RESUMO
BACKGROUND AND PURPOSE: Most mitochondrial disorders with onset in early childhood are progressive and involve multiple organs. The m.3250T>C mutation in MTTL1 has previously been described in a few individuals with a possibly riboflavin-responsive myopathy and an association with sudden infant death syndrome was suspected. We describe a large family with this mutation and evaluate the effect of riboflavin treatment. METHODS: Medical data were collected with the help of a standardized data collection form. Sanger sequencing was used to screen for variants in mitochondrial DNA and the proportion of the mutation was analyzed in different tissues. Biochemical and muscle morphological investigations of muscle tissue were performed in two individuals. The effect of riboflavin treatment was evaluated in two individuals. RESULTS: Thirteen family members experienced exercise intolerance with fatigue and weakness. Inheritance was maternal with 100% penetrance. The course was either static or showed improvement over time. There was no evidence of other organ involvement except for a possible mild transient cardiac enlargement in one child. Muscle investigations showed isolated complex I deficiency and mitochondrial proliferation. The level of m.3250T>C was apparently 100%, i.e. homoplasmic, in all examined tissues. Riboflavin treatment showed no effect in any treated family member and there have been no cases of sudden infant death in this family. CONCLUSIONS: This study illustrates the importance of considering mitochondrial disorders in the work-up of individuals with exercise intolerance and provides a better understanding of the phenotype associated with the m.3250T>C mutation in MTTL1.
Assuntos
DNA Mitocondrial/genética , Tolerância ao Exercício/genética , Miopatias Mitocondriais/genética , Mutação , RNA de Transferência/genética , Adulto , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Mitocôndrias/genética , Miopatias Mitocondriais/tratamento farmacológico , Linhagem , Fenótipo , Riboflavina/uso terapêutico , Complexo Vitamínico B/uso terapêutico , Adulto JovemRESUMO
UNLABELLED: BACKGROUND, OBJECTIVE AND METHODS: We describe a female patient with a mitochondrial encephalopathy, lactic acidosis and stroke-like episodes syndrome. As a child, she developed epilepsy and stroke-like episodes giving cognitive impairment and ataxia but no hearing impairment. At the age of 44 years, she suffered a cerebral sinus thrombosis which was warfarin treated. One month later, she developed an episode of severe acidosis associated with encephalopathy and myelopathy. RESULTS: She was found to harbour a 7512T>C mutation in the mitochondrial encoded tRNA(Ser(UCN)) gene (MTTS1). The mutation load was 91% in muscle and 24% in blood. Enzyme histochemical analysis of the muscle tissue showed numerous cytochrome c oxidase (COX)-negative fibres. Restriction fragment length polymorphism (RFLP) analysis of single muscle fibres showed significantly higher level (median 97%, range: 94-99%) of the mutation in the COX-negative fibres compared with COX-positive fibres (median 36%, range: 12-91%), demonstrating the pathogenic effect of the mutation. Different levels of heteroplasmy (range 34-61%) were detected in hair shafts analysed by RFLP. CONCLUSION: This case adds to the spectrum of clinical presentations, i.e. sinus thrombosis, in patients having MTTS1 mutations.
Assuntos
Síndrome MELAS/genética , Mutação Puntual/genética , RNA de Transferência de Serina/genética , Adulto , Análise Mutacional de DNA/métodos , Feminino , Humanos , Síndrome MELAS/patologia , Síndrome MELAS/fisiopatologiaRESUMO
Limb-girdle muscular dystrophy (LGMD) type 2I, caused by mutations in the fukutin-related protein gene (FKRP), is one of the most common forms of LGMD in childhood. We describe two patients with LGMD2I and a Duchenne-like phenotype. In addition to the common L276I mutation, both patients had a new mutation in FKRP, L169P and P89L, respectively. Clinical onset was triggered by viral upper respiratory tract infections. In addition to the common dystrophic pattern with a weak immune histochemical staining for alpha-dystroglycan, muscle biopsy showed inflammatory changes. This was especially striking in one of the patients with up-regulation of MHC class 1 antigen, suggestive of myositis. Both patients showed a good clinical response to treatment with prednisolone, which was initiated at daily dosage of 0.35 mg/kg/day. Our results provide evidence for an inflammatory involvement in the pathological expression of LGMD2I and open up the possibility that this disorder could be treatable with corticosteroids.
