenotypes in Reunion Island and French metropolitan communities. Neuromuscul Disord 1996;6:447–53. [6] Mercuri E, Counsell S, Allsop J, et al. Selective muscle involvement on magnetic resonance imaging in autosomal dominant Emery–Dreifuss muscular dystrophy. Neuropediatrics 2002;33: 10–14. [7] Mercuri E, Cini C, Counsell S, et al. Muscle MRI findings in a threegeneration family affected by Bethlem myopathy. Eur J Paediatr Neurol 2002;6:309–14. [8] Mercuri E, Talim B, Moghadaszadeh B, et al. Clinical and imaging findings in 6 cases of congenital muscular dystrophy with rigid spine syndrome linked to chromosome 1p (RSMD1). Neuromuscul Disord 2002;12:631–8. [9] Mercuri E, Yuva Y, Brown SC, et al. Collagen VI involvement in Ullrich Syndrome: a clinical, genetic and immunohistochemical study. Neurology 2002;58:1354–9. [10] Topaloglou H, Dincer P, Richard I, et al. Calpain-3 deficiency causes a mild muscular dystrophy in childood. Neuropediatrics 1997;28: 212–6. [11] Bonne G, di Barletta MR, Varnous S, et al. Mutation in the gene enconding lamin A/C cause autosomal dominant Emery–Dreifuss muscular dystrophy. Nat Genet 1992;21:285–8. [12] Bonne G, Mercuri E, Muchir A, et al. Clinical and molecular genetic spectrum of autosomal dominant Emery–Dreifuss muscular dystrophy due to mutations of the lamin A/C gene. Ann Neurol 2000;48:170–80. [13] Bonne G, Ben Yaou R, Beroud C, et al. 108th ENMC international workshop, EUROMEN, 7th international Emery–Dreifuss muscular dystrophy (EDMD) workshop, 13–15 September 2002, Naarden, The Netherlands. Neuromuscul Disord 2003;13:508–15. [14] Jobsis GJ, Boers JM, Barth PG, de Visser M. Bethlem myopathy: a slowly progressive congenital muscular dystrophy with contractures. Brain 1999;122:649–55.
Muscle MRI findings in patients with limb girdle muscular dystrophy with calpain 3 deficiency (LGMD2A) and early contractures Eugenio Mercuria,b,*, Kate Bushbyc, Enzo Riccid, Daniel Birchalle, Marika Paneb, Maria Kinalia, Joanna Allsopf, Vincenzo Nigrog, Amets Sa´enzh, Annachiara Nascimbenii, Luigi Fulizioi, Corrado Angelinii, Francesco Muntonia a
Department of Paediatrics and Neonatal Medicine, Dubowitz Neuromuscular Centre, Imperial College London, Hammersmith Hospital, Du Cane Road, London W12 0NN, UK b Pediatric Neurology Unit, Catholic University Hospital, Rome, Italy c Institute of Human Genetics, International Centre for Life, University of Newcastle, Newcastle upon Tyne, UK d Institute of Neurology, Catholic University Hospital, Rome, Italy e Department of Radiology, University of Newcastle, Newcastle upon Tyne, UK f MRI Robert Steiner Unit, Imperial College London, UK g Telethon Institute of Genetics and Medicine, Naples, Italy h Grupo Neurogene´tica, Unidad Experimental Hospital Donostia, San Sebastia´n, Spain i Department of Neuroscience, University of Padua, Padua, Italy Received 28 July 2004; received in revised form 11 October 2004; accepted 14 October 2004
Abstract Limb girdle muscular dystrophy 2A is a common variant secondary to mutations in the calpain 3 gene. A proportion of patients has early and severe contractures, which can cause diagnostic difficulties with other conditions. We report clinical and muscle magnetic resonance imaging findings in seven limb girdle muscular dystrophy 2A patients (four sporadic and three familial) who had prominent and early contractures. All patients showed a striking involvement of the posterior thigh muscles. The involvement of the other thigh muscles was variable and was related to clinical severity. Young patients with minimal functional motor impairment showed a predominant involvement of the adductors and semimembranosus muscles while patients with restricted ambulation had a more diffuse involvement of the posterolateral muscles of the thigh and of the vastus intermedius with relative sparing of the vastus lateralis, sartorius and gracilis. At calf level all patients showed involvement of the soleus muscle and of the medial head of the gastrocnemius with relative sparing of the lateral head. MRI findings were correlated to those found in two patients with the phenotype of limb girdle