Muscle coenzyme Q10 in mitochondrial encephalomyopathies.

Coenzyme Q₁₀ (CoQ) content was measured in isolated muscle mitochondria from 25 patients with mitochondrial encephalomyopathies (MEM), most of whom had mitochondrial DNA mutations. The CoQ level was significantly lower in MEM patients than in controls. CoQ levels varied widely from patient to patient, especially in those with chronic progressive external ophthalmoplegia including Kearns-Sayre syndrome, which may explain, at least in part, the variable response of patients to CoQ administration.     

Skeletal muscle CK-B activity in neurogenic muscular atrophies

Summary Creatine kinase isoenzymes were determined in skeletal muscle biopsy specimens of 34 patients suffering from neurogenic muscular atrophies. The findings were compared: (1) with those of 38 control muscle samples and (2) with those in 41 muscular dystrophies and other myopathic conditions. The measurements were made by electrophoretic separation and elution of the isoenzymes and by immunoinhibition assay. The results showed that the total and specific CK activity were significantly decreased (P<0.005) in neurogenic atrophies in contrast to myopathic conditions where no differences from control levels were observed. This decrease was due to a decrease of the CK-M subunit activity, while the CK-B subunit was elevated. The muscle CK-MB activity was considerably elevated in muscular dystrophies (P<0.02) and myositis (P<0.001), but it was also slightly elevated in neurogenic conditions. The similarity of the muscle CK isoenzyme pattern in neurogenic atrophies and myotonic dystrophy was noted. These findings could possibly reflect considerable difference in the regeneration process of neurogenic atrophies and muscular dystrophies.     

Clinical and biochemical correlations in mitochondrial myopathies treated with coenzyme Q10

We tested the efficacy of coenzyme Q10 (ubidecarenone, CoQ10) therapy in patients with Kearns-Sayre syndrome and other mitochondrial myopathies with chronic progressive external ophthalmoplegia (CPEO). We treated seven patients for 1 year with daily oral administration of 120 mg of CoQ10. Throughout the treatment most of our patients showed a progressive reduction of serum lactate and pyruvate levels following standard muscle exercise and generally improved neurologic functions. The ECG and echocardiogram showed no significant changes in our patients. None of our patients showed any improvement in ptosis and CPEO.     

Partial deficiency of complexes I and IV of the mitochondrial respiratory chain in skeletal muscle of two patients with mitochondrial myopathy

Summary Respiratory chain enzymes were studied in isolated mitochondria of two patients with mitochrondrial myopathy. Both patients had been suffering from chronic progressive external ophthalmoplegia and abnormal muscular fatigability since late childhood. One of the patients exhibited the complete triad of symptoms characteristic of Kearns-Sayre syndrome. Venous lactate levels at rest and during minimal exercise were increased in both patients. Histochemical examination of muscle revealed ragged red fibres and intermingled fibres negative for cytochrome c oxidase. Biochemical studies showed decreased activities of complex I and complex IV of the respiratory chain in both patients. Reduced minus oxidized spectra of mitochondrial cytochromes revealed a decreased content of cytochrome aa₃ in only one patient, but a normal content in the other. A combined deficiency of complexes I and IV in muscle might either be due to a deficiency of a single subunit common to both complexes or to a coincidental deficiency of both complexes expressed either in the same or in different fibres.     

Exogenous coenzyme Q (coq) fails to increase coq in skeletal muscle of two patients with mitochondrial myopathies

