- Email: [email protected]
Peripheral Neuropathy in Genetic Mitochondrial Diseases
Peripheral Neuropathy in Genetic Mitochondrial Diseases David E. Stickler, MD*, Edward Valenstein, MD*, Richard E. Neiberger, MD, PhD†, Leigh Ann Perkins, ARNP‡, Paul R. Carney, MD†, Jonathan J. Shuster, PhD§储, Douglas W. Theriaque, MS储, and Peter W. Stacpoole, PhD, MD‡储¶ Peripheral neuropathy is an underrecognized but common occurrence in genetic mitochondrial disorders. To gain insight into the frequency and clinical presentation of this complication, nerve conduction studies were performed on 43 subjects with congenital lactic acidosis enrolled in a controlled clinical trial of oral dichloroacetate. Median and peroneal motor conduction studies and median and sural sensory conduction studies were performed on each patient. The mean amplitude of the peroneal motor nerve (P < 0.001) and the conduction velocities of the median (P < 0.001) and peroneal (P < 0.001) motor nerves were uniformly lower in our subjects than in healthy literature control subjects. There were no significant differences in sensory nerve conduction studies. A generalized reduction in motor nerve conduction velocity was the dominant electrophysiological abnormality in the patients in this study and was independent of age, sex, or congenital mitochondrial disorder. We postulate that cellular energy failure is the most likely common cause of peripheral neuropathy in patients with genetic mitochondrial diseases, owing to the high demand for adenosine triphosphate via aerobic carbohydrate metabolism by nerve tissue. © 2006 by Elsevier Inc. All rights reserved. Stickler D, Valenstein E, Neiberger R, Perkins L, Carney P. Peripheral Neuropathy in Genetic Mitochondrial Diseases. Pediatr Neurol 2006;34:127-131.
from mutations in either mitochondrial or nuclear deoxyribonucleic acid. Neuropathy in these disorders is extremely variable and is typically observed in conjunction with more classic clinical manifestations, including, but not limited to, encephalopathy, myopathy, seizures, retinal degeneration, ophthalmoplegia, cardiomyopathy, ataxia, and myoclonus. The spectrum ranges from acute neuropathy, resembling Guillain-Barré syndrome [1], to chronic sensory-motor polyneuropathy, which is the most common clinical neuropathic manifestation of genetic mitochondrial diseases [2-5]. Nerve conduction studies have reported the presence of axonal [3,6,7], demyelinating [4,5,8-10], and mixed neuropathies [11-13]. Nerve biopsies have demonstrated features of both of axonal [2,7,1416] and demyelinating [2,8,14,17] processes, and mitochondrial structural abnormalities have been observed by electron microscopy [2,4,5,16,18]. Most descriptions of mitochondrial peripheral neuropathies derive from individual case reports or from small series. The present report describes the electrophysiologic examination of nerve conduction in 43 children with congenital causes of lactic acidosis to gain insight into the frequency and clinical presentation of this underrecognized but common complication of genetic mitochondrial disorders.
Materials and Methods Patients
Inborn errors of mitochondrial metabolism encompass a heterogeneous group of multisystem disorders that result
Subjects were enrolled in a controlled clinical trial of oral dichloroacetate conducted in the General Clinical Research Center at the University of Florida. A summary of the primary outcome of the clinical trial is in press [19]. The study was approved by the Health Science Center Institutional Review Board, and all patients or guardians gave informed consent or assent to participate.
From the *Departments of Neurology, †Pediatrics, ‡Medicine, § Biostatistics, ¶Biochemistry and Molecular Biology and 储the General Clinical Research Center, University of Florida, Gainesville, Florida. Abstract presented at the American Academy of Neurology annual meeting, San Francisco, California, April 2004.
Communications should be addressed to: Dr. Stickler; MUSC, 96 Jonathan Lucas St.; Clinical Services Building, Ste. 307; Charleston, SC 29425. E-mail: [email protected] Received April 12, 2005; accepted August 2, 2005.
Introduction
© 2006 by Elsevier Inc. All rights reserved. doi:10.1016/j.pediatrneurol.2005.08.006 ● 0887-8994/06/$—see front matter
Stickler et al: Neuropathy in Mitochondrial Diseases 127
Entry criteria included (1) biochemical or molecular genetic proof of a defect in the pyruvate dehydrogenase complex or in one or more complexes of the respiratory chain or a pathologic mutation in mitochondrial deoxyribonucleic acid; (2) any combination of at least three lactate concentrations measured over 1 month and within 6 months before entry into the study of at least 2 mmol/L (arterial blood) or 2.5 mmol/L (venous blood or cerebrospinal fluid), obtained at least 4 hours after the last meal, or an increase in venous blood lactate of at least 1 mmol/L, measured within 2 hours after a standardized high carbohydrate meal; and (3) ability to maintain a blood glucose concentration of at least 60 mg/dL after fasting for 4 hours (if less than 2 years of age) or 8 hours (if greater than 2 years of age). Biochemical proof of a deficiency in the activity of the pyruvate dehydrogenase complex or of one or more complexes of the respiratory chain or molecular genetic evidence of a mutation in mitochondrial deoxyribonucleic acid was made by noncommercial diagnostic laboratories in North America or Australia recognized as diagnostic referral centers for mitochondrial diseases. These findings were reviewed by the chairman and the vice-chairman of the Steering and Planning Committee for the trial and by the chairman of the Data and Safety Monitoring Board, and a unanimous decision about the appropriateness of the diagnosis was required for the subject to be eligible for enrollment. Patients with hyperlactatemia associated with proven biotinidase deficiency or with enzyme deficiencies of gluconeogenesis were excluded, as were patients with primary disorders of amino acid or fatty acid metabolism, malabsorption syndromes associated with d-lactic acidosis, renal insufficiency, or primary hepatic disease unrelated to congenital lactic acidosis.