muscular dystrophy 2A without early contractures and the pattern observed was quite similar. However, the pattern observed in limb girdle muscular dystrophy 2A is different from that reported in other muscle diseases such as Emery–Dreifuss muscular dystrophy and Bethlem myopathy which have a significant clinical overlap with limb girdle muscular dystrophy 2A once early contractures are present. Our results suggest that muscle MRI may help in recognising patients with limb girdle muscular dystrophy 2A even when the clinical presentation overlaps with other conditions, and may therefore, be used as an additional investigation to target the appropriate biochemical and genetic tests. q 2004 Elsevier B.V. All rights reserved. Keywords: Calpain; MRI; Muscle; Contractures
1. Introduction * Corresponding author. Address: Department of Paediatrics and Neonatal Medicine, Imperial College London, Hammersmith Hospital, Du Cane Road, London W12 0NN, UK. Tel.: C44 20 83833295; fax: C44 20 87408281. E-mail address: [email protected] (E. Mercuri). 0960-8966/$ - see front matter q 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.nmd.2004.10.008
Limb girdle muscular dystrophies are a clinically and geneticallyheterogeneous group of disorders which shareprogressive muscle weakness and degenerative muscle changes. The form of limb girdle muscular dystrophy secondary to
E. Mercuri et al. / Neuromuscular Disorders 15 (2005) 164–171
calpain 3 deficiency (LGMD2A) is one of the most common forms with a reported frequency ranging from 9 to 40% of limb girdle muscular dystrophies. Calpain-3 is a muscle specific intracellular protease. LGMD2A was the first type of muscular dystrophy shown to be caused by an enzymatic defect rather than a defect in a structural muscle protein. From a diagnostic point of view, the definitive diagnosis of LGMD2A relies on the identification of mutations in the calpain 3 gene, located on 15q15-q21. Although a deficiency of calpain 3 expression on western blot analysis can be demonstrated in most cases of LGMD2A, a secondary reduction of calpain 3 has been reported in other forms of recessive limb girdle muscular dystrophy such as LGMD2B or LGMD2J in which the primary defects are, respectively, in dysferlin and titin. Furthermore, recent reports suggest that immunoblot for calpain 3 can be normal in some patients carrying missense mutations in the calpain 3 gene [1]. The original paper reporting clinical and genetic findings in Reunion Island patients with LGMD2A described limb girdle distribution of wasting, scapular winging and hip girdle weakness which sparing of the hip abductors [2]. Contractures were often limited to the ankles in the early phases of the diseases and became more diffuse in patients on a wheelchair. Although similar clinical findings have been reported in other patients by several subsequent studies [3,4], variations from this phenotype have also been reported. Fardeau et al. in 1996 described two French metropolitan families with LGMD2A, early onset of contractures, especially of the elbows, and more obvious involvement of the distal muscles [5]. This phenotype, also characterised by early elbow, finger and hip flexor contractures has been reported in a few more cases [4]. In these patients, the diagnosis based on clinical ground only can be difficult as the distribution of muscle wasting, weakness and contractures is similar to Emery–Dreifuss muscular (EDMD) dystrophy and overlaps also with Bethlem myopathy. In the last few years, several studies have reported the value of muscle magnetic resonance imaging (MRI) in identifying patterns of muscle involvement in genetically distinct muscle disorders [6–9]. The aim of this study was to report clinical and MR findings in seven patients with LGMD2A and early contractures. More specifically, we wished to establish whether muscle MRI can identify a specific pattern of muscle involvement in patients with such phenotype and whether MR findings are different from those found in patients with LGMD2A and the ‘non-contracted’ phenotype, and from other conditions such as Bethlem myopathy and EDMD.