Assuntos
Inflamação/tratamento farmacológico , Distrofia Muscular do Cíngulo dos Membros/tratamento farmacológico , Esteroides/uso terapêutico , Adolescente , Criança , Análise Mutacional de DNA , Distroglicanas/metabolismo , Humanos , Inflamação/etiologia , Inflamação/genética , Inflamação/patologia , Estudos Longitudinais , Masculino , Distrofia Muscular do Cíngulo dos Membros/complicações , Distrofia Muscular do Cíngulo dos Membros/genética , Distrofia Muscular do Cíngulo dos Membros/patologia , Mutação , Pentosiltransferases , Proteínas/genéticaRESUMO
We describe a second patient with the 583G>A mutation in the tRNA(phe) gene of mitochondrial DNA (mtDNA). This 17-year-old girl had a mitochondrial myopathy with exercise intolerance and an asymptomatic retinopathy. Muscle investigations showed occasional ragged red fibers, 30% cytochrome c oxidase (COX)-negative fibers, and reduced activities of complex I+IV in the respiratory chain. The mutation was heteroplasmic (79%) in muscle but undetectable in other tissues. Analysis of single muscle fibers revealed a significantly higher level of mutated mtDNA in COX-negative fibers. Our study indicates that the 583G>A mutation is pathogenic and expands the clinical spectrum of this mutation.
Assuntos
Miopatias Mitocondriais/genética , Mutação/genética , RNA de Transferência de Fenilalanina/genética , RNA/genética , Doenças Retinianas/genética , Adolescente , Análise Mutacional de DNA , Transporte de Elétrons/genética , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Tolerância ao Exercício/genética , Feminino , Humanos , Miopatias Mitocondriais/complicações , Miopatias Mitocondriais/fisiopatologia , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/patologia , Debilidade Muscular/genética , Debilidade Muscular/fisiopatologia , RNA Mitocondrial , Retina/patologia , Retina/fisiopatologia , Artéria Retiniana/patologia , Artéria Retiniana/fisiopatologia , Doenças Retinianas/complicações , Doenças Retinianas/fisiopatologiaRESUMO
OBJECTIVE: The epidemiology of giant cell arteritis (GCA) may indicate a pathogenetic relationship between GCA and female sex hormone metabolism; GCA is two to four times more common in women compared with men. Our previous analyses gave no support for the hypothesis that the pathogenesis of GCA should be related to somatic mutations in the estrogen receptor alpha (ERalpha) gene. The object of the present study was to investigate the size of the estrogen receptor beta (ERBeta), and the size and nucleotide sequence of the ERBeta gene in temporal arteries in GCA. METHODS: The ERBeta protein was analyzed by Western blot technique and the ERBeta gene by RT-PCR and direct sequencing of the PCR product. RESULTS: Western blot analysis revealed an ERBeta of normal size. There were no aberrations in size or nucleotide sequence in the ERBeta gene in the GCA patients. CONCLUSION: The present observations gave no support for the hypothesis that somatic mutations in the ERBeta gene should be involved in the pathogenesis of GCA.
