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Limb-girdle myasthenia: clinical, electrophysiological and morphological features in familial and autoimmune cases
Neuromuscular Disorders 12 (2002) 964–969 www.elsevier.com/locate/nmd
Limb-girdle myasthenia: clinical, electrophysiological and morphological features in familial and autoimmune cases C. Rodolico*, A. Toscano, M. Autunno, S. Messina, C. Nicolosi, M. Aguennouz, M. Laura`, P. Girlanda, C. Messina, G. Vita Department of Neurosciences, Psychiatry and Anaesthesiology, University of Messina, Policlinico Universitario, Via C. Valeria 98125, Messina, Italy Received 7 February 2002; received in revised form 20 May 2002; accepted 6 June 2002
Abstract Limb-girdle myasthenia is an uncommon disease and includes familial and autoimmune forms. Patients present proximal muscle weakness and wasting, and sometimes fatigability, without cranial nerve involvement and fluctuations. We observed, during a 15-year period, nine subjects with limb-girdle myasthenia, (24–55 years; 8 males, 1 female) who constituted 3.2% of 281 myasthenic patients attending our department. All had previously received a diagnosis different from myasthenia. Diagnosis of limb-girdle myasthenia was established by clinical, muscle biopsy and electrophysiological assessment including repetitive nerve stimulation and single fiber electromyography. Five patients had the familial form with tubular aggregates in skeletal muscle; four patients had the autoimmune form. Patients with the familial form had a good response to acetylcholinesterase inhibitors, and the patients with the autoimmune form responded to immunotherapy. Our findings reinforce the opportunity to suspect limb-girdle myasthenia in unclassifiable proximal myopathies and to differentiate familial from autoimmune cases, especially for therapeutic implications. q 2002 Elsevier Science B.V. All rights reserved. Keywords: Myasthenia; Limb-girdle weakness; Tubular aggregates
1. Introduction The term ‘myasthenic myopathy’ was used, for the first time, by Walton and Nattrass in 1954, to define patients with proximal muscle weakness and wasting, and sometimes fatigability, who responded to acetylcholinesterase inhibitors (AChEI) [1]. Recently, this definition has been replaced by the term ‘limb-girdle myasthenia’ (LGM), including familial and autoimmune cases [2–8]. Familial LGM is a rare condition, inherited by autosomal recessive mechanism. Genetic alteration is still unknown but autoimmunity is not believed to play a pathogenic role. Clinically it is characterized by a progressive history of muscular fatigability with proximal muscle weakness and wasting, usually beginning in the first decade of life. Extraocular muscles are not involved. Serum creatine kinase (CK) level is often increased. There are not thymic alterations and antibodies against acetylcholine receptors (AChR-Ab) are negative. Repetitive nerve stimulation test (RNS) on proximal muscles shows a decremental response. Single-fiber electromyography (SFEMG) evidences a clear-cut neuromuscular transmission defect.
Muscle biopsy is characterized by the presence of tubular aggregates. Patients generally respond to AChEI. There is a debate regarding the existence of an autoimmune form of LGM. Oh and Kuruoglu [2] have described 12 cases with a pure limb-girdle muscle weakness and wasting as the unusual presentation of myasthenia gravis, constituting 3.8% of their myasthenic population during a 20-year investigation. The patients had a late onset, frequent association with thymoma, positivity of AChR-Ab, absence of tubular aggregates in skeletal muscle, poor response to AChEI but long benefit from immunotherapy. In order to further delineate such a disease we describe here the clinical, electrophysiological and morphological findings of nine LGM patients. 2. Patients and methods We studied nine patients (3.2% of 281 myasthenic patients who accessed our department during a 15-year period) affected by LGM. Age ranged from 23 to 55 years (8 M, 1 F). 2.1. Electrophysiological investigations
* Corresponding author. Tel.: 139-090-2212757; fax: 139-090-2212789. E-mail address: [email protected] (C. Rodolico).
A
0960-8966/02/$ - see front matter q 2002 Elsevier Science B.V. All rights reserved. PII: S09 60- 8966(02)0013 7-2
standard
concentric
needle
electromyography
C. Rodolico et al. / Neuromuscular Disorders 12 (2002) 964–969
(CNEMG) was performed in biceps brachii, first dorsal interosseous, quadriceps femoris and tibialis anterior muscles. Quantitative analysis of motor unit potentials was performed in right vastus lateralis muscle; the mean duration, mean amplitude and number of polyphasic potentials were determined, collecting 20 different motor unit potentials (MUPs) in four different needle insertions. All results were compared with normal age-adjusted data of our laboratory; we considered as abnormal a mean duration which differed ^20% from normal, a mean amplitude which deviated ^80% and a percentage of polyphasic potentials exceeding 15% [9]. SFEMG was performed on the right extensor digitorum communis (EDC) muscle, which is the muscle more commonly used for SFEMG study on neuromuscular transmission disturbances [10]. In each examination, 20 potential pairs were recorded. In each of 20 pairs, the mean value of consecutive differences (MCD) was determined and subsequently the mean MCD was calculated. The percentage of recordings with jitter exceeding 55 ms and the percentages of recordings with neuromuscular blocking were also evaluated. We considered the test abnormal when mean MCD was longer than 34 ms, or more than 10% of recordings had blocking or a jitter greater than 55 ms. RNS test was performed on a proximal muscle (deltoid or trapezius) using standard technique. We tested the variation of compound muscle action potential (CMAP) amplitude at low and high rate of stimulation; the test was considered positive for myasthenia if there was a reproducible decrement more than 10% between the first and the fifth response. Moreover 3 Hz RNS was repeated 3 min after 30 s of maximal effort. 2.2. Morphological investigations A vastus lateralis muscle biopsy was performed in each patient. For histochemical study, portions were frozen in isopentane cooled in liquid nitrogen and 10 mm serial cross-section were cut in a cryostat. Sections were stained
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with hematoxylin–eosin, modified trichrome stain, NADHtetrazolium reductase (NADH-TR), succinic dehydrogenase (SDH), cytocrome c oxidase, myofibrillar ATPase at pH 9.4, 4.6, 4.3, adenylate deaminase, alkaline and acid phosphatase, PAS, sudan black and crystal violet. Other segments were fixed in phosphate-buffered 2.5% glutaraldehyde, dehydrated and embedded in Durcupan, for ultrastructural examination.