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and rigidity of the spine were included in the study. In six out of seven, the diagnosis of LGMD2A was confirmed by genetic analysis showing a mutation in the CAPN3 gene while in the remaining one the diagnosis was suggested not only by the clinical picture but also the complete absence of calpain 3 on Western blot. Mutations in the LMNA gene, responsible for the dominant variant of EDMD were excluded in all. We systematically collected data regarding age and mode of onset, muscle power, contractures, functional abilities, progression of the disease, cardiac and respiratory function. Data on muscle biopsy were also collected. Clinical and immunohistochemical details of three patients have already been reported [4]. Details of histological, histochemical and immunohistochemial procedures have already been described in the same paper [4]. 2.2. Control group Another two patients followed at the Catholic University in Rome, with genetically confirmed diagnosis of LGMD2A and with the phenotype without early contractures were also included in the study as controls for the MRI study. 2.3. Muscle magnetic resonance imaging All the patients were fully co-operative and no sedation or general anaesthesia was required. Muscle MRI was performed on a 1.0-tesla Picker HPQ system, using conventional T1 weighted spin echo. Non-contrast transverse images were obtained in the legs, selecting the axial plane with respect to the long axis of the body. This involved two sequential scans. The slices were 5 mm thick and the gap between slices was 50 mm. TR was 525 msec, TE 14 msec. The scans were examined by two independent observers, blinded to clinical and genetic data, looking for normal and abnormal muscle bulk (atrophy) and for normal and abnormal signal intensity within the different muscles. Sections were generally analysed within the mid-upper section of the thighs and lower legs, for the reason that muscle bulk is greatest at this level and that muscle abnormality could be more clearly visualised as a result. Care was taken to review the remainder of the muscle bulk in order to confirm that the analysed section was representative of the whole muscle. The abnormal signal was classified as mild if only traces of increased signal intensity could be observed in an otherwise well preserved muscle, moderate if less than 50% of the muscle showed increased signal intensity, and severe if at least 50% of it showed increased signal intensity. The muscles examined were:
2. Subjects and methods 2.1. Study group Seven patients (four sporadic and three familial cases) with a diagnosis of LGMD2A, early elbow contractures
anterior thigh: sartorius, gracilis, quadriceps femoris. posterior thigh: semimembranosus, semitendinosus, biceps femoris, adductor muscles. lower leg: gastrocnemius, soleus, tibialis anterior, tibialis posterior, extensor allucis and extensor digitorum longum.
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3. Results
4. Muscle MRI
3.1. Age of onset
4.1. Thigh muscles
All patients had normal motor milestones and first clinical signs were noted between 2 and 15 years, consisting in toe walking due to Achilles tendon contractures and in one case, in muscle pain and cramps following exercise. 3.2. Maximal functional ability All our patients are still able to walk independently but the four oldest patients have restricted ambulation (details on Table 1).
4.1.1. Study group All seven patients showed a striking involvement of the adductor magnus, associated with variable involvement of the adductor longus and semimembranosus (Fig. 2(a)–(c))in the three youngest children. The oldest patients showed a more severe and diffuse involvement of thigh muscles with prominent involvement of adductors, vastus intermedius and semimembranosus and relative sparing of sartorius and gracilis and partly of the vastus lateralis. (Fig. 2(d) and 3(a,b)).