Assuntos
Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/fisiologia , Arterite de Células Gigantes/etiologia , Arterite de Células Gigantes/genética , Idoso , Idoso de 80 Anos ou mais , Sequência de Aminoácidos , Western Blotting , Receptor beta de Estrogênio/análise , Receptor beta de Estrogênio/química , Feminino , Arterite de Células Gigantes/fisiopatologia , Humanos , Masculino , Pessoa de Meia-Idade , Mutação/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Caracteres Sexuais , Artérias Temporais/químicaRESUMO
Inclusion body myositis (IBM) is an autoimmune, inflammatory myopathy where morphological changes of muscle, including ragged red fibers, have indicated mitochondrial dysfunction in some muscle fibers. In this study enzyme histochemical analysis showed that cytochrome c oxidase (COX)-deficient muscle fibers were present at a frequency ranging from 0.5 to 5% of the muscle fibers in a series of 20 IBM patients. In age-matched controls, only occasional COX-deficient muscle fibers were present. Polymerase chain reaction (PCR) analysis of DNA extracted from muscle tissue of the IBM patients showed multiple mtDNA deletions. PCR analysis of isolated, single muscle fibers showed presence of mtDNA with only one type of deletion and deficiency of wild-type mtDNA in each COX-deficient muscle fiber. This finding was supported by results from in situ hybridization using different mtDNA probes on consecutive sections. A 5 kb deletion was identified in all 20 IBM patients. DNA sequencing of the breakpoint region showed that this deletion was the so-called "common deletion." Most but not all of the investigated deletion breakpoints were flanked by direct repeats. COX-deficient fibers were more frequent among fibers with positive immunostaining with antibodies directed toward a regeneration marker, the Leu-19 antigen, than in the entire fiber population. These results show that COX deficiency in muscle fiber segments in IBM is associated with deletions of mtDNA. Clonal expansion of mtDNA with deletions may take place in regenerating muscle fibers following segmental necrosis.
Assuntos
Deficiência de Citocromo-c Oxidase , DNA Mitocondrial/genética , Miopatias Mitocondriais/genética , Fibras Musculares Esqueléticas/química , Proteínas Musculares/deficiência , Deleção de Sequência , Adulto , Idoso , Complexo IV da Cadeia de Transporte de Elétrons/genética , Humanos , Hibridização In Situ , Corpos de Inclusão/patologia , Pessoa de Meia-Idade , Miopatias Mitocondriais/patologia , Fibras Musculares Esqueléticas/enzimologia , Fibras Musculares Esqueléticas/patologia , Proteínas Musculares/genética , Dor/enzimologia , Dor/genética , RegeneraçãoRESUMO
OBJECTIVE: To relate signs and symptoms to morphologic changes and presence of multiple mitochondrial DNA (mtDNA) deletions in a patient with autosomal dominant progressive external ophthalmoplegia (adPEO) and mitochondrial myopathy. BACKGROUND: An etiologic association between the somatic multiple mtDNA deletions in adPEO and clinical manifestations other than the myopathy has so far not been demonstrated. METHODS: The authors investigated a patient with adPEO and multiorgan system manifestations including levodopa-responsive parkinsonism. She died at age 61 years of pancreatic carcinoma. Autopsy tissue specimens were investigated for morphologic alterations and occurrence of mtDNA deletions by Southern blot and long-extension PCR analyses. RESULTS: The patient had carcinoma of the pancreas with metastases to liver, lymph nodes, and bone marrow. The brain revealed slight gliosis of the gray and white matter and degeneration of the substantia nigra. The myocardium showed focal areas with loss and atrophy of myocytes and fibrosis. Analysis of mtDNA revealed multiple deletions in different regions of the brain, skeletal muscle, and myocardium. Twenty-five different mtDNA deletions were identified. Most of these were flanked by large direct-sequence repeats. Six identical deletions were found in muscle and brain. CONCLUSIONS: These findings indicate that somatic multiple mtDNA deletions are associated with degenerative tissue changes and clinical manifestations in adPEO.
Assuntos
Encéfalo/patologia , DNA Mitocondrial/genética , Oftalmoplegia Externa Progressiva Crônica/genética , Oftalmoplegia Externa Progressiva Crônica/patologia , Deleção de Sequência , Antiparkinsonianos/uso terapêutico , Córtex Cerebral/patologia , Feminino , Genes Dominantes , Humanos , Levodopa/uso terapêutico , Pessoa de Meia-Idade , Músculo Esquelético/patologia , Doença de Parkinson/complicações , Doença de Parkinson/tratamento farmacológico , Sequências Repetitivas de Ácido NucleicoRESUMO
We investigated the distribution in skeletal muscle of mitochondrial DNA (mtDNA) with the tRNA(Lys) A8344G mutation, which is associated with myoclonus epilepsy and ragged red fibres (MERRF) syndrome. Isolated muscle fibre segments (n = 144) from six individuals of two different families carrying the mutation were studied. Two of these individuals were affected by MERRF while four had no or minor clinical symptoms. In one individual with a low overall level of mutated mtDNA (mean 18%) the variation in the proportion of mutated mtDNA between individual muscle fibres ranged from 0 to 80%. This result demonstrates that segregation of the tRNA(Lys) A8344G mutation within a tissue may lead to very marked variation of the level of mutated mtDNA between individual cells. There was a very high apparent threshold level of mutated mtDNA (95.3-97.7%) for expression of histochemical cytochrome c oxidase (COX) deficiency in individual muscle fibres. The results indicated that this apparent threshold level varied slightly between patients. Ultrastructural examination revealed that an appreciable proportion of the mitochondria in COX-positive muscle fibres lacked COX activity. Variation in intercellular and interorganellar distribution of mutated mtDNA in addition to the absolute mtDNA copy number may explain differences in clinical phenotypes in patients with high levels of the tRNA(Lys) A8344G mutation.
