Cytochrome c Oxidase Deficiency and Long-Chain Acyl Coenzyme A Dehydrogenase Deficiency with Leigh’s Subacute Necro tizing Encephalomyelopathy Heinz Reichmann,* Helmut Scheel,? Bert Bier,$ Uwe-Peter Ketelsen,§ and Siegfried Zabranskyt
A female infant was seen at the age of 2 months because of hypotonia, delayed motor development, and lactic acidosis, and she died at age 13 months due to respiratory failure. In a muscle specimen taken at 11 months and in a liver specimen obtained 1.5 hours postmortem, we found decreased activities of cytochrome c oxidase and longchain acyl coenzyme A dehydrogenase. Neuropathological changes were typical for Leigh’s subacute necrotizing encephalomyelopathy. To our knowledge, this is the first report of a combined defect of complex IV of the respiratory chain and of the long-chain specific acyl coenzyme A dehydrogenase of @-oxidation in muscle and liver. Reichmann H , Scheel H, Bier B, Ketelsen U-P, Zabransky S. Cytochrome c oxidase deficiency and long-chain acyl coenzyme A dehydrogenase deficiency with Leigh’s subacute necrotizing encephalomyelopathy. Ann Neurol 1992;31:107-109
Since the first description of mitochondrial myopathy {l], a body of literature has emerged concerning defects of mitochondrial energy metabolism (for review, see [2-4]), mainly due to defects of the respiratory chain. Because the brain is often also affected by the metabolic defect, the term mitochondrial encephalomyopathy was introduced by Shapira and colleagues [ S ] (for review, see [6,7)).Leigh’s disease, an encephalomyelopathy is often associated with cytochrome c oxidase (COX) deficiency {S-1 11. Although COX activity is decreased in various tissues, there are reports of
From the *Department of Neurology, University of Wiirzburg, Wurzburg; Departments of ?Pediatrics and $Pathology, University of Homburg/Saar, Homburg; and SDeparrment of Pediatrics, University of Freiburg, Freiburg, Germany. Received Mar 29, 1991, and in revised form Jun 24 and Jul 26. Accepted for publication Jul 27, 1991. Address correspondence ro Dr Reichmann, Professor of Neurology, Department of Neurology, University of Wiirzburg, Josef-Schneider-Strasse 11, D-8700 Wurzburg, Germany.
patients with normal COX activity in liver [27, contrasting with decreased activity in muscle and other tissues. In the present report, we describe a patient with a biochemical defect of the respiratory chain and of (3-oxidation presenting as Leigh’s disease.
Patient Report Our patient was the first child of healthy nonconsanguineous parents. Pregnancy and delivery were uneventful; birth weight was 3,080 gm. At the age of 7 months, she was admitted because of petechial hemorrhages on her face and trunk. She had rhinitis and a mild chronic bronchitis. She was lethargic and showed motor retardation (she had no head control and could not roll over from back to front or sit without support); she had atrophy in the lumbar and gluteal region. Deep tendon reflexes were increased in the lower extremities, where brief rnyoclonus could also be induced. Metabolic acidosis (pH 7.25) with respiratory compensation was found. Lactate was up to 5.22 mmol/L, pyruvate to 240 nmoliL, and alanine to 60 fmol/L. After an oral glucose tolerance test, there was excessive rise in lactate, pyruvate, and alanine. Blood triglycerides were increased to 480 m g q with a rise of very low density lipoproteins. Screening for amino acids, ketone bodies, reducing substances, and mucopolysaccharides was negative. A slightly elevated excretion of ethylrnaIonic, oxalic, adipic, and glutaric acids was found. Total serum carnitine levels were normal; however, acylcarnitines were increased and free carnitine was decreased. Ammonium, ascorbic acid, transaminases, thyroid hormones, cross-reacting protein; rheumatoid factor, coagulation status, thrombocyte function, urinary steroid excretion, and immunoglobulins were normal. Urinalyses revealed erythrocyturia. Cranial computed tomography showed moderate enlargement of the ventricles and cisterns. Electroencephalogram (EEG) and electromyogram were normal. Liver and kidneys were slightly enlarged. The renal scintigram was normal. Slight hypertrophy of the septum and left ventricular wall was observed. A skin biopsy indicated vasculitis without evidence of connective tissue disorder. In spite of supplementation of thiamine and