Just Accepted by International Journal of Neuroscience
Electrodiagnosis and muscle biopsy in asymptomatic hyperckemia P. Kokotis, G.K. Papadimas, V. Zouvelou, T. Zambelis, P. Manta, N. Karandreas doi:10.3109/00207454.2015.1038534
Int J Neurosci Downloaded from informahealthcare.com by Nyu Medical Center on 05/26/15 For personal use only.
ABSTRACT Purpose/Aim of the study: An increased serum level of creatine kinase in asymptomatic individuals is a diagnostic challenge, as it may be associated with either physiological conditions, such as exercise or even signal an ominous neuromuscular disease at a presymptomatic stage. The electromyogram and the muscle biopsy play a key role in the evaluation of asymptomatic hyperckemia. The objective of this study was to investigate asymptomatic individuals with increased creatine kinase levels. Materials and Methods: We comparatively studied electromyogram, quantitative electromyogram and muscle biopsy in asymptomatic clinically normal individuals with repeatedly increased creatine kinase levels. Results: Conventional electromyogram was abnormal in 76% of patients, while quantitative electromyogram showed abnormal results in 88.9%. Muscle biopsy was diagnostic in 28%, one patient had neurogenic findings, 40% showed non specific changes and 28% had normal results. Conclusions: Electromyogram and especially quantitative electromyogram are highly sensitive in detecting subclinical neuromuscular diseases, whereas muscle biopsy may better contribute in the final diagnosis. No strong correlations were found between histological abnormalities and electrophysiological data, but further research is needed.
© 2015 Informa Healthcare USA, Inc. This provisional PDF corresponds to the article as it appeared upon acceptance. Fully formatted PDF and full text (HTML) versions will be made available soon. DISCLAIMER: The ideas and opinions expressed in the journal’s Just Accepted articles do not necessarily reflect those of Informa Healthcare (the Publisher), the Editors or the journal. The Publisher does not assume any responsibility for any injury and/or damage to persons or property arising from or related to any use of the material contained in these articles. The reader is advised to check the appropriate medical literature and the product information currently provided by the manufacturer of each drug to be administered to verify the dosages, the method and duration of administration, and contraindications. It is the responsibility of the treating physician or other health care professional, relying on his or her independent experience and knowledge of the patient, to determine drug dosages and the best treatment for the patient. Just Accepted have undergone full scientific review but none of the additional editorial preparation, such as copyediting, typesetting, and proofreading, as have articles published in the traditional manner. There may, therefore, be errors in Just Accepted articles that will be corrected in the final print and final online version of the article. Any use of the Just Accepted articles is subject to the express understanding that the papers have not yet gone through the full quality control process prior to publication.
Publisher: Taylor & Francis Journal: International Journal of Neuroscience DOI: http://dx.doi.org/10.3109/00207454.2015.1038534
and
muscle
biopsy
hyperckemia
in
asymptomatic
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Electrodiagnosis
EP
Department of Neurology, University of Athens, School of Medicine, Aeginition Hospital, Athens, Greece
AC
C
Responsible author’s address: George K. Papadimas, MD University of Athens, School of Medicine 1st Department of Neurology Eginition Hospital 74, Vas. Sophias Ave. 11528 Athens-Greece Tel: ++302107289152 Fax: ++302107216474 E- mail: [email protected]
ST
ABSTRACT
Purpose/Aim of the study: An increased serum level of creatine kinase in asymptomatic individuals is a diagnostic challenge, as it may be associated with either physiological
JU
Int J Neurosci Downloaded from informahealthcare.com by Nyu Medical Center on 05/26/15 For personal use only.
P. Kokotis, G.K. Papadimas, V. Zouvelou, T. Zambelis, P. Manta, N. Karandreas
conditions, such as exercise or even signal an ominous neuromuscular disease at a presymptomatic stage. The electromyogram and the muscle biopsy play a key role in the evaluation of asymptomatic hyperckemia. The objective of this study was to investigate asymptomatic individuals with increased creatine kinase levels. Materials and Methods: We comparatively studied electromyogram, quantitative electromyogram and muscle biopsy in asymptomatic clinically normal individuals with repeatedly increased creatine kinase
1
levels. Results: Conventional electromyogram was abnormal in 76% of patients, while quantitative electromyogram showed abnormal results in 88.9%. Muscle biopsy was diagnostic in 28%, one patient had neurogenic findings, 40% showed non specific changes
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and 28% had normal results. Conclusions: Electromyogram and especially quantitative electromyogram are highly sensitive in detecting subclinical neuromuscular diseases,
whereas muscle biopsy may better contribute in the final diagnosis. No strong correlations
EP
research is needed.
ST
AC
C
Key words: muscle biopsy, electromyogram, hyperckemia
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Int J Neurosci Downloaded from informahealthcare.com by Nyu Medical Center on 05/26/15 For personal use only.
were found between histological abnormalities and electrophysiological data, but further
2
INTRODUCTION The increased levels of serum creatine kinase (CK) may be the sole initial manifestation of an underlying neuromuscular disease or may be secondarily elevated after
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medication use, especially statins, or either associated with other systemic disorders such as viral infections, endocrinopathies and connective tissue diseases [1]. It is also well
established that CK serum levels may be influenced by several physiological parameters
EP
designation of diagnostic algorithms in the investigation of asymptomatic subjects with
constantly high CK plasma levels [1]. EMG (electomyogram) and muscle biopsy provide complementary information and constitute an integral part of the diagnostic work-up either
C
for individuals with asymptomatic hyperckaemia or for patients with suspected muscle diseases [1,3,4].