3.3. Clinical examination None of our patients had facial weakness. All but one showed wasting of the pectoralis and biceps with a striking differential involvement between biceps and triceps. In five cases, the biceps appeared a thin and fibrotic string (Fig. 1). All patients had generalised weakness involving trunk and limbs (proximal more than distal) and the weakest muscle were the hip adductors and extensors which always had subgravity power. Hip abductors were invariably better preserved with antigravity power. All but one also had mild distal weakness in the lower limbs. All patients had TA tightness, which was always the presenting sign but in addition developed marked elbow and hamstrings contractures and spine rigidity which was severe in three cases mostly affecting the lumbar and lower thoracic spine. Five of the seven also had long finger contractures. 3.4. Serum creatine kinase Serum CK was always markedly elevated (15–20 times the normal limits). 3.5. Respiratory function FVC was always relatively well preserved (70–90% of the predicted values) compared to the proximal muscle power and no patients had frequent chest infection or symptoms of nocturnal hypoventilation. 3.6. Cardiac function All patients had serial ECG which were always normal. All seven also had echocardiograms (age 13–37) which were also normal. 3.7. Progression Motor difficulties were progressive after the onset of clinical signs. Although none of our patients has lost independent ambulation they have all experienced increasing difficulties and progressive restriction of walking.
4.1.2. Control group The two patients with the non-contracted phenotype showed a diffuse involvement of the postero-lateral thigh muscles with prominent involvement of adductors, vastus intermedius and semimembranosus and relative sparing of sartorius and gracilis and partly of the vastus lateralis (Fig. 3(c) and (d)). 4.2. Calf muscles 4.2.1. Study group All seven patients showed a relatively normal anterior compartment with relative sparing of the tibialis anterior (7/7) and peroneals muscles (6/7). In the posterior compartment all seven showed a predominant involvement of the medial head of the gastrocnemius, with relative sparing of the lateral head of the gastrocnemius (Fig. 4(a)– (d)), associated with a variable involvement of the soleus. 4.2.2. Control group Both patients with the non-contracted phenotype showed a relatively normal anterior compartment and a predominant involvement of the gastrocnemius, with relative sparing of the lateral head of the gastrocnemius in one of the two and involvement of the soleus.
5. Correlation between clinical severity and MRI changes 5.1. Study group Three of the seven patients were still able to walk independently for relatively long distances. All three showed a selective impairment of adductors muscles and semimembranosus. The other four patients had more restricted ambulation and could only walk short distances with support. All of these patients had a more diffuse involvement of the postero-lateral thigh muscles. Changes in the calf muscles were similar in all seven patients, regardless of severity.
Table 1 Clinical findings Onset
Max functional ability
Age examination (years)
Progress max ability when last seen
Scapular winging
Wasting
Weakness
Contractures
Rigidity of spine
Scoliosis
FVC (%)
Cardiac
CK
1
6 yrs TA contractures, 7 yrs weakness
Walking at 1 year
15
Walking
Yes
YesCC
K
94
ECG: N, Echo: N
X 30
10 yrs TA contractures
Walking at 1 year
13
Walking
Yes
Ankles, hamstrings, fingers, elbows
YesC
K
71
ECG: N, Echo: N
X 15
3
8 yrs: TA contractures, scapular winging 15yrs: pain and cramps, TA and elbow contractures 2 yrs: TA contractures
Walking at 1 year
16
Walking
Yes
Proximal Odistal, adductors subgravity, abductors antigr with res, some distal weakness Proximal Odistal, adductors subgravity, abductors antigr with res, some distal weakness ProximalOdistal, hip add subgravity, abductors antigr with res
Ankles, hamstrings, elbow
2
Pectoralis, biceps, trapezius. medial calves Pectoralis, biceps, hamstrings, medial calves Pectoralis, biceps, deltoid