Recently, several studies were published on therapy with coenzyme Q (CoQ) in patients with mitochondrial myopathies without biochemically established muscular deficiency of CoQ. Two patients with mitochondrial myopathies presenting as oculocraniosomatic syndromes were treated with coenzyme Q (CoQ). The muscle biopsy of both patients showed ragged-red fibers and single muscle fibers without histochemical reaction for cytochrome c oxidase. Biochemical analysis revealed normal activities of the respiratory chain complexes in muscle and normal levels of CoQ in serum and muscle. After one year of treatment CoQ in serum of both patients had increased 1.4-fold and 2.0-fold, respectively. In muscle, however, there was no increase of CoQ in either patient. In both patients the activities of citrate synthase and of the respiratory chain complexes I + III and IV, and in 1 patient also of complex II + III, were lower in the second biopsy compared with the first biopsy. In both patients there was no improvement of maximal isometric muscle strength assessed by a quantitative electronic strain gauge. The exercise-induced pathological rise of lactate in 1 patient remained essentially unchanged during therapy. The data indicate that orally administered CoQ fails to increase total CoQ in muscle of patients with mitochondrial myopathies but without muscular CoQ deficiency.     

Lipid peroxidation and superoxide dismutase activity in muscle and erythrocytes in adult muscular dystrophies and neurogenic atrophies

Summary Lipid peroxidation (LP) and superoxide dismutase (SOD) activity were determined in erythrocytes and skeletal muscle obtained from patients with limb-girdle and facioscapulohumeral muscular dystrophies, neurogenic atrophies and from age-matched control subjects. Neither lipid peroxidation nor SOD activity in erythrocytes of patients differed from control values. SOD activity and LP in muscle specimens were also normal in types of neurogenic atrophy. Lipid peroxidation in the muscle from patients with adult types of muscular dystrophy had a tendency to be increased. The values were widely scattered, the highest being obtained in the older patients with long duration of disease.     

Mitochondrial myopathies: clinical and biochemical features of 30 patients with major deletions of muscle mitochondrial DNA

Analysis of mitochondrial DNA (mtDNA) in muscle and blood from 72 patients with mitochondrial myopathy showed that 30 had major deletions of a variable proportion of muscle mtDNA. All of these 30 patients presented with progressive external ophthalmoplegia and limb weakness, and 8 had the additional features of the Kearns-Sayre syndrome. Of the 42 patients without detectable muscle mtDNA deletions, 10 had progressive external ophthalmoplegia and limb weakness, 2 had the Kearns-Sayre syndrome, 11 had limb weakness without extraocular involvement, and 19 had multisystem disorders predominantly affecting the central nervous system. Only 2 patients with mtDNA deletions had clinically affected relatives, compared with 10 of those without deletions. In the 4 patients with polarographic defects exclusively involving complex I (NADH coenzyme Q reductase), the deleted protein-coding genes were confined to those for complex I subunits. Thirteen other patients with apparently identical deletions had variable clinical and biochemical features. Immunoblots of complex I polypeptides from patients with deletions were either indistinguishable from controls or showed only a mild generalized decrease in all identifiable subunits.     

Cytochrome c oxidase activity in single muscle fibers: Assay techniques and diagnostic applications

Microphotometric enzyme assay was used to study cytochrome c oxidase activity in single human skeletal muscle fibers. The assay techniques combine the precise localization of enzyme activity provided by histochemical methodology with the precise quantitation of a sensitive assay system. Abnormalities of cytochrome c oxidase were investigated using microphotometric enzyme assay in 12 patients with Kearns-Sayre syndrome, chronic progressive external ophthalmoplegia, or Leigh's syndrome. Control values were obtained using muscle biopsy specimens from 20 juvenile and 18 adult subjects with no evidence of neuromuscular disease. In the patients with Leigh's syndrome due to cytochrome c oxidase deficiency, the abnormality was found to be expressed uniformly throughout the muscle fiber population. In contrast, patients with Kearns-Sayre syndrome or chronic progressive external ophthalmoplegia showed abnormal heterogeneity of cytochrome c oxidase activity. In many cases, extreme degrees of variability were seen, with fibers containing high activity adjacent to fibers with no detectable activity. Mitochondrial DNA analysis showed that most of the patients with Kearns-Sayre syndrome and chronic progressive external ophthalmoplegia had major rearrangements of mitochondrial DNA. It was concluded that the extreme variability of cytochrome c oxidase activity detected using microphotometric enzyme assay was an indicator of a probable abnormality of mitochondrial DNA. Conversely, cytochrome c oxidase defects in muscle which show a homogeneous distribution are more likely to be associated with defects of the nuclear genome.     