AC
Although EMG does not substantially contribute in the differentiation of the various forms of myopathy, it can definitely detect myopathic changes, especially with the application of quantitative methods initially introduced by Buchthal and Clemmensen which
ST
have importantly increased the diagnostic yield [3,5]. The duration and amplitude of MUP (motor unit potential), as well as the percentage of polyphasic potentials, are the main conventional
parameters
usually
analysed
during
the
QEMG
(quantitative
electromyography) [3]. MUP duration shortening which results from muscle fiber loss is the
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Int J Neurosci Downloaded from informahealthcare.com by Nyu Medical Center on 05/26/15 For personal use only.
such as gender and race [1,2]. The multifactorial causes of hypercckemia necessitated the
most reliable feature of a myopathic condition [6]. EMG provides also invaluable information about the extent and pattern of muscle involvement and in combination with the clinical examination can guide the selection of muscle for biopsy [7]. The utility of open muscle biopsy in the evaluation of a suspected myopathy has been already confirmed and its role for the investigation of most forms of acquired or inherited muscle diseases remains still crucial [4].
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The aim of the present study is a comparative study between EMG, QEMG and muscle biopsy findings in individuals with asymptomatic hyperckaemia.
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MATERIALS AND METHODS Patients
We reviewed EMG findings, QEMG measurements and muscle biopsy specimens of
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Hospital for neuromuscular evaluation between 2005 and 2011. All the participants who
included 19 males and 6 females (age range: 18-76yrs), should have been asymptomatic and met the following criteria: a) hyperckemia (CK more than 1.5 times the upper limit of
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normal) in at least two different measurements (normal value below 180 U/L), while the individuals were clearly advised to abstain from heavy physical activity, b) normal clinical
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examination, c) normative nerve conduction studies on electrophysiological testing, d) no evidence of known causes of CK increase, such as specific medication use, e) no episode of rhabdomyolysis, even if the interictal CK measurements were normal.
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Electromyogram
All the patients have been submitted to EMG performed by the same physician, before the muscle biopsy. Nerve conduction studies included two sensory and two motor nerves in the upper extremities and one sensory and one motor nerve in the lower extremity.
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consecutive individuals who referred to the Department of Neurology of the Aeginition
An EMG apparatus (Medtronic Functional Diagnostics, Keypoint 31A02, Skovlunde, Denmark) with MUAP analysis software (Medtronic KeypointVR . NET, version 4.3.505.0) and concentric needle electrodes (Alpine Biomed, Aps, Skovlunde, Denmark, type DCNTM25), 25 mm in length and 0.46 mm in diameter with a recording surface of 0.07 mm2, were used; filter settings were 5 Hz to 10 kHz. During weak contraction, 1 to 4 MUAPs were automatically recorded through a template and averaging program; duration,
4
amplitude, rise time, number of phases, turns, and area were measured automatically according to the automatic multi-MUP method described by Stalberg et al. [8]. The examining room temperature remained above 24 oC during all procedures. At least 2
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puncture sites were used. The needle was then displaced to 2–3 locations in each site, in steps of at least 5 mm. MUAP sampling was done during weak muscle contractions to avoid
overlapping of the detected MUAPs and only when potentials with a crisp sound and sharp
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neighboring areas were rejected. At least 20 MUAPs were collected from each tested
muscle, including polyphasic potentials. The onset and the end of each MUAP were determined on well-defined, clear-cut deflections of the potentials, making sure to include
present in more than 4 potentials.
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components of the MUAP with brief duration and small amplitude that were consistently
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Myopathy according to the EMG criteria was defined as a needle study revealing polyphasic motor units potentials of short duration and low amplitude, usually with early recruitment and occasionally with abnormal spontaneous activity [9].
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Quantitative electromyography has not been completed in six patients and the results are not included in the study.
The muscles usually tested for QEMG were tibialis anterior, bicheps brachialis and iliopsoas, for which there are the lower normal limits for amplitude and duration from our EMG
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morphology were present in the site. MUAPs with the same morphology, recorded from
laboratory normative data. Specifically, iliopsoas was selected on the basis of the usually proximal muscle involvement in most myopathic diseases and was examined with an already described reliable electromyographic technique [10]. Muscle biopsy
Muscle specimens were obtained by open biopsy, under local anesthesia, from the left quadriceps femoris. A portion of the muscle sample was snap frozen in liquid-nitrogen-
5
cooled isopentane. Cryostat sections were cut at 6-μm thickness and used for histological, histochemical, and immunohistochemical studies, using conventional techniques. Muscle morphology was demonstrated with hematoxylin–eosin (HE) and Gomori’s modified
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trichrome stains. Histochemical reactions included NADH-TR, SDH, cytochrome oxidase (COX), SDH-COX, ATPase (pH 9.4, 4.3, 4.6), phosphorylase, AMPDA, NSE, periodic acid
Schiff (PAS), diastase-PAS, and Oil red O. Immunohistochemical study included the
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DYS-2: C-terminus; DYS-3: N-terminus (1:10, Novocastra); anti- α, β, γ, δ sarcoglycan (1:100, 1:50, 1:100, 1:25, Novocastra); anti-caveolin (1:50, Santa Cruz); anti-dysferlin NCL-
Hamlet (1:10, Novocastra); anti-merosin (1:2000, Chemicon); anti-sarcoplasmic reticulum
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Ca2+ ATPase (SERCA2-ATPase, 1:100, Novocastra); anti-emerin (1:20, Novocastra); and anti-Lamin A antibody (1:5, Chemicon). Sections were incubated for 1 h at room
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temperature with the primary antibody and immunostained with biotin-extravidin (Extra-2, Sigma, St. Louis, Mo.). Color was developed with amino-ethyl-carbazole (AEC). All the patients had also a normal western blot for calpain.
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The clinical examination of the patients and the interpretation of muscle biopsies were performed by the same physician with expertise in neuromuscular disorders. Ethical approval for this study was not required given it was a retrospective chart review.