ITB, ankles, elbow
C
K
94
ECG: N, Echo: N
X 20
Walking at 1 year
38
Walking with support (stick)
Yes
CC
K
80
ECG: N, Echo: N
X 20
14
Yes
Paraspinal, finger, ankle
K
C
80
ECG: N, Echo: N
X 45
6
6 yrs: TA contractures
Walking at 1 year
17
Walking with support, wheelchair part-time since age 13 years Walking with support, climbs stairs with railing
Proximal Odistal, adductors subgravity, abductors antigr with res, some distal weakness Proximal Odistal, adductors subgravity, abductors antigr with res, ankle dorsiflexion weaker than plantar flexion
Ankles, fingers, hamstrings, elbows
Walking at 1 year
Pectoralis, biceps, trapezius. medial calves Pectoralis, biceps, medial calves
Yes
Pectoralis, biceps, medial calves
Paraspinal, neck, ankle, wrist, elbow, finger
K
K
84
ECG: N, Echo: N
7
9 yrs: pain, TA contractures
Walking at 1 year
38
Walking with support, climbs stairs both hands on banister
Yes
Pectoralis, biceps, medial calves
Proximal Odistal, adductors antigr with res, abductors antigr with res, ankle dorsiflexion weaker than plantar flexion Proximal Odistal, adductors subgravity, abductors antigr with res, ankle dorsiflexion weaker than plantar flexion
Ankle
CC
K
90
ECG: N, Echo: N
4
5
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Case
X6
yrs, years; antigr, antigravity; res, resistence; TA, tendon Achilles; N, normal. 167
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Fig. 1. Thin and fibrotic biceps (patient 1).
6. Discussion The seven cases with LGMD2A presented in this paper have calf hypertrophy, rigidity of the spine and early contractures affecting Achilles tendons, elbow and long finger flexors. The distribution of muscle weakness and contractures in these patients is in keeping with what
previously described by Fardeau et al. in 1996 as a variant of the phenotype originally described in patients with LGMD2A. A similar ‘contracted’ phenotype has been previously reported in a few other patients with LGMD2A [4] but it is not well recognised in the literature. As a significant number of our patients presented with these features, we want to draw attention to this phenotype, which has a significant clinical overlap with other conditions, in particular with Emery–Dreifuss muscular dystrophy (EDMD) and the severe end of the Bethlem myopathy spectrum. Using muscle MRI we have been able to identify a distinct pattern of muscle involvement. The severity of the involvement was variable and was related to the clinical severity of motor impairment. The three younger patients who were able to walk independently for relatively long distances had a selective impairment of the adductor magnus and of the semimembranosus. In the four oldest patients, who all had restricted ambulation, there was a similar involvement of the adductor magnus, but this was associated with a more diffuse involvement of the posterior thigh muscles and vasti intermedius and a with selective sparing of sartorius and gracilis muscles and vastus lateralis. Although none of the previous studies has systematically evaluated muscle imaging in patients with LGMD2A,
Fig. 2. Transverse T1 weighted images through thigh muscles in four patients with contracted phenotype (cases 1–4). Note the predominant involvement of the adductors muscles and the early signs of involvement of the semimembranosus in the three young patients (cases 1–3, panels a,b,c) and the more diffuse involvement in the oldest patient (case 4, panel d) with relative sparing of the sartorius.
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Fig. 3. Transverse T1 weighted images through thigh muscles in two patients with contracted phenotype (cases 5, 6, panel a and b) and in two patients with the non-contracted phenotype (panel c and d). Note the predominant involvement of the posterior muscles and of the vastus intermedius and medialis with relative sparing of the sartorius, gracilis and partly of the vastus lateralis and of the rectus femoris in all four patients regardless of their clinical phenotype.