Assuntos
Fibras Musculares Esqueléticas/metabolismo , Mutação , RNA de Transferência de Lisina/biossíntese , RNA de Transferência de Lisina/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , DNA Mitocondrial/genética , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Feminino , Histocitoquímica , Humanos , Masculino , Fibras Musculares Esqueléticas/enzimologia , Linhagem , Reação em Cadeia da PolimeraseRESUMO
We report a novel heteroplasmic T-->C mutation at nt position 582 within the mitochondrial tRNA(Phe) gene of a 70-year-old woman with mitochondrial myopathy. No other family members were affected, suggesting that our patient was a sporadic case. The muscle showed frequent ragged red fibers and 43% cytochrome c oxidase deficient fibers. The mutation alters a conserved base pairing in the aminoacyl acceptor stem. The mutation load was 70% in muscle homogenate and varied from 0 to 95% in individual muscle fiber segments. Cytochrome c oxidase-negative fibers showed significantly higher levels of mutated mtDNA (>75%) than Cytochrome c oxidase-positive fibers (<55%). This mutation adds to the previously described four pathogenic mutations in the tRNA(Phe) gene.
Assuntos
DNA Mitocondrial/genética , Mitocôndrias/genética , Miopatias Mitocondriais/genética , Músculo Esquelético/metabolismo , Mutação Puntual/genética , RNA de Transferência de Fenilalanina/genética , Idoso , Substituição de Aminoácidos/genética , Aminoacil-tRNA Sintetases/genética , Respiração Celular/genética , Análise Mutacional de DNA , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Feminino , Humanos , Mitocôndrias/metabolismo , Miopatias Mitocondriais/diagnóstico , Miopatias Mitocondriais/patologia , Músculo Esquelético/patologia , Homologia de Sequência do Ácido NucleicoRESUMO
Inclusion body myositis (IBM) is the most common muscle disease affecting individuals over 50 years of age. The inflammatory reaction is characterized by cell infiltrates predominated by CD8+ cytotoxic T cells. To analyze clonality of muscle infiltrating lymphocytes, we studied the complementarity determining region 3 (CDR3) length distribution of the T cell receptor (TCR). Muscle infiltrating lymphocytes were studied in three IBM patients and compared with peripheral blood lymphocytes (PBL) in two of these patients. The study was performed by reverse transcription polymerase chain reaction (RT-PCR) of RNA extracted from muscle tissue and PBL followed by analysis of fragment length distribution of the CDR3 region in each of 24 different Vbeta families. There was a restricted usage of TCR Vbeta gene families in muscle infiltrating T cells in all three patients. Some of the TCR Vbeta gene families showed oligoclonal expansions but polyclonal patterns were dominating. The CDR3 distribution of most Vbeta families differed between muscle infiltrating lymphocytes and PBL indicating that T cells have expanded locally or selectively accumulated in muscle.