An autosomal recessive syndrome with myopathy and central and peripheral nervous system involvement (author's transl)

Three of 11 children, offspring of a consanguineous marriage, presented a progressive myopathy and seizures, associated with symptoms suggesting both central and peripheral nervous system involvement. The ultrastructural muscular lesions were not specific. The association of severe impairment of muscle tissue and of central nervous system is rare, being described in centronuclear myopathy, cerebromuscular dystrophy, Kearns-Sayre syndrome and in a few isolated cases. Clinically only these isolated observations and especially the Kearns-Sayre syndrome demonstrate analogies to our observations. These data lead us to the discussion of the specificity of ultrastructural lesions, especially mitochondrial abnormalities. Some authors consider these abnormalities to be the biochemical hallmark for ophthalmoplegia plus, whereas for others, especially Drachman, they are an inconstant and nonspecific finding, merely the consequence and not the cause of this disease. These observations argue for the relationship between muscular pathology and nervous system dysfunction.     

Limb girdle syndromes. Clinical, morphological and electrophysiological studies

The clinical syndrome of slowly progressive proximal limb and limb girdle muscular weakness and atrophy, or limb girdle syndromes (LGS), has a diverse aetiology. Several of the congenital, mitochondrial and other metabolic myopathies and spinal muscular atrophies are recently recognized causes of LGS. Thus the position of limb girdle muscular dystrophy (LGMD) as a discrete entity in the nosology of muscle disease deserves reappraisal. We have therefore reevaluated our experience of 33 patients in this light. Detailed clinical, electrophysiological, and pathological studies including autopsies in 2 cases, were performed. As a result we are confident that LGMD does exist as a sporadic or autosomal dominant (2 families) or recessive condition (2 families). There are therefore probably at least 2 distinct genotypes. Typical LGMD (18 patients in our series) is characterized by slowly progressive symmetrical proximal upper and lower limb girdle weakness and atrophy, elevation of the serum creatine kinase at some stage, dystrophic or less severe myopathic muscle lesions on biopsy, and myopathic EMG findings. Two minor subgroups of LGMD were identified in our series with similar clinical and laboratory features but distinguishable by the development of either facial (4 patients) or by distal limb muscle involvement (3 patients). A further group of patients with sporadic LGS (5 patients) had slowly progressive proximal symmetrical upper and lower limb-girdle weakness and atrophy with myopathic or neurogenic features on either EMG or muscle biopsy so that the precise characterization was difficult. Two of these patients had distal limb muscle involvement and contractures. One patient had upper limb-girdle muscle atrophy with normal lower limbs. A disorder affecting muscle, nerve or both remains a possibility in these cases.     

Clinical forms of inclusion body myositis: 12 cases

Inclusion body myositis is now a well-known disease but its incidence is underestimated. We report 12 cases with clinical heterogeneity. Three groups of patients could be described. The first one corresponded to asymmetrical muscle involvement and distribution with a slow clinical course (4 cases). The second was characterized by a polymyositis-like syndrome (3 cas), but steroid therapy was ineffective. The last group mimicked a chronic spinal muscular atrophy (4 cases). One patient showed a scapuloperoneal syndrome. Both myopathic and neurogenic EMG patterns were present in 6 patients; a neurogenic pattern was found in 4 cases and a myopathic pattern in 2 cases. In all patients, muscle biopsies showed rimmed vacuoles with eosinophilic inclusions. In 9 cases ultrastructural studies displayed abnormal filaments of 15-18 nm in diameter in the vacuoles. Intranuclear filaments were rarely observed. The significance of the filaments is unknown and their specificity is doubtful because they are present in other myopathies with rimmed vacuoles (some distal myopathies and oculopharyngeal muscular dystrophies). Finally a rich inflammatory exudate was present in 8 patients only.     