biotin, lactic acidosis had not improved when the patient was seen at 11 months for a muscle biopsy (musculus quadriceps femoris). Motor function was not improved. Myoclonic seizures were observed. The EEG showed multifocal epileptic discharges. When myoclonic seizures occurred daily, valproate was started, whereupon the seizure activity in the EEG quickly subsided, and the attacks occurred only infrequently. Transaminases remained within normal levels. Methylene blue was administered intravenously and carnitine, ascorbic acid, and vitamin K, were given orally without any noticeable biochemical or clinical improvement. At 13 months, her general condition unexpectedly deteriorated. Respiratory insufficiency worsened, the acidosis became intractable, and she died within a few hours. One-half hour post mortem, specimens of liver and of the abdominal musculature were obtained. Postmortem examination of the brain revealed macroscopically a slight degree of polymicrogyria, extensive gliosis and reduction in the number of ganglion cells, pronounced capillary proliferation in the paraventricular and brainstem regions, severe demyelination in the
Copyright 0 1992 by the American Neurological Association
107
anomalies or focal myofibrillar degeneration. The matrix of most mitochondria was condensed, but showed no pathological inclusions. A few scattered mitochondria had marginal lipid droplets. The sarcoplasmic reticulum system was normal. The glycogen content was slightly increased; there were no lysosomal glycogen accumulations and no sarcoplasmic or nuclear inclusions. In contrast, electron microscopic examinations performed on postmortem tissue showed ultrastructural changes typical of mitochondriopathy. The liver cells were enlarged and contained abnormal mitochondria with lipid inclusions and irregular cristae. Muscle mitochondria displayed gross variation in size and shape and also contained lipid inclusions.
capsula interna, capillary calcification in the brainstem ganglia, and severe gliosis in the nuclei of the pons and medulla oblongata, wasting of the myelin sheaths, and capillary proliferation. There was hepatomegaly with centrolobular, finedroplet fatty infiltr,ition and a slight degree of muscle fiber atrophy.
Materials and Methods Routine histochemical methods were applied as outlined in Dubowitz and Brooke [ 121. Muscle was homogenized as previously described [13]. Activity of enzymes of the respiratory chain was analyzed as described [ 141; activity measurements of all enzymes from 8-oxidation were performed as previously described [I?;, 15, 161. Control values were obtained from biopsies that finally were shown to not represent a neurornuscular disorder. Liver biopsies were obrained from autopsies within the first 2 hours post mortem.
Enzymes of the Respiratory Chain The results of the biochemical investigation are listed in Tables 1 and 2. T h e activity of cytochrome c oxitlase was markedly decreased both in the muscle biopsy (19% of normal) and in the liver specimen obtained postmortem (7% of normal). The activities of complex I, 11, and 111 and of citrate synthase, a key enzyme of the citric acid cycle, were normal, and histochemical investigations revealed a normal stain for succinate Jehydrogenase and a normal adenosine triphosphatase reaction. T h e carnitine content was low normal. In the liver specimen, the activities of complex I, 11, and I11
Results Hzj.tologicul und El'ectron Mirro.tcopicul Findings The histological and electron microscopical investigations of the muscle biopsy obtained at 11 months showed no signs of fiber atrophy or of necrosis, no inflammatory infiltration, and no changes in the blood vessels. The distribution of type I and type I1 fibers was normal. Oil red 0 staining showed a slight increase of intracellular neutral fat droplets. Gomori trichrome stain showed no ragged-red fibers. Electron microscopy showed no evidence of rnyofibrillar structure Table 1 Biothemiral Analyses of Enzymes ofthe Respiratory Chain Enzyme
Muscle
Muscle Controls (n)
Liver ~
0.53 42.1 1.84 2.63 1.11 5.6
Cytochrome c oxidase N A D H dehydrogenase NADH-cytochrome c reductase Succinate dehydrogenase Succinate cytochrome c reductase citrate synthetase
2.8 t 0.9 (113) 4 8 k 9.4 (121) 3.2 t 1.5 (98) 2.4 1.1 (160) 1.64 ? 0.6 (125) 5.3 t 2.1 (49)
Liver Controls ( n ) ~~
~
0.5 73.4 15.7 3.2 2.4 ...