Nerve Conduction Studies Baseline nerve conduction studies were performed before treatment with dichloroacetate or placebo, and the results of those are reported here. Nerve conduction studies were performed on a single upper and lower extremity of each patient, and median and peroneal motor conduction studies and median and sural sensory conduction studies were performed. A single technician performed all studies, and the results were interpreted by a single clinician (E.V.) at the time of testing. The median motor studies were performed with supramaximal stimulation at the wrist and elbow with the G1 electrode over the abductor pollicis brevis. The median sensory potential was recorded using ring electrodes placed on the index finger with stimulation at the wrist. The peroneal motor nerve conduction studies involved stimulation over the dorsum of the foot near the ankle, across the fibular head, and above the knee. The G1 electrode was placed on the extensor digitorum brevis. Sural sensory responses were measured with the recording electrode below and immediately behind the lateral malleolus with a stimulation distance of 100 mm on the mid-calf.
Statistical Analysis Patient data were compared with three separate groups of published sets of nerve conduction findings obtained in healthy children [20-22]. Patients were segregated chronologically according to identical age strata derived from literature sources. Patients whose age did not match the age range of a specific publication were excluded from comparison with the published values in that article. Each age stratum of a particular study was weighted according to the number of years in the stratum, irrespective of the number of patients entered. This procedure standardized the analysis to a uniform age distribution by age, namely the natural population pediatric age distribution in the general population. To control studywise error, we declared statistical significance only if all series produced a significant difference in the same direction. All P values are based upon two-sided stratified z tests.
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Results Nerve Conduction Table 1 summarizes diagnostic and clinical characteristics of the patients. The 21 males and 22 females ranged from 9 months to 19 years at the time of entry into the trial. The mean age was 5 ⫾ 5 years. There were 38 Caucasians, three Hispanics, and two Asians. The most common diagnosis was pyruvate dehydrogenase complex deficiency (11 patients). Twenty-five patients manifested one or more defects in respiratory chain complexes, and seven patients manifested congenital lactic acidosis caused by a point mutation in mitochondrial deoxyribonucleic acid. Patients were administered a standardized clinical neurologic examination that evaluated cognition, stance and gait when possible, muscle tone and strength, and reflexes. All patients had hypotonia, and all but two patients exhibited psychomotor delay. Cortical atrophy and structural brain abnormalities were observed by brain magnetic resonance imaging in the majority of subjects in whom this was performed. Because of cognitive disabilities or chronic muscle contractures, or both, in most patients, an adequate examination of the biceps, brachioradialis, patellar, and ankle deep tendon reflexes could be made in only two children with pyruvate dehydrogenase complex deficiency and 10 children with a respiratory chain defect, whereas examination of only some reflex groups was made in three other patients with pyruvate dehydrogenase complex deficiency. Most patients had no demonstrable deep tendon reflexes, whereas hyperreflexia with clonus was present in three children. The nerve conduction studies demonstrated abnormalities consistent with primary demyelination or mixed demyelinating and axonal changes in the majority of studies, with pure axonal changes present in only 4 of the 43 studies (Table 1). To evaluate the relationship between clinical finding of neuropathy and quantitative nerve conduction results, we compared means for amplitude and velocity parameters of the median motor, peroneal, and sensory nerves between the patients in whom complete tendon reflexes could be obtained and the other 30 children in our study. No significant differences were detected, using a two sample t test. Next, nerve conduction medians and ranges were examined within each clinical grouping (Absent, Normal, Hyperreflexive/clonus) in an attempt to evaluate potential trends. However, the number of subjects was small, and a similar range of electrical values was found for each of the subgroups. The stratified means between our patient data and normative values reported in the literature were compared. Parano et al. [22] examined 155 healthy children from Italy, age 7 days to 14 years, to establish normal pediatric nerve conduction data. When compared with these values, the nerve conduction velocities in our patients were uniformly slower. Motor, but not sensory, amplitudes were also significantly lower in our patients. Cai and Zhang [20]
Table 1.
Clinical characteristics
Diagnosis
Age Range (yr)
Sex (M/F)
Psychomotor Delay (Y/N)
Hypotonia (Y/N)
Seizures (Y/N)
Brain MRI (nl/abn/nd)
Nerve Conduction Results (ax/d/m/nl)
PDC Complex I Complex II Complex III Complex IV Complex I, IV Complex I, II, IV Complex I, III, IV Complex II, III, IV OXPHOS MERRF MELAS Composite
1-6 1-19 1-2 1-2 1-9 1-7 4 1
3/8 3/3 2/0 0/2 3/2 5/1 1/0 0/1
11/0 5/1 2/0 2/0 5/0 6/0 1/0 1/0
11/0 6/0 2/0 2/0 5/0 6/0 1/0 1/0
8/3 5/1 1/1 1/1 2/3 3/3 0/1 0/1
1/10/0 0/5/1 1/0/1 1/1/0 1/4/0 0/6/0 0/1/0 0/1/0
1/5/5/0 1/1/4/0 1/0/1/0 0/0/2/0 0/1/4/0 1/2/3/0 0/0/1/0 0/1/0/0
2
1/0
1/0
1/0
0/1
0/1/0
0/0/1/0
1 4 5-19 1-19
1/0 0/1 2/4 21/22
1/0 1/0 5/1 41/2
1/0 1/0 6/0 43/0
1/0 1/0 4/2 26/17
0/1/0 1/0/0 2/4/0 7/34/2
0/1/0/0 0/0/1/0 0/1/4/1 4/12/26/1
Abbreviations: abn ⫽ Abnormal ax ⫽ Axonal CNS ⫽ Central nervous system d ⫽ Demyelinating m ⫽ Mixed axonal and demyelinating MELAS ⫽ Mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes MERRF ⫽ Myoclonus epilepsy with ragged red fibers MRI ⫽ Magnetic resonance imaging n ⫽ Normal nd ⫽ Not done OXPHOS ⫽ Generalized decrease in respiratory chain enzyme activity PDC ⫽ Pyruvate dehydrogenase complex
studied 168 healthy Chinese individuals, age 24 hours to 30 years, to establish a set of normal values that were also compared with our findings (Table 2). Significant slowing of median and peroneal motor conduction velocities was observed in the patients compared with the normative data. In contrast, sural sensory and median sensory conduction velocities were higher in our patients compared with the results reported by Cai and Zhang [20]. The mean amplitude of the peroneal nerve was reduced in the patients, compared with the healthy children. Finally, we compared our data with findings recorded by Gamstorp [21] in 86 Table 2.