3. Results Clinical findings are summarized in Table 1. Five patients (cases 1–5) were diagnosed having the familial form of LGM: cases 1 and 2 are brothers, sons of first cousins parents; the others are unrelated cases, each of them with consanguineous parents too. Age of disease onset ranged from 7 to 12 years, with a duration before diagnosis from 29 to 45 years. The major symptoms referred were proximal muscle wasting and weakness; wasting was particularly severe in patients with longer duration of symptomatology. Two patients also complained of muscular cramps and one patient had muscular fatigability. CK levels were normal or slightly increased. Signs of dysimmunity, such as increased AChR-Ab, thymoma or association with autoimmune disorders, were absent. CNEMG evidenced MUPs of low amplitude and short duration in four patients and was normal in one (Table 2). Patients who presented cramps did not show fibrillations or fasciculations. Baseline CMAP amplitudes were in the normal range but RNS revealed a decremental response in every case. SFEMG evidenced in all cases an impaired neuromuscular transmission with recordings with increased jitter, blocks, and increased mean jitter value. Muscle biopsy revealed the presence of tubular aggregates. Basophilic material, bright red with the Gomori trichrome stain, located especially in subsarcolemmal regions, had an intense reaction for NADH-TR and negative for SDH. Myofibrillar ATPase staining was negative in these areas
Table 1 Clinical and laboratory features a Case
Age (yrs)/sex Onset (yrs) Symptoms
CK b
AChR-Ab c
Thymoma
Treatment
1 2 3 4 5 6 7 8 9
36/M 55/M 52/M 52/M 38/M 24/M 43/M 32/M F/30
250 160 302 280 316 498 95 302 134
Neg. Neg. Neg. Neg. Neg. Neg. 8 mmol/l Neg. Neg.
No No No No No Yes No Yes Yes
AChEI AChEI AChEI AChEI AChEI Thymectomy, steroid, AChEI Thymectomy, IVIgG, AChEI Thymectomy, steroid, AChEI Thymectomy, steroid, AChEI
a b c
7 10 12 8 9 14 42 27 15
PWW PWW PWW, cramps PWW, fatigability PWLL, cramps PWW PWW PWW PWW, fatigability 1 Hashimotos’s thyroiditis
PWW, proximal wasting and weakness; PWLL, proximal weakness at lower limbs. n.v.: 25–200 IU/l. n.v.: ,0.8 nmol/l.
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Table 2 Electrophysiological and morphological findings a Case
1 2 3 4 5 6 7 8 9
CNEMG
SSMUP SSMUP Normal SSMUP SSMUP SSMUP SSMUP SSMUP SSMUP a
RNS
Decr. resp. Decr. resp. Decr. resp. Decr. resp. Decr. Resp. Decr. resp. Decr. resp. Decr. resp. Decr. resp.
SFEMG
Muscle biopsy
% Jitter "
% Recordings with blocks
MJV (MCD) (ms)
85 30 30 35 40 80 40 30 35
30 15 18 25 20 30 25 20 18
112 48.3 52 68 48.2 95 55.6 66 53
Tubular aggregates Tubular aggregates Tubular aggregates Tubular aggregates Tubular aggregates Increased connective, centralized nuclei, atrophic fibers Type 2b fiber atrophy Increased connective, centralized nuclei, atrophic fibers Type 2b fibers atrophy
SSMUP, small-short motor unit potentials; % Jitter " , percentage of recordings with abnormally increased jitter; MJV, mean jitter value.
and revealed a localization of these structures in a percentage of fibers ranging from 35 to 65%, predominantly in type 2 fibers. On ultrastructural examination, the abnormal areas contained tubules, located usually at the periphery of the muscle fibers, 60–80 nm in diameter, having a parallel, double-wall arrangement and packing in hexagonal arrays. No other significant ultrastructural alteration was present. All these patients had a good response to AChEI. In four patients the autoimmune form of LGM was diagnosed (cases 6–9, Table 1). The onset of the symptomatology ranged from 14 to 42 years and the duration of the disease, before the diagnosis, ranged from 1 to 15 years. Symptoms were represented by proximal muscle weakness and wasting. Only one patient also had muscular fatigability. CK was mildly increased in two cases. One patient had increased AChR-Ab levels (8 nmol/l). It was not possible to perform assays for other muscle autoantibodies. Thymoma was found in three of them. Histological features were consistent with a non-invasive lymphoepithelial thymoma. CNEMG showed a myogenic pattern (MUPs of low amplitude and short duration) in each patient (Table 2). RNS revealed a decremental response and SFEMG confirmed, in all cases, an impaired neuromuscular transmission with increased jitter, recording with blocks and increased mean jitter value. Muscle biopsy revealed type 2b fiber atrophy in two cases, whereas the other two patients, having increased CK levels, presented a ‘myopathic pattern’, with increased connective tissue, many atrophic fibers, and centralized nuclei. Isolated treatment with AChEI was ineffective. All four patients underwent thymectomy, but it did not modify the clinical course. They responded to a combined AChEI and steroid therapy. Case 7, with increased AChR-Ab levels, had a partial recovery after IgG e.v. infusion. Because of the diagnostic challenge, two illustrative cases are reported in detail. 3.1. Case 1 A 36-year-old man, born from consanguineous parents, complained of progressive weakness at arms and legs
muscles with difficulty in climbing stairs, since the age of 7 years. The 55-year-old brother (case 2) referred the same symptomatology, since his early childhood. Neurological examination evidenced waddling gait with positive Gowers’ manoeuvre, limb-girdle muscle weakness and wasting, and reduced deep tendon reflexes. Eyelid ptosis and ophthalmoparesis were absent. Serum CK level was slightly increased (250 IU/l). At CNEMG motor unit potentials (MUP) with low amplitude and short duration were recorded at four limbs. A provisional diagnosis of limb-girdle muscular dystrophy was made and the patient underwent vastus lateralis muscle biopsy. Fifty-five percent of muscle fibers evidenced the presence of basophilic subsarcolemmal material, bright red at Gomori thricrome stain, with an intense reactivity at NADH-TR (Fig. 1A). Ultrastructural examination confirmed the presence of tubular aggregates. (Fig. 1B). This finding led us to investigate neuromuscular junction with RNS test and SFEMG. RNS examination revealed a decremental response. SFEMG confirmed the abnormal function of neuromuscular junction: 85% of fibers with abnormal increased jitter, 30% of recordings with blocks, and a mean MCD of 112 ms, were recorded (Fig. 2). These features were consistent with the diagnosis of familial LGM. Treatment with AChEI was started with a consistent improvement of disturbances. 3.2. Case 6 A 24-year-old man came to our observation because of the onset of progressive proximal weakness and wasting at upper limbs since the age of 14 years. At the age of 20 years he started to notice also weakness at lower limbs. This symptomatology did not present fluctuations during the day. The patient did not complain of diplopia, dysphagia and other disturbances suggesting cranial nerves involvement. Neurological examination revealed bilateral deltoid and suprascapular muscles weakness, with a moderate wasting of the same muscles. Weakness of quadriceps muscles was also present. Deep tendon reflexes were absent at upper limbs. CK level was increased up to 498 IU/l. CNEMG