*
-
~
~~
5 t 1.8 ( 4 ) 101 t 32 (4) 1.3.1 t 1.7 ( 4 ) lr.7 ? 1.8 (4) 3.1 ? 1.0 (4) ...
Values of the enzyme activities are the mean f SD and arc expressed in units per gram of tissue ( I unir = 1 pmol ofsiibstr~tteturnowrimin). Muscle was obtained at 11 months; liver was obtained 2.5 hours post mortem. N A D H = reduced nicotinamide adenine dinucleotide.
Table 2. Enqme.r of ,&Oxidation Enzyme
Muscle
Muscle Controls ( n )
Liver
Liver Controls (n
Palmitoyl CoA DH Octanoyl CoA DH Butyryl CoA DH Enoyl CoA hydratase Hydroxyacyl CoA DH Thiolase C 10 Ketone body metabolism Thiolase C4
0.29 0.54 1.0 8.1 10.6 2.11
1.02 0.36 (65) 0.83 ? 0.31 (52) 0.84 ? 0.33 (63) 13.3 k 3.1 (41) 12.6 ? 3.43 (58) 2.12 t 1.47 (44)
0 0.82 0.72 12.6 26.6 12.31
1.52
1.66
4.66 2 2.38 (54)
4.55
*
=
3)
? 0.43 0.9 2 0.23 1.4 ? 0.27 20.5 2 4.2 26.1 ? 1 . 7 12.3 2 5.5
25.2
* 7.2
Values o f enzyme activities are the mean SD and are expressed in units per gram of muscle o r liver ( 1 unit = 1 pmol of substrate turnover/ min). Acyl CoA dehydrmogenase activity was measured according to Fong and Schulz 116). CoA = coenzyme A; DH = dehydrogenase. _f
108 Annals of Neurology
Vol 31
No 1 January 1992
were normal, whereas cytochrome c oxidase activity was reduced. Enzymes of Poxidation The activity of long-chain (palmitoyl) acyl coenzyme A dehydrogenase was reduced in the muscle biopsy and totally absent in the liver specimen obtained at autopsy (see Table 2). In the muscle biopsy, the activity of octanoyl CoA dehydrogenase, butyryl CoA dehydrogenase, thiolase C 10, and hydroxyacyl CoA dehydrogenase was normal, but the activities of enoyl CoA hydratase and thiolase C4 were slightly decreased. Carnitine and pyruvate dehydrogenase were also normal in muscle. In postmortem liver, the activities of octanoyl CoA dehydrogenase and butyryl CoA dehydrogenase and all other enzymes of P-oxidation were normal.
Discussion In our young patient, cytochrome c oxidase deficiency was documented in muscle and liver at 11 months and confirmed post mortem at 13 months. In both muscle and liver, we also found a deficiency in palmitoyl CoA dehydrogenase. Although the defect of palmitoyl CoA dehydrogenase was expressed in the first muscle specimen, the decision to treat the patient with valproate was made when the enzyme defect had not yet been detected and the clinical situation made anticonvulsant therapy unavoidable, despite the impairment in the respiratory chain. Administration of valproate may well have worsened the palmitoyl CoA dehydrogenase deficiency, but cannot have caused it because the deficiency already existed at 11 months. How, then, can palmitoyl CoA dehydrogenase deficiency be explained? It is not likely that a general breakdown of the respiratory chain would impair ETF (electron transport flavoprotein) metabolism and affect selectively the long-chain acyl CoA dehydrogenase as we observed in our patient. Rather, we suggest an independent mitochondrial enzyme defect in the P-oxidation pathway, not secondary to the enzyme defect in the respiratory chain. The concurrent clinical and pathological signs of severe disturbance of fat metabolism (massive hypertriglyceridemia and fatty changes of the liver), which have not been observed in other patients with cytochrome c oxidase deficiency, also support the concept of an independent defect of p-oxidation. Miyabayashi and colleagues “)I were able to confirm cytochrome c oxidase deficiency in the brain cells of a patient afflicted with Leigh‘s disease. The therapeutic trials with thiamine, biotin, methylene blue, ascorbic acid, and vitamin K, were aimed at supplying artificial electron transporters to “unload” the defective respiratory chain. As in other reports, our attempts did not succeed. Many questions remain unanswered. We believe that this was a rare occurrence of two independent mitochondrial enzyme defects. Secondary carnitine de-
ficiency was described in patients with respiratory chain defects, but measurements of P-oxidation have been performed infrequently so far. Systematic studies of P-oxidation enzymes would be of interest to clarify the question raised by this patient’s condition of a possible relationship between defects of the respiratory chain and of P-oxidation. Only recently, Watmough and colleagues [17] reported on lipid accumulation in biopsies from patients with a complex I defect of the respiratory chain. This study was supported by the Deutsche Forschungsgmeinschaft (Re 265/5-3). H . Reichmann is grateful to S. Seufert for expert technical assistance and B. Gobel for secretarial work.