healthy infants from Sweden, age 1 day to 16 years, in whom ulnar, median, and peroneal motor conduction velocities were measured. Both median (50.9 ⫾ 1.0 m/s vs 60.1 ⫾ 0.8 m/s) and peroneal (42.9 ⫾ 1.2 m/s vs 57.0 ⫾ 1.0 m/s) motor conduction velocities were lower (both P ⬍ 0.001) in our patients. We determined which conduction velocity results obtained in our patients were significantly different in the same direction from all the literature data sets in which a particular measurement was obtained. Using this criterion, the mean amplitude of the peroneal motor nerve (P ⬍
Stratified means comparison of study patients vs literature
Measure Amplitude Median motor Median sensory Peroneal Sural Velocity Median motor Median sensory Peroneal Sural
Study Mean ⴞ S.E.
Parano Mean [22]
Cai [20]
8.4 ⫾ 0.9 mV 35.7 ⫾ 4.9 V 2.7 ⫾ 0.4 mV 31.7 ⫾ 6.8 V
11.4 ⫾ 0.7 mV* 26.0 ⫾ 3.6 V 7.5 ⫾ 0.6 mV† 24.8 ⫾ 0.9 V
7.7 ⫾ 0.3 mV 20.1 ⫾ 0.5 V† 7.0 ⫾ 0.2 (mV†) 18.5 ⫾ 0.6 V
48.4 ⫾ 1.8 m/s 49.2 ⫾ 1.7 m/s 42.0 ⫾ 1.5 m/s 44.3 ⫾ 1.8 m/s
55.0 ⫾ 0.5 m/s† 52.4 ⫾ 0.5 m/s 56.2 ⫾ 0.6 m/s† 53.4 ⫾ 0.6 m/s†
55.7 ⫾ 0.6 (m/s†) 41.5 ⫾ 0.5 (m/s†) 49.4 ⫾ 0.5 (m/s†) 39.2 ⫾ 0.7 (m/s†)
Data are expressed as mean ⫾ S.E. Statistically significant differences between study and literature results are: * P ⫽ 0.007; † P ⬍ 0.001. Other comparisons not significantly different (P ⬎ 0.05).
Stickler et al: Neuropathy in Mitochondrial Diseases 129
0.001) and the conduction velocities of the median (P ⬍ 0.001) and peroneal (P ⬍ 0.001) motor nerves were uniformly lower in our subjects than in the healthy literature control individuals. Thus, significant slowing of conduction velocity in one motor nerve predicted diffuse pathologic involvement. Discussion A generalized reduction in motor nerve conduction velocity was the dominant electrophysiologic abnormality in our patients and was independent of age, sex, or congenital mitochondrial disorder. Many of these individuals had central nervous system complications and moderate to severe neuromuscular degeneration, which made difficult or impossible a reliable evaluation of peripheral nervous system status by clinical examination alone. The findings presented here emphasize the extremely high frequency of peripheral nerve involvement in congenital lactic acidosis when evaluated by standard nerve conduction techniques, compared with the relatively poor sensitivity of clinical assessment. It is interesting that the conduction velocities of the median sensory and sural sensory nerves were increased when compared with those reported in Chinese children [20]. Furthermore, considerable variability was evident in values for lower extremity nerve conductions among the three normative data sets, which originated from geographically disparate regions (China, Italy, and Sweden). The method of recording nerve conduction was similar among the studies, and the patients in the present study most closely resembled the children examined by Parano et al. [22] and Gamstorp [21]. This finding may not be surprising, given the ethnic composition of the patients in this study. We found no literature that compared pediatric populations of different ethnic or racial backgrounds, although one study of peripheral nerve function in adults found no significant racial differences [23]. Thus, it is doubtful that ethnicity and race are important variables to consider when interpreting these results. No previous study of peripheral nerve function in patients with genetic mitochondrial diseases has compared nerve conduction findings in pediatric patients with the extent of normative data employed in the present analysis. However, prior reports of peripheral neuropathy in patients with pyruvate dehydrogenase complex deficiency [14,24], mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) [11], myoclonus epilepsy with ragged red fibers (MERRF) [3,7,15,25-27], and respiratory chain defects [8,17] are consistent with our findings. Limited published biopsy data include evidence of axonal degeneration [3,6,10,16,28,29], diminished size or numbers of myelinated fibers with onion bulb formations [8,16,27], and mitochondrial dysmorphism [30]. One limitation of the present study is that an adequate clinical assessment of peripheral nerve function was difficult or impossible to obtain in most subjects, due mainly
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to their psychomotor disabilities. Thus, clinical and electrical findings of peripheral nerve function in the study group could not readily be compared. This limitation further illustrates the unreliability of clinical examination alone in diagnosing peripheral neuropathy in young children severely compromised by mitochondrial neurodegenerative disorders. Another shortcoming of the study is that nerve biopsies were not obtained in any patient. The nearly uniform presence of diminished motor nerve conduction velocity is most consistent with a primary demyelinating pathogenesis. Whereas slight nerve conduction slowing can also be the result of severe axonal loss, only four of the 42 studies demonstrated pure axonal pathology [31]. Although the relative preservation of nerve conduction amplitudes suggests primary dysfunction of the myelin sheath, the absence of histologic confirmation and the potentially wide range of