Assuntos
Corpos de Inclusão/imunologia , Miosite/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Idoso , Idoso de 80 Anos ou mais , Biópsia , Primers do DNA , DNA Complementar , Feminino , Humanos , Região Variável de Imunoglobulina/genética , Região Variável de Imunoglobulina/imunologia , Corpos de Inclusão/patologia , Masculino , Pessoa de Meia-Idade , Miosite/patologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/química , Linfócitos T/imunologiaRESUMO
Inclusion body myositis (IBM) is the most common muscle disease affecting individuals over 50 years of age. An important feature of IBM is invasion of muscle fibers by T cells. The muscle infiltrating T cells show a restricted usage of variable (V) alpha/beta gene families. In this study we have investigated the clonality of T cells using two of the predominant V beta families i.e. V beta 3 and V beta 8 in three patients with IBM. The study was performed by reverse transcription and polymerase chain reaction (RT-PCR) analysis, followed by cloning and sequencing of the T cell receptor complementarity determining region 3. We found oligoclonal expansion of V beta 3 bearing muscle infiltrating T cells in two patients and of V beta 8 in one patient, supporting the concept that antigen stimulated T cells are important in the pathogenesis of IBM. Results of HLA typing indicated a genetic predisposition for the disease by the presence of DR3, DR52 and DQB1*0201/0202 in all three patients.
Assuntos
Clonagem Molecular , Músculos/patologia , Miosite de Corpos de Inclusão/patologia , Linfócitos T/patologia , Idoso , Sequência de Aminoácidos , Feminino , Humanos , Região Variável de Imunoglobulina/genética , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Família Multigênica , Miosite de Corpos de Inclusão/genética , Reação em Cadeia da Polimerase , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Linfócitos T/metabolismo , Transcrição GênicaRESUMO
OBJECTIVE: Giant cell arteritis (GCA) predominantly affects postmenopausal women. Estrogen receptor alpha (ER alpha) accumulates in the cytoplasm of smooth muscle cells, activated mononuclear inflammatory cells and giant cells in the temporal arteries of GCA patients, as well as in smooth muscle cells in arteries from non-GCA controls. The aim of this study was to analyse whether this accumulation is related to structural aberrations in the ER alpha mRNA leading to a change in protein structure. METHODS: Total RNA was extracted from inflamed temporal artery tissue in two GCA patients and from non-inflamed arteries in two non-GCA controls. Products from the nested RT-PCR of the cDNA were cloned and plasmid inserts of 20 different clones from each case were investigated using nucleotide sequence analysis. RESULTS: A total of eight different types of transcripts lacking parts of the ER alpha mRNA were detected. Seven of these could be explained by alternative splicing. There were no significant differences between the GCA patients and the non-GCA controls in terms of the number of transcript variants. CONCLUSION: The accumulated cytoplasmic ER alpha in temporal arterial tissue from elderly persons appears mainly to be of wild type. The main structural changes in the ER alpha mRNA may be due to alternative splicing. Somatic mutations of the ER alpha gene appear to be rare and it is therefore unlikely that they are involved in the pathogenesis of GCA.
Assuntos
Arterite de Células Gigantes/genética , Receptores de Estrogênio/genética , Idoso , Idoso de 80 Anos ou mais , Processamento Alternativo , Análise Mutacional de DNA , Receptor alfa de Estrogênio , Éxons , Feminino , Humanos , Masculino , Mutação Puntual , Reação em Cadeia da PolimeraseRESUMO
The mitochondrial DNA (mtDNA) depletion syndrome is a genetically heterogeneous group of diseases caused by nuclear gene mutations and secondary reduction in mtDNA copy number. We describe a patient with progressive muscle weakness and increased creatine kinase and lactate levels. Muscle weakness was first noted at age 1.5 years and he died of respiratory failure and bronchopneumonia at age 3.5 years. The muscle biopsy showed dystrophic features with ragged red fibers and numerous cytochrome c oxidase (COX)-negative fibers. qPCR analysis demonstrated depletion of mtDNA and sequence analysis of the mitochondrial thymidine kinase 2 (TK2) gene revealed two novel heterozygous variants, c.332C > T, p.(T111I) and c.156 + 5G > C. Quantitative analysis of mtDNA in single muscle fibers demonstrated that COX-deficient fibers showed more pronounced depletion of mtDNA when compared with fibers with residual COX activity (P < 0.01, n = 25). There was no evidence of manifestations from other organs than skeletal muscle although there was an apparent reduction of mtDNA copy number also in liver. The patient showed a pronounced, albeit transient, improvement in muscle strength after onset of treatment with coenzyme Q10, asparaginase, and increased energy intake, suggesting that nutritional modulation may be a therapeutic option in myopathic mtDNA depletion syndrome.