References 1. Luft R, Ikkos D, Palmieri G, et al. A case of severe hypermetabolism of nonthyroid origin with a defect in the maintenance of mitochondrial respiratory control: a correlated clinical, biochemical and morphological study. J Clin Invest 1962;41: 1776-1804 2. DiMauro S, Bonilla E, Zeviani M, e t al. Mitochondrial myopathies. Ann Neurol 1985;17:521-538 3. Sengers RCA, Stadhouders AMM, Trijbels JMF. Mitochondrial myopathies. Eur J Pediatr 1984;141:192-207 4. Morgan-Hughes JA. The mitochondrial myopathies. In: Banker AG, Banker BQ, eds. Myology. New York: McGraw-Hill, 1986:1709-1743 5. Shapira Y , Hare1 S, Russel A. Mitochondrial encephalomyopathies: a group of neuromuscular disorders with defects in oxidative metabolism. Isr J Med Sci 1977;13:161-164 6. Goebel HH, Bornemann A, Reichmann H. Mitochondriarelated encephalomyopathies. Neuropathol Appl Neurobiol 1989; 1597-1 19 7. Lombes A, Bonilla E, DiMauro S.Mitochondrial encephalomyopathies. Rev Neurol (Paris) 1989;145:67 1-689 8. Willems JL, Monnens LAH, Trijbels JMF, e t al. Leigh’s encephalomyelopathy in a patient with cytochrome c oxidase deficiency in muscle tissue. Pediatrics 1977;60:850-857 9. Miyabayashi S, Narisawa K, Tada K, et al. Two siblings with cytochrome c oxidase deficiency. J Inherited Metab Dis 1983; 6: 12 1-122 0. DiMauro S, Servidei S, Zeviani M, et al. Cytochrome c oxidase deficiency in Leigh syndrome. Ann Neurol 1987;22:498-506 1. Arts WFM, Scholte H R , Loonen MCB, e t al. cytochrome c oxidase deficiency in subacute necrorizing encephalopathy. J Neurol Sci 1987;77:103-115 2. Dubowitz V, Brooke MH. Muscle biopsy: a modern approach. Philadelphia: W B Saunders, 1973 3. Reichmann H, Maltese WA, DeVivo DC. Enzymes of fatty acid P-oxidation in developing brain. J Neurochem 1988;51:339344 14. Reichmann H , Rohkamm R, Zeviani M, et al. Mitochondrial myopathy due to complex I11 deficiency with normal reducible cytochrome b concentration. Arch Neurol 1985:43:957-361 15. Trevisan CP, Reichmann H , DeVivo DC, DiMauro S. Betaoxidation enzymes in normal human muscle and in muscle from a patient with an unusual form of myopathic carnitine deficiency. Muscle Nerve 1985;8:672-675 16. Fong JC, Schulz H. On the rate determining step of fatty acid oxidation in heart. J Biol Chem 1978;253:6917-6922 17. Watmough NJ, Bindoff LA, Birch-Machin MA, et al. Impaired mitochondrial P-oxidation in a patient with an abnormaliry of the respiratory chain. J Clin Invest 1990;85:177-184
Brief Communication: Reichmann et al: Combined Enzymatic Defect in Leigh’s Disease
109
A female infant was seen at the age of 2 months because of hypotonia, delayed motor development, and lactic acidosis, and she died at age 13 months due to respiratory failure. In a muscle specimen taken at 11 months and in a liver specimen obtained 1.5 hours postmortem, we found decreased activities of cytochrome c oxidase and longchain acyl coenzyme A dehydrogenase. Neuropathological changes were typical for Leigh’s subacute necrotizing encephalomyelopathy. To our knowledge, this is the first report of a combined defect of complex IV of the respiratory chain and of the long-chain specific acyl coenzyme A dehydrogenase of @-oxidation in muscle and liver. Reichmann H , Scheel H, Bier B, Ketelsen U-P, Zabransky S. Cytochrome c oxidase deficiency and long-chain acyl coenzyme A dehydrogenase deficiency with Leigh’s subacute necrotizing encephalomyelopathy. Ann Neurol 1992;31:107-109