normal sensory amplitudes make a definitive diagnosis impossible. Despite these limitations, the results of this and previous reports demonstrate that peripheral neuropathy is a strikingly common and early manifestation of mitochondrial diseases, regardless of underlying genotype, and suggests that a common mechanism underlies this complication. The efficient conversion of substrate fuel (primarily carbohydrate) into energy requires the integrity of both nuclear and mitochondrial genomes, but this process is perturbed in patients with loss-of-function mutations in the pyruvate dehydrogenase complex or respiratory chain complexes [32,33]. The common pathologic consequences of these defects are the abnormal accumulation of lactate and protons and the inhibition of adenosine triphosphate production. In the case of mutations involving the electron transport system (complexes I-IV), accumulation of reactive oxygen species may also ensue and be a potential cause of cell damage [29,34,35]. We postulate that cellular energy failure is the most likely common cause of peripheral neuropathy in patients with pyruvate dehydrogenase complex or respiratory chain complex deficiency, regardless of how the neuropathy is manifested clinically or histologically. Although mitochondrial diseases may affect any cell, the most susceptible tissues are the nervous system, skeletal muscle, heart, and kidney cortex, which are extremely dependent on high rates of mitochondrial oxidative metabolism for energy and normal function. Lactic acidosis may be a surrogate marker for underlying mitochondrial energy failure [33], but it alone may not be sufficient to cause significant cell damage in the absence of a critical fall in adenosine triphosphate concentration. Myelination of central and peripheral nerve axons requires energy [36,37] and is inhibited under experimental conditions in which adenosine triphosphate synthesis is decreased [36]. In summary, objective electrophysiologic evaluation of nerve conduction discloses a high frequency of peripheral neuropathy among children (and probably adults) with genetic mitochondrial diseases that give rise to congenital forms of lactic acidosis. The resulting inhibition of oxida-
tive phosphorylation leads to pathologic accumulation of lactate, protons, and free radicals and reduced synthesis of adenosine triphosphate. To variable degrees, these biochemical events may impair myelination and other vital processes of peripheral nerve function. Quantitation of the amplitude and velocity of peripheral nerve conduction should be considered essential to a thorough neurologic evaluation of patients with mitochondrial diseases. This study was supported by grants FD-R-002013, RO1 ES007355, and MO1 RR00082. We thank the patients and their families for their participation, the staff of the General Clinical Research Center for its dedication, the pilots and staff of Mercy Medical Airlift and Angel Flight of Florida for facilitating patient transport, the physicians who referred patients for this study, and Ms. Candace L. Caputo and Lesa R. Gilbert, RN, for administrative assistance.
References [1] Coker S. Leigh Disease presenting as Guillain-Barré Syndrome. Pediatr Neurol 1993;9:61-3. [2] Bouillot S, Martin-Negrier ML, Vital A, et al. Peripheral neuropathy associated with mitochondrial disorders: 8 cases and review of the literature. J Peripher Nerv Syst 2002;7:213-20. [3] Chu CC, Huang CC, Fang W, Chu NS, Pang CY, Wei YH. Peripheral neuropathy in mitochondrial encephalopathies. Eur Neurol 1997;37:110-5. [4] Goebel HH, Bardosi A, Friede RL, Kohlschutter A, Albani M, Siemes H. Sural nerve biopsy in Leigh’s subacute necrotizing encephalomyelopathy. Muscle Nerve 1986;9:165-73. [5] Yiannikas C, McLeod J, Pollard J, Baverstock J. Peripheral neuropathy associated with mitochondrial myopathy. Ann Neurol 1986; 20:249-57. [6] Ciafaloni E, Ricci E, Servidei S, et al. Widespread tissue distribution of a tRNALeu(UUR) mutation in the mitochondrial DNA of a patient with MELAS syndrome. Neurology 1991;41:1663-5. [7] Naumann M, Kiefer R, Toyka KV, Sommer C, Seibel P, Reichmann H. Mitochondrial dysfunction with myoclonus epilepsy and ragged red fibers point mutation in nerve, muscle and adipose tissue of a patient with multiple symmetric lipomatosis. Muscle Nerve 1997;20: 833-9. [8] Mizusawa H, Watanabe M, Kanazawa I, et al. Familial mitochondrial myopathy associated with peripheral neuropathy: Partial deficiencies of complex I and IV. J Neurol Sci 1988;86:171-84. [9] Moosa A. Motor nerve conduction velocities in Leigh’s encephalomyelopathy. Arch Dis Child 1978;53:62-5. [10] Rusanen H, Majamaa K, Tolonen U, Remes AM, Myllyla R, Hassinen IE. Demyelinating polyneuropathy in a patient with the tRNALeu(UUR) mutation at the base pair 3243 of the mitochondrial DNA. Neurology 1995;45:1188-92. [11] Kärppä M, Syrjälä P, Tolonen U, Majamaa K. Peripheral neuropathy in patients with the 3243 A⬎G mutation in mitochondrial DNA. J Neurol 2003;250:216-21. [12] Peyronnard J, Charron L, Bellavance A, Marchand L. Neuropathy and mitochondrial myopathy. Ann Neurol 1980;7:262-8. [13] Pezeshkpour G, Krarup C, Buchthal F, DiMauro S, Bresolin N, McBurney J. Peripheral neuropathy in mitochondrial disease. J Neurol Sci 1987;77:285-304. [14] Bonne G, Benelli C, De Meirleir L, et al. E1 pyruvate dehydrogenase deficiency in a child with motor neuropathy. Pediatr Res 1990;33:284-8. [15] Byrne E, Dennett X, Trounce I, Burdon J. Mitochondrial myoneuropathy with respiratory failure and myoclonic epilepsy. J Neurol Sci 1985;71:273-81. [16] Huang CC, Chu CC, Pang CY, Wei YH. Tissue mosaicism in
