Novel heteroplasmic mutation in the anticodon stem of mitochondrial tRNA(Lys) associated with dystonia and stroke-like episodes.

OBJECTIVES: We report a novel heteroplasmic mitochondrial tRNA(Lys) mutation associated with dystonia, stroke-like episodes, sensorineural hearing loss and epilepsy in a Hungarian family. MATERIAL AND METHODS: A 16-year-old boy, his brother and mother were investigated. Thorough clinical investigation as well as electrophysiological, neuroradiological and myopathological examinations were performed. Molecular studies included the analysis of the DYT1, DDP1/TIMM8A (deafness-dystonia peptid-1) genes and mitochondrial DNA (mtDNA). RESULTS: The mtDNA analysis of the proband revealed a heteroplasmic A8332G substitution in the anticodon stem of the tRNA(Lys) gene. The mutation segregated in all affected family members. Besides this mutation 16 further mtDNA polymorphisms were detected. Complex I activity of the patient's fibroblast cultures showed decreased activity confirming mitochondrial dysfunction. CONCLUSION: The novel A8332G heteroplasmic mutation is most likely a new cause of dystonia and stroke-like episodes due to mitochondrial encephalopathy. The synergistic effect of the G8697A, A11812G and T10463C single nucleotide polymorphisms may modify the phenotype.

Challenges for the genetic screening in dysferlin deficiency--report of an instructive case and review of the literature.

The homozygous or compound heterozygous mutation of the alleles of DYSF gene causes dysferlinopathy resulting in limb girdle muscular dystrophy Type 2B (LGMD 2B) or Miyoshi myopathy. However, patients with only 1 (heterozygous) mutation on 1 allele are increasingly recognized. Based on the Leiden database (www.dmd.nl) among 257 different mutations resulting in dysferlin-deficient muscular dystrophy, pathogenic mutations were detected only on 1 allele in 45 cases, while the exons of the other allele did not show any pathological alterations. The relatively high number of these so-called heterozygous cases raises the question if present routine molecular techniques are sufficient alone for confirming the diagnosis of dysferlin deficiency. In fact, the heterogenous genetic background of the disease makes it impossible to make the correct diagnosis without Western blot of the muscle dysferlin. This paper presents the clinical, myopathological and molecular genetic results of a 30-year-old male with dysferlinopathy as an instructive case. The cDNA sequencing of the dysferlin gene revealed a single C5302T heterozygous mutation resulting in Arg1768Trp exchange. The paper highlights the importance of the protein analysis in the diagnosis of dysferlin deficiency, discusses the difficulties of the complete genomic analysis of the dysferlin gene alterations and the possible etiopathogenetic role of the noncoding DNA sequence of the dysferlin gene in dysferlin deficiencies.

Myotonic dystrophy in two European grey wolves (Canis lupus).

Two related European Grey wolves (Canis lupus) with the history of muscle stiffness beginning at 2 weeks of age were examined in this study. Muscle tone and muscle mass were increased in both animals. Muscle stiffness was worsened by stress so that the animals fell into lateral recumbency. Blood chemistry revealed mildly increased serum creatine kinase activity. Abnormal potentials typical of myotonic discharges were recorded by electromyography. Cataract, first-degree atrioventricular (AV) block and inhomogeneous myocardial texture by ultrasound suggested extramuscular involvement. Myopathology demonstrated dystrophic signs in the muscle biopsy specimen. The presumptive diagnosis based on the in vivo findings was myotonic dystrophy. Immunochemistry of the striated muscles revealed focal absence of dystrophin 1 and beta-dystroglycan in both cases. Cardiac and ophthalmologic involvement suggested a disorder very similar to a human form of myotonic dystrophy. This is the first description of myotonic dystrophy in wolves.