Since the first description of mitochondrial myopathy {l], a body of literature has emerged concerning defects of mitochondrial energy metabolism (for review, see [2-4]), mainly due to defects of the respiratory chain. Because the brain is often also affected by the metabolic defect, the term mitochondrial encephalomyopathy was introduced by Shapira and colleagues [ S ] (for review, see [6,7)).Leigh’s disease, an encephalomyelopathy is often associated with cytochrome c oxidase (COX) deficiency {S-1 11. Although COX activity is decreased in various tissues, there are reports of
From the *Department of Neurology, University of Wiirzburg, Wurzburg; Departments of ?Pediatrics and $Pathology, University of Homburg/Saar, Homburg; and SDeparrment of Pediatrics, University of Freiburg, Freiburg, Germany. Received Mar 29, 1991, and in revised form Jun 24 and Jul 26. Accepted for publication Jul 27, 1991. Address correspondence ro Dr Reichmann, Professor of Neurology, Department of Neurology, University of Wiirzburg, Josef-Schneider-Strasse 11, D-8700 Wurzburg, Germany.
patients with normal COX activity in liver [27, contrasting with decreased activity in muscle and other tissues. In the present report, we describe a patient with a biochemical defect of the respiratory chain and of (3-oxidation presenting as Leigh’s disease.
Patient Report Our patient was the first child of healthy nonconsanguineous parents. Pregnancy and delivery were uneventful; birth weight was 3,080 gm. At the age of 7 months, she was admitted because of petechial hemorrhages on her face and trunk. She had rhinitis and a mild chronic bronchitis. She was lethargic and showed motor retardation (she had no head control and could not roll over from back to front or sit without support); she had atrophy in the lumbar and gluteal region. Deep tendon reflexes were increased in the lower extremities, where brief rnyoclonus could also be induced. Metabolic acidosis (pH 7.25) with respiratory compensation was found. Lactate was up to 5.22 mmol/L, pyruvate to 240 nmoliL, and alanine to 60 fmol/L. After an oral glucose tolerance test, there was excessive rise in lactate, pyruvate, and alanine. Blood triglycerides were increased to 480 m g q with a rise of very low density lipoproteins. Screening for amino acids, ketone bodies, reducing substances, and mucopolysaccharides was negative. A slightly elevated excretion of ethylrnaIonic, oxalic, adipic, and glutaric acids was found. Total serum carnitine levels were normal; however, acylcarnitines were increased and free carnitine was decreased. Ammonium, ascorbic acid, transaminases, thyroid hormones, cross-reacting protein; rheumatoid factor, coagulation status, thrombocyte function, urinary steroid excretion, and immunoglobulins were normal. Urinalyses revealed erythrocyturia. Cranial computed tomography showed moderate enlargement of the ventricles and cisterns. Electroencephalogram (EEG) and electromyogram were normal. Liver and kidneys were slightly enlarged. The renal scintigram was normal. Slight hypertrophy of the septum and left ventricular wall was observed. A skin biopsy indicated vasculitis without evidence of connective tissue disorder. In spite of supplementation of thiamine and biotin, lactic acidosis had not improved when the patient was seen at 11 months for a muscle biopsy (musculus quadriceps femoris). Motor function was not improved. Myoclonic seizures were observed. The EEG showed multifocal epileptic discharges. When myoclonic seizures occurred daily, valproate was started, whereupon the seizure activity in the EEG quickly subsided, and the attacks occurred only infrequently. Transaminases remained within normal levels. Methylene blue was administered intravenously and carnitine, ascorbic acid, and vitamin K, were given orally without any noticeable biochemical or clinical improvement. At 13 months, her general condition unexpectedly deteriorated. Respiratory insufficiency worsened, the acidosis became intractable, and she died within a few hours. One-half hour post mortem, specimens of liver and of the abdominal musculature were obtained. Postmortem examination of the brain revealed macroscopically a slight degree of polymicrogyria, extensive gliosis and reduction in the number of ganglion cells, pronounced capillary proliferation in the paraventricular and brainstem regions, severe demyelination in the