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following primary mouse monoclonal antibodies: anti-dystrophin: DYS-1: rod domain;
RESULTS This study involved a total number of 25 patients aged from 18 to 76yrs who have
been submitted to EMG and skeletal muscle biopsy (Table 1). All the patients included had an unremarkable neuromuscular examination and their nerve conduction studies were within the reference range of normal. The main referral cause was the repetitive increase of CK in consecutive measurements, which had been initially detected in a routine laboratory test. The
6
range of patients’ serum CK levels in consecutive measurements was from mildly elevated (min value: 388U/L) up to 5300U/L. Nineteen of the 25 patients included in the study had abnormal findings on
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conventional EMG which were suggestive of a myopathy, a neurogenic disorder or a mixed myopathic and neurogenic process. Based on EMG data analysis for the 18 patients who
were submitted to quantitative EMG, sixteen were found to have an abnormally reduced
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showed normal results on conventional EMG, but abnormal findings in muscle biopsy. Two patients had normal EMG with both methods and normal muscle biopsy (Figure 1). In regards to the muscle biopsy, four patients had mitochondrial myopathy, one
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patient had inflammatory myopathy, one patient had glycogen storage disease type V (Mc Ardle’s disease), one patient was diagnosed with LGMD1C (caveolinopathy) and one patient
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had neurogenic changes. Muscle biopsy findings in patients with McArdle’s disease and LGMD1C are shown in Figures 2 and 3, respectively. In ten patients, only subtle nonspecific myoapthic changes were described. The most prominent histopathological non
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specific myopathic features included an increased number of centrally located nuclei, some scattered atrophic fibers and an increased fiber size variation. Rarer findings were the presence of few ring fibers and fiber splitting in one case and a small number of scattered moth-eaten fibers and core-like structures in another patient. Seven subjects showed no
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mean MUP duration in the brachial biceps and/or tibialis anterior muscle, while two of them
abnormal microscopic findings on tissue biopsy.
DISCUSSION The aim of the present study was to correlate conventional EMG, QEMG and muscle biopsy findings in individuals with asymptomatic hyperckemia. Quantitative EMG showed abnormal measurements in sixteen out of 18 subjects (88.8%), while conventional EMG was
7
abnormal in fourteen of them. The discordant results were obtained in one patient with mild myositis and in another one with mild myopahic changes on muscle biopsy suggesting that the quantitative measurements may have a higher sensitivity than the conventional EMG in
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detecting a myopathy. Overall, conventional EMG was abnormal in 19 (76%) patients, while the muscle biopsy showed myopathic findings in 17 (68%) participants with increased CK.
The above observations support the important role of electrodiagnosis as a first step
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patients with a normal muscle biopsy interpretation. These patients had an inconclusive diagnosis and their thorough investigation did not yield evidence for any underlying
pathology, possibly meaning that although EMG seems to be very sensitive in detecting
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myopathic changes, it may have some phenomenally positive results compared to muscle biopsy. To further support the aforementioned finding, EMG was normal only in one patient
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with evidence of mitochondrial dysfunction on muscle biopsy. Even in this case this discrepancy might be possibly avoided if the patient had been submitted to quantitative EMG.
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Despite the remarkable sensitivity of EMG in revealing even subclinical evidence of a myopathic process, it is well known to have a restricted usefulness in differentiating the various forms of myopathy [3]. On the contrary, muscle biopsy may better contribute to a specific diagnosis and its usefulness in the evaluation of neuromuscular diseases is well
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towards diagnosis. On the other hand, according to our results, EMG was abnormal in 4
established. Nevertheless, it may give some false negative results possibly providing a further explanation for the inconsistency in some cases. A common cause of false negative results on muscle biopsy is attributed to a patchy distribution of abnormal changes in some myopathies, such as the inflammatory myopathies which may exhibit regional variability of the histologic abnormalities [4]. Therefore the need for a careful selection of muscle to be biopsied is of critical importance in order to reach a correct diagnosis and it is already known
8
that even within the same muscle, there may be some variability in the presence of pathological findings [11-13]. In our study a definite diagnosis had been established at a presymptomatic stage in 7
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patients (28%), providing an etiologic explanation for the increased CK levels. The relatively low probability of reaching a final diagnosis has been also confirmed in a previous retrospective study which included individuals with asymptomatic or even oligosymptomatic
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diagnoses, such as the presence of ragged red fibers in the cases of mitochondrial myopathy or the subsarcolemnal vacuoles due to glycogen accumulation in the patient with McArdle’s
disease, the main abnormalities of the myopathic biopsies were non-specific and consisted of
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an increase in internal nuclei number and in fiber size variation. Less common findings included few regenerated and necrotic muscle fibers, some scattered angulated atrophied
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fibers, an occasional presence of lobulated and ring fibers, rare pyknotic nuclear clumps and sparse fibers with internal splitting. In case that some of the above abnormalities are combined and profuse may be quite diagnostic, but when infrequent and isolated they are
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usually unrevealing and not diagnostic.
It is worth mentioned that the results of the present study should be interpreted with
caution because of the small number of the sample and the lack of an age control group. Nevertheless, although some myopathic findings such as the increased variability in muscle
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hyperckemia [14]. Apart from the typical histopathologic findings related to conclusive
fiber size and the internal nucleation seem to be the most sensitive histological features, they cannot be reliably correlated with EMG in the detection of a myopathic condition. This is in accordance with previous studies that have similarly failed to prove a correlation between histological findings and EMG parameters, with just few exceptions, such as a possible association of long duration polyphasic potentials with regenerating fibers in polymyositis [15].
