Neuropathology of myoclonus epilepsy associated with ragged-red fibers (Fukuhara's disease)

Summary The post-mortem findings are reported of two patients with myoclonus epilepsy associated with ragged-red fibers (MERRF, Fukuhara's disease), whose clinical findings have been described in detail previously. In addition to the mitochondrial myopathy, both patients had consistent lesions in the central and peripheral nervous systems: (1) degeneration of the dentatorubral and pallidoluysian systems, (2) spinal cord lesions resembling Friedreich's ataxia, and (3) degeneration of the substantia nigra, cerebellar cortex, inferior olivary nucleus, locus ceruleus, gracile and cuneate nuclei, and the pontine tegmentum. The nature and distribution of the lesions are different not only from the other mitochondrial encephalomyopathies but also from other known diseases. It is concluded that MERRF is a disease entity.Supported in part by grants from the Ministry of Health and Weifare of Japan     

The Ramsay Hunt syndrome revisited: Mediterranean myoclonus versus mitochondrial encephalomyopathy with ragged-red fibers and Baltic myoclonus

Among progressive myoclonus epilepsies (PME), the nosography of the Ramsay Hunt syndrome (RHS) has been much debated. The authors report on a homogeneous group of 43 patients originating from around the western Mediterranean, with a large number of northern African subjects, who were followed up for a mean period of 11.6 years. Onset is between 6 and 17 years (mean: 11.2) and the transmission appears to be recessive. The clinical features include: action myoclonus, generalized epileptic seizures, mild cerebellar signs and lack of dementia. EEG features include normal background activity, spontaneous fast generalized spike-wave discharges, photosensitivity, lack of activation during nREM sleep and vertex/rolandic spikes during REM sleep. The prognosis is variable, even within families, but the progression seems to be slow in a majority of patients. This condition can be distinguished from mitochondrial encephalomyopathy and is less severe than Baltic myoclonus. The authors propose that this form of PME, formerly reported as RHS, be more properly described as Mediterranean myoclonus.     

肌阵挛癫痫合并破碎红纤维综合征的线粒体DNA突变特点

目的　探讨肌阵挛癫痫伴破碎红纤维综合征 ( MERRF)的分子遗传学特点。方法　用聚合酶链反应-限制片段长度多态性 ( PCR- RFL P)方法检测 6例 MERRF患者及其部分母系亲属的肌肉和 (或 )外周血细胞的mt DNA的 A8344 G点突变 ,并进行突变型 mt DNA的定量分析。结果　在 2例患者的肌肉和外周血细胞中检测到A8344 G点突变。但其母亲的外周血细胞中未能检测到此突变。这 2例 A8344 G阳性标本中 ,肌肉组织的突变型mt DNA的比例分别为 79.0 %和 86 .8%,而在外周血细胞中分别为 5 9.7%和 72 .9%,突变型 m t DNA的比例在肌肉组织中高于外周血细胞中。结论　在 MERRF患者不同组织中检测到 mt DNA A8344 G点突变 ,与国外报道一致。但国外报道 MERRF多为母系遗传 ,而我们的病例未能有此发现 ,须扩大样本量进一步分析。     

Gene dosage effect in one family with myoclonic epilepsy and ragged-red fibers (MERRF)

Objectives - We analyzed the percentage of mitochondrial DNA (mtDNA) heteroplasmy in blood samples of 13 individuals belonging to a three family generation of myoclonic epilepsy with ragged-red fibers (MERRF) and compared the 5 affected patients and the 8 unaffected relatives. Material and methods - DNA was extracted from blood and muscle of the proband and from blood of 12 maternal relatives. A PCR restriction analysis method was used to detect the mutation. Results - The proband had the complete MERRF phenotype. The phenotype in three other individuals in the maternal lineage was consistent with the MERRF syndrome. The remaining were asymptomatic. The np 8344 mutation was observed in muscle and blood of the proband, and in blood from every one of 12 maternal relatives, ranging from 44% to 83% of mutated genomes. Symptomatic individuals had higher levels ( P < 0.001) of mutated mtDNA than asymptomatic maternal relatives. However, high proportions of mutant genomes (up to 63%) were found in asymptomatic relatives. Conclusions - Although there seems to be a gene dosage effect in MERRF, we found no absolute relationship between the relative proportion of mutant genomes in blood and clinical severity. Factors other than gene dosage in blood may account for the differences in clinical phenotype.     