Unusual presentation of Kearns-Sayre syndrome in early childhood

Congenital glaucoma and insulin-dependent diabetes mellitus were the predominant presenting signs in a patient with Kearns-Sayre syndrome. Thereafter, he developed short stature, pigmentary retinopathy, progressive external ophthalmoplegia, and ataxia. The diagnosis was confirmed by detecting a deletion of mitochondrial DNA in muscle, thus demonstrating that Kearns-Sayre syndrome can have the unusual presenting signs described above.     

Neurogenic muscle involvement in myasthenia gravis: A clinical and histopathological study

An investigation was made into the occurrence of muscular atrophy and muscular pathology in a series of 170 patients with myasthenia gravis. The results can be summarized as follows: (1) Of the 148 patients with generalized myasthenia gravis, 14 showed local muscular atrophies. Of 10 biopsies from atrophic muscles, eight showed neurogenic changes, with or without lymphocytic infiltrations. One biopsy showed lymphocytic infiltrations only, and one showed type II-fibre atrophy (Table 1). No relationship was demonstrable between the presence of clilnical muscular atrophy and age, sex, duration of the disease, severity of the disease, presence of a thymoma, or drug resistant ophthalmoplegia. (2) In this group of patients 61 biopsies were examined from 46 individuals; 40 of these biopsies were taken from the quadriceps muscle. A thymoma was present in 17 patients. Examination disclosed neurogenic changes in 17 biopsies, lymphocytic infiltrates in 21, and myositis in one biopsy (Table 2). A distinct correlation was established between the presence of a thymoma and lymphocytic infiltrates, but none was demonstrable between thymoma and neurogenic changes (Table 3). (3) An enzyme-histochemical study was carried out in 35 cases, including 12 with neurogenic changes. A normal differentiation of type I- and type II-fibres was observed in eight instances, type grouping of type II-fibres in three, and type II-fibre atrophy in two cases. (4) In 21 patients and 19 controls, the smallest mean diameter was determined in the quadriceps muscle. Both type I- and type II-fibres proved to have a smaller mean diameter in the female patients than in the controls. In the male patients this could not be proven. (5) Of the eight patients who had died without disorders of ventilation, 90 muscle specimens were examined postmortem. Four of these patients had a thymoma. Lymphocytic infiltrations, found in 32 biopsy specimens, were mostly observed in the presence of a thymoma. Neurogenic changes were apparently unrelated to the presence of a thymoma (Tables 5 and 6). The post mortem examination included the spinal cord in five, and peripheral nerves in three cases. No abnormalities were found. (6) The muscular atrophy found in patients with myasthenia is not a myopathy but an affection of the lower motor neurone. Neurogenic changes were regularly found in the muscles of patients with myasthenia, even without muscular atrophy. The finding of these changes is no reason to reject the diagnosis. It is postulated that denervation occurs at the neuromuscular junction as a result of permanent absence of acetylcholine.     

Isoelectric focusing and electrophoresis of cerebrospinal fluid proteins in muscular dystrophies and spinal muscular atrophies.

In the very few previous investigations of the CSF-proteins in muscular dystrophies the results have generally been reported as normal. In spinal muscular atrophies a barrier-damage pattern of CSF-proteins has been found in amyotrophic lateral sclerosis (ALS). In the present investigation the CSF-proteins were examined by isoelectric focusing and quantitative paper electrophoresis in 13 patients with muscular dystrophies and in 11 patients with spinal muscular atrophies. On isoelectric focusing, CSF-protein abnormalities were found in 85% of the cases with muscular dystrophies and in all patients with spinal muscular atrophies. Differences in the CSF-protein patterns were observed within the group of muscular dystrophies and between these and the cases of spinal muscular atrophies. In ALS and in myotonic dystrophy, abnormal CSF-protein fractions occurred mainly in the alkaline pH-range, while in limb-girdle dystrophy and the patient with facioscapulohumeral dystrophy, aberrant fractions appeared mainly in the acidic region. CSF-protein abnormalities were found in both the alkaline fractions (HAFs) with pI 9.2-9.6 and a fraction with PI 7.1 were found in half of the patients with myotonic dystrophy. The CSF electrophoresis in myotonic dystrophy showed increased levels of beta1-globulin in all cases examined. Signs of barrier-damage were commonly encountered in ALS in contrast to the muscular dystrophies, except for myotonic dystrophy. The results are discussed in terms of possible diagnostic value and with regard to pathogenetic significance, particularly in relation to the current hypothesis of a neural involvement in muscular disorders.     