9
CONCLUSION The present study showed that asymptomatic hyperckemia may hint at a possible
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underlying myopathic disease that has not been manifested yet. On the other hand, the elevated CK levels seemed to have a very high positive predictive value in detecting myopathic changes on EMG and in revealing non specific histologic abnormalities on
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Overall, periodic re-evaluation and systematic follow-up of those subjects with asymptomatic CK elevation is recommended to allow for an early detection of a possible
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later onset neuromuscular disease or some systemic disorder.
Declaration of interest: The authors report no conflict of interest. The authors alone are
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responsible for the content and writing of the paper
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muscle biopsy, although the clinical significance of the above findings remains elusive.
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REFERENCES 1. Morandi L, Angelini C, Prelle A, et al. High plasma creatine kinase : review of the literature and proposal fo a diagnostic algorithm. Neurol Sci. 2006;27:303-311.
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2. Wong ET, Cobb C, Humehara MK, et al. Heterogeneity of serum creatine kinase activity among racial and gender groups of the population. Am J Clin Pathol. 1983;79:582–586.
3. Fuglsang-Frederiksen A. The role of different EMG methods in evaluating myopathy.
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4. Lai CH, Melli G, Chang YJ, et al. Open muscle biopsy in suspected myopathy: diagnostic yield and clinical utility. Eur J Neurol. 2010;17:136-142.
5. Buchthal F, Clemmensen S. On the differentiation of muscle atrophy by
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electromyography. Acta Psychiatr Neurol. 1941;16:143–181.
6. Stewart CR, Nandedkar SD, Massey JM, Gilchrist JM, Barkhaus PE, Sanders DB.
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Evaluation of an automatic method of measuring features of motor unit action potentials. Muscle Nerve. 1989;12:141–148.
7. Mastaglia FL, Laing NG. Investigation of muscle disease. J Neurol Neurosurg Psych.
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1996;60:256-274.
8. Stalberg E, Falck B, Sonoo M, Stalberg S, Astrom M. Multi-MUP EMG analysis-a two year experience in daily clinical work. Electroencephalogr Clin Neurophysiol. 1995;97:145–154.
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Clin Neurophysiol. 2006;117:1173-1189.
9. Buchthal F. Electromyography in the evaluation of muscle diseases. Neurol Clin. 1985;3:573-598.
10. Katsavrias E, Primetis E, Karandreas N. Iliopsoas: a new electromyographic technique and normal motor unit action potential values. Clin Neurophysiol. 2005;116:2528-2532.
11. Prayson RA. Diagnostic yield associated with multiple simultaneous skeletal muscle biopsies. Am J Clin Pathol. 2006;126:843-848.
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12. Wilbourn AJ. The electrodiagnostic examination with myopathies. J Clin Neurophysiol. 1993;10:132-148. 13. Black JT, Bhatt GP, Dejesus PV, Schotland DL, Rowland LP. Diagnostic accuracy of
disease. A study of 105 cases. J Neurol Sci. 1974;21:59-70.
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clinical data, quantitative electromyography and histochemistry in neuromuscular
14. Prelle A, Tancredi L, Sciacco M, et al. Retrospective study of a large population of
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2002;249:305–311.
15. Uncini A, Lange DJ, Lovelace RE, Solomon M, Hays AP. Long-duration polyphasic motor unit potentials in myopathies: a quantitative study with pathological correlation.
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AC
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Muscle Nerve. 1990;13:263-267.
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patients effeceted with asymptomatic or paucisymptomatic hyperCKemia. J Neurol.
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Muscle biopsy
M
myopathic
F
44
normal
myopathic
3
M
49
myopathic
myopathic
4 5
M M
18 48
myopathic normal
myopathic myopathic
6
M
44
myopathic
myopathic
7
F
68
myopathic
myopathic
8
M
36
myopathic
myopathic
9
M
56
myopathic
myopathic
10 11
M F
54 28
normal myopathic
normal myopathic
12
M
42
myopathic
13 14
M M
25 23
15 16
M M
22 38
mitochondrial dysfunction inflammatory infiltrates myopathic changes normal myopathic changes mitochondrial dysfunction myopathic changes phosphorylase deficiency myopathic changes normal myopathic changes mitochondrial dysfunction normal myopathic changes normal mitochondrial dysfunction myopathic changes normal myopathic changes neurogenic
2
EP
1
myopathic
myopathic myopathic
myopathic myopathic
Normal normal
normal -
17
F
76
myopathic
-
18 19
F M
66 49
myopathic myopathic
myopathic
20
M
63
-
21
F
41
myopathic + neurogenic myopathic
22
M
36
neurogenic
-
23
M
23
24 25
M M
48 58
myopathic + neurogenic normal myopathic
myopathic + neurogenic myopathic
-
13
Final diagnosis Mitochondrial myopathy Inflammatory myopathy -
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QEMG
44
EMG (conventional) myopathic
C
Age
AC
Sex
ST
Patient
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Table 1: Electrophysiologic findings, biopsy features and clinical diagnoses
-
Mitochondrial myopathy McArdle’s disease -
Mitochondrial myopathy Mitochondrial myopathy -
myopathic changes myopathic changes normal
LGMD1C
normal myopathic changes
-
-
ST
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C
AC
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EMG: electromyogram, QEMG:quantitative EMG, LGMD1C: limb girdle muscular dystrophy 1C,
14
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abnormal muscle biopsy, QEMG and conventional EMG, respectively.
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EMG: Conventional Electromygraphy, QEMG: Quantitative Electromyography, BN: Muscle biopsy normal, BA: Muscle biopsy abnormal
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Figure 1: Categorized histogram shows the number of observations with normal and
15
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ST JU Figure 2: Histological findings from patient with McArdle’s disease showing subsarcolemmal vacuoles in many muscle fibers on hematoxylin-eosin (HE) staining (2a) with abnormal accumulation of glycogen on PAS staining (2b). 16
17
ST
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C
AC
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Figure 3: Muscle biopsy from patient with LGMD1C revealing significantly reduced immunohistochemical expression of caveolin-3 in muscle fibers (3a) compared to control
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(3b).