C. Rodolico et al. / Neuromuscular Disorders 12 (2002) 964–969
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Fig. 1. Muscle biopsy of Case 1. (A) Large amount of subsarcolemmal material with an intense reactivity to NADH-TR. Magnification £ 280. (B) Electron micrograph reveals that accumulated material consists of aggregates of sarcoplasmic reticulum-derived tubules, often arranged in hexagonal arrays. Magnification £ 100,000.
revealed a myogenic pattern (low amplitude and short duration of MUPs). A vastus lateralis muscle biopsy showed the presence of a great number of centralized nuclei, slightly increased connective tissue and some atrophic fibers. Immunocytochemical study for dystrophin, a, b, g, d sarcoglycans and caveolin 3 was normal. Western-blot for calpain-3 and dysferlin resulted normal. A provisional diagnosis of limb-girdle muscular dystrophy was made. In the following months, for the occurrence of persistent cough and recurrent fever, the patient was submitted to chest CT scan which evidenced the presence of a thymoma. Therefore he was readmitted to the hospital and investigated for myasthenia. RNS showed a decremental pattern, and SFEMG evidenced the presence of 80% of fibers with increased jitter, 30% of recordings with blocks and a mean MCD of 95 ms. Serum AChR-Ab dosage resulted negative. The patient was diagnosed having an autoimmune LGM; treatment with AChEI was started with poor results. The patient underwent thymoma excision; he did not improve, and 2 months later steroid therapy (75 mg/day of prednisone) was added to AChEI therapy. There was a progressive amelioration of clinical conditions and at 6 months follow-up the patient had a normal neurological examination.
4. Discussion Our observations confirm that, in agreement with previous reports, there is a population of myasthenic patients who present a persistent limb-girdle involvement, without cranial nerves impairment and with absence of the typical clinical fluctuations of symptomatology. For these cases, as recommended by other authors [2], the term limbgirdle myasthenia should be used. In our experience these patients constitute 3.2% of 281 myasthenic patients observed in a 15-year period. Five of our patients had the familial form with tubular aggregates, and four the autoimmune form. All the patients represented a diagnostic chal-
Fig. 2. SFEMG of Case 1: recordings with increased jitter and blocks of neuromuscular transmission.
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lenge. For them different diagnoses rather than myasthenia had been previously proposed, including polymyositis, limb-girdle muscle dystrophy, Lambert–Eaton syndrome, metabolic myopathy, and spinal muscular atrophy. The diagnostic difficulty for LGM is high and it is evident that, in consideration of therapeutic implications, the disease requires a strong index of suspicion. Familial LGM is a rare disorder inherited by autosomic recessive mechanism but genetic basis of the disease is still unknown [2–8]. All our patients with the familial form had a good response to AChEI. We stress here the concept that the presence of tubular aggregates in the muscle should suggest to the clinician the investigation of neuromuscular junction with RNS and SFEMG. Tubular aggregates are thought to represent an adaptive mechanism aiming at regulating an increased intracellular level of calcium, in order to prevent the muscle fibers from hypercontraction and necrosis. They are derived from the sarcoplasmic reticulum as shown by electron microscopy and by the use of anti-Ca 21 and sarcoplasmic reticulum antibodies [11]. Several patients with muscle pain, cramps and stiffness have been reported to have tubular aggregates in muscle biopsy [12–16]. Interestingly, cases 3 and 5 of our group referred cramps and had the most high percentage of fibers containing tubular aggregates. Oh and Kuruoglu [2] reported 12 cases with a pure limbgirdle muscle weakness as the unusual presentation of myasthenia gravis, constituting 3.8% of a myasthenic population during a 20-year investigation. One patient had hyperthyroidism, one had epilepsy and one was affected by systemic lupus erythematosus and chloroquine-precipitated myasthenia. Five out of seven patients had increased titres of Ab-AChR. Three out of 12 patients had thymic hyperplasia. In all patients there was AChEI response, although the degree of responsiveness was variable. In our study 4/9 patients with LGM presented similar findings. All of them had predominant weakness in the proximal muscles, in two in the upper extremities. Three of these patients were classified as mild (modified Osserman classification II A), and one (case 7) as II B. The onset of the disease was in the adult age. These patients constitute 1.4% of the myasthenic population observed in a 15-year period. Clear signs of autoimmunity were evident in all: 3/4 had thymoma and one was seropositive for Ab-AchR. One patient also had autoimmune hypothyroidism. Muscle biopsy revealed type 2b fiber atrophy in two patients. In two cases muscle abnormalities suggested a primitive muscular disorder. These findings are quite unusual in LGM, but it is reasonable that they are related with the long duration of the disease [2,17,18]. In our cases, combination of thymectomy, AChEI and steroid/IVIgG induced good improvement both on clinical ground and electrophysiological findings with a significant decrease of mean jitter value. No patient accepted to undergo a further muscle biopsy. In a 4-year follow-up all patients experienced a stabilization of their clinical status.