the skeletal muscle and sural nerve biopsies in the MELAS syndrome. Acta Neurol Scand 1999;99:125-9. [17] Shepherd IM, Birch-Machin MA, Johnson MA, et al. Cytochrome oxidase deficiency: Immunological studies of skeletal muscle mitochondrial fractions. J Neurol Sci 1988;87:265-74. [18] Schröder J, Sommer C. Mitochondrial abnormalities in human sural nerves: Fine structural evaluation of cases with mitochondrial myopathy, hereditary and non-hereditary neuropathies, and a review of the literature. Acta Neuropathol 1991;82:471-82. [19] Stacpoole PW, Kerr DS, Barnes C, et al. A controlled clinical trial of dichloroacetate for treatment of congenital lactic acidosis in children. Pediatrics. In press. [20] Cai F, Zhang J. Study of nerve conduction and late responses in normal Chinese infants, children and adults. J Child Neurol 1997;12: 13-8. [21] Gamstorp I. Normal conduction velocity of ulnar, median and peroneal nerves in infancy, childhood and adolescence. Acta Paediatr 1963;146:68-76. [22] Parano E, Uncini A, DeVivo D, Lovelace R. Electrophysiologic correlates of peripheral nervous system maturation in infancy and childhood. J Child Neurol 1993;8:336-8. [23] Buschbacher RM, Koch J. Race effect on nerve conduction studies: A comparison between 50 blacks and 50 whites. Arch Phys Med Rehabil 1999;80:536-9. [24] Chabrol B, Mancini J, Benelli C, Gire C, Munnich A. Leigh syndrome: Pyruvate dehydrogenase defect. A case with peripheral neuropathy. J Child Neurol 1994;9:52-5. [25] Arenas J, Campos Y, Bornstein B, et al. A double mutation (A8296G and G8353A) in the mitochondrial DNA tRNALys gene associated with myoclonus epilepsy with ragged-red fibers. Neurology 1999;52:377-82. [26] Calabresi P, Silvestri G, DiMauro S, Griggs R. Ekbom’s syndrome: Lipomas, ataxia, and neuropathy with MEERF. Muscle Nerve 1994;17:943-5. [27] Nagashima T, Kato H, Maguchi S, et al. A mitochondrial encephalo-myo-neuropathy with a nucleotide position 3271 (T-C) point mutation in the mitochondrial DNA. Neuromuscul Disord 2001;11: 470-6. [28] Barak Y, Arnon S, Wolach B, et al. MELAS syndrome: Peripheral neuropathy and cytochrome c-oxidase deficiency: A case report and review of the literature. Isr J Med Sci 1995;31:224-9. [29] Chang SH, Garcia J, Melendez JA, Kilberg M, Agarwal A. Haem oxygenase 1 gene induction by glucose deprivation is mediated by reactive oxygen species via the mitochondrial electron transport chain. Biochem J 2003;371:877-85. [30] Schröder J. Neuropathy associated with mitochondrial disorders. Brain Pathol 1993;3:177-90. [31] Logigian EL, Kelley JJ, Adelman LS. Nerve conduction and biopsy correlation in over 100 consecutive patients with suspected polyneuropathy. Muscle Nerve 1994;17:1010-20. [32] Enns GM. The contribution of mitochondria to common disorders. Mol Genet Metab 2003;80:11-26. [33] Stacpoole PW. Lactic acidosis and other mitochondrial disorders. Metabolism 1997;46:306-21. [34] Ischiropoulos H, Beckman JS. Oxidative stress and nitration in neurodegeneration: Cause, effect, or association? J Clin Invest 2003;11: 163-9. [35] Scortegagna M, Dinsg K, Oktay Y, et al. Multiple organ pathology, metabolic abnormalities and impaired homeostasis of reactive oxygen species in Epas 1-/- mice. Nat Genet 2003;35:331-40. [36] Fern R, Davis P, Waxman SG, Ransom BR. Axon conduction and survival in CNS white matter during energy deprivation: A developmental study. J Neurophysiol 1998;79:95-105. [37] Zhou C-J, Inagaki N, Pleasure SJ, Zhao L-X, Kikuyama S, Shioda S. ATP-binding in the developing spinal cord and PNS during myelination. J Comp Neurol 2002;451:334-45.
Stickler et al: Neuropathy in Mitochondrial Diseases 131
from mutations in either mitochondrial or nuclear deoxyribonucleic acid. Neuropathy in these disorders is extremely variable and is typically observed in conjunction with more classic clinical manifestations, including, but not limited to, encephalopathy, myopathy, seizures, retinal degeneration, ophthalmoplegia, cardiomyopathy, ataxia, and myoclonus. The spectrum ranges from acute neuropathy, resembling Guillain-Barré syndrome [1], to chronic sensory-motor polyneuropathy, which is the most common clinical neuropathic manifestation of genetic mitochondrial diseases [2-5]. Nerve conduction studies have reported the presence of axonal [3,6,7], demyelinating [4,5,8-10], and mixed neuropathies [11-13]. Nerve biopsies have demonstrated features of both of axonal [2,7,1416] and demyelinating [2,8,14,17] processes, and mitochondrial structural abnormalities have been observed by electron microscopy [2,4,5,16,18]. Most descriptions of mitochondrial peripheral neuropathies derive from individual case reports or from small series. The present report describes the electrophysiologic examination of nerve conduction in 43 children with congenital causes of lactic acidosis to gain insight into the frequency and clinical presentation of this underrecognized but common complication of genetic mitochondrial disorders.