18
Electrodiagnosis and muscle biopsy in asymptomatic hyperckemia P. Kokotis, G.K. Papadimas, V. Zouvelou, T. Zambelis, P. Manta, N. Karandreas doi:10.3109/00207454.2015.1038534
Int J Neurosci Downloaded from informahealthcare.com by Nyu Medical Center on 05/26/15 For personal use only.
ABSTRACT Purpose/Aim of the study: An increased serum level of creatine kinase in asymptomatic individuals is a diagnostic challenge, as it may be associated with either physiological conditions, such as exercise or even signal an ominous neuromuscular disease at a presymptomatic stage. The electromyogram and the muscle biopsy play a key role in the evaluation of asymptomatic hyperckemia. The objective of this study was to investigate asymptomatic individuals with increased creatine kinase levels. Materials and Methods: We comparatively studied electromyogram, quantitative electromyogram and muscle biopsy in asymptomatic clinically normal individuals with repeatedly increased creatine kinase levels. Results: Conventional electromyogram was abnormal in 76% of patients, while quantitative electromyogram showed abnormal results in 88.9%. Muscle biopsy was diagnostic in 28%, one patient had neurogenic findings, 40% showed non specific changes and 28% had normal results. Conclusions: Electromyogram and especially quantitative electromyogram are highly sensitive in detecting subclinical neuromuscular diseases, whereas muscle biopsy may better contribute in the final diagnosis. No strong correlations were found between histological abnormalities and electrophysiological data, but further research is needed.
© 2015 Informa Healthcare USA, Inc. This provisional PDF corresponds to the article as it appeared upon acceptance. Fully formatted PDF and full text (HTML) versions will be made available soon. DISCLAIMER: The ideas and opinions expressed in the journal’s Just Accepted articles do not necessarily reflect those of Informa Healthcare (the Publisher), the Editors or the journal. The Publisher does not assume any responsibility for any injury and/or damage to persons or property arising from or related to any use of the material contained in these articles. The reader is advised to check the appropriate medical literature and the product information currently provided by the manufacturer of each drug to be administered to verify the dosages, the method and duration of administration, and contraindications. It is the responsibility of the treating physician or other health care professional, relying on his or her independent experience and knowledge of the patient, to determine drug dosages and the best treatment for the patient. Just Accepted have undergone full scientific review but none of the additional editorial preparation, such as copyediting, typesetting, and proofreading, as have articles published in the traditional manner. There may, therefore, be errors in Just Accepted articles that will be corrected in the final print and final online version of the article. Any use of the Just Accepted articles is subject to the express understanding that the papers have not yet gone through the full quality control process prior to publication.
Publisher: Taylor & Francis Journal: International Journal of Neuroscience DOI: http://dx.doi.org/10.3109/00207454.2015.1038534
and
muscle
biopsy
hyperckemia
in
asymptomatic
TE D
Electrodiagnosis
EP
Department of Neurology, University of Athens, School of Medicine, Aeginition Hospital, Athens, Greece
AC
C
Responsible author’s address: George K. Papadimas, MD University of Athens, School of Medicine 1st Department of Neurology Eginition Hospital 74, Vas. Sophias Ave. 11528 Athens-Greece Tel: ++302107289152 Fax: ++302107216474 E- mail: [email protected]
ST
ABSTRACT
Purpose/Aim of the study: An increased serum level of creatine kinase in asymptomatic individuals is a diagnostic challenge, as it may be associated with either physiological
JU
Int J Neurosci Downloaded from informahealthcare.com by Nyu Medical Center on 05/26/15 For personal use only.
P. Kokotis, G.K. Papadimas, V. Zouvelou, T. Zambelis, P. Manta, N. Karandreas
conditions, such as exercise or even signal an ominous neuromuscular disease at a presymptomatic stage. The electromyogram and the muscle biopsy play a key role in the evaluation of asymptomatic hyperckemia. The objective of this study was to investigate asymptomatic individuals with increased creatine kinase levels. Materials and Methods: We comparatively studied electromyogram, quantitative electromyogram and muscle biopsy in asymptomatic clinically normal individuals with repeatedly increased creatine kinase
1
levels. Results: Conventional electromyogram was abnormal in 76% of patients, while quantitative electromyogram showed abnormal results in 88.9%. Muscle biopsy was diagnostic in 28%, one patient had neurogenic findings, 40% showed non specific changes
TE D
and 28% had normal results. Conclusions: Electromyogram and especially quantitative electromyogram are highly sensitive in detecting subclinical neuromuscular diseases,
whereas muscle biopsy may better contribute in the final diagnosis. No strong correlations
EP
research is needed.
ST
AC
C
Key words: muscle biopsy, electromyogram, hyperckemia
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Int J Neurosci Downloaded from informahealthcare.com by Nyu Medical Center on 05/26/15 For personal use only.
were found between histological abnormalities and electrophysiological data, but further
2
INTRODUCTION The increased levels of serum creatine kinase (CK) may be the sole initial manifestation of an underlying neuromuscular disease or may be secondarily elevated after
TE D
medication use, especially statins, or either associated with other systemic disorders such as viral infections, endocrinopathies and connective tissue diseases [1]. It is also well
established that CK serum levels may be influenced by several physiological parameters
EP
designation of diagnostic algorithms in the investigation of asymptomatic subjects with
constantly high CK plasma levels [1]. EMG (electomyogram) and muscle biopsy provide complementary information and constitute an integral part of the diagnostic work-up either
C
for individuals with asymptomatic hyperckaemia or for patients with suspected muscle diseases [1,3,4].