The reason why LGM patients have a selective impairment of limb-girdle muscles mimicking a myopathy is unknown. It is easily presumable that the pathogenesis of the two forms, familial and autoimmune, is different. A normal appearance of neuromuscular junctions with regularly arranged junctional folds and normal distribution of the acetylcholine receptors has been demonstrated for familial LGM [4]. Therefore, a functional alteration of neuromuscular transmission may be hypothesized, but in vitro electrophysiological studies of neuromuscular transmission are not available. Conversely, the pathogenetic mechanism leading to selective proximal muscle weakness and wasting in autoimmune LGM may be related to the presence of unknown autoantibodies or specific immunologic profile. In our study all patients showed abnormalities both of RNS and SFEMG. RNS was consistent with the diagnosis of myasthenia and, as ascertained by other authors, resulted to be useful for the differential diagnosis with Lambert–Eaton myasthenic syndrome, which may mimic LGM [19,20] and therefore can be suggested as the first choice electrophysiological test in LGM.
Acknowledgements We are grateful to Professor Amelia Evoli of the Institute of Neurology, Catholic University, Rome, Italy, for the AChR antibodies assays. This work was financially supported by grant from the Project on ‘Molecular, Cellular, Diagnostic and Epidemiological Analysis of Pediatric and Neurological Diseases’ (Cluster 04) of the Italian Ministry of Education, University and Research (MIUR).
References [1] Walton JM, Nattrass FJ. On the classification, natural history and treatment of the myopathies. Brain 1954;77:169–184. [2] Oh SJ, Kuruoglu R. Chronic limb-girdle myasthenia gravis. Neurology 1992;42:1153–1156. [3] Mc Quillen. Familial limb-girdle myasthenia. Brain 1966;89:121– 132. [4] Furui E, Fukushima K, Sakashita T, Matsubara SS, Takamori M. Familial limb-girdle myasthenia with tubular aggregates. Muscle Nerve 1997:599–603. [5] Azulay JPh, Pouget J, Figarella-Branger D, Colamarino R, Pellisser JF, Serratrice G. Faiblesse musculaire proximale isole`e re`ve`latrice d’un syndrome myasthe`nique. Rev Neurol 1994;150(5):377–381. [6] Vasant A, Taly AB, Sathynarayanaswamy H. Limb girdle myastenia: a study of familial and sporadic cases. J Assoc Physicians India 1994;42(8):601–603. [7] Sieb JP, Tolksdorf K, Dengler R, Jerusalem F. An autosomal-recessive congenital myasthenic syndrome with tubular aggregates in a Libyan family. Neuromuscul Disord 1996;6(2):115–119. [8] Morgan-Hughes JA, Lecky BR, Landon DN, Murray NM. Alterations in the number and affinity of junctional acetylcholine receptors in a myopathy with tubular aggregates. A newly recognized receptor defect. Brain 1981;104(2):279–295. [9] Buchthal F, Kamieniecka Z. The diagnostic yield of quantified elec-
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[10] [11]
[12]
[13]
[14]
tromyography and quantified muscle biopsy in neuromuscular disorders. Muscle Nerve 1982;5(4):265–280. Stalberg E, Hasson O. Single fibre EMG in juvenile myasthenia gravis. Neuropediatrie 1973;4(1):20–29. Salviati G, Pierobon-Bormioli S, Betto R, et al. Tubular aggregates: sarcoplasmatic reticulum origin, calcium storage ability, and functional implications. Muscle Nerve 1985;8:299–306. Vissing J, Schmalbruch H, Haller RG, Clausen T. Muscle phosphoglycerate mutase deficiency with tubular aggregates: effect of dantrolene. Ann Neurol 1999;46(2):274–277. Martin JJ, Ceuterick C, van Goethem G. On a dominantly inherited myopathy with tubular aggregates. Neuromuscul Disord 1997;7:512– 520. Danon MJ, Carpenter S, Harati Y. Muscle pain associated with tubular aggregates and structures resembling cylindrical spirals. Muscle Nerve 1989;12:265–272.
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[15] Martin JE, Mather K, Swash M, Gray AB. Expression of heat shock protein in tubular aggregates. Muscle Nerve 1991;14:219–225. [16] Lazaro RP, Fenichel GM, Kilroy AW, Saito A, Fleisher S. Cramps, muscle pain and tubular aggregates. Arch Neurol 1980;37:715– 717. [17] Engel AG. Myasthenic syndromes. In: Engel AG, FranziniArmstrong C, editors. Myology, 2nd ed. New York, NY: McGrawHill, 1994. pp. 1798–1835. [18] Husain F, Ryan NJ, Hogan GR. Concurrence of limb-girdle muscle dystrophy and myasthenia gravis. Arch Neurol 1989;46(1):101–102. [19] Oh SJ, Head T, Fesenmeier J, Claussen G. Peroneal nerve repetitive nerve stimulation test: its value in diagnosis of myasthenia gravis and Lambert–Eaton syndrome. Muscle Nerve 1995;18:867– 873. [20] Oh SJ. Diagnostic sensitivities of laboratory tests in myasthenia gravis. Muscle Nerve 1989;12:720–724.
Limb-girdle myasthenia: clinical, electrophysiological and morphological features in familial and autoimmune cases C. Rodolico*, A. Toscano, M. Autunno, S. Messina, C. Nicolosi, M. Aguennouz, M. Laura`, P. Girlanda, C. Messina, G. Vita Department of Neurosciences, Psychiatry and Anaesthesiology, University of Messina, Policlinico Universitario, Via C. Valeria 98125, Messina, Italy Received 7 February 2002; received in revised form 20 May 2002; accepted 6 June 2002
Abstract Limb-girdle myasthenia is an uncommon disease and includes familial and autoimmune forms. Patients present proximal muscle weakness and wasting, and sometimes fatigability, without cranial nerve involvement and fluctuations. We observed, during a 15-year period, nine subjects with limb-girdle myasthenia, (24–55 years; 8 males, 1 female) who constituted 3.2% of 281 myasthenic patients attending our department. All had previously received a diagnosis different from myasthenia. Diagnosis of limb-girdle myasthenia was established by clinical, muscle biopsy and electrophysiological assessment including repetitive nerve stimulation and single fiber electromyography. Five patients had the familial form with tubular aggregates in skeletal muscle; four patients had the autoimmune form. Patients with the familial form had a good response to acetylcholinesterase inhibitors, and the patients with the autoimmune form responded to immunotherapy. Our findings reinforce the opportunity to suspect limb-girdle myasthenia in unclassifiable proximal myopathies and to differentiate familial from autoimmune cases, especially for therapeutic implications. q 2002 Elsevier Science B.V. All rights reserved. Keywords: Myasthenia; Limb-girdle weakness; Tubular aggregates
1. Introduction The term ‘myasthenic myopathy’ was used, for the first time, by Walton and Nattrass in 1954, to define patients with proximal muscle weakness and wasting, and sometimes fatigability, who responded to acetylcholinesterase inhibitors (AChEI) [1]. Recently, this definition has been replaced by the term ‘limb-girdle myasthenia’ (LGM), including familial and autoimmune cases [2–8]. Familial LGM is a rare condition, inherited by autosomal recessive mechanism. Genetic alteration is still unknown but autoimmunity is not believed to play a pathogenic role. Clinically it is characterized by a progressive history of muscular fatigability with proximal muscle weakness and wasting, usually beginning in the first decade of life. Extraocular muscles are not involved. Serum creatine kinase (CK) level is often increased. There are not thymic alterations and antibodies against acetylcholine receptors (AChR-Ab) are negative. Repetitive nerve stimulation test (RNS) on proximal muscles shows a decremental response. Single-fiber electromyography (SFEMG) evidences a clear-cut neuromuscular transmission defect.
Muscle biopsy is characterized by the presence of tubular aggregates. Patients generally respond to AChEI. There is a debate regarding the existence of an autoimmune form of LGM. Oh and Kuruoglu [2] have described 12 cases with a pure limb-girdle muscle weakness and wasting as the unusual presentation of myasthenia gravis, constituting 3.8% of their myasthenic population during a 20-year investigation. The patients had a late onset, frequent association with thymoma, positivity of AChR-Ab, absence of tubular aggregates in skeletal muscle, poor response to AChEI but long benefit from immunotherapy. In order to further delineate such a disease we describe here the clinical, electrophysiological and morphological findings of nine LGM patients. 2. Patients and methods We studied nine patients (3.2% of 281 myasthenic patients who accessed our department during a 15-year period) affected by LGM. Age ranged from 23 to 55 years (8 M, 1 F). 2.1. Electrophysiological investigations
* Corresponding author. Tel.: 139-090-2212757; fax: 139-090-2212789. E-mail address: [email protected] (C. Rodolico).
A
0960-8966/02/$ - see front matter q 2002 Elsevier Science B.V. All rights reserved. PII: S09 60- 8966(02)0013 7-2
standard
concentric
needle
electromyography
C. Rodolico et al. / Neuromuscular Disorders 12 (2002) 964–969
(CNEMG) was performed in biceps brachii, first dorsal interosseous, quadriceps femoris and tibialis anterior muscles. Quantitative analysis of motor unit potentials was performed in right vastus lateralis muscle; the mean duration, mean amplitude and number of polyphasic potentials were determined, collecting 20 different motor unit potentials (MUPs) in four different needle insertions. All results were compared with normal age-adjusted data of our laboratory; we considered as abnormal a mean duration which differed ^20% from normal, a mean amplitude which deviated ^80% and a percentage of polyphasic potentials exceeding 15% [9]. SFEMG was performed on the right extensor digitorum communis (EDC) muscle, which is the muscle more commonly used for SFEMG study on neuromuscular transmission disturbances [10]. In each examination, 20 potential pairs were recorded. In each of 20 pairs, the mean value of consecutive differences (MCD) was determined and subsequently the mean MCD was calculated. The percentage of recordings with jitter exceeding 55 ms and the percentages of recordings with neuromuscular blocking were also evaluated. We considered the test abnormal when mean MCD was longer than 34 ms, or more than 10% of recordings had blocking or a jitter greater than 55 ms. RNS test was performed on a proximal muscle (deltoid or trapezius) using standard technique. We tested the variation of compound muscle action potential (CMAP) amplitude at low and high rate of stimulation; the test was considered positive for myasthenia if there was a reproducible decrement more than 10% between the first and the fifth response. Moreover 3 Hz RNS was repeated 3 min after 30 s of maximal effort. 2.2. Morphological investigations A vastus lateralis muscle biopsy was performed in each patient. For histochemical study, portions were frozen in isopentane cooled in liquid nitrogen and 10 mm serial cross-section were cut in a cryostat. Sections were stained
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with hematoxylin–eosin, modified trichrome stain, NADHtetrazolium reductase (NADH-TR), succinic dehydrogenase (SDH), cytocrome c oxidase, myofibrillar ATPase at pH 9.4, 4.6, 4.3, adenylate deaminase, alkaline and acid phosphatase, PAS, sudan black and crystal violet. Other segments were fixed in phosphate-buffered 2.5% glutaraldehyde, dehydrated and embedded in Durcupan, for ultrastructural examination.
