Skip Navigation



Rheumatology Advance Access published online on August 7, 2008
Rheumatology, doi:10.1093/rheumatology/ken325
This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
47/10/1539    most recent
ken325v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Suzuki, S.
Right arrow Articles by Kuwana, M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Suzuki, S.
Right arrow Articles by Kuwana, M.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© The Author 2008. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org

Clinical utility of anti-signal recognition particle antibody in the differential diagnosis of myopathies

S. Suzuki1, T. Satoh2, S. Sato2, M. Otomo3, Y. Hirayama4, H. Sato1, M. Kawai3, T. Ishihara5, N. Suzuki1 and M. Kuwana2

1Department of Neurology 2Division of Rheumatology, Department of Internal Medicine, Keio University School of Medicine, Tokyo 3Department of Neurology, Higashisaitama National Hospital, Saitama 4Department of Pediatric Neurology, Tokyo Metropolitan Higashiyamato Medical Center for the Severely Disabled, Tokyo and 5Department of Neurology, Hakone National Hospital, Kanagawa, Japan.

Correspondence to: S. Suzuki, Department of Neurology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan. E-mail: shigeaki{at}sc.itc.keio.ac.jp


   Abstract
Top
 Abstract
Introduction
 Patients, materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Objective. Auto-antibodies to signal recognition particle (SRP) are known to be specific to PM among rheumatic disorders, but the specificity in myopathic diseases remains unclear. The clinical utility of anti-SRP antibody in the differential diagnosis of myopathies has not been studied. The aim of the present study was to elucidate whether detection of anti-SRP antibody can discriminate of PM from muscular dystrophy (MD).

Methods. We report a patient with a childhood onset myopathy, in whom it was clinically difficult to make a differential diagnosis of PM or MD for 21 yrs, despite repeated muscle biopsies. Myositis-specific auto-antibodies to RNA-associated antigens were screened in this particular case as well as in 105 serum samples from various types of MD and 84 from PM patients using RNA immunoprecipitation. The MD and PM serum samples were obtained from different institutions. The presence of anti-SRP antibody was confirmed by RNA immunoprecipitation combined with immunodepletion of SRP from the antigen.

Results. Anti-SRP antibody was positive in the present patient, supporting the diagnosis of PM. Anti-SRP antibody was detected in seven (8.3%) patients with PM, but in none of the patients with MD. Myositis-specific auto-antibodies were not detected in any of the patients with MD.

Conclusion. Anti-SRP antibody is useful for discriminating PM from MD among patients with myopathies.

KEY WORDS: Anti-signal recognition particle antibody, Polymyositis, Muscular dystrophy


   Introduction
Top
 Abstract
 Introduction
Patients, materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
The inflammatory myopathies are a heterogeneous group of systemic diseases characterized by muscle weakness, elevated serum creatine kinase (CK) values, electromyographic abnormalities and inflammatory infiltrates in skeletal muscle [1]. Myositis-specific auto-antibodies include those directed against aminoacyl-tRNA synthetases (ARS), signal recognition particle (SRP) and nuclear helicase Mi-2. Anti-Jo-1 antibody, one of the anti-ARS antibodies, is closely related to PM and DM with a high frequency of intestinal lung disease (ILD) [2]. On the other hand, anti-SRP antibody is clinically associated with pure PM [3–6]. SRP, one of the most abundant and best characterized RNP particles, regulates the translocation of proteins across the endoplasmic reticulum during protein synthesis [3, 4]. Patients with anti-SRP antibodies most often present with severe muscle involvement characterized by rapidly developing proximal weakness that culminates in severe disability, and often by a poor response to steroid therapy [6].

Muscular dystrophy (MD), a group of hereditary and sporadic progressive diseases, each with unique phenotypic and genetic features, is the most common and representative myopathy [7]. It is sometimes difficult to distinguish anti-SRP-positive PM patients from MD patients for the following reasons [8]. First, since anti-SRP-positive PM patients have a low incidence of pulmonary fibrosis, skin rash, arthritis and RP, it is difficult to differentiate them from MD patients based on the clinical manifestations alone. Second, severe necrosis of skeletal muscle without lymphocytic infiltration is a histological finding common to both anti-SRP-positive PM and MD [9, 10]. In similar clinical settings, it has not been elucidated whether the detection of auto-antibodies using RNA immunoprecipitation is reliable for the differential diagnosis of myopathies, especially between MD and PM. To address this question, we screened the serum of MD patients and PM patients for myositis-specific auto-antibodies, including anti-SRP antibody.


   Patients, materials and methods
Top
 Abstract
 Introduction
 Patients, materials and methods
Results
 Discussion
 Acknowledgements
 References
 
Case report
A 32-yr-old Japanese man was admitted to Keio University Hospital in 2004 for evaluation of long-standing myopathy. He had a 21-yr history of severe weakness in the trunk, arms and legs, but no familial history of neuromuscular disorders. He was normal until the age of 10 yrs of age, when he had difficulty in running fast and hanging from a horizontal bar. When he first visited the other hospital at age 11 yrs, he showed scapulohumeral dominant muscle atrophy without facial muscle involvement. Serum CK was 4180 IU/l (normal, <198 IU/l), and the electromyography showed myopathic features. The first muscle biopsy of the left biceps brachii muscle revealed a prominent variation in muscle fibre size, but no perifascicular atrophy. Necrotic muscle fibres were observed with evidence of regeneration, but there was no lymphocytic infiltration in the perimysial or perivascular region. Endomysial connective tissue was increased (Fig. 1A and B). Under the tentative diagnosis of scapulohumeral MD, oral predonisolone (1 mg/kg/day) therapy was prescribed for 3 months, but the patient did not respond to the treatment. His weakness worsened over the next 2 yrs, and he eventually lost ambulation and had difficulty in blowing, swallowing and eating at the age of 13 yrs. After the bulbar symptoms continued for ~6 months, his symptoms gradually began to improve. However, he could not walk and required a wheelchair. The serum CK levels decreased to 1640 IU/l at the age of 14 yrs and 614 IU/l at the age of 16 yrs. The second muscle biopsy of the quadriceps femoris muscle was performed at 16 yrs of age. The muscle fibres had largely been replaced by adipose tissue, but there still was no lymphocytic infiltration (Fig. 1C). There have been no changes in his muscle weakness since then.


Figure 1
View larger version (102K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 1. Histological findings of the present patient. The histological appearance of muscle biopsy specimens obtained at 11 yrs of age (A and B) and 16 yrs of age (C). Haematoxylin–eosin staining. Bar = 100 µm.

 
The neurological examination on admission revealed severe symmetrical proximal-dominant weakness, with Medical Research Council scale grade at 3/5 in the lower extremities and 4/5 in the upper extremities, but there was no facial muscle involvement. Muscle atrophy was remarkable in the lower trunk and proximal muscles of the legs. Deep tendon reflex was absent. The CK levels had retuned to the normal range. T1-weighted muscle magnetic resonance images of the thighs showed evidence of severe diffuse muscle atrophy.

Patients and sera
Serum samples were obtained from the patient described above, and from 105 Japanese patients with MD (82 males and 23 females) who were seen consecutively between January 2005 and June 2006 at Higashisaitama National Hospital. The mean age of the MD patients at the time of serum collection was 30.3 ± 17.2 yrs. The MD cases included those with Duchenne MD (n = 58), Becker MD (n = 6), myotonic MD (n = 19), limb-girdle MD (n = 6), facioscapulohumeral MD (n = 4), Fukuyama-type congenital MD (n = 7) and unclassified types of MD (n = 5), and the diagnosis of each was based on clinical, pathological and genetic features [7]. In contrast, stored serum for a cohort of 84 Japanese patients with PM (25 males and 59 females) was available at Keio University Hospital. All sera were obtained at diagnosis before initiation of immunosuppresive therapy. The mean age of the PM patients at the time of serum collection was 51.4 ± 13.4 yrs. The diagnosis of probable or definite PM was based on the criteria of Bohan and Peter [11]. Patients with DM, IBM and overlap syndrome were excluded from study entry. We obtained MD and PM serum samples from different institutions because most MD patients, especially those with Duchenne MD, stayed at the national hospital and sanatorium for the severely disabled. All blood samples and clinical information were obtained after patients provided informed consent and ethical approval. The study was approved by the institutional review boards.