AC
Although EMG does not substantially contribute in the differentiation of the various forms of myopathy, it can definitely detect myopathic changes, especially with the application of quantitative methods initially introduced by Buchthal and Clemmensen which
ST
have importantly increased the diagnostic yield [3,5]. The duration and amplitude of MUP (motor unit potential), as well as the percentage of polyphasic potentials, are the main conventional
parameters
usually
analysed
during
the
QEMG
(quantitative
electromyography) [3]. MUP duration shortening which results from muscle fiber loss is the
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such as gender and race [1,2]. The multifactorial causes of hypercckemia necessitated the
most reliable feature of a myopathic condition [6]. EMG provides also invaluable information about the extent and pattern of muscle involvement and in combination with the clinical examination can guide the selection of muscle for biopsy [7]. The utility of open muscle biopsy in the evaluation of a suspected myopathy has been already confirmed and its role for the investigation of most forms of acquired or inherited muscle diseases remains still crucial [4].
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The aim of the present study is a comparative study between EMG, QEMG and muscle biopsy findings in individuals with asymptomatic hyperckaemia.
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MATERIALS AND METHODS Patients
We reviewed EMG findings, QEMG measurements and muscle biopsy specimens of
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Hospital for neuromuscular evaluation between 2005 and 2011. All the participants who
included 19 males and 6 females (age range: 18-76yrs), should have been asymptomatic and met the following criteria: a) hyperckemia (CK more than 1.5 times the upper limit of
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normal) in at least two different measurements (normal value below 180 U/L), while the individuals were clearly advised to abstain from heavy physical activity, b) normal clinical
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examination, c) normative nerve conduction studies on electrophysiological testing, d) no evidence of known causes of CK increase, such as specific medication use, e) no episode of rhabdomyolysis, even if the interictal CK measurements were normal.
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Electromyogram
All the patients have been submitted to EMG performed by the same physician, before the muscle biopsy. Nerve conduction studies included two sensory and two motor nerves in the upper extremities and one sensory and one motor nerve in the lower extremity.
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consecutive individuals who referred to the Department of Neurology of the Aeginition
An EMG apparatus (Medtronic Functional Diagnostics, Keypoint 31A02, Skovlunde, Denmark) with MUAP analysis software (Medtronic KeypointVR . NET, version 4.3.505.0) and concentric needle electrodes (Alpine Biomed, Aps, Skovlunde, Denmark, type DCNTM25), 25 mm in length and 0.46 mm in diameter with a recording surface of 0.07 mm2, were used; filter settings were 5 Hz to 10 kHz. During weak contraction, 1 to 4 MUAPs were automatically recorded through a template and averaging program; duration,
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amplitude, rise time, number of phases, turns, and area were measured automatically according to the automatic multi-MUP method described by Stalberg et al. [8]. The examining room temperature remained above 24 oC during all procedures. At least 2
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puncture sites were used. The needle was then displaced to 2–3 locations in each site, in steps of at least 5 mm. MUAP sampling was done during weak muscle contractions to avoid
overlapping of the detected MUAPs and only when potentials with a crisp sound and sharp
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neighboring areas were rejected. At least 20 MUAPs were collected from each tested
muscle, including polyphasic potentials. The onset and the end of each MUAP were determined on well-defined, clear-cut deflections of the potentials, making sure to include
present in more than 4 potentials.
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components of the MUAP with brief duration and small amplitude that were consistently
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Myopathy according to the EMG criteria was defined as a needle study revealing polyphasic motor units potentials of short duration and low amplitude, usually with early recruitment and occasionally with abnormal spontaneous activity [9].
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Quantitative electromyography has not been completed in six patients and the results are not included in the study.
The muscles usually tested for QEMG were tibialis anterior, bicheps brachialis and iliopsoas, for which there are the lower normal limits for amplitude and duration from our EMG
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morphology were present in the site. MUAPs with the same morphology, recorded from
laboratory normative data. Specifically, iliopsoas was selected on the basis of the usually proximal muscle involvement in most myopathic diseases and was examined with an already described reliable electromyographic technique [10]. Muscle biopsy
Muscle specimens were obtained by open biopsy, under local anesthesia, from the left quadriceps femoris. A portion of the muscle sample was snap frozen in liquid-nitrogen-
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cooled isopentane. Cryostat sections were cut at 6-μm thickness and used for histological, histochemical, and immunohistochemical studies, using conventional techniques. Muscle morphology was demonstrated with hematoxylin–eosin (HE) and Gomori’s modified
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trichrome stains. Histochemical reactions included NADH-TR, SDH, cytochrome oxidase (COX), SDH-COX, ATPase (pH 9.4, 4.3, 4.6), phosphorylase, AMPDA, NSE, periodic acid
Schiff (PAS), diastase-PAS, and Oil red O. Immunohistochemical study included the
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DYS-2: C-terminus; DYS-3: N-terminus (1:10, Novocastra); anti- α, β, γ, δ sarcoglycan (1:100, 1:50, 1:100, 1:25, Novocastra); anti-caveolin (1:50, Santa Cruz); anti-dysferlin NCL-
Hamlet (1:10, Novocastra); anti-merosin (1:2000, Chemicon); anti-sarcoplasmic reticulum
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Ca2+ ATPase (SERCA2-ATPase, 1:100, Novocastra); anti-emerin (1:20, Novocastra); and anti-Lamin A antibody (1:5, Chemicon). Sections were incubated for 1 h at room
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temperature with the primary antibody and immunostained with biotin-extravidin (Extra-2, Sigma, St. Louis, Mo.). Color was developed with amino-ethyl-carbazole (AEC). All the patients had also a normal western blot for calpain.
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The clinical examination of the patients and the interpretation of muscle biopsies were performed by the same physician with expertise in neuromuscular disorders. Ethical approval for this study was not required given it was a retrospective chart review.