Copyright 0 1992 by the American Neurological Association
107
anomalies or focal myofibrillar degeneration. The matrix of most mitochondria was condensed, but showed no pathological inclusions. A few scattered mitochondria had marginal lipid droplets. The sarcoplasmic reticulum system was normal. The glycogen content was slightly increased; there were no lysosomal glycogen accumulations and no sarcoplasmic or nuclear inclusions. In contrast, electron microscopic examinations performed on postmortem tissue showed ultrastructural changes typical of mitochondriopathy. The liver cells were enlarged and contained abnormal mitochondria with lipid inclusions and irregular cristae. Muscle mitochondria displayed gross variation in size and shape and also contained lipid inclusions.
capsula interna, capillary calcification in the brainstem ganglia, and severe gliosis in the nuclei of the pons and medulla oblongata, wasting of the myelin sheaths, and capillary proliferation. There was hepatomegaly with centrolobular, finedroplet fatty infiltr,ition and a slight degree of muscle fiber atrophy.
Materials and Methods Routine histochemical methods were applied as outlined in Dubowitz and Brooke [ 121. Muscle was homogenized as previously described [13]. Activity of enzymes of the respiratory chain was analyzed as described [ 141; activity measurements of all enzymes from 8-oxidation were performed as previously described [I?;, 15, 161. Control values were obtained from biopsies that finally were shown to not represent a neurornuscular disorder. Liver biopsies were obrained from autopsies within the first 2 hours post mortem.
Enzymes of the Respiratory Chain The results of the biochemical investigation are listed in Tables 1 and 2. T h e activity of cytochrome c oxitlase was markedly decreased both in the muscle biopsy (19% of normal) and in the liver specimen obtained postmortem (7% of normal). The activities of complex I, 11, and 111 and of citrate synthase, a key enzyme of the citric acid cycle, were normal, and histochemical investigations revealed a normal stain for succinate Jehydrogenase and a normal adenosine triphosphatase reaction. T h e carnitine content was low normal. In the liver specimen, the activities of complex I, 11, and I11
Results Hzj.tologicul und El'ectron Mirro.tcopicul Findings The histological and electron microscopical investigations of the muscle biopsy obtained at 11 months showed no signs of fiber atrophy or of necrosis, no inflammatory infiltration, and no changes in the blood vessels. The distribution of type I and type I1 fibers was normal. Oil red 0 staining showed a slight increase of intracellular neutral fat droplets. Gomori trichrome stain showed no ragged-red fibers. Electron microscopy showed no evidence of rnyofibrillar structure Table 1 Biothemiral Analyses of Enzymes ofthe Respiratory Chain Enzyme
Muscle
Muscle Controls (n)
Liver ~
0.53 42.1 1.84 2.63 1.11 5.6
Cytochrome c oxidase N A D H dehydrogenase NADH-cytochrome c reductase Succinate dehydrogenase Succinate cytochrome c reductase citrate synthetase
2.8 t 0.9 (113) 4 8 k 9.4 (121) 3.2 t 1.5 (98) 2.4 1.1 (160) 1.64 ? 0.6 (125) 5.3 t 2.1 (49)
Liver Controls ( n ) ~~
~
0.5 73.4 15.7 3.2 2.4 ...