RNA immunoprecipitaion
RNA immunoprecipitation assay was performed using extracts from leukaemia cell line K562, as previously described [12]. Briefly, a 10 µl volume of patient serum was mixed with 2 mg of protein A-Sepharose CL-4B (Pharmacia Biotech AB, Uppsala, Sweden) in 500 µl of immunoprecipitation buffer (10 mM Tris–HCl, pH 8.0, 500 mM NaCl, 0.1% Nonidet P40), and after incubation for 2 h, was washed three times with immunoprecipitation buffer. Antibody-bound Sepharose beads were mixed with 100 µl of K562 cell extract (6 x 106 cell equivalents per sample) for 2 h, and 30 µl of 3 M sodium acetate, 30 µl of 10% SDS and 300 µl of phenol:chloroform:isoamyl alcohol (50:50:1, containing 0.1% 8-hydroxyquinoline) were added to extract bound RNA. After ethanol precipitation, the RNA was resolved on a 7 M urea–10% polyacrylamide gel, and the gel was silver-stained (Bio-Rad, Hercules, CA, USA).

Immunodepletion experiments
The immunodepletion studies were undertaken using prototype sera obtained from two anti-SRP-positive PM patients (patients #1 and #2). A 50 µl volume of the prototype serum #2 and normal serum as a negative control was mixed with 10 mg of protein A-Sepharose CL-4B in 500 µl of immunoprecipitation buffer, and incubated for 2 h. After washing three times with immunoprecipitation buffer, the antibody-bound Sepharose beads were mixed with 100 µl of K562 cell extract for 2 h to fully the deplete antigens recognized by prototype serum #2 or normal serum. Then, the supernatant was further incubated with Sepharose beads pre-conjugated with the present patient serum and the prototype serum #1. After washing for five times, the RNA immunoprecipitation assay was analysed as described above.

Statistic analysis
The frequencies of MD patients and PM patients who were positive for each auto-antibody were compared. Categorical variables were compared by the {chi}2-test.


   Results
Top
 Abstract
 Introduction
 Patients, materials and methods
 Results
Discussion
 Acknowledgements
 References
 
Our patient's serum immunoprecipitated RNA located in the 7S RNA lesion, as did the prototype serum #1 obtained from an anti-SRP-positive PM patient (Fig. 2, lanes 2 and 5). Immunodepletion treatment with the other prototype #2 serum obtained from an anti-SRP-positive PM patient successfully depleted the SRP-antigen complex from the extract (lane 3), but normal serum did not (lane 4). Immunoprecipitation of 7S RNA in our patient was cancelled when SRP-antigen depleted extract was used (lane 6). These findings confirmed that our patient's serum was positive for anti-SRP antibody.


Figure 2
View larger version (33K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 2. Analysis of immunoprecipitates from K562 cell extracts by 7 M urea–10% PAGE and silver staining. The results of the prototype serum #1 of the anti-signal recognition particle (SRP)-positive PM patient (lanes 2–4) and the present patient serum (lanes 5–7) are compared. Immunoprecipitates were obtained from no treatment (lanes 2 and 5), after immunodepletion using prototype serum #2 from an anti-SRP-positive patient (lanes 3 and 6), and serum from a healthy control (lanes 4 and 7).

 
We screened auto-antibodies to RNA-associated auto-antigens in sera from 105 MD patients and 84 PM patients by the RNA immunoprecipitation assay. The frequencies of myositis-specific auto-antibodies detected by RNA immunoprecipitation in MD patients and PM patients are summarized in Table 1. Anti-SRP antibodies were detected in 7 PM patients (8.3%), but in none of the MD patients (P = 0.008). Similarly, auto-antibodies to ARS, such as those to Jo-1, PL-7, PL-12, EJ and OJ, were detected in PM patients, but not in MD patients. Myositis-specific auto-antigens were not detected in MD patients.


View this table:
[in this window]
[in a new window]

  		TABLE 1. Frequencies of myositis-specific auto-antibody detection by RNA immunoprecipitation in patients with MD and those with PM

 
Table 2 shows the clinical and histological findings of the present case and for seven PM patients with anti-SRP antibodies. The eight PM patients with anti-SRP antibodies had common clinical features including muscle weakness, levels of serum CK > 3000 IU/l (except for one case) and myopathic findings in the EMG. Although three patients had ILD, none had skin rash, arthritis or RP. Malignancy was found in two patients. Response to high-dose steroid therapy was generally poor. Most patients showed refractory muscle weakness and required other immunosuppressive agents. Histological findings disclosed severe necrosis with regeneration of skeletal muscle in all patients, and lymphocytic infiltration in half.


View this table:
[in this window]
[in a new window]

  		TABLE 2. Clinical and histological findings from PM patients with anti-SRP antibodies

 

   Discussion
Top
 Abstract
 Introduction
 Patients, materials and methods
 Results
 Discussion
Acknowledgements
 References
 
In this article, we describe a case of myopathy with childhood onset, in whom it was difficult to make a differential diagnosis between PM and MD despite repeated muscle biopsy. The patient was found to be positive for anti-SRP antibody 21 yrs after onset. Screening the serum of MD patients and PM patients by RNA immunoprecipitation assay for auto-antibodies revealed anti-SRP antibodies in seven (8.3%) PM patients, but not in MD patients. However, the comparison of frequencies of these auto-antibodies may not be appropriate, since the MD and PM patients were selected from medical centres with different settings. Anti-SRP antibody was found to be specific to PM and useful in excluding MD in patients with myopathies.

Anti-SRP antibody was detected in 8.3% in our cohort of Japanese PM patients, similar to the rate in European patients [13]. Although anti-SRP antibody is known to be PM-specific, it has also been found in patients with DM, IBM and SSc [6, 13]. However, the specificity of anti-SRP antibody in myopathic diseases is not fully elucidated. Perurena et al. [14] and Hengstman et al. [15] reported that anti-SRP antibody was negative in 17 and 48 patients with MD, respectively. Combining these results with our findings, anti-SRP antibody is not exclusively present in a total of 170 patients with MD. These findings suggest that production of anti-SRP antibody does not result from non-specific muscle injuries.

Although muscle biopsy is the most accurate method of establishing a diagnosis of PM, but its interpretation is sometimes difficult [1]. van der Meulen et al. [16] have reported that PM is an overdiagnosed entity using strict diagnostic criteria of muscle biopsy. To avoid misdiagnosis of PM, Dalakas and Hohlfeld [1] have advocated the use of new histological criteria based on CD8+ and MHC-I immunohistochemistry. Examination of the muscle biopsy specimens obtained from anti-SRP-positive patients have usually shown active necrosis and little or no inflammation [9, 10], and the same histological findings were observed in our patient. Based on the histological findings alone, anti-SRP-positive myopathy may be included in a different category from ‘histological’ PM. In fact, some investigators have considered that anti-SRP antibody is associated with a ‘necrotizing myopathy’ that differs from ‘histological’ PM [8–10]. The clinical diagnosis of PM, however, is not based on histological findings alone. The diagnostic criteria proposed by Bohan and Peter [11] are known to be clinically practical, sensitive, and specific, and they have served the community well for nearly three decades [17]. We therefore conclude that ‘anti-SRP-positive’ should be classified as a particular form of PM.

We concluded that our patient had anti-SRP-positive ‘probable PM’. Anti-SRP-positive PM usually affects middle-aged people, however, and our patient is the youngest reported in the literature [5, 6,8–10]. More importantly, anti-SRP antibody is associated with treatment-resistant and refractory PM, although the clinical features of anti-SRP-positive PM patients show heterogeneity and some of them have a favourable prognosis [9]. It should be realized that anti-SRP-positive PM has sometimes been misdiagnosed as other types of myopathies [8].

Formula


   Acknowledgements
Top
 Abstract
 Introduction
 Patients, materials and methods
 Results
 Discussion
 Acknowledgements
References
 
We thank Dr Ichizo Nishino (National Institute of Neuroscience, National Center of Neurology and Psychiatry) for valuable comments on muscle biopsy and Ms Matsuko Ishida for assisting with the RNA immunoprecipitation assays.

Funding: This work was supported by a grant from the Japanese Ministry of Education, Science, Sports, and Culture, a research grant on intractable diseases from the Japanese Ministry of Health, Labour, and Welfare, and a Neuroimmunological Disease Research Committee grant from the Japanese Ministry of Health, Labour, and Welfare.

Disclosure statement: The authors have declared no conflicts of interest.


   References
Top
 Abstract
 Introduction
 Patients, materials and methods
 Results
 Discussion
 Acknowledgements
 References
 

  1. Dalakas MC, Hohlfeld R. Polymyositis and dermatomyositis. Lancet (2003) 362:971–82.[CrossRef][Web of Science][Medline]
  2. Targoff IN. Myositis specific autoantibodies. Curr Rheumatol Rep (2006) 8:196–203.[CrossRef][Medline]
  3. Reeves WH, Nigam SK, Blobel G. Human autoantibodies reactive with the signal-recognition particle. Proc Natl Acad Sci USA (1986) 83:9507–11.[Abstract/Free Full Text]
  4. Okada N, Mimori T, Mukai R, et al. Characterization of human autoantibodies that selectively precipitate the 7SL RNA component of the signal recognition particle. J Immunol (1987) 138:3219–23.[Abstract]
  5. Targoff IN, Johnson AE, Miller FW. Antibody to signal recognition particle in polymyositis. Arthritis Rheum (1990) 33:1361–70.[Web of Science][Medline]
  6. Kao AH, Lacomis D, Lucas M, et al. Anti-signal recognition particle autoantibody in patients with and patients without idiopathic inflammatory myopathy. Arthritis Rheum (2004) 50:209–15.[CrossRef][Web of Science][Medline]
  7. Engel AG. Muscular dystrophies. In: Myology: basic and clinical—Engel AG, Franzini-Armstrong C, eds. (2004) New York: McGraw-Hill. 961–1256.
  8. Dimitri D, Andre C, Roucoules J, et al. Myopathy associated with anti-signal recognition peptide antibodies: clinical heterogeneity contrasts with stereotyped histopathology. Muscle Nerve (2007) 35:389–95.[CrossRef][Web of Science][Medline]
  9. Hengstman GJ, ter Laak HJ, Vree Egberts WT, et al. Anti-signal recognition particle autoantibodies: marker of a necrotising myopathy. Ann Rheum Dis (2006) 65:1635–8.[Abstract/Free Full Text]
  10. Miller T, Al-Lozi MT, Lopate G, et al. Myopathy with antibodies to the signal recognition particle: clinical and pathological features. J Neurol Neurosurg Psychiatr (2002) 73:420–8.[Abstract/Free Full Text]
  11. Bohan A, Peter JB. Polymyositis and dermatomyositis (second of two parts). N Engl J Med (1975) 292:403–7.[Web of Science][Medline]
  12. Forman MS, Nakamura M, Mimori T, et al. Detection of antibodies to small nuclear ribonucleoproteins and small cytoplasmic ribonucleoproteins using unlabeled cell extracts. Arthritis Rheum (1985) 28:1356–61.[Web of Science][Medline]
  13. Brouwer R, Hengstman GJ, Vree Egberts W, et al. Autoantibody profiles in the sera of European patients with myositis. Ann Rheum Dis (2001) 60:116–23.[Abstract/Free Full Text]
  14. Perurena O, Targoff IN, Bobele G, et al. Lack of myositis-specific autoantibodies in Duchenne's muscular dystrophy (abstract). Ann Neurol (1993) 34:308.
  15. Hengstman GJ, van Brenk L, Vree Egberts WT, et al. High specificity of myositis specific autoantibodies for myositis compared with other neuromuscular disorders. J Neurol (2005) 252:534–7.[CrossRef][Web of Science][Medline]
  16. van der Meulen MF, Bronner IM, Hoogendijk JE, et al. Polymyositis: an overdiagnosed entity. Neurology (2003) 61:316–21.[Abstract/Free Full Text]
  17. Miller FW, Rider LG, Plotz PH, et al. Diagnostic criteria for polymyositis and dermatomyositis. Lancet (2003) 362:1762–3.[Medline]
Submitted 17 March 2008; revised version accepted 9 July 2008.
Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 Arch NeurolHome page
S. Suzuki, K. Utsugisawa, H. Yoshikawa, M. Motomura, S. Matsubara, K. Yokoyama, Y. Nagane, T. Maruta, T. Satoh, H. Sato, et al.
Autoimmune Targets of Heart and Skeletal Muscles in Myasthenia Gravis
Arch Neurol, November 1, 2009; 66(11): 1334 - 1338.
[Abstract] [Full Text] [PDF]

Home page
 Rheumatology (Oxford)Home page
T. Liewluck
Comment on: Clinical utility of anti-signal recognition particle antibody in the differential diagnosis of myopathies
Rheumatology, January 1, 2009; 48(1): 90 - 90.
[Full Text] [PDF]

Home page
 Rheumatology (Oxford)Home page
S. Suzuki, N. Suzuki, and M. Kuwana
Comment on: Clinical utility of anti-signal recognition particle antibody in the differential diagnosis of myopathies: reply
Rheumatology, January 1, 2009; 48(1): 90 - 91.
[Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
47/10/1539    most recent
ken325v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Suzuki, S.
Right arrow Articles by Kuwana, M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Suzuki, S.
Right arrow Articles by Kuwana, M.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?