the picture of muscle MRI [10] and CT [2,5] previously reported show similar findings at thigh level. The pattern of muscle involvement in the calves consisted in a selective involvement of the medial head of the gastrocnemius muscle compared to the lateral head, which was always relatively spared. The involvement of the gastrocnemius was associated with variable involvement of the soleus muscle. It is of interest that a similar pattern of muscle involvement was also found in the control group of patients with LGMD2A but with the non-contracted phenotype. Although we had only two patients with the non-contracted phenotype who had a scan, when patients with the same age and level of function were compared, the pattern observed was very similar with prominent involvement of the posterior thigh muscles and vastus intermedius and with relative sparing of vastus lateralis, sartorius and gracilis. These findings, therefore, suggest that, despite the differences in muscle wasting and in contractures, both forms of LGMD2A have a similar pattern of muscle involvement on muscle MR, which appears to vary in relation to the severity of clinical functional impairment. This is in keeping with our studies in a number of genetically distinct forms of muscular dystrophies including EDMD, a variety of congenital myopathies and in Bethlem myopathy and neurogenic disorder, in which we
were able to recognise a disease specific pattern of selective muscle involvement. The pattern of involvement on muscle MRI observed in LGMD2A was, in contrast, different from those previously reported in other forms of muscle disorders with overlapping clinical phenotype, such as Emery–Dreifuss muscular dystrophy. Although the differential involvement of the medial and lateral head of the gastrocnemius observed in the calf muscles in patients with LGMD2A was similar to that reported in patients with the dominant form of EDMD [6], the pattern of muscle involvement at thigh level was different. While patients with LGMD2A had a more striking involvement of the adductor muscles and relative sparing of the vastus lateralis, the vasti muscles were selectively affected in the dominant form of EDMD. The pattern of selective muscle involvement observed in patients with LGMD2A was also different from that observed in the X-linked form of Emery–Dreifuss muscular dystrophy, characterised by a more prominent involvement of the soleus and relative sparing of the gastrocnemii muscles [6]. This information can be important at the time of directing mutation analysis as the two forms have a significant clinical overlap. In both forms the age of onset is variable but the most common presentation is TA tightness at school age. Other common features are scapuloperoneal wasting and scapular winging, rigidity of the spine, elbow
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Fig. 4. Transverse T1 weighted images through calf muscles in four patients with contracted phenotype (cases 1–4). Note the differential involvement between medial and lateral head of the gastrocnemius and the variable involvement of the soleus. Perneus muscles were only mildly affected in case 1 (panel a).
contractures and, in some cases, a striking wasting of biceps muscles in the arms with relatively preserved triceps muscles. Both the dominant form of EDMD and LGMD2A show some wasting in the medial aspects of the calves but there are some differences in the distribution of muscle wasting in the thighs. While patients with LGMD2A have a striking wasting of the medial and posterior aspects of the thighs with a relative hypertrophy of the quadriceps, patients with the dominant form of EDMD generally have more diffuse wasting [11–13]. The absent cardiac involvement in LGMD2A is a major difference compared to what found in EDMD. However, in EDMD cardiac involvement is found invariably only in patients above the age of 30 years and as all our patients came to our observation before the age of 20, a normal cardiac assessment cannot exclude the diagnosis. Both forms also have dystrophic changes on muscle biopsy and markedly increased CK, although in the dominant form of EDMD CK levels are more variable and overall tend to be lower than in our cases with LGMD2A [12,13]. The phenotype with early contractures also has some clinical overlap with Bethlem myopathy [12,14], especially if patients are in the first decade or in their teens. CK levels, however, are generally lower in Bethlem myopathy and the changes on muscle biopsy are generally non-dystrophic.
However, serum CK of several thousands have been reported in patients with proven BM and this is also our experience. Muscle MRI can help in the differential diagnosis as patients with Bethlem myopathy show a typical pattern of muscle involvement with a concentric atrophy of the rectus and vastus lateralis [7]. Our results suggest that muscle MRI may help in the differential diagnosis between patients with LGMD2A and ADED and Bethlem myopathy. These results are most helpful when patients are young and less affected and patterns of selective involvement are better observed. This is typically the time when the various diagnostic investigations are requested, and the identification of a specific MRI pattern might help to target the appropriate gene. The finding of a selective pattern of skeletal muscle involvement in LGMD2A despite their unusual phenotype will also help to persevere in the search of mutations in the calpain 3 gene in cases with apparently normal protein expression on western blot. We also predict that it will help the study of cases with partial reduction of calpain 3, in whom the deficiency could be either primary or represent a secondary defect due to mutations in other genes. In this study, we used a short protocol, suitable for pediatric age [6–9] with standard T1transverse sequences.