Assuntos
DNA Mitocondrial/metabolismo , Miopatias Mitocondriais/genética , Miopatias Mitocondriais/fisiopatologia , Fibras Musculares Esqueléticas/metabolismo , Timidina Quinase/genética , Pré-Escolar , Análise Mutacional de DNA , Evolução Fatal , Humanos , Fígado/patologia , Masculino , Mitocôndrias Musculares/genética , Mitocôndrias Musculares/metabolismo , Miopatias Mitocondriais/tratamento farmacológico , Miopatias Mitocondriais/patologia , Fibras Musculares Esqueléticas/patologia , Debilidade Muscular/genética , MutaçãoRESUMO
AIM: To describe ophthalmological phenotypes in patients with mitochondrial disease and known genotypes. METHODS: A retrospective study was performed on 59 patients (29 male, 30 female) with a mean age of 11.8 years who had mitochondrial disease with known DNA mutations. Fifty-seven of the 59 subjects underwent a detailed ophthalmological examination including visual acuity (VA), eye motility, refraction, slit-lamp examination, ophthalmoscopy and, in almost one-half of the cases, a full-field electroretinogram (ERG). RESULTS: Forty-six (81%) of the patients had one or more ophthalmological findings such as ptosis (n = 16), reduced eye motility (n = 22) including severe external ophthalmoplegia (n = 9), strabismus (n = 4), nystagmus (n = 9), low VA (n = 21), refractive errors (n = 26), photophobia (n = 4), and partial or total optic atrophy (n = 25). Pigmentation in the macula and/or periphery was noted in 16 patients. In 10/27 investigated individuals with full field ERG, retinal dystrophy was recorded in six different genotypes representing Kearns-Sayre syndrome (n = 5), Leigh syndrome (n = 1), Mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) (n = 1), Myoclonus epilepsy with red ragged fibres (MERRF) (n = 1), Leber hereditary optic neuropathy (n = 1) and mitochondrial myopathy (n = 1). CONCLUSION: The results show that a majority of patients with mitochondrial disorders have ophthalmological abnormalities. We recommend that an ophthalmological examination, including ERG, be performed on all children and adolescents who are suspected of having a mitochondrial disease.
Assuntos
Oftalmopatias/etiologia , Doenças Mitocondriais/complicações , Adolescente , Adulto , Blefaroptose/etiologia , Criança , Pré-Escolar , DNA Mitocondrial/genética , Eletrorretinografia , Feminino , Genótipo , Humanos , Hiperpigmentação/etiologia , Lactente , Masculino , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/genética , Mutação , Transtornos da Motilidade Ocular/etiologia , Atrofia Óptica/etiologia , Fenótipo , Erros de Refração/etiologia , Estudos Retrospectivos , Adulto JovemRESUMO
Complex I of the oxidative phosphorylation system is composed of at least 45 subunits, seven of which are encoded by mitochondrial DNA (mtDNA). In this study we have investigated two children with complex I deficiency in muscle mitochondria. Patient 1 had cerebellar ataxia from early infancy and an abnormal MRI of the brain compatible with Leigh syndrome (LS). The course was rapidly progressive with frequent exacerbations and death at 2 years and 10 months of age. Patient 2 had a lactic acidosis in the newborn period and had a severe psychomotor developmental retardation. In her teens she developed hypertrophic cardiomyopathy and died at 26 years of age because of cardiac insufficiency. Sequencing analysis of mitochondrial encoded ND genes (MTND) showed two DE NOVO mutations in MTND1 in both patients. Patient 1 had a novel heteroplasmic G3890A mutation, R195Q. Patient 2 had a heteroplasmic G3481A mutation, E59K. The G3890A mutation in patient 1 is the first identified mutation in MTND1 in association with LS and complex I deficiency. The findings in this patient as well as in patient 2 demonstrate new clinical expressions of mutations in MTND1. The findings in patient 2 also illustrates that MTND mutations may be pathogenic even at a low percentage.
Assuntos
Complexo I de Transporte de Elétrons/deficiência , Complexo I de Transporte de Elétrons/genética , Encefalomiopatias Mitocondriais/genética , Mutação de Sentido Incorreto , NADH Desidrogenase/genