Late onset muscular dystrophy with cerebral white matter changes due to partial merosin deficiency

PERGAMON

Neuromuscular

Disorders 7 (1997) 85-89

Late onset muscular dystrophy with cerebral white matter changes due to partial merosin deficiency Ersin Tan ‘,*, Haluk Topaloglu b, Caroline Sewry ‘, Yasar Zorlu ‘, Isam Naom ‘, Sevim Erdem a, Mariella D’Alessandro ‘, Francesco Muntoni ‘, Victor Dubowitz ’ a Neuromuscular Diseases Research Laboratory, Department of’Adult Neurology, Hacettepe University Hospitals, Ankara, Turke) b Neuromuscular Diseases Research Laboratory, Department of Pediatric Neurology, Hacettepe, Ankara, Turke! ’ Department of‘Paediatrics and Neonatal Medicine, Neuromuscular Unit Royal Postgraduate Medical School. Hammersmith Hospital, London, UK d I:mir Tepecik Social Insurance Hospital. Izmir, Turke)

Received 30 August 1996: received in revised form 21 November 1996: accepted 4 December 1996

Abstract Merosin-deficient congenital muscular dystrophy (CMD) is an autosomal recessive condition usually with onset at birth or within the first months of life. Affected children are severely disabled and usually do not achieve the ability to walk without support. They invariably have white matter abnormalities on brain magnetic resonance imaging (MRI). We report a 29-year-old man with a late childhood onset limb-girdle type muscular dystrophy and cerebral white matter changes on MRI. lmmunocytochemical studies of the patient’s muscle biopsy showed a reduction in expression of the laminin c(? chain of merosin. The patient had three affected siblings, and microsatellite genotyping confirmed linkage to the laminin CQlocus (LAMA2) on chromosome 6q2

in this family. This case probably represents a milder allelic variant of classical merosin-deficient CMD. Merosin status should be assessed in patients with late-onset limb girdle muscular dystrophy. 0 1997 Elsevier Science B.V. Keywords: Congenital

muscular dystrophy;

Leukoencephalopathy;

1. Introduction Congenital muscular dystrophy (CMD) is a severe autosomal recessive congenital myopathy characterised by muscle weakness and hypotonia at birth, or within the first few months of life, and dystrophic changes on muscle biopsy [l]. These inclusion criteria have been agreed at a recent workshop on CMD [2]. Approximately half of the children with the classical form of CMD have recently been shown to have a deficiency of the laminin CI*chain, a component of the extracellular matrix protein laminin-2 (merosin) [3,4]. Affected children have severe hypotonia, normal intelli* Corresponding author. Present address: Cobanyildizi sokak 7A/ 15, Cankaya Ankara 06680 Turkey. Tel.: +90 312 4682261; fax: + 90 312 4275511. 0960-8966/97i$l7.00 0 1997 Elsevier Science B.V. All rights reserved. PI1 SO960-8966(96)00421-X

Merosin

and central nervous system white matter gence, changes. They have very poor functional achievement and usually never walk independently. Linkage to chromosome 6q2 has been established and some mutations in the LAMA gene coding for the laminin a2 chain have been identified [5,6]. The diagnosis of this form of CMD is made from analysis of the expression of merosin in muscle or skin biopsies, and affected individuals show an absence or only traces of the protein [3,4,7]. In contrast to the severe, neonatal phenotype of CMD with a deficiency of merosin, we present a case of late childhood onset muscular dystrophy with cerebral white matter changes who was found to have a primary, partial deficiency of merosin. This suggests the presence of a milder allelic variant of the merosin-deficient CMD.

86

E. Tan et al. , Nrurornuscular

D65407

3 CM

C

NO

LOCUS

D651620

D65407

D6S1705

D6S1705

D6Sl620

0

1CM

0651572 D651572

DSS262

I

D65457

D65262

1 CM

7 (1997) 85-89

Markers

Genetic distance

0

Disortlers

065457

1 CM

D6S1656 0

I

1 D6S413 1cM D6S472 2 CM D6S975 1 CM D6SZ70 2 CM

D6S1626

0

I DSS292 Fig. 1. Family of the figure.