Materials and Methods Patients
Inborn errors of mitochondrial metabolism encompass a heterogeneous group of multisystem disorders that result
Subjects were enrolled in a controlled clinical trial of oral dichloroacetate conducted in the General Clinical Research Center at the University of Florida. A summary of the primary outcome of the clinical trial is in press [19]. The study was approved by the Health Science Center Institutional Review Board, and all patients or guardians gave informed consent or assent to participate.
From the *Departments of Neurology, †Pediatrics, ‡Medicine, § Biostatistics, ¶Biochemistry and Molecular Biology and 储the General Clinical Research Center, University of Florida, Gainesville, Florida. Abstract presented at the American Academy of Neurology annual meeting, San Francisco, California, April 2004.
Communications should be addressed to: Dr. Stickler; MUSC, 96 Jonathan Lucas St.; Clinical Services Building, Ste. 307; Charleston, SC 29425. E-mail: [email protected] Received April 12, 2005; accepted August 2, 2005.
Introduction
© 2006 by Elsevier Inc. All rights reserved. doi:10.1016/j.pediatrneurol.2005.08.006 ● 0887-8994/06/$—see front matter
Stickler et al: Neuropathy in Mitochondrial Diseases 127
Entry criteria included (1) biochemical or molecular genetic proof of a defect in the pyruvate dehydrogenase complex or in one or more complexes of the respiratory chain or a pathologic mutation in mitochondrial deoxyribonucleic acid; (2) any combination of at least three lactate concentrations measured over 1 month and within 6 months before entry into the study of at least 2 mmol/L (arterial blood) or 2.5 mmol/L (venous blood or cerebrospinal fluid), obtained at least 4 hours after the last meal, or an increase in venous blood lactate of at least 1 mmol/L, measured within 2 hours after a standardized high carbohydrate meal; and (3) ability to maintain a blood glucose concentration of at least 60 mg/dL after fasting for 4 hours (if less than 2 years of age) or 8 hours (if greater than 2 years of age). Biochemical proof of a deficiency in the activity of the pyruvate dehydrogenase complex or of one or more complexes of the respiratory chain or molecular genetic evidence of a mutation in mitochondrial deoxyribonucleic acid was made by noncommercial diagnostic laboratories in North America or Australia recognized as diagnostic referral centers for mitochondrial diseases. These findings were reviewed by the chairman and the vice-chairman of the Steering and Planning Committee for the trial and by the chairman of the Data and Safety Monitoring Board, and a unanimous decision about the appropriateness of the diagnosis was required for the subject to be eligible for enrollment. Patients with hyperlactatemia associated with proven biotinidase deficiency or with enzyme deficiencies of gluconeogenesis were excluded, as were patients with primary disorders of amino acid or fatty acid metabolism, malabsorption syndromes associated with d-lactic acidosis, renal insufficiency, or primary hepatic disease unrelated to congenital lactic acidosis.
Nerve Conduction Studies Baseline nerve conduction studies were performed before treatment with dichloroacetate or placebo, and the results of those are reported here. Nerve conduction studies were performed on a single upper and lower extremity of each patient, and median and peroneal motor conduction studies and median and sural sensory conduction studies were performed. A single technician performed all studies, and the results were interpreted by a single clinician (E.V.) at the time of testing. The median motor studies were performed with supramaximal stimulation at the wrist and elbow with the G1 electrode over the abductor pollicis brevis. The median sensory potential was recorded using ring electrodes placed on the index finger with stimulation at the wrist. The peroneal motor nerve conduction studies involved stimulation over the dorsum of the foot near the ankle, across the fibular head, and above the knee. The G1 electrode was placed on the extensor digitorum brevis. Sural sensory responses were measured with the recording electrode below and immediately behind the lateral malleolus with a stimulation distance of 100 mm on the mid-calf.
Statistical Analysis Patient data were compared with three separate groups of published sets of nerve conduction findings obtained in healthy children [20-22]. Patients were segregated chronologically according to identical age strata derived from literature sources. Patients whose age did not match the age range of a specific publication were excluded from comparison with the published values in that article. Each age stratum of a particular study was weighted according to the number of years in the stratum, irrespective of the number of patients entered. This procedure standardized the analysis to a uniform age distribution by age, namely the natural population pediatric age distribution in the general population. To control studywise error, we declared statistical significance only if all series produced a significant difference in the same direction. All P values are based upon two-sided stratified z tests.
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Results Nerve Conduction Table 1 summarizes diagnostic and clinical characteristics of the patients. The 21 males and 22 females ranged from 9 months to 19 years at the time of entry into the trial. The mean age was 5 ⫾ 5 years. There were 38 Caucasians, three Hispanics, and two Asians. The most common diagnosis was pyruvate dehydrogenase complex deficiency (11 patients). Twenty-five patients manifested one or more defects in respiratory chain complexes, and seven patients manifested congenital lactic acidosis caused by a point mutation in mitochondrial deoxyribonucleic acid. Patients were administered a standardized clinical neurologic examination that evaluated cognition, stance and gait when possible, muscle tone and strength, and reflexes. All patients had hypotonia, and all but two patients exhibited psychomotor delay. Cortical atrophy and structural brain abnormalities were observed by brain magnetic resonance imaging in the majority of subjects in whom this was performed. Because of cognitive disabilities or chronic muscle contractures, or both, in most patients, an adequate examination of the biceps, brachioradialis, patellar, and ankle deep tendon reflexes could be made in only two children with pyruvate dehydrogenase complex deficiency and 10 children with a respiratory chain defect, whereas examination of only some reflex groups was made in three other patients with pyruvate dehydrogenase complex deficiency. Most patients had no demonstrable deep tendon reflexes, whereas hyperreflexia with clonus was present in three children. The nerve conduction studies demonstrated abnormalities consistent with primary demyelination or mixed demyelinating and axonal changes in the majority of studies, with pure axonal changes present in only 4 of the 43 studies (Table 1). To evaluate the relationship between clinical finding of neuropathy and quantitative nerve conduction results, we compared means for amplitude and velocity parameters of the median motor, peroneal, and sensory nerves between the patients in whom complete tendon reflexes could be obtained and the other 30 children in our study. No significant differences were detected, using a two sample t test. Next, nerve conduction medians and ranges were examined within each clinical grouping (Absent, Normal, Hyperreflexive/clonus) in an attempt to evaluate potential trends. However, the number of subjects was small, and a similar range of electrical values was found for each of the subgroups. The stratified means between our patient data and normative values reported in the literature were compared. Parano et al. [22] examined 155 healthy children from Italy, age 7 days to 14 years, to establish normal pediatric nerve conduction data. When compared with these values, the nerve conduction velocities in our patients were uniformly slower. Motor, but not sensory, amplitudes were also significantly lower in our patients. Cai and Zhang [20]
Table 1.