Oculoskeletal myopathy with abnormal mitochondria

A clinical, electrophysiological and pathological review of 14 patients having oculoskeletal myopathy with abnormal mitochondria was undertaken. These patients present with ophthalmoplegia, and mild skeletal muscle weakness. The clinical course is slowly progressive. Electromyographic examination shows myopathic changes. Serum enzymes are normal. The diagnosis is confirmed by skeletal muscle biopsy which shows abnormal mitochondria, including crystalloid inclusions on electron microscopy. These patients form a distinct clinical group in which the risk of sudden cardiac death is much less than it is in the Kearns-Sayre syndrome.     

Phosphomannosyl receptors of lysosomal enzymes of skeletal muscle in neuromuscular diseases

The phosphomannosyl receptor system is responsible for both the receptor-mediated endocytosis and the intracellular transport of lysosomal enzymes. In the present study this receptor system was examined in affected muscles of patients with various neuromuscular diseases. The total activity of beta-N-acetyl-glucosaminidase, a marker enzyme of lysosomal hydrolases, was significantly elevated in the patients with myopathies (polymyositis and muscular dystrophies) but only slightly increased in those with neurogenic muscle atrophies (amyotrophic lateral sclerosis, polyneuropathy or other neurogenic muscle disease). The increase was most prominent in the group of polymyositis. The content of phosphomannosyl receptors was increased in the patients with myogenic muscle disease but not in those with neurogenic disease. The receptor binding of lysosomal enzymes was saturable and inhibited with mannose 6-phosphate showing the typical characteristics of phosphomannosyl receptors. The characteristics of the receptors were very similar both to control and to diseased muscle samples. When surveying all the material, the content of phosphomannosyl receptors correlated highly significantly with the muscular activity of beta-N-acetylglucosaminidase, muscle atrophy index, and serum creatine kinase activity.     

Chronic progressive external ophthalmoplegia: A correlative study of mitochondrial DNA deletions and their phenotypic expression in muscle biopsies

Deleted mitochondrial DNA (mtDNA) has been shown to coexist with normal mtDNA (heteroplasmy) in muscles from chronic progressive external ophthalmoplegia, including Kearns-Sayre syndrome. In this study, we correlated heteroplasmic mtDNA abnormality with clinical, biochemical and histological findings with the following results: (1) large deletions ranging from 1.8 to 8.8 kb in 22 muscle specimens from 28 patients who had ophthalmoplegia clinically and focal cytochromec oxidase (CCO) deficiency by histochemistry, (2) no difference in clinical and biochemical findings between patients with and without mtDNA deletions, (3) no relationship between the size, site or populations of deleted mtDNA and respiratory chain enzyme activities in muscles, (4) positive correlation between the number of CCO-deficient fibers and the populations of deleted mtDNA, and (5) higher incidence of CCO-negative fibers in patients with deleted mtDNA than in those with no deletion of mtDNA. These results suggest that deleted mtDNA is, at least in part, responsible for focal CCO deficiency as a phenotypic expression and that the investigation on pathogenetic mechanism of focal CCO deficiency may provide a clue to understanding the underlying pathophysiology in this disorder.     

Possible neurogenic factor in muscular dystrophy: its similarity to denervation atrophy

Muscle biopsy specimens from 179 cases of muscular dystrophies and from 140 cases of anterior horn cell disorders (from a total of 1,348 biopsied patients) were examined histologically. There were 72 cases of Duchenne type muscular dystrophy (DMD), five of Becker type MD, four girls with myopathy resembling DMD, 40 with limb-girdle, 10 with facioscapulohumeral, seven with late onset, 13 with congenital, and 28 with unclassifiable muscular dystrophies. Groups of small atrophied muscle fibres were encountered in 42 (23%) of the cases in this group, most frequently in patients with limb-girdle, facioscapulohumeral, and least frequently with DM dystrophy. In the second group there were 25 cases of infantile, 38 of juvenile, and 39 of adult spinal muscular atrophy (SMA); there were 21 patients with motor neurone disease (MND), six with poliomyelitis, and 11 with an unclassifiable type of anterior horn cell disorder. Pseudomyopathic changes were encountered in 43 (30%) of all cases in this group. They were most frequently present among patients with juvenile and adult SMA and in those with MND. The presence of group atrophy in muscular dystrophy is considered significant myopathological evidence of a denervation process. On the other hand, pseudomyopathic changes, variation in fibre size, rounding, central nuclei, and increase in connective tissue occurring in various anterior horn cell disorders are seen not to be specific `myopathic'' changes. Thus there was an overlap of pathological reactions in muscles from the dystrophies and the neurogenic atrophies. Comparably atrophied fibres (much less than 2 SDs below the normal mean diameter) and hypertrophied fibres (much more than 2 SDs above the normal mean diameter) were encountered in both dystrophy and neurogenic atrophy, considering the large muscles of the limb. Likewise, the mean fibre diameters were comparable in DMD and in juvenile SMA. The fourth evidence of a neurogenic factor in muscular dystrophy was derived from an examination of SDH preparations of muscle. There was a preponderance of type I muscle fibres in dystrophic muscles compared with specimens from controls, suggesting depletion of type II fibres. It appears that the concept of muscular dystrophy as a primary muscle disease needs to be re-examined.     

Deletions of mitochondrial DNA in Kearns-Sayre syndrome and ocular myopathies: genetic, biochemical and morphological studies.

Genetic, biochemical and morphological investigations were conducted on skeletal muscle mitochondria from 6 cases of ocular myopathy: 4 cases with Kearns-Sayre syndrome (KSS) and 2 with chronic progressive external ophthalmoplegia. All of these 6 cases showed mitochondrial DNA (mtDNA) deletions in addition to normal sized DNA in the quadriceps muscle. The deletions ranging from 3 to 8 kbp were also mapped between nucleotides 5500 and 16000 by Southern blot. The deleted genes encoded for some subunits of complexes I, IV, V and 5-10 tRNAS. The boundaries of the deletions have been sequenced in three patients. Five patients had mitochondrial respiratory chain deficiency in complex I as shown by the low oxygen consumption in isolated mitochondria using three NAD(+)-linked substrates. Mitochondria with an abnormal ultrastructure were also observed in 2 cases. A good relationship between the cytochrome c oxidase deficiency and the amount of deleted mtDNA was shown in our present investigations.     

Mitochondrial DNA deletion in a child with megaloblastic anemia and recurrent encephalopathy

A 3 1/2-year-old boy presented with megaloblastic anemia and recurrent episodes of severe lactic acidosis and coma. At age 4 years, he developed sepsis and died; postmortem examination failed to show any gross abnormality in any tissue. Biochemical analysis of muscle showed decreased activities for all respiratory chain enzymes except complex II. Muscle histochemistry revealed diffuse cytochrome c oxidase deficiency. Southern blot analysis of mitochondrial DNA from muscle, liver, and blood showed a heteroplasmic single mitochindrial DNA deletion of 2.4 kb, which removed the genes for cytochrome c oxidase I and II and the transfer ribonucleic acid genes for serine and aspartic acid. Single large-scale deletions in mitochondrial DNA have been associated with Pearson's syndrome, Kearns-Sayre syndrome, and progressive external ophthalmoplegia. This patient's presentation is unusual and suggests an overlap between Pearson's syndrome and Kearns-Sayre syndrome.