*
-
~
~~
5 t 1.8 ( 4 ) 101 t 32 (4) 1.3.1 t 1.7 ( 4 ) lr.7 ? 1.8 (4) 3.1 ? 1.0 (4) ...
Values of the enzyme activities are the mean f SD and arc expressed in units per gram of tissue ( I unir = 1 pmol ofsiibstr~tteturnowrimin). Muscle was obtained at 11 months; liver was obtained 2.5 hours post mortem. N A D H = reduced nicotinamide adenine dinucleotide.
Table 2. Enqme.r of ,&Oxidation Enzyme
Muscle
Muscle Controls ( n )
Liver
Liver Controls (n
Palmitoyl CoA DH Octanoyl CoA DH Butyryl CoA DH Enoyl CoA hydratase Hydroxyacyl CoA DH Thiolase C 10 Ketone body metabolism Thiolase C4
0.29 0.54 1.0 8.1 10.6 2.11
1.02 0.36 (65) 0.83 ? 0.31 (52) 0.84 ? 0.33 (63) 13.3 k 3.1 (41) 12.6 ? 3.43 (58) 2.12 t 1.47 (44)
0 0.82 0.72 12.6 26.6 12.31
1.52
1.66
4.66 2 2.38 (54)
4.55
*
=
3)
? 0.43 0.9 2 0.23 1.4 ? 0.27 20.5 2 4.2 26.1 ? 1 . 7 12.3 2 5.5
25.2
* 7.2
Values o f enzyme activities are the mean SD and are expressed in units per gram of muscle o r liver ( 1 unit = 1 pmol of substrate turnover/ min). Acyl CoA dehydrmogenase activity was measured according to Fong and Schulz 116). CoA = coenzyme A; DH = dehydrogenase. _f
108 Annals of Neurology
Vol 31
No 1 January 1992
were normal, whereas cytochrome c oxidase activity was reduced. Enzymes of Poxidation The activity of long-chain (palmitoyl) acyl coenzyme A dehydrogenase was reduced in the muscle biopsy and totally absent in the liver specimen obtained at autopsy (see Table 2). In the muscle biopsy, the activity of octanoyl CoA dehydrogenase, butyryl CoA dehydrogenase, thiolase C 10, and hydroxyacyl CoA dehydrogenase was normal, but the activities of enoyl CoA hydratase and thiolase C4 were slightly decreased. Carnitine and pyruvate dehydrogenase were also normal in muscle. In postmortem liver, the activities of octanoyl CoA dehydrogenase and butyryl CoA dehydrogenase and all other enzymes of P-oxidation were normal.
Discussion In our young patient, cytochrome c oxidase deficiency was documented in muscle and liver at 11 months and confirmed post mortem at 13 months. In both muscle and liver, we also found a deficiency in palmitoyl CoA dehydrogenase. Although the defect of palmitoyl CoA dehydrogenase was expressed in the first muscle specimen, the decision to treat the patient with valproate was made when the enzyme defect had not yet been detected and the clinical situation made anticonvulsant therapy unavoidable, despite the impairment in the respiratory chain. Administration of valproate may well have worsened the palmitoyl CoA dehydrogenase deficiency, but cannot have caused it because the deficiency already existed at 11 months. How, then, can palmitoyl CoA dehydrogenase deficiency be explained? It is not likely that a general breakdown of the respiratory chain would impair ETF (electron transport flavoprotein) metabolism and affect selectively the long-chain acyl CoA dehydrogenase as we observed in our patient. Rather, we suggest an independent mitochondrial enzyme defect in the P-oxidation pathway, not secondary to the enzyme defect in the respiratory chain. The concurrent clinical and pathological signs of severe disturbance of fat metabolism (massive hypertriglyceridemia and fatty changes of the liver), which have not been observed in other patients with cytochrome c oxidase deficiency, also support the concept of an independent defect of p-oxidation. Miyabayashi and colleagues “)I were able to confirm cytochrome c oxidase deficiency in the brain cells of a patient afflicted with Leigh‘s disease. The therapeutic trials with thiamine, biotin, methylene blue, ascorbic acid, and vitamin K, were aimed at supplying artificial electron transporters to “unload” the defective respiratory chain. As in other reports, our attempts did not succeed. Many questions remain unanswered. We believe that this was a rare occurrence of two independent mitochondrial enzyme defects. Secondary carnitine de-
ficiency was described in patients with respiratory chain defects, but measurements of P-oxidation have been performed infrequently so far. Systematic studies of P-oxidation enzymes would be of interest to clarify the question raised by this patient’s condition of a possible relationship between defects of the respiratory chain and of P-oxidation. Only recently, Watmough and colleagues [17] reported on lipid accumulation in biopsies from patients with a complex I defect of the respiratory chain. This study was supported by the Deutsche Forschungsgmeinschaft (Re 265/5-3). H . Reichmann is grateful to S. Seufert for expert technical assistance and B. Gobel for secretarial work.