Clinical characteristics
Diagnosis
Age Range (yr)
Sex (M/F)
Psychomotor Delay (Y/N)
Hypotonia (Y/N)
Seizures (Y/N)
Brain MRI (nl/abn/nd)
Nerve Conduction Results (ax/d/m/nl)
PDC Complex I Complex II Complex III Complex IV Complex I, IV Complex I, II, IV Complex I, III, IV Complex II, III, IV OXPHOS MERRF MELAS Composite
1-6 1-19 1-2 1-2 1-9 1-7 4 1
3/8 3/3 2/0 0/2 3/2 5/1 1/0 0/1
11/0 5/1 2/0 2/0 5/0 6/0 1/0 1/0
11/0 6/0 2/0 2/0 5/0 6/0 1/0 1/0
8/3 5/1 1/1 1/1 2/3 3/3 0/1 0/1
1/10/0 0/5/1 1/0/1 1/1/0 1/4/0 0/6/0 0/1/0 0/1/0
1/5/5/0 1/1/4/0 1/0/1/0 0/0/2/0 0/1/4/0 1/2/3/0 0/0/1/0 0/1/0/0
2
1/0
1/0
1/0
0/1
0/1/0
0/0/1/0
1 4 5-19 1-19
1/0 0/1 2/4 21/22
1/0 1/0 5/1 41/2
1/0 1/0 6/0 43/0
1/0 1/0 4/2 26/17
0/1/0 1/0/0 2/4/0 7/34/2
0/1/0/0 0/0/1/0 0/1/4/1 4/12/26/1
Abbreviations: abn ⫽ Abnormal ax ⫽ Axonal CNS ⫽ Central nervous system d ⫽ Demyelinating m ⫽ Mixed axonal and demyelinating MELAS ⫽ Mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes MERRF ⫽ Myoclonus epilepsy with ragged red fibers MRI ⫽ Magnetic resonance imaging n ⫽ Normal nd ⫽ Not done OXPHOS ⫽ Generalized decrease in respiratory chain enzyme activity PDC ⫽ Pyruvate dehydrogenase complex
studied 168 healthy Chinese individuals, age 24 hours to 30 years, to establish a set of normal values that were also compared with our findings (Table 2). Significant slowing of median and peroneal motor conduction velocities was observed in the patients compared with the normative data. In contrast, sural sensory and median sensory conduction velocities were higher in our patients compared with the results reported by Cai and Zhang [20]. The mean amplitude of the peroneal nerve was reduced in the patients, compared with the healthy children. Finally, we compared our data with findings recorded by Gamstorp [21] in 86 Table 2.
healthy infants from Sweden, age 1 day to 16 years, in whom ulnar, median, and peroneal motor conduction velocities were measured. Both median (50.9 ⫾ 1.0 m/s vs 60.1 ⫾ 0.8 m/s) and peroneal (42.9 ⫾ 1.2 m/s vs 57.0 ⫾ 1.0 m/s) motor conduction velocities were lower (both P ⬍ 0.001) in our patients. We determined which conduction velocity results obtained in our patients were significantly different in the same direction from all the literature data sets in which a particular measurement was obtained. Using this criterion, the mean amplitude of the peroneal motor nerve (P ⬍
Stratified means comparison of study patients vs literature
Measure Amplitude Median motor Median sensory Peroneal Sural Velocity Median motor Median sensory Peroneal Sural
Study Mean ⴞ S.E.
Parano Mean [22]
Cai [20]
8.4 ⫾ 0.9 mV 35.7 ⫾ 4.9 V 2.7 ⫾ 0.4 mV 31.7 ⫾ 6.8 V
11.4 ⫾ 0.7 mV* 26.0 ⫾ 3.6 V 7.5 ⫾ 0.6 mV† 24.8 ⫾ 0.9 V
7.7 ⫾ 0.3 mV 20.1 ⫾ 0.5 V† 7.0 ⫾ 0.2 (mV†) 18.5 ⫾ 0.6 V
48.4 ⫾ 1.8 m/s 49.2 ⫾ 1.7 m/s 42.0 ⫾ 1.5 m/s 44.3 ⫾ 1.8 m/s
55.0 ⫾ 0.5 m/s† 52.4 ⫾ 0.5 m/s 56.2 ⫾ 0.6 m/s† 53.4 ⫾ 0.6 m/s†
55.7 ⫾ 0.6 (m/s†) 41.5 ⫾ 0.5 (m/s†) 49.4 ⫾ 0.5 (m/s†) 39.2 ⫾ 0.7 (m/s†)
Data are expressed as mean ⫾ S.E. Statistically significant differences between study and literature results are: * P ⫽ 0.007; † P ⬍ 0.001. Other comparisons not significantly different (P ⬎ 0.05).