3. Results Clinical findings are summarized in Table 1. Five patients (cases 1–5) were diagnosed having the familial form of LGM: cases 1 and 2 are brothers, sons of first cousins parents; the others are unrelated cases, each of them with consanguineous parents too. Age of disease onset ranged from 7 to 12 years, with a duration before diagnosis from 29 to 45 years. The major symptoms referred were proximal muscle wasting and weakness; wasting was particularly severe in patients with longer duration of symptomatology. Two patients also complained of muscular cramps and one patient had muscular fatigability. CK levels were normal or slightly increased. Signs of dysimmunity, such as increased AChR-Ab, thymoma or association with autoimmune disorders, were absent. CNEMG evidenced MUPs of low amplitude and short duration in four patients and was normal in one (Table 2). Patients who presented cramps did not show fibrillations or fasciculations. Baseline CMAP amplitudes were in the normal range but RNS revealed a decremental response in every case. SFEMG evidenced in all cases an impaired neuromuscular transmission with recordings with increased jitter, blocks, and increased mean jitter value. Muscle biopsy revealed the presence of tubular aggregates. Basophilic material, bright red with the Gomori trichrome stain, located especially in subsarcolemmal regions, had an intense reaction for NADH-TR and negative for SDH. Myofibrillar ATPase staining was negative in these areas
Table 1 Clinical and laboratory features a Case
Age (yrs)/sex Onset (yrs) Symptoms
CK b
AChR-Ab c
Thymoma
Treatment
1 2 3 4 5 6 7 8 9
36/M 55/M 52/M 52/M 38/M 24/M 43/M 32/M F/30
250 160 302 280 316 498 95 302 134
Neg. Neg. Neg. Neg. Neg. Neg. 8 mmol/l Neg. Neg.
No No No No No Yes No Yes Yes
AChEI AChEI AChEI AChEI AChEI Thymectomy, steroid, AChEI Thymectomy, IVIgG, AChEI Thymectomy, steroid, AChEI Thymectomy, steroid, AChEI
a b c
7 10 12 8 9 14 42 27 15
PWW PWW PWW, cramps PWW, fatigability PWLL, cramps PWW PWW PWW PWW, fatigability 1 Hashimotos’s thyroiditis
PWW, proximal wasting and weakness; PWLL, proximal weakness at lower limbs. n.v.: 25–200 IU/l. n.v.: ,0.8 nmol/l.
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Table 2 Electrophysiological and morphological findings a Case
1 2 3 4 5 6 7 8 9
CNEMG
SSMUP SSMUP Normal SSMUP SSMUP SSMUP SSMUP SSMUP SSMUP a
RNS
Decr. resp. Decr. resp. Decr. resp. Decr. resp. Decr. Resp. Decr. resp. Decr. resp. Decr. resp. Decr. resp.
SFEMG
Muscle biopsy
% Jitter "
% Recordings with blocks
MJV (MCD) (ms)
85 30 30 35 40 80 40 30 35
30 15 18 25 20 30 25 20 18
112 48.3 52 68 48.2 95 55.6 66 53
Tubular aggregates Tubular aggregates Tubular aggregates Tubular aggregates Tubular aggregates Increased connective, centralized nuclei, atrophic fibers Type 2b fiber atrophy Increased connective, centralized nuclei, atrophic fibers Type 2b fibers atrophy
SSMUP, small-short motor unit potentials; % Jitter " , percentage of recordings with abnormally increased jitter; MJV, mean jitter value.
and revealed a localization of these structures in a percentage of fibers ranging from 35 to 65%, predominantly in type 2 fibers. On ultrastructural examination, the abnormal areas contained tubules, located usually at the periphery of the muscle fibers, 60–80 nm in diameter, having a parallel, double-wall arrangement and packing in hexagonal arrays. No other significant ultrastructural alteration was present. All these patients had a good response to AChEI. In four patients the autoimmune form of LGM was diagnosed (cases 6–9, Table 1). The onset of the symptomatology ranged from 14 to 42 years and the duration of the disease, before the diagnosis, ranged from 1 to 15 years. Symptoms were represented by proximal muscle weakness and wasting. Only one patient also had muscular fatigability. CK was mildly increased in two cases. One patient had increased AChR-Ab levels (8 nmol/l). It was not possible to perform assays for other muscle autoantibodies. Thymoma was found in three of them. Histological features were consistent with a non-invasive lymphoepithelial thymoma. CNEMG showed a myogenic pattern (MUPs of low amplitude and short duration) in each patient (Table 2). RNS revealed a decremental response and SFEMG confirmed, in all cases, an impaired neuromuscular transmission with increased jitter, recording with blocks and increased mean jitter value. Muscle biopsy revealed type 2b fiber atrophy in two cases, whereas the other two patients, having increased CK levels, presented a ‘myopathic pattern’, with increased connective tissue, many atrophic fibers, and centralized nuclei. Isolated treatment with AChEI was ineffective. All four patients underwent thymectomy, but it did not modify the clinical course. They responded to a combined AChEI and steroid therapy. Case 7, with increased AChR-Ab levels, had a partial recovery after IgG e.v. infusion. Because of the diagnostic challenge, two illustrative cases are reported in detail. 3.1. Case 1 A 36-year-old man, born from consanguineous parents, complained of progressive weakness at arms and legs
muscles with difficulty in climbing stairs, since the age of 7 years. The 55-year-old brother (case 2) referred the same symptomatology, since his early childhood. Neurological examination evidenced waddling gait with positive Gowers’ manoeuvre, limb-girdle muscle weakness and wasting, and reduced deep tendon reflexes. Eyelid ptosis and ophthalmoparesis were absent. Serum CK level was slightly increased (250 IU/l). At CNEMG motor unit potentials (MUP) with low amplitude and short duration were recorded at four limbs. A provisional diagnosis of limb-girdle muscular dystrophy was made and the patient underwent vastus lateralis muscle biopsy. Fifty-five percent of muscle fibers evidenced the presence of basophilic subsarcolemmal material, bright red at Gomori thricrome stain, with an intense reactivity at NADH-TR (Fig. 1A). Ultrastructural examination confirmed the presence of tubular aggregates. (Fig. 1B). This finding led us to investigate neuromuscular junction with RNS test and SFEMG. RNS examination revealed a decremental response. SFEMG confirmed the abnormal function of neuromuscular junction: 85% of fibers with abnormal increased jitter, 30% of recordings with blocks, and a mean MCD of 112 ms, were recorded (Fig. 2). These features were consistent with the diagnosis of familial LGM. Treatment with AChEI was started with a consistent improvement of disturbances. 3.2. Case 6 A 24-year-old man came to our observation because of the onset of progressive proximal weakness and wasting at upper limbs since the age of 14 years. At the age of 20 years he started to notice also weakness at lower limbs. This symptomatology did not present fluctuations during the day. The patient did not complain of diplopia, dysphagia and other disturbances suggesting cranial nerves involvement. Neurological examination revealed bilateral deltoid and suprascapular muscles weakness, with a moderate wasting of the same muscles. Weakness of quadriceps muscles was also present. Deep tendon reflexes were absent at upper limbs. CK level was increased up to 498 IU/l. CNEMG
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Fig. 1. Muscle biopsy of Case 1. (A) Large amount of subsarcolemmal material with an intense reactivity to NADH-TR. Magnification £ 280. (B) Electron micrograph reveals that accumulated material consists of aggregates of sarcoplasmic reticulum-derived tubules, often arranged in hexagonal arrays. Magnification £ 100,000.
revealed a myogenic pattern (low amplitude and short duration of MUPs). A vastus lateralis muscle biopsy showed the presence of a great number of centralized nuclei, slightly increased connective tissue and some atrophic fibers. Immunocytochemical study for dystrophin, a, b, g, d sarcoglycans and caveolin 3 was normal. Western-blot for calpain-3 and dysferlin resulted normal. A provisional diagnosis of limb-girdle muscular dystrophy was made. In the following months, for the occurrence of persistent cough and recurrent fever, the patient was submitted to chest CT scan which evidenced the presence of a thymoma. Therefore he was readmitted to the hospital and investigated for myasthenia. RNS showed a decremental pattern, and SFEMG evidenced the presence of 80% of fibers with increased jitter, 30% of recordings with blocks and a mean MCD of 95 ms. Serum AChR-Ab dosage resulted negative. The patient was diagnosed having an autoimmune LGM; treatment with AChEI was started with poor results. The patient underwent thymoma excision; he did not improve, and 2 months later steroid therapy (75 mg/day of prednisone) was added to AChEI therapy. There was a progressive amelioration of clinical conditions and at 6 months follow-up the patient had a normal neurological examination.
4. Discussion Our observations confirm that, in agreement with previous reports, there is a population of myasthenic patients who present a persistent limb-girdle involvement, without cranial nerves impairment and with absence of the typical clinical fluctuations of symptomatology. For these cases, as recommended by other authors [2], the term limbgirdle myasthenia should be used. In our experience these patients constitute 3.2% of 281 myasthenic patients observed in a 15-year period. Five of our patients had the familial form with tubular aggregates, and four the autoimmune form. All the patients represented a diagnostic chal-
Fig. 2. SFEMG of Case 1: recordings with increased jitter and blocks of neuromuscular transmission.
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lenge. For them different diagnoses rather than myasthenia had been previously proposed, including polymyositis, limb-girdle muscle dystrophy, Lambert–Eaton syndrome, metabolic myopathy, and spinal muscular atrophy. The diagnostic difficulty for LGM is high and it is evident that, in consideration of therapeutic implications, the disease requires a strong index of suspicion. Familial LGM is a rare disorder inherited by autosomic recessive mechanism but genetic basis of the disease is still unknown [2–8]. All our patients with the familial form had a good response to AChEI. We stress here the concept that the presence of tubular aggregates in the muscle should suggest to the clinician the investigation of neuromuscular junction with RNS and SFEMG. Tubular aggregates are thought to represent an adaptive mechanism aiming at regulating an increased intracellular level of calcium, in order to prevent the muscle fibers from hypercontraction and necrosis. They are derived from the sarcoplasmic reticulum as shown by electron microscopy and by the use of anti-Ca 21 and sarcoplasmic reticulum antibodies [11]. Several patients with muscle pain, cramps and stiffness have been reported to have tubular aggregates in muscle biopsy [12–16]. Interestingly, cases 3 and 5 of our group referred cramps and had the most high percentage of fibers containing tubular aggregates. Oh and Kuruoglu [2] reported 12 cases with a pure limbgirdle muscle weakness as the unusual presentation of myasthenia gravis, constituting 3.8% of a myasthenic population during a 20-year investigation. One patient had hyperthyroidism, one had epilepsy and one was affected by systemic lupus erythematosus and chloroquine-precipitated myasthenia. Five out of seven patients had increased titres of Ab-AChR. Three out of 12 patients had thymic hyperplasia. In all patients there was AChEI response, although the degree of responsiveness was variable. In our study 4/9 patients with LGM presented similar findings. All of them had predominant weakness in the proximal muscles, in two in the upper extremities. Three of these patients were classified as mild (modified Osserman classification II A), and one (case 7) as II B. The onset of the disease was in the adult age. These patients constitute 1.4% of the myasthenic population observed in a 15-year period. Clear signs of autoimmunity were evident in all: 3/4 had thymoma and one was seropositive for Ab-AchR. One patient also had autoimmune hypothyroidism. Muscle biopsy revealed type 2b fiber atrophy in two patients. In two cases muscle abnormalities suggested a primitive muscular disorder. These findings are quite unusual in LGM, but it is reasonable that they are related with the long duration of the disease [2,17,18]. In our cases, combination of thymectomy, AChEI and steroid/IVIgG induced good improvement both on clinical ground and electrophysiological findings with a significant decrease of mean jitter value. No patient accepted to undergo a further muscle biopsy. In a 4-year follow-up all patients experienced a stabilization of their clinical status.
The reason why LGM patients have a selective impairment of limb-girdle muscles mimicking a myopathy is unknown. It is easily presumable that the pathogenesis of the two forms, familial and autoimmune, is different. A normal appearance of neuromuscular junctions with regularly arranged junctional folds and normal distribution of the acetylcholine receptors has been demonstrated for familial LGM [4]. Therefore, a functional alteration of neuromuscular transmission may be hypothesized, but in vitro electrophysiological studies of neuromuscular transmission are not available. Conversely, the pathogenetic mechanism leading to selective proximal muscle weakness and wasting in autoimmune LGM may be related to the presence of unknown autoantibodies or specific immunologic profile. In our study all patients showed abnormalities both of RNS and SFEMG. RNS was consistent with the diagnosis of myasthenia and, as ascertained by other authors, resulted to be useful for the differential diagnosis with Lambert–Eaton myasthenic syndrome, which may mimic LGM [19,20] and therefore can be suggested as the first choice electrophysiological test in LGM.
Acknowledgements We are grateful to Professor Amelia Evoli of the Institute of Neurology, Catholic University, Rome, Italy, for the AChR antibodies assays. This work was financially supported by grant from the Project on ‘Molecular, Cellular, Diagnostic and Epidemiological Analysis of Pediatric and Neurological Diseases’ (Cluster 04) of the Italian Ministry of Education, University and Research (MIUR).
References [1] Walton JM, Nattrass FJ. On the classification, natural history and treatment of the myopathies. Brain 1954;77:169–184. [2] Oh SJ, Kuruoglu R. Chronic limb-girdle myasthenia gravis. Neurology 1992;42:1153–1156. [3] Mc Quillen. Familial limb-girdle myasthenia. Brain 1966;89:121– 132. [4] Furui E, Fukushima K, Sakashita T, Matsubara SS, Takamori M. Familial limb-girdle myasthenia with tubular aggregates. Muscle Nerve 1997:599–603. [5] Azulay JPh, Pouget J, Figarella-Branger D, Colamarino R, Pellisser JF, Serratrice G. Faiblesse musculaire proximale isole`e re`ve`latrice d’un syndrome myasthe`nique. Rev Neurol 1994;150(5):377–381. [6] Vasant A, Taly AB, Sathynarayanaswamy H. Limb girdle myastenia: a study of familial and sporadic cases. J Assoc Physicians India 1994;42(8):601–603. [7] Sieb JP, Tolksdorf K, Dengler R, Jerusalem F. An autosomal-recessive congenital myasthenic syndrome with tubular aggregates in a Libyan family. Neuromuscul Disord 1996;6(2):115–119. [8] Morgan-Hughes JA, Lecky BR, Landon DN, Murray NM. Alterations in the number and affinity of junctional acetylcholine receptors in a myopathy with tubular aggregates. A newly recognized receptor defect. Brain 1981;104(2):279–295. [9] Buchthal F, Kamieniecka Z. The diagnostic yield of quantified elec-
C. Rodolico et al. / Neuromuscular Disorders 12 (2002) 964–969
[10] [11]
[12]
[13]
[14]
tromyography and quantified muscle biopsy in neuromuscular disorders. Muscle Nerve 1982;5(4):265–280. Stalberg E, Hasson O. Single fibre EMG in juvenile myasthenia gravis. Neuropediatrie 1973;4(1):20–29. Salviati G, Pierobon-Bormioli S, Betto R, et al. Tubular aggregates: sarcoplasmatic reticulum origin, calcium storage ability, and functional implications. Muscle Nerve 1985;8:299–306. Vissing J, Schmalbruch H, Haller RG, Clausen T. Muscle phosphoglycerate mutase deficiency with tubular aggregates: effect of dantrolene. Ann Neurol 1999;46(2):274–277. Martin JJ, Ceuterick C, van Goethem G. On a dominantly inherited myopathy with tubular aggregates. Neuromuscul Disord 1997;7:512– 520. Danon MJ, Carpenter S, Harati Y. Muscle pain associated with tubular aggregates and structures resembling cylindrical spirals. Muscle Nerve 1989;12:265–272.
969
[15] Martin JE, Mather K, Swash M, Gray AB. Expression of heat shock protein in tubular aggregates. Muscle Nerve 1991;14:219–225. [16] Lazaro RP, Fenichel GM, Kilroy AW, Saito A, Fleisher S. Cramps, muscle pain and tubular aggregates. Arch Neurol 1980;37:715– 717. [17] Engel AG. Myasthenic syndromes. In: Engel AG, FranziniArmstrong C, editors. Myology, 2nd ed. New York, NY: McGrawHill, 1994. pp. 1798–1835. [18] Husain F, Ryan NJ, Hogan GR. Concurrence of limb-girdle muscle dystrophy and myasthenia gravis. Arch Neurol 1989;46(1):101–102. [19] Oh SJ, Head T, Fesenmeier J, Claussen G. Peroneal nerve repetitive nerve stimulation test: its value in diagnosis of myasthenia gravis and Lambert–Eaton syndrome. Muscle Nerve 1995;18:867– 873. [20] Oh SJ. Diagnostic sensitivities of laboratory tests in myasthenia gravis. Muscle Nerve 1989;12:720–724.