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following primary mouse monoclonal antibodies: anti-dystrophin: DYS-1: rod domain;
RESULTS This study involved a total number of 25 patients aged from 18 to 76yrs who have
been submitted to EMG and skeletal muscle biopsy (Table 1). All the patients included had an unremarkable neuromuscular examination and their nerve conduction studies were within the reference range of normal. The main referral cause was the repetitive increase of CK in consecutive measurements, which had been initially detected in a routine laboratory test. The
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range of patients’ serum CK levels in consecutive measurements was from mildly elevated (min value: 388U/L) up to 5300U/L. Nineteen of the 25 patients included in the study had abnormal findings on
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conventional EMG which were suggestive of a myopathy, a neurogenic disorder or a mixed myopathic and neurogenic process. Based on EMG data analysis for the 18 patients who
were submitted to quantitative EMG, sixteen were found to have an abnormally reduced
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showed normal results on conventional EMG, but abnormal findings in muscle biopsy. Two patients had normal EMG with both methods and normal muscle biopsy (Figure 1). In regards to the muscle biopsy, four patients had mitochondrial myopathy, one
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patient had inflammatory myopathy, one patient had glycogen storage disease type V (Mc Ardle’s disease), one patient was diagnosed with LGMD1C (caveolinopathy) and one patient
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had neurogenic changes. Muscle biopsy findings in patients with McArdle’s disease and LGMD1C are shown in Figures 2 and 3, respectively. In ten patients, only subtle nonspecific myoapthic changes were described. The most prominent histopathological non
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specific myopathic features included an increased number of centrally located nuclei, some scattered atrophic fibers and an increased fiber size variation. Rarer findings were the presence of few ring fibers and fiber splitting in one case and a small number of scattered moth-eaten fibers and core-like structures in another patient. Seven subjects showed no
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mean MUP duration in the brachial biceps and/or tibialis anterior muscle, while two of them
abnormal microscopic findings on tissue biopsy.
DISCUSSION The aim of the present study was to correlate conventional EMG, QEMG and muscle biopsy findings in individuals with asymptomatic hyperckemia. Quantitative EMG showed abnormal measurements in sixteen out of 18 subjects (88.8%), while conventional EMG was
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abnormal in fourteen of them. The discordant results were obtained in one patient with mild myositis and in another one with mild myopahic changes on muscle biopsy suggesting that the quantitative measurements may have a higher sensitivity than the conventional EMG in
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detecting a myopathy. Overall, conventional EMG was abnormal in 19 (76%) patients, while the muscle biopsy showed myopathic findings in 17 (68%) participants with increased CK.
The above observations support the important role of electrodiagnosis as a first step
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patients with a normal muscle biopsy interpretation. These patients had an inconclusive diagnosis and their thorough investigation did not yield evidence for any underlying
pathology, possibly meaning that although EMG seems to be very sensitive in detecting
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myopathic changes, it may have some phenomenally positive results compared to muscle biopsy. To further support the aforementioned finding, EMG was normal only in one patient
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with evidence of mitochondrial dysfunction on muscle biopsy. Even in this case this discrepancy might be possibly avoided if the patient had been submitted to quantitative EMG.
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Despite the remarkable sensitivity of EMG in revealing even subclinical evidence of a myopathic process, it is well known to have a restricted usefulness in differentiating the various forms of myopathy [3]. On the contrary, muscle biopsy may better contribute to a specific diagnosis and its usefulness in the evaluation of neuromuscular diseases is well
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towards diagnosis. On the other hand, according to our results, EMG was abnormal in 4
established. Nevertheless, it may give some false negative results possibly providing a further explanation for the inconsistency in some cases. A common cause of false negative results on muscle biopsy is attributed to a patchy distribution of abnormal changes in some myopathies, such as the inflammatory myopathies which may exhibit regional variability of the histologic abnormalities [4]. Therefore the need for a careful selection of muscle to be biopsied is of critical importance in order to reach a correct diagnosis and it is already known
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that even within the same muscle, there may be some variability in the presence of pathological findings [11-13]. In our study a definite diagnosis had been established at a presymptomatic stage in 7
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patients (28%), providing an etiologic explanation for the increased CK levels. The relatively low probability of reaching a final diagnosis has been also confirmed in a previous retrospective study which included individuals with asymptomatic or even oligosymptomatic
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diagnoses, such as the presence of ragged red fibers in the cases of mitochondrial myopathy or the subsarcolemnal vacuoles due to glycogen accumulation in the patient with McArdle’s
disease, the main abnormalities of the myopathic biopsies were non-specific and consisted of
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an increase in internal nuclei number and in fiber size variation. Less common findings included few regenerated and necrotic muscle fibers, some scattered angulated atrophied
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fibers, an occasional presence of lobulated and ring fibers, rare pyknotic nuclear clumps and sparse fibers with internal splitting. In case that some of the above abnormalities are combined and profuse may be quite diagnostic, but when infrequent and isolated they are
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usually unrevealing and not diagnostic.
It is worth mentioned that the results of the present study should be interpreted with
caution because of the small number of the sample and the lack of an age control group. Nevertheless, although some myopathic findings such as the increased variability in muscle
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hyperckemia [14]. Apart from the typical histopathologic findings related to conclusive
fiber size and the internal nucleation seem to be the most sensitive histological features, they cannot be reliably correlated with EMG in the detection of a myopathic condition. This is in accordance with previous studies that have similarly failed to prove a correlation between histological findings and EMG parameters, with just few exceptions, such as a possible association of long duration polyphasic potentials with regenerating fibers in polymyositis [15].
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CONCLUSION The present study showed that asymptomatic hyperckemia may hint at a possible
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underlying myopathic disease that has not been manifested yet. On the other hand, the elevated CK levels seemed to have a very high positive predictive value in detecting myopathic changes on EMG and in revealing non specific histologic abnormalities on
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Overall, periodic re-evaluation and systematic follow-up of those subjects with asymptomatic CK elevation is recommended to allow for an early detection of a possible
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later onset neuromuscular disease or some systemic disorder.