pedigree

showing

the identical

haplotypes

in four affected

2. Case report The patient was a 29-year old man who had been diagnosed with proximal weakness at the age of 12 years. He has five sisters, three of whom are also affected (Fig. 1). His oldest sister became wheelchair bound at the age of 28 years. There is no consanguinity or other relevant family history. His intrauterine movements had been normal. He was born at term with a birthweight of 3350 g. There were no neonatal problems. He sat without support at 6 months, pulled to standing at 13 months and walked at 16 months. Serum creatine kinase activity was markedly elevated at 2417 IUjl (upper limit of normal 200 IUjl). Neurological examination revealed normal intelligence and normal cranial nerve function. He had a waddling gait with a lordotic posture. Muscle strength was symmetrically diminished in proximal muscle groups and tendon reflexes and vibration and pin sensation were normal. EMG showed a myopathic pattern with normal nerve conduction velocities. Brain MRI showed a marked increase in periventricular signal (Fig. 2). His somatosensory and visual evoked potentials recordings were normal. 2.1. Muscle biopsy Muscle biopsy from the left biceps brachii showed dystrophic features, with marked variability in muscle fiber size, increased endomysial and perimysial connective tissue, fiber splitting and occasional fiber degeneration. lmmunohistochemical studies showed normal expression of dystrophin (DYS 1, 2 and 3; Novocastra) a,-sarcoglycan and P-dystroglycan components of the

siblings.

The localisation

of LAMA

locus is indicated

in the left side

dystrophin associated glycoprotein complex (50 DAG, 43 DAG; Novocastra). Expression of the laminin a2 chain using a commercial antibody that recognises the 80 kDa fragment (Chemicon) was only very slightly reduced but with an antibody to the 300 kDa fragment [7,8] there was a pronounced reduction (Fig. 3). lmmunolabeling of the laminin cc? chain with both antibodies in a variety of disease-control patients, including children affected by Duchenne and Becker muscular dystrophy, adhalin deficiency (a,-sarcoglycan) limb-girdle muscular dystrophy, Emery-Dreifuss muscular dystrophy, facioscapulohumeral muscular dystrophy and juvenile dermatomyositis was normal. The specificity of 300 kDa antibody has been verified in CMD patients with partial or complete merosin deficiency in muscle and skin [7]. 2.2. Linkuggr anulvsis Genomic DNA was isolated from whole blood by the salt-chloroform method [9]. Thirteen highly informative CA-repeat polymorphisms spanning an area of 16 CM on chromosome 6q2 were selected [lo]. The microsatellite markers were typed in all available family members using the radionuclide end-labeling method described previously [I 11. A two-point LOD score was calculated using the LINK program of the linkage package (version 5.2), assuming a recessive disease frequency of 0.001 with complete penetrance. The four affected members in this family all showed identical haplotypes (Fig. 1). A recombination in the maternally derived haplotype of member 1l-3 (unaffected) was found between markers D6S407 and D6S1620. This is in keeping with the recent localisation

E. Tun et al. /Neuromuscular

Fig. 2 T 2W-SE

(TR/TE:2001/80

ms) transverse

MRI

Disorders

scan shows symmetric

of the LAMA locus in a 3 cm interval between these two markers [12]. A maximum LOD score of 2.052 was obtained at O:O.OOl with the D6S407 marker, strongly supporting linkage to the LAMA locus.

3. Discussion Congenital muscular dystrophy usually presents at birth or within the first few months of life with hypotonia and is characterised by dystrophic changes on muscle biopsy. Recently, a defect in the expression of the x2 chain of merosin has been documented in approximately 40&50% of children with classical CMD [3,4]. These patients are a relatively homogeneous clinical group with severe neonatal hypotonia associated with joint stiffness and contractures, delayed motor milestones, elevated CK and MRI, changes consistent with abnormalities of white matter myelination. Merosin deficiency has so far only been reported in classical CMD and in the Fukuyama form of CMD, prevalent in Japan. The latter form, however, links to chromosome 9q, and not to the locus for the laminin CI~ gene on chromosome 6q22-23, indicating that the merosin defect in Fukuyama CMD is a secondary phenomenon [ 131. The abnormality in merosin expression in the muscle, and linkage to the LAMA locus on chromosome 6q2, indicate that the primary defect in the family presented is a mutation that only partially disrupts the expression of the laminin c(~ chain. Affected members in this family have white matter changes in the central nervous system similar to those seen in with the severe form of the disease but have a milder involvement of skeletal mus-

87

7 (1997) 85589

periventricular

white matter

hyperintensities.