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This short protocol provides reliable information on the presence and the degree of abnormal signal within individual muscles in the legs but has some limitations related to the short duration of the exam. Further information could be achieved by imaging the upper limbs and the paravertebral muscles. A more quantitative analysis may also provide further information on the degree of atrophy and of abnormal signal which are difficult to obtain with our short protocol.
Acknowledgements This study was supported by the MDC centre grant, United Kingdom, Banca Telethon GTF02009, ’EurobioEurobiobank Network (EC Fifth Framework Programme (QLRI-CT-2002-02769) and by the Italian Ministry of Health. The authors thank Caroline Sewry, Louise Anderson and Lucy Feng for their help with muscle biopsies.
References [1] Fanin M, Nascimbeni AC, Fulizio L, et al. Loss of calpain-3 autocatalytic activity in LGMD2A patients with normal protein expression. Am J Pathol 2003;163:1929–36. [2] Fardeau M, Hillaire D, Mignard C, et al. Juvenile limb-girdle muscular dystrophy. Clinical, histopatological and genetic data from a small community living in the Reunion Island. Brain 1996;119: 295–308. [3] Dincer P, Leturcq F, Richard I, et al. A biochemical, genetic, and clinical survey of autosomal recessive limb girdle muscular dystrophies in Turkey. Ann Neurol 1997;42:222–9.
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[4] Pollit C, Anderson LVB, Pogue P, et al. The phenotype of calpainopathy: diagnosis based on a multidisciplinary approach. Neuromuscul Disord 2001;11:287–96. [5] Fardeau M, Eymard B, Mignard C, et al. Chromosome 15-linked limb-girdle muscular dystrophy: clinical phenotypes in Reunion Island and French metropolitan communities. Neuromuscul Disord 1996;6:447–53. [6] Mercuri E, Counsell S, Allsop J, et al. Selective muscle involvement on magnetic resonance imaging in autosomal dominant Emery–Dreifuss muscular dystrophy. Neuropediatrics 2002;33: 10–14. [7] Mercuri E, Cini C, Counsell S, et al. Muscle MRI findings in a threegeneration family affected by Bethlem myopathy. Eur J Paediatr Neurol 2002;6:309–14. [8] Mercuri E, Talim B, Moghadaszadeh B, et al. Clinical and imaging findings in 6 cases of congenital muscular dystrophy with rigid spine syndrome linked to chromosome 1p (RSMD1). Neuromuscul Disord 2002;12:631–8. [9] Mercuri E, Yuva Y, Brown SC, et al. Collagen VI involvement in Ullrich Syndrome: a clinical, genetic and immunohistochemical study. Neurology 2002;58:1354–9. [10] Topaloglou H, Dincer P, Richard I, et al. Calpain-3 deficiency causes a mild muscular dystrophy in childood. Neuropediatrics 1997;28: 212–6. [11] Bonne G, di Barletta MR, Varnous S, et al. Mutation in the gene enconding lamin A/C cause autosomal dominant Emery–Dreifuss muscular dystrophy. Nat Genet 1992;21:285–8. [12] Bonne G, Mercuri E, Muchir A, et al. Clinical and molecular genetic spectrum of autosomal dominant Emery–Dreifuss muscular dystrophy due to mutations of the lamin A/C gene. Ann Neurol 2000;48:170–80. [13] Bonne G, Ben Yaou R, Beroud C, et al. 108th ENMC international workshop, EUROMEN, 7th international Emery–Dreifuss muscular dystrophy (EDMD) workshop, 13–15 September 2002, Naarden, The Netherlands. Neuromuscul Disord 2003;13:508–15. [14] Jobsis GJ, Boers JM, Barth PG, de Visser M. Bethlem myopathy: a slowly progressive congenital muscular dystrophy with contractures. Brain 1999;122:649–55.