References 1. Luft R, Ikkos D, Palmieri G, et al. A case of severe hypermetabolism of nonthyroid origin with a defect in the maintenance of mitochondrial respiratory control: a correlated clinical, biochemical and morphological study. J Clin Invest 1962;41: 1776-1804 2. DiMauro S, Bonilla E, Zeviani M, e t al. Mitochondrial myopathies. Ann Neurol 1985;17:521-538 3. Sengers RCA, Stadhouders AMM, Trijbels JMF. Mitochondrial myopathies. Eur J Pediatr 1984;141:192-207 4. Morgan-Hughes JA. The mitochondrial myopathies. In: Banker AG, Banker BQ, eds. Myology. New York: McGraw-Hill, 1986:1709-1743 5. Shapira Y , Hare1 S, Russel A. Mitochondrial encephalomyopathies: a group of neuromuscular disorders with defects in oxidative metabolism. Isr J Med Sci 1977;13:161-164 6. Goebel HH, Bornemann A, Reichmann H. Mitochondriarelated encephalomyopathies. Neuropathol Appl Neurobiol 1989; 1597-1 19 7. Lombes A, Bonilla E, DiMauro S.Mitochondrial encephalomyopathies. Rev Neurol (Paris) 1989;145:67 1-689 8. Willems JL, Monnens LAH, Trijbels JMF, e t al. Leigh’s encephalomyelopathy in a patient with cytochrome c oxidase deficiency in muscle tissue. Pediatrics 1977;60:850-857 9. Miyabayashi S, Narisawa K, Tada K, et al. Two siblings with cytochrome c oxidase deficiency. J Inherited Metab Dis 1983; 6: 12 1-122 0. DiMauro S, Servidei S, Zeviani M, et al. Cytochrome c oxidase deficiency in Leigh syndrome. Ann Neurol 1987;22:498-506 1. Arts WFM, Scholte H R , Loonen MCB, e t al. cytochrome c oxidase deficiency in subacute necrorizing encephalopathy. J Neurol Sci 1987;77:103-115 2. Dubowitz V, Brooke MH. Muscle biopsy: a modern approach. Philadelphia: W B Saunders, 1973 3. Reichmann H, Maltese WA, DeVivo DC. Enzymes of fatty acid P-oxidation in developing brain. J Neurochem 1988;51:339344 14. Reichmann H , Rohkamm R, Zeviani M, et al. Mitochondrial myopathy due to complex I11 deficiency with normal reducible cytochrome b concentration. Arch Neurol 1985:43:957-361 15. Trevisan CP, Reichmann H , DeVivo DC, DiMauro S. Betaoxidation enzymes in normal human muscle and in muscle from a patient with an unusual form of myopathic carnitine deficiency. Muscle Nerve 1985;8:672-675 16. Fong JC, Schulz H. On the rate determining step of fatty acid oxidation in heart. J Biol Chem 1978;253:6917-6922 17. Watmough NJ, Bindoff LA, Birch-Machin MA, et al. Impaired mitochondrial P-oxidation in a patient with an abnormaliry of the respiratory chain. J Clin Invest 1990;85:177-184
Brief Communication: Reichmann et al: Combined Enzymatic Defect in Leigh’s Disease
109