Peripheral nerve and skeletal muscle involvement in CADASIL.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is characterized by degeneration of vascular smooth muscle cells (VSMC) of nearly all tissues studied so far. The clinical phenotype of CADASIL shows great variability. The disease is caused by mutations of the Notch3 gene encoding the transmembrane receptor Notch3, which is expressed predominantly in VSMC. In some patients, neuromuscular symptoms have been described. To investigate the fine structural features of peripheral nerve and muscle biopsy specimens in more cases and greater detail, seven electron microscopically confirmed CADASIL patients showing a variable amount of granular osmiophilic material on the surface of VSMC were included in this study. Pathogenic mutations within the cluster region (exon 3 and 4) of the Notch3 gene were identified in six cases. Degeneration and regeneration of nerve fibers in the sural nerves, studied in four cases, was present, although moderate, in all nerve biopsy specimens, whereas an intramuscular nerve fascicle showed more severe changes. Enlarged mitochondria with needle-like calcium precipitates were repeatedly seen. In muscle biopsy specimens, some degree of neurogenic atrophy was apparent in addition to myopathic changes, including occasional ragged red fibers with abnormally large mitochondria, focal tubular aggregates, abnormal terminal cisternae, and myofibrillary abnormalities. Automated sequence analysis of the whole mitochondrial DNA performed in one patient revealed several nucleotide polymorphisms, which were not considered pathogenic. The findings suggest that in CADASIL degeneration of small blood vessels is initiated by defects of the surface membrane of VSMC. Dysfunction of these blood vessels may cause low-grade chronic ischemia with secondary hypoxidosis and a large variety of structural changes noted in skeletal muscle and peripheral nerves, although a primary influence of the underlying genetic defect can not be excluded.

Marked phenotypic variation in a family with a new myelin protein zero mutation.

Myelin protein zero (MPZ) is a member of the immunoglobulin gene superfamily, which has a role in myelin compaction. MPZ gene mutations cause mostly demyelinating neuropathies of the Charcot-Marie-Tooth 1B type (CMT1B), but axonal CMT have been described as well. There is a broad spectrum of phenotypic manifestation of neuropathies caused by MPZ mutations. Some mutations of MPZ cause severe early-onset neuropathies such as Dejerine-Sottas disease, while others cause the classical CMT phenotype with normal early milestones but development of disability during the first two decades of life. We describe a family in which five members of three consecutive generations had a heterozygous mutation in nucleotide position 143 with a T-C transition in exon 2 of the MPZ gene. The resulting substitution of Leu48 with proline has not been previously described. The age of onset of symptoms varied from 8 months to 41 years. The marked variation of the age of disease onset and clinical phenotype in this one family, related to the same MPZ mutation, suggests that in addition to the type and intragenic location of the mutation, other putative modifying gene(s) are regulating MPZ gene expression, mRNA stability and posttranslational protein modification and may have an important effect on the ultimate clinical phenotype.

Cerebral blood flow and glucose metabolism in mitochondrial disorders.

OBJECTIVE: To investigate cerebral metabolism by 2-[18F]fluorodeoxy-d-glucose (FDG) uptake using PET and cerebrovascular reverse capacity by transcranial Doppler sonography (TCD) in different mitochondrial diseases (mitochondrial myopathy; mitochondrial encephalopathy, lactacidosis, and stroke-like episodes [MELAS]; and chronic external ophthalmoplegia). BACKGROUND: Previous studies on individual patients with mitochondriopathies revealed abnormal accumulations of mitochondria in endothelium, smooth muscle cells, and pericytes of blood vessels in different parts of the nervous system (cerebrum, cerebellum, sural nerve) and skeletal muscle. On this basis, some investigators suggested a pathogenic role of vascular involvement in the MELAS syndrome and other encephalopathies. smhd1 DESIGN/METHODS: The authors investigated neuronal metabolism and cerebrovascular involvement with PET in 5 cases and with TCD with acetazolamide stimulation in 15 cases. The patients were divided into three groups: 1) interictal MELAS (n = 4); 2) progressive external ophthalmoplegia (n = 6); and 3) pure mitochondrial myopathy and neuropathy (n = 5). The results were compared with those from matched normal control subjects. The diagnoses were based on clinical phenotype as well as histopathologic and molecular analysis. RESULTS: Cerebral glucose uptake was impaired in all patients, both with and without CNS symptoms, particularly in the occipital and temporal lobes. The vasoreactivity of the small arterioles to acetazolamide did not differ significantly between the patients and healthy control subjects or between the different groups of mitochondrial disorders. CONCLUSIONS: MELAS does not appear to be a functional disturbance of arterioles leading to an ischemic vascular event. The clinical symptoms in MELAS are not the result of a mitochondrial angiopathy but are the consequences of a mitochondrial cytopathy affecting neurons or glia. There is no correlation between the decreased glucose metabolism and the duration of the disease.