Declaration of interest: The authors report no conflict of interest. The authors alone are
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responsible for the content and writing of the paper
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muscle biopsy, although the clinical significance of the above findings remains elusive.
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REFERENCES 1. Morandi L, Angelini C, Prelle A, et al. High plasma creatine kinase : review of the literature and proposal fo a diagnostic algorithm. Neurol Sci. 2006;27:303-311.
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2. Wong ET, Cobb C, Humehara MK, et al. Heterogeneity of serum creatine kinase activity among racial and gender groups of the population. Am J Clin Pathol. 1983;79:582–586.
3. Fuglsang-Frederiksen A. The role of different EMG methods in evaluating myopathy.
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4. Lai CH, Melli G, Chang YJ, et al. Open muscle biopsy in suspected myopathy: diagnostic yield and clinical utility. Eur J Neurol. 2010;17:136-142.
5. Buchthal F, Clemmensen S. On the differentiation of muscle atrophy by
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electromyography. Acta Psychiatr Neurol. 1941;16:143–181.
6. Stewart CR, Nandedkar SD, Massey JM, Gilchrist JM, Barkhaus PE, Sanders DB.
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Evaluation of an automatic method of measuring features of motor unit action potentials. Muscle Nerve. 1989;12:141–148.
7. Mastaglia FL, Laing NG. Investigation of muscle disease. J Neurol Neurosurg Psych.
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1996;60:256-274.
8. Stalberg E, Falck B, Sonoo M, Stalberg S, Astrom M. Multi-MUP EMG analysis-a two year experience in daily clinical work. Electroencephalogr Clin Neurophysiol. 1995;97:145–154.
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Clin Neurophysiol. 2006;117:1173-1189.
9. Buchthal F. Electromyography in the evaluation of muscle diseases. Neurol Clin. 1985;3:573-598.
10. Katsavrias E, Primetis E, Karandreas N. Iliopsoas: a new electromyographic technique and normal motor unit action potential values. Clin Neurophysiol. 2005;116:2528-2532.
11. Prayson RA. Diagnostic yield associated with multiple simultaneous skeletal muscle biopsies. Am J Clin Pathol. 2006;126:843-848.
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12. Wilbourn AJ. The electrodiagnostic examination with myopathies. J Clin Neurophysiol. 1993;10:132-148. 13. Black JT, Bhatt GP, Dejesus PV, Schotland DL, Rowland LP. Diagnostic accuracy of
disease. A study of 105 cases. J Neurol Sci. 1974;21:59-70.
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clinical data, quantitative electromyography and histochemistry in neuromuscular
14. Prelle A, Tancredi L, Sciacco M, et al. Retrospective study of a large population of
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2002;249:305–311.
15. Uncini A, Lange DJ, Lovelace RE, Solomon M, Hays AP. Long-duration polyphasic motor unit potentials in myopathies: a quantitative study with pathological correlation.
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Muscle Nerve. 1990;13:263-267.
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patients effeceted with asymptomatic or paucisymptomatic hyperCKemia. J Neurol.
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Muscle biopsy
M
myopathic
F
44
normal
myopathic
3
M
49
myopathic
myopathic
4 5
M M
18 48
myopathic normal
myopathic myopathic
6
M
44
myopathic
myopathic
7
F
68
myopathic
myopathic
8
M
36
myopathic
myopathic
9
M
56
myopathic
myopathic
10 11
M F
54 28
normal myopathic
normal myopathic
12
M
42
myopathic
13 14
M M
25 23
15 16
M M
22 38
mitochondrial dysfunction inflammatory infiltrates myopathic changes normal myopathic changes mitochondrial dysfunction myopathic changes phosphorylase deficiency myopathic changes normal myopathic changes mitochondrial dysfunction normal myopathic changes normal mitochondrial dysfunction myopathic changes normal myopathic changes neurogenic
2
EP
1
myopathic
myopathic myopathic
myopathic myopathic
Normal normal
normal -
17
F
76
myopathic
-
18 19
F M
66 49
myopathic myopathic
myopathic
20
M
63
-
21
F
41
myopathic + neurogenic myopathic
22
M
36
neurogenic
-
23
M
23
24 25
M M
48 58
myopathic + neurogenic normal myopathic
myopathic + neurogenic myopathic
-
13
Final diagnosis Mitochondrial myopathy Inflammatory myopathy -
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QEMG
44
EMG (conventional) myopathic
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Age
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Sex
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Patient
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Table 1: Electrophysiologic findings, biopsy features and clinical diagnoses
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Mitochondrial myopathy McArdle’s disease -
Mitochondrial myopathy Mitochondrial myopathy -
myopathic changes myopathic changes normal
LGMD1C
normal myopathic changes
-
-
ST
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C
AC
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EMG: electromyogram, QEMG:quantitative EMG, LGMD1C: limb girdle muscular dystrophy 1C,
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abnormal muscle biopsy, QEMG and conventional EMG, respectively.
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EMG: Conventional Electromygraphy, QEMG: Quantitative Electromyography, BN: Muscle biopsy normal, BA: Muscle biopsy abnormal
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Figure 1: Categorized histogram shows the number of observations with normal and
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ST JU Figure 2: Histological findings from patient with McArdle’s disease showing subsarcolemmal vacuoles in many muscle fibers on hematoxylin-eosin (HE) staining (2a) with abnormal accumulation of glycogen on PAS staining (2b). 16
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C
AC
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Figure 3: Muscle biopsy from patient with LGMD1C revealing significantly reduced immunohistochemical expression of caveolin-3 in muscle fibers (3a) compared to control
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(3b).
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