Functional respiratory chain studies in mitochondrial cytopathies. Support for mitochondrial DNA heteroplasmy in myoclonus epilepsy and ragged red fibers (MERRF) syndrome

Summary Mitochondrial respiratory chain function was investigated with polarographic and enzymatic studies, and correlated with immunoblot studies using a battery of probes against respiratory chain holocomplexes in a series of patients with myoclonus epilepsy and ragged red fibers (MERRF) syndrome. State III respiration rates in intact skeletal muscle mitochondria were normal in two cases, suggested site I deficiency in one case and a midrespiratory defect in another. Immunological studies of complex I showed reduced levels of several subunits with the apparent absence of two bands (which at 45 and 42 kDa, coincide with the predicted electrophoretic mobility of the ND5 gene product) in one case. Complex I, III and IV composition was normal in the other three cases indicating no major disruption of complex assembly. A differing severity of skeletal muscle respiratory chain impairment in a group of unrelated patients with severe cerebral clinical involvement is best explained by uneven tissue distribution between brain and muscle of a heteroplasmic mtDNA mutation. The relationship between MERRF and mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) encephalopathies is reappraised by extension of this hypothesis.     

Sporadic MERRF/MELAS overlap syndrome associated with the 3243 tRNALeu(UUR) mutation of mitochondrial DNA

We studied a patient with a mitochondrial encephalomyopathy characterized by the presence of all the cardinal features of both myoclonic epilepsy and ragged-red fibers (MERRF) and mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes (MELAS) syndromes. Muscle biopsy showed ragged-red fibers (RRF). Some RRF were cytochrome c oxidase (COX)-negative, while some others stained positive for COX. Muscle biochemistry revealed defects of complexes I and IV of the respiratory chain. Both muscle and blood mitochondrial DNA from the patient showed the presence of the mutation at nucleotide position 3243 in the tRNA^Leu(UUR) gene and the absence of point mutations related to MERRF syndrome. The proportions of mutant mtDNA were 70% in muscle and 30% in blood. The mutation was absent in blood from all maternal relatives, in hair follicles from the mother, and in muscle from one sister of the proband. Therefore, there was no evidence of maternal inheritance. © 1996 John Wiley & Sons, Inc.     

Myoclonus epilepsy associated with ragged-red fibers: A G-to-A mutation at nucleotide pair 8363 in mitochondrial tRNALys in two families

In addition to well-known mutations at nucleotide pair 8344 and 8356 in mitochondrial DNA in patients with myoclonus epilepsy associated with ragged-red fibers (MERRF), we found a new G-to-A point mutation at nucleotide 8363 in two Japanese families. The probands had the typical clinical characteristics of MERRF. Since the 8363 mutation was present in a heteroplasmic state, and seen in none of 92 patients with other mitochondrial diseases or 50 normal individuals, this mutation is thought to be disease-related and probably specific to MERRF. As seen in muscle biopsies with the previous two mutations, focal cytochrome c oxidase (CCO) deficiency was the most characteristic finding. With single fiber analysis, the CCO-negative fibers contained a higher percentage of mutant DNA (88.4 ± 6.6%) than CCO-positive fibers (65.1 ± 8.0%). These findings suggest that mutations in tRNA^Lys coding region are related to the MERRF phenotype and are responsible for the reduced CCO activity. © 1997 John Wiley & Sons, Inc. Muscle Nerve, 20, 271–278, 1997.     

Myoclonus epilepsy and ragged-red fibers: blood mitochondrial DNA heteroplasmy in affected and asymptomatic members of a family

By a rapid PCR-based method to assess the 8344 mtDNA mutation associated with MERRF disease, we have studied DNA from blood samples of 10 individuals belonging to a family spanning four generations in which one patient showed the complete MERRF phenotype, three other members were less severely affected, while the remaining were unaffected. The percentage of mutant mtDNA was quantified by laser-densitometric scanning of the negative photographic sheets of the agarose gels. The results showed that the MERRF patient had 53% of mutated mtDNA while the two less affected patients had 62% and 14% of mutated mtDNA, respectively. However, a high percentage of mutated genomes (up to 64%) was also found in some unaffected relatives. These results show that although on one hand the mutation is probably the primary cause of the disease, on the other hand the relative amount of mutated mtDNA in blood samples is not indicative of its clinical expression.     

Analysis of the tissue distribution and inheritance of heteroplasmic mitochondrial DNA point mutation by denaturing gradient gel electrophoresis in merrf syndrome

MERRF (Myoclonic Epilepsy and Ragged-Red Fibres) syndrome is one of the maternally inherited diseases for which a mitochondrial DNA (mtDNA) point mutation has recently been identified. The mutation is always heteroplasmic, that is normal and mutant mtDNA coexist within the same individual. We studied mtDNA heteroplasmy in two families with MERRF syndrome, using a denaturing gradient gel electrophoresis technique that avoids the errors in the evaluation of wild/mutant mtDNA ratios caused by restriction enzyme cutting in the situation of amplification of a heteroplasmic DNA. In two patients, the proportion of muscle mutant mtDNA was in agreement with the severity of muscle mitochondrial proliferation, energy defect and fibre type I predominance. In nine patients from three generations of one family, mutant mtDNA proportion in leukocytes was in relative agreement with the clinical severity of the disease. Transmission of mutant mtDNA through these three generations did not show any tendency toward homoplasmy. Homogeneity of the mutant mtDNA proportion among different tissues from one patient was demonstrated in brain, liver, muscle and heart but a possibility of divergence of the mutant mtDNA proportion during mitosis was documented in cultured skin fibroblasts.     

Mitochondrial dysfunction with myoclonus epilepsy and ragged-red fibers point mutation in nerve,muscle, and adipose tissue of a patient with multiple isymmetric lipomatosis

We report a 64-year-old man presenting with multiple symmetric lipomatosis (MSL) and mitochondrial encephalomyoneuropathy. The diagnosis of a mitochondrial cytopathy was based on the typical clinical symptoms and signs, including chronic progressive external ophthalmoplegia, hearing impairment, cerebellar ataxia, proximal myopathy, and polyneuropathy, and on molecular genetic and histological examinations. As a unique finding, the A → G⁽⁸³⁴⁴⁾ myoclonus epilepsy and ragged-red fibers point mutation was found in peripheral nerve, muscle, and adipose tissue. Muscle biopsy revealed multiple ragged-red fibers and other morphological signs of a mitochondrial myopathy. Sural nerve biopsy demonstrated a mixed axonal and demyelinating neuropathy with extensive loss of myelinated fibers and conspicuous onion bulb formations, as well as structural mitochondrial abnormalities on electron microscopy. These findings clearly demonstrate mitochondrial dysfunction in muscle, adipose tissue, and for the first time also in nervous tissue of an MSL patient, and strongly support the concept of mitochondrial cytopathy as one of the possible causes of multiple symmetric lipomatosis. © 1997 John Wiley & Sons, Inc. Muscle Nerve 20: 833–839, 1997     

MERRF/MELAS overlap syndrome associated with 3243 tRNALeu(UUR) mutation of mitochondrial DNA

We report a case of myoclonus epilepsy associated with ragged-red fibers (MERRF)/mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episode (MELAS) overlap syndrome with hearing loss, external ophthalmoplegia, and myoclonus epilepsy in addition to stroke-like episode and diabetes mellitus. Pathologically, there was degeneration in the dentate nuclei, substantia nigra, red nucleus, and subthalamic nucleus which has been reported as characteristic of MERRF, as well as necrotic lesions of various stages in the cerebral cortex, characteristic of MELAS. The gene study disclosed 3243 mutation in the tRNA^Leu(UUR) gene of mitochondrial DNA. This case is the first neuropathological report of MERRF/MELAS overlap syndrome verified by gene analyses.     

The T-C(8356) mitochondrial DNA mutation in a Japanese family

A rare point mutation at nucleotide position 8356 in the transfer RNA gene in mitochondrial DNA was found in a Japanese family. Our proband had migraine and dementia associated with lactic acidosis in addition to myoclonic epilepsy with ataxia and ragged-red fibres in a muscle biopsy specimen consistent with the clinical characteristics of myoclonic epilepsy with ragged-red fibres (MERRF). His mother, who had the same point mutation, also had migraine but without myoclonus or ataxia. His aunt, who had the same point mutation and migraine, developed diabetes mellitus, encephalomyopathy and several stroke-like episodes associated with lactic acidosis (MELAS). This is the third family with the rare mutation seen in American and Italian families. The mutation may not be specific to Caucasians, and is probably closely related to the MERRF/MELAS overlap syndrome.     

MERRF family with 8344 mutation in tRNA (lys). Evidence of a mitochondrial vasculopathy in muscle biopsies

This article reports a new MERRF family. The mother, regarded as suffering from Ramsay-Hunt Syndrome, and her three daughters, had the same clinical pattern: myoclonic epilepsy and ataxia. Two daughters were studied on morphological, biochemical and molecular genetic levels. Muscle biopsies showed ragged-red fibres and mitochondrial vasculopathy. Arterioles were strongly SDH-reactive and COX-negative. By electron microscopy, abnormal mitochondria were observed in skeletal muscle fibres, in smooth muscle fibres of intramuscular vessels and in sweat gland epithelium. The study of the respiratory chain showed complex IV and I + IV deficiency, respectively. Mitochondrial tRNA (lys) mutation at position 8344 was pointed out as previously reported in the MERRF syndrome.     

Cytochrome c oxidase activity is deficient in blood vessels of patients with myoclonus epilepsy with ragged-red fibers

Summary More than half of the intramuscular blood vessels in muscle biopsies from five patients with myoclonus epilepsy with ragged-fibers (MERRF) who had a point mutation in mitochondrial DNA at the tRNA^Lys region were darkly stained with succinate dehydrogenase (SDH) stain, showing the morphologic characteristics of strongly SDH-reactive blood vessels (SSV), but they had no cytochrome c oxidase (CCO) activity. By electron cytochemistry, the mitochondria in the smooth muscle cells of SSV had no CCO activity. On the other hand, SSV in muscle biopsies from patients with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) had normal CCO activity as shown by light and electron microscopy. The defect in CCO activity in the arteriolar smooth muscle cells and in muscle fibers suggests that CCO deficiency is related to the pathophysiology of MERRF.     

MELAS syndrome associated with a tandem duplication in the D-loop of mitochondrial DNA

We describe a family with two cases of adult-onset mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome. Interestingly, the proband also had non-insulin dependent diabetes mellitus and hyperthyroidism. Endocrinological studies demonstrated a high titer of TSH receptor antibody in the proband and elevated levels in her maternal relatives. Analysis of mitochondrial DNA (mtDNA) showed an A-to-G transition at nucleotide position 3243 in the tRNA^Leu(UUR) gene (A3243G) in the three generations of the family. Furthermore, a previously described ~ 260 bp tandem duplication in the D-loop region of mtDNA was also found in the proband and her maternal relatives. To our knowledge, such kind of duplication has never before been reported in the MELAS syndrome. The proportions of mtDNA with the ~260 bp tandem duplication and A3243G point mutation were 12.5% and 82% in the muscle, respectively, and 1.6% and 35% in the blood cells, respectively, of the proband. We conclude that the hyperthyroidism in this MELAS patient may be related to the tandem duplication in the D-loop of mtDNA. This study further substantiates the importance of searching for additional genetic mutations in mitochondrial encephalomyopathic patients with new clinical phenotypes.     

Simple detection of tRNA(Lys) mutation in myoclonus epilepsy associated with ragged-red fibers (MERRF) by polymerase chain reaction with a mismatched primer.

We developed a simple method for the detection of a tRNA(Lys) mutation in myoclonus epilepsy associated with ragged-red fibers (MERRF) by polymerase chain reaction with use of a mismatched primer. Although the tRNA(Lys) mutation does not alter recognition sequences for commercially available restriction enzymes, we have successfully changed two nucleotides flanking the A to G mutation at nucleotide position 8344 in a tRNA(Lys) gene of a mitochondrial genome. As a result, the mutation can be detected as a Nae I restriction fragment length polymorphism. With this method, all eight MERRF patients and an asymptomatic mother of a MERRF patient, from six independent families, had the same tRNA(Lys) mutation. Our method is simple and should also be useful for the quantitation of heteroplasmies.     

Tissue distribution and disease manifestations of the tRNALys A»G(8344) mitochondrial DNA mutation in a case of myoclonus epilepsy and ragged red fibres

This man with myoclonus epilepsy and ragged red fibres (MERRF) syndrome due to the tRNA^Lys A»G⁽⁸³⁴⁴⁾ mutation of mitochondrial DNA (mtDNA) died of bronchopneumonia at 18 years of age. He had progressive clinical symptoms from 6 months of age manifesting as ataxia, myoclonic seizures, and muscle weakness. A postmortem examination revealed 91–99% mutated mtDNA in all 32 examined tissue samples, including various organs and different brain regions. The brain appeared without macroscopic changes, but microscopic examination showed degeneration with loss of nerve cells and gliosis affecting the globus pallidus, substantia nigra, red nucleus, dentate nucleus, inferior olivary nucleus, cerebellar cortex, and the spinal cord. Skeletal muscle showed cytochrome c oxidase deficient muscle fibres with proliferation of mitochondria. In addition to pathological changes of muscle and brain there were few morphological changes that could be attributed to his mitochondrial disease. These data support the concept that in patients with the tRNA^Lys A»G⁽⁸³⁴⁴⁾ mutation who are manifesting disease there are high levels of mutated mtDNA in all tissues, but only some tissues and brain regions are vulnerable.     

Bilateral putaminal necrosis associated with the mitochondrial DNA A8344G myoclonus epilepsy with ragged red fibers (MERRF) mutation: an infantile case

Myoclonus epilepsy with ragged red fibers (MERRF) is one of the major mitochondrial encephalomyopathies. Its main clinical features are myoclonus epilepsy, ataxia, and myopathy with ragged red fibers. Whereas there is a close correlation between MERRF syndrome and the A8344G mutation of mitochondrial DNA, the reverse is not true. In fact, this mutation is also responsible for various other syndromes, such as Leigh syndrome, spinocerebellar degeneration, atypical Charcot-Marie-Tooth disease, and multiple truncal lipomas. We describe a child with the A8344G mutation of mitochondrial DNA and an unusual clinical, neuroradiologic, and biochemical phenotype, characterized by early-onset, nonprogressive cerebellar ataxia, and subclinical myoclonias in association with bilateral putaminal necrosis on magnetic resonance imaging and a reduction in complex V activity. Our case confirms the existence of a relationship between alteration in adenosine triphosphatase activity and basal ganglia involvement. We recommend that the possibility of a mitochondrial pathology should always be taken into consideration in the presence of bilateral symmetric lesions of the basal ganglia, even when the typical clinical picture is lacking. (J Child Neurol 2006;21:79-82).     

The 8,344 mutation in mitochondrial DNA: A comparison between the proportion of mutant DNA and clinico-pathologic findings

Ten patients, two men and eight women with mitochondrial encephalomyopathy, had an A-G mutation at nucleotide pair 8,344 in the mitochondrial DNA, the most common genetic defect in myoclonus epilepsy with ragged-red fibers (MERRF). Eight patients had the clinical and pathologic characteristics of MERRF including myoclonus, seizures, cerebellar ataxia and myopathy with ragged-red fibers. Two patients had atypical symptoms such as early onset of fatal cardiac failure and late onset of rapid mental deterioration, respectively. The striking feature in our patients with the 8,344 mutation was that four of 10 patients had cardiac involvement and two developed progressive heart failure. In the typical MERRF patients, the proportion of mutant mitochondrial DNA in their skeletal muscles, quantified by a single strand conformation polymorphism analysis, was above 85%. However, there was no significant correlation between clinical severity, histopathological findings and the proportion of mutant mtDNA in muscle biopsy samples, suggesting that non-ragged-red fibers play an important role in the phenotypic expression of the mutants.