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0.001) and the conduction velocities of the median (P ⬍ 0.001) and peroneal (P ⬍ 0.001) motor nerves were uniformly lower in our subjects than in the healthy literature control individuals. Thus, significant slowing of conduction velocity in one motor nerve predicted diffuse pathologic involvement. Discussion A generalized reduction in motor nerve conduction velocity was the dominant electrophysiologic abnormality in our patients and was independent of age, sex, or congenital mitochondrial disorder. Many of these individuals had central nervous system complications and moderate to severe neuromuscular degeneration, which made difficult or impossible a reliable evaluation of peripheral nervous system status by clinical examination alone. The findings presented here emphasize the extremely high frequency of peripheral nerve involvement in congenital lactic acidosis when evaluated by standard nerve conduction techniques, compared with the relatively poor sensitivity of clinical assessment. It is interesting that the conduction velocities of the median sensory and sural sensory nerves were increased when compared with those reported in Chinese children [20]. Furthermore, considerable variability was evident in values for lower extremity nerve conductions among the three normative data sets, which originated from geographically disparate regions (China, Italy, and Sweden). The method of recording nerve conduction was similar among the studies, and the patients in the present study most closely resembled the children examined by Parano et al. [22] and Gamstorp [21]. This finding may not be surprising, given the ethnic composition of the patients in this study. We found no literature that compared pediatric populations of different ethnic or racial backgrounds, although one study of peripheral nerve function in adults found no significant racial differences [23]. Thus, it is doubtful that ethnicity and race are important variables to consider when interpreting these results. No previous study of peripheral nerve function in patients with genetic mitochondrial diseases has compared nerve conduction findings in pediatric patients with the extent of normative data employed in the present analysis. However, prior reports of peripheral neuropathy in patients with pyruvate dehydrogenase complex deficiency [14,24], mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) [11], myoclonus epilepsy with ragged red fibers (MERRF) [3,7,15,25-27], and respiratory chain defects [8,17] are consistent with our findings. Limited published biopsy data include evidence of axonal degeneration [3,6,10,16,28,29], diminished size or numbers of myelinated fibers with onion bulb formations [8,16,27], and mitochondrial dysmorphism [30]. One limitation of the present study is that an adequate clinical assessment of peripheral nerve function was difficult or impossible to obtain in most subjects, due mainly
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to their psychomotor disabilities. Thus, clinical and electrical findings of peripheral nerve function in the study group could not readily be compared. This limitation further illustrates the unreliability of clinical examination alone in diagnosing peripheral neuropathy in young children severely compromised by mitochondrial neurodegenerative disorders. Another shortcoming of the study is that nerve biopsies were not obtained in any patient. The nearly uniform presence of diminished motor nerve conduction velocity is most consistent with a primary demyelinating pathogenesis. Whereas slight nerve conduction slowing can also be the result of severe axonal loss, only four of the 42 studies demonstrated pure axonal pathology [31]. Although the relative preservation of nerve conduction amplitudes suggests primary dysfunction of the myelin sheath, the absence of histologic confirmation and the potentially wide range of normal sensory amplitudes make a definitive diagnosis impossible. Despite these limitations, the results of this and previous reports demonstrate that peripheral neuropathy is a strikingly common and early manifestation of mitochondrial diseases, regardless of underlying genotype, and suggests that a common mechanism underlies this complication. The efficient conversion of substrate fuel (primarily carbohydrate) into energy requires the integrity of both nuclear and mitochondrial genomes, but this process is perturbed in patients with loss-of-function mutations in the pyruvate dehydrogenase complex or respiratory chain complexes [32,33]. The common pathologic consequences of these defects are the abnormal accumulation of lactate and protons and the inhibition of adenosine triphosphate production. In the case of mutations involving the electron transport system (complexes I-IV), accumulation of reactive oxygen species may also ensue and be a potential cause of cell damage [29,34,35]. We postulate that cellular energy failure is the most likely common cause of peripheral neuropathy in patients with pyruvate dehydrogenase complex or respiratory chain complex deficiency, regardless of how the neuropathy is manifested clinically or histologically. Although mitochondrial diseases may affect any cell, the most susceptible tissues are the nervous system, skeletal muscle, heart, and kidney cortex, which are extremely dependent on high rates of mitochondrial oxidative metabolism for energy and normal function. Lactic acidosis may be a surrogate marker for underlying mitochondrial energy failure [33], but it alone may not be sufficient to cause significant cell damage in the absence of a critical fall in adenosine triphosphate concentration. Myelination of central and peripheral nerve axons requires energy [36,37] and is inhibited under experimental conditions in which adenosine triphosphate synthesis is decreased [36]. In summary, objective electrophysiologic evaluation of nerve conduction discloses a high frequency of peripheral neuropathy among children (and probably adults) with genetic mitochondrial diseases that give rise to congenital forms of lactic acidosis. The resulting inhibition of oxida-
tive phosphorylation leads to pathologic accumulation of lactate, protons, and free radicals and reduced synthesis of adenosine triphosphate. To variable degrees, these biochemical events may impair myelination and other vital processes of peripheral nerve function. Quantitation of the amplitude and velocity of peripheral nerve conduction should be considered essential to a thorough neurologic evaluation of patients with mitochondrial diseases. This study was supported by grants FD-R-002013, RO1 ES007355, and MO1 RR00082. We thank the patients and their families for their participation, the staff of the General Clinical Research Center for its dedication, the pilots and staff of Mercy Medical Airlift and Angel Flight of Florida for facilitating patient transport, the physicians who referred patients for this study, and Ms. Candace L. Caputo and Lesa R. Gilbert, RN, for administrative assistance.
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