cle. To our knowledge, this is the first report of latechildhood onset of merosin deficiency. There is only one case report that describes 3 siblings with adult onset of CMD with leucoencephalopathy but there is no information on the merosin status. The authors concluded it was a newly recognised autosomal recessive syndrome and it might be due to an allelic mutation at the putative CMD locus [14]. Hermann et al. [15], recently defined a milder phenotype of CMD in two patients with partial expression of the laminin x2 chain. The age of onset was less than 2 years in both patients and their disease followed a milder course compared to CMD patients with an absence of laminin c(, expression. The mechanisms that lead to the pathological features in merosin-deficiency are not certain. Merosin is expressed in the basement membrane of each muscle fiber and of Schwann cells in the peripheral nervous system and is know to have diverse biological functions. These include mediation of cell attachment, promotion of neurite outgrowth and Schwann cell migration. In the brain, recent evidence has shown that the laminin c(? chain is only found in the blood brain barrier [16] but the reasons for the white matter changes are not known. We conclude that the spectrum of severity due to merosin-deficiency is wider than previously thought and encompasses not only the typical, early onset and severe form of congenital muscular dystrophy, but also a late childhood onset limb girdle dystrophy. Late onset autosomal recessive muscular dystrophy with merosin deficiency should therefore be considered amongst the differential diagnoses in patients presenting with a limb girdle type of muscular dystrophy, especially if accom-

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E. Tan et al. /Neuromuscular

Disorders 7 (1997) 85-89

Fig. 3. Immunolabeling of laminin G(~chain in control muscle (A),(B) compared with the case presented (C),(D) using an antibody to the 80 kDa fragment (A),(C) and an antibody to the 300 kDa fragment (B),(D) of the laminin x2 chain. Note the near normal labelling with the 80 kDa antibody (C) but very reduced labeling with the 300 kDa antibody (D) in the patient ( x 170).

panied by white matter changes on MRI. Merosin expression should therefore be studied in all children with limb girdle muscular dystrophy, after exclusion of other defects such as sarcoglycan abnormalities.

Acknowledgements

The authors thank Dr Lydia Sorokin (Germany) for the 300 kDa. This study was supported, by a grant from the Muscular Dystrophy of Great Britain and Northern Ireland and by the Medical Research Council (CAS).

References [II Dubowitz

V. Muscle disorders of childhood, second ed. London: Saunders, 1995. I21Dubowitz V. Workshop report on 22nd ENMC sponsored meeting on congenital muscular dystrophy. Baarn, Holland, May 1993. Neuromuscular Disord. 1994;4:75-81. 131Tome FMS, Evengelista T, Leclerc A et al. Congenital muscular dystrophy with merosin deficiency. C R Acad Sci Paris Life Sci 1994:317:351-7. D, Wilson LA, Muntoni F. 141Sewry CA, Philpot J, Mahony Dubowitz V. Expression of laminin subunits in congenital muscular dystrophy. Neuromuscular Disord. 1995;5:307-16.

[5] Hillaire D, Leclerc A, Fame S et al. Localisation of merosin-negative congenital muscular dystrophy to chromosome 6q2 by homozygosity mapping. Hum. Mol. Genet. 1994;3:1657-61. [6] Helbling-Leclerc A, Topaloglu H, Tome FMS et al. Readjusting the localisation of merosin (laminin a,-chain) deficient congenital muscular dystrophy locus on chromosome 6q2. C R Acad Sci Paris Life Sci 1995;318:1245-52. [7] Sewry C, Philpot J, Sorokin LM, et al. Diagnosis of merosin (laminin alpha-2) deficient congenital muscular dystrophy by skin biopsy. Lancet 1996;347:582-4. [8] Schuler F, Sorokin LM. Expression of laminin isoforms in mouse myogenic cells in vitro and in vivo. J. Cell Sci. 1995;108:3795-805. [9] Mullenback R, Lagoda PJL, We Her C. An efficient salt-chloroform extraction of DNA from blood and tissues. Trends Genet. 1989;5:391. [IO] Helbling Leclerc A, Zhank X, Topaloglu H et al. Mutations in the laminin al-chain gene (LAMAZ) cause merosin-deficient congenital muscular dystrophy. Nature Genet. 1995;11:216-8. [l l] Naom IS. Mathew CG, Town M. Genetic mapping with microsatellites. In: Richwood D, Hames BD, editors. DNA cloning: complex genomes, second ed. London: Oxford University Press, 1995;195-217. [12] Naom I, D’Alessandro M, Topaloglu H et al. Refinement of the laminin az chain locus to chromosome 6q2 in severe and mild merosin deficient congenital muscular dystrophy. J. Med. Genet. 1997:in press. [13] Toda T, Segawa M, Nomura Y et al. Localisation of a gene for Fukuyama type congenital muscular dystrophy to chromosome 9q 31-33. Nature Genet 1993;5:28336. [14] van Engelen BGM. Leyten QH, Bernsen PLJA et al. Familial adult-onset muscular dystrophy with leukoencephalopathy. Ann. Neural. 1992;32:577780.

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[16] Villanova M, Malandrini A, Toti P et al. Localisation of merosin in the normal human brain: implications for congenital muscular dystrophy with merosin deficiency. J. Submicrosc. Cytol. Pathol. 1996;28:l--4.