New phenotypic diversity associated with the mitochondrial tRNA(SerUCN) gene mutation.

We performed detailed clinical, histopathological, biochemical, in vitro translation and molecular genetic analysis in patients from two unrelated families harbouring the tRNA(SerUCN) 7472C-insertion mutation. Proband 1 developed a progressive neurodegenerative phenotype characterised by myoclonus, epilepsy, cerebellar ataxia and progressive hearing loss. Proband 2 had a comparatively benign phenotype characterised by isolated myopathy with exercise intolerance. Both patients had the 7472C-insertion mutation in identical proportions and they exhibited a similar muscle biochemical and histopathological phenotype. However, proband 2 also had a previously unreported homoplasmic A to C transition at nucleotide position 7472 in the tRNA(SerUCN) gene. This change lengthens further the homopolymeric C run already expanded by the 7472C-insertion. These data extend the phenotypic range associated with the 7472C-insertion to include isolated skeletal myopathy, as well as a MERRF-like phenotype.

A novel mutation in the mitochondrial tRNA(TYr) gene associated with exercise intolerance.

The authors report a novel A5874G mutation in the mitochondrial tRNA tyrosine (tRNA(TYr)) gene associated with exercise intolerance, limb weakness, and complex III deficiency. The mutation was absent in blood from the patient and all maternal family members, indicating that it may be a spontaneous somatic mutation in muscle. This is the first point mutation in the tRNA(TYr) gene associated with human disease and is further evidence that exercise intolerance associated with complex III deficiency is genetically heterogeneous.

The mitochondrial DNA G13513A transition in ND5 is associated with a LHON/MELAS overlap syndrome and may be a frequent cause of MELAS.

We report on 4 male patients with clinical, radiological, and muscle biopsy findings typical of the mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes (MELAS) phenotype. Skeletal muscle mitochondrial DNA (mtDNA) analysis showed that all patients harbored a heteroplasmic G13513A mutation in the ND5 subunit gene. One of these cases (Patient 1) presented with symptoms characteristic of Leber's hereditary optic neuropathy (LHON) 2 years before the first stroke-like episode. Quantitative analysis in several postmortem tissue sections showed that the relative proportions of mutant mtDNA were generally lower than those reported with other pathogenic mtDNA mutations. Single-fiber polymerase chain reaction studies demonstrated significantly higher amounts of mutant mtDNA in ragged red fibers (RRFs) compared with non-RRFs. This study indicates that the G13513A transition is likely to be pathogenic, that it can cause an LHON/MELAS overlap syndrome, and that it may be a more frequent cause of MELAS than previously recognized.

Exercise intolerance due to mutations in the cytochrome b gene of mitochondrial DNA.

BACKGROUND: The mitochondrial myopathies typically affect many organ systems and are associated with mutations in mitochondrial DNA (mtDNA) that are maternally inherited. However, there is also a sporadic form of mitochondrial myopathy in which exercise intolerance is the predominant symptom. We studied the biochemical and molecular characteristics of this sporadic myopathy. METHODS: We sequenced the mtDNA cytochrome b gene in blood and muscle specimens from five patients with severe exercise intolerance, lactic acidosis in the resting state (in four patients), and biochemical evidence of complex III deficiency. We compared the clinical and molecular features of these patients with those previously described in four other patients with mutations in the cytochrome b gene. RESULTS: We found a total of three different nonsense mutations (G15084A, G15168A, and G15723A), one missense mutation (G14846A), and a 24-bp deletion (from nucleotide 15498 to 15521) in the cytochrome b gene in the five patients. Each of these mutations impairs the enzymatic function of the cytochrome b protein. In these patients and those previously described, the clinical manifestations included progressive exercise intolerance, proximal limb weakness, and in some cases, attacks of myoglobinuria. There was no maternal inheritance and there were no mutations in tissues other than muscle. The absence of these findings suggests that the disorder is due to somatic mutations in myogenic stem cells after germ-layer differentiation. All the point mutations involved the substitution of adenine for guanine, but all were in different locations. CONCLUSIONS: The sporadic form of mitochondrial myopathy is associated with somatic mutations in the cytochrome b gene of mtDNA. This myopathy is one cause of the common and often elusive syndrome of exercise intolerance.

Transthyretin Leu12Pro is associated with systemic, neuropathic and leptomeningeal amyloidosis.

We report a middle-aged woman with a novel transthyretin (TTR) variant, Leu12Pro. She had extensive amyloid deposition in the leptomeninges and liver as well as the involvement of the heart and peripheral nervous system which characterizes familial amyloid polyneuropathy caused by variant TTR. Clinical features attributed to her leptomeningeal amyloid included radiculopathy, central hypoventilation, recurrent subarachnoid haemorrhage, depression, seizures and periods of decreased consciousness. MRI showed a marked enhancement throughout her meninges and ependyma, and TTR amyloid deposition was confirmed by meningeal biopsy. The simultaneous presence of extensive visceral amyloid and clinically significant deposits affecting both the peripheral and central nervous system extends the spectrum of amyloid-related disease associated with TTR mutations. The unusual association of severe peripheral neuropathy with symptoms of leptomeningeal amyloid indicates that leptomeningeal amyloidosis should be considered part of the syndrome of TTR-related familial amyloid polyneuropathy.

Mitochondrial encephalomyopathies: the enigma of genotype versus phenotype.

Over the past decade a large body of evidence has accumulated implicating defects of human mitochondrial DNA in the pathogenesis of a group of disorders known collectively as the mitochondrial encephalomyopathies. Although impaired oxidative phosphorylation is likely to represent the final common pathway leading to cellular dysfunction in these diseases, fundamental issues still remain elusive. Perhaps the most challenging of these is to understand the mechanisms which underlie the complex relationship between genotype and phenotype. Here we examine this relationship and discuss some of the factors which are likely to be involved.

MELAS: a new disease associated mitochondrial DNA mutation and evidence for further genetic heterogeneity.

OBJECTIVES: To define the molecular genetic basis of the MELAS phenotype in five patients without any known mutation of mitochondrial DNA. METHODS: Systematic automated mitochondrial DNA sequencing of all mitochondrial transfer RNA and cytochrome c oxidase genes was undertaken in five patients who had the MELAS phenotype. RESULTS: A novel heteroplasmic mitochondrial DNA mutation was identified in the transfer RNA gene for phenylalanine in one case (patient 3). This mutation was not detected in the patient's blood or in her mother's blood. No pathogenic mutations were identified in the other four patients. CONCLUSIONS: This is the first point mutation in the transfer RNA gene for phenylalanine to be associated with MELAS. The absence of mutations in the remaining four patients suggests that there is further genetic heterogeneity associated with this mitochondrial phenotype.

Salbutamol treatment in a patient with hyperkalaemic periodic paralysis due to a mutation in the skeletal muscle sodium channel gene (SCN4A).

A 35 year old woman with clinical features of hyperkalaemic periodic paralysis confirmed on provocative exercise testing was investigated. DNA sequence analysis of the gene for the alpha-subunit of the skeletal muscle voltage gated sodium channel (SCN4A) on chromosome 17q23 identified a point mutation at nucleotide position 2188. This results in a threonine to methionine substitution at amino acid position 704. The patient was intolerant of diuretic medication but showed a striking clinical and electrophysiological improvement with salbutamol therapy. Treatment with beta-adrenergic agents should be considered in patients with hyperkalaemic periodic paralysis who are intolerant of, or resistant to, diuretic medications.

Fifty year follow-up of a patient with central core disease shows slow but definite progression.

The follow-up of a patient with central core disease (CCD) over 50 years showed that although initially the condition was moderately non-progressive, progression of a significant degree did eventually occur. Histopathological and electron microscopic data were available from muscle biopsies carried out at the ages of 19 and 55 years, and show a marked predominance of type 1 fibres with central cores in most fibres at both ages. The four mutations within the RYR1 gene described in association with CCD and three of the more common malignant hyperthermia-associated mutations within RYR1 were not present.

Cytochrome c oxidase deficiency associated with the first stop-codon point mutation in human mtDNA.

We have identified the first stop-codon point mutation in mtDNA to be reported in association with human disease. A 36-year-old woman experienced episodes of encephalopathy accompanied by lactic acidemia and had exercise intolerance and proximal myopathy. Histochemical analysis showed that 90% of muscle fibers exhibited decreased or absent cytochrome c oxidase (COX) activity. Biochemical studies confirmed a severe isolated reduction in COX activity. Muscle immunocytochemistry revealed a pattern suggestive of a primary mtDNA defect in the COX-deficient fibers and was consistent with either reduced stability or impaired assembly of the holoenzyme. Sequence analysis of mtDNA identified a novel heteroplasmic G-->A point mutation at position 9952 in the patient's skeletal muscle, which was not detected in her leukocyte mtDNA or in that of 120 healthy controls or 60 additional patients with mitochondrial disease. This point mutation is located in the 3' end of the gene for subunit III of COX and is predicted to result in the loss of the last 13 amino acids of the highly conserved C-terminal region of this subunit. It was not detected in mtDNA extracted from leukocytes, skeletal muscle, or myoblasts of the patient's mother or her two sons, indicating that this mutation is not maternally transmitted. Single-fiber PCR studies provided direct evidence for an association between this point mutation and COX deficiency and indicated that the proportion of mutant mtDNA required to induce COX deficiency is lower than that reported for tRNA-gene point mutations. The findings reported here represent only the second case of isolated COX deficiency to be defined at the molecular genetic level and reveal a new mutational mechanism in mitochondrial disease.

Growth hormone therapy may benefit protein metabolism in mitochondrial encephalomyopathy.

Mitochondrial encephalomyopathy is a genetic disorder for which there is at present no cure. Conventional treatment regimes may not be effective in preventing weight loss and muscle wasting in many patients. Recombinant human GH has been shown to have anabolic effects on protein metabolism and to reduce muscle wasting in various diseases. We have treated a patient known to have myoclonus, epilepsy with ragged red fibres (MERRF) with a high protein diet for 1 month followed by a high protein diet and GH therapy for 1 month. To assess the benefit of these treatments the patient underwent whole body protein turnover, myometric and body composition studies at baseline, following the high protein diet (100 g/day) and following GH therapy. Whole body protein synthesis (and protein breakdown) increased following a high protein intake and was further enhanced by treatment with GH and in a high protein diet. Body composition did not change significantly following treatment with either the high protein diet or GH but there was an improvement in muscle performance following GH treatment. Mitochondrial encephalomyopathy, a wasting disorder, may be a disease in which the known protein anabolic effect of GH may have a therapeutic benefit.

Nemaline myopathy: an unusual cause of ophthalmoparesis.

Ophthalmoparesis and ptosis are extremely rare in nemaline myopathy. A 45-year-old man with a long history of bilateral ptosis and a 1-year history of diplopia is reported. Leg and arm weakness and wasting had been present since childhood, with a very slow deterioration over time. On examination, there was nonfatigueable bilateral ptosis that was more marked on the right. There was diplopia on left gaze. Extraocular movements showed limitation of elevation and adduction of the right eye. There was bilateral facial weakness, as well as proximal and distal wasting and weakness in the arms and legs. Electromyography (EMG) showed a combination of myopathic and neurogenic changes. Triceps muscle biopsy showed small multiple collections of rod-like structures in > 50% of fibers. This patient presented with a clinical picture that did not primarily suggest nemaline myopathy. This case illustrates the heterogeneity of this disorder and the need for muscle biopsy to make an accurate diagnosis in patients with ptosis and progressive external ophthalmoparesis.

A rapidly progressive adolescent-onset oculopharyngeal somatic syndrome with rimmed vacuoles in two siblings.

We describe 2 Greek siblings who developed a rapidly progressive oculopharyngeal somatic syndrome, at the ages of 11 and 14 years, with muscle biopsies showing rimmed vacuoles and, in 1 case, cytoplasmic and intranuclear tubulofilamentous inclusions 25 nm in diameter. Although a similar pattern of muscle involvement with rimmed vacuoles is described in autosomal dominant oculopharyngeal muscular dystrophy, the age of onset, the rapid progression of the symptoms, and the nature of the tubulofilaments distinguish this as a separate entity.

Mitochondrial encephalopathy with multiple mitochondrial DNA deletions: a report of two families and two sporadic cases with unusual clinical and neuropathological features.

A mitochondrial myopathy associated with multiple deletions of mitochondrial DNA has been identified in pedigrees showing an autosomal dominant mode of inheritance. We report the first two British kindreds with this disorder, and two sporadic cases. The families exhibited some unusual clinical features, including pigmentary retinopathy and tremor; the latter was levodopa-responsive and associated with rigidity and micrographia in one family. Members of one pedigree and both sporadic patients had a peripheral neuropathy and nerve biopsy showed marked axonal degeneration. Post-mortem examination of one patient without parkinsonism showed severe neuronal loss in the substantia nigra.

Affinity of antigen-specific IgG distinguishes multiple sclerosis from encephalitis.

The characteristics of antigen-specific IgG in patients with multiple sclerosis and patients with encephalitis have been compared. Both groups of patients showed antigen-specific oligoclonal bands locally synthesised in the CSF. When the affinity distribution of the antigen-specific IgG was measured there was a marked difference between the two groups. Encephalitis patients had high affinity antibody against the causative antigen. This was consistent with the antibody undergoing affinity maturation as a result of the immune system fighting a primary infection. Multiple sclerosis patients lacked high affinity response. This lack of high affinity antibody was also seen in those encephalitis patients when antigens other than the causative antigen were studied.

Kearns-Sayre syndrome associated with mitochondrial DNA deletion or duplication: a molecular genetic and pathological study.

The neuropathological findings in 2 patients with Kearns-Sayre syndrome and mitochondrial DNA (mtDNA) rearrangements, one a predominant deletion and the other a predominant duplication, were remarkably similar, showing diffuse vacuolation of white matter. There were some of the pathological features of Leigh's syndrome in the spinal cord of the patient with a duplication. In the patient with a predominant deletion, rearranged mtDNA was undetectable in blood, spleen, and testis, and present in highest amounts in muscle and the brain, but relatively low in cerebellum, reflecting the ratio seen, albeit in much smaller amounts, in normal aged brains. MtDNA rearrangements in this patient were largely deletions or deletion dimers; duplicated mtDNA was present in only trace amounts in some tissues and there was none in skeletal muscle. The patient with a predominant duplication of mtDNA had higher amounts of rearranged mtDNA in blood (mainly duplicated) than muscle (mainly deleted). Correlation of these data with tissue dysfunction is probably complicated by the replicative behaviour of deleted, duplicated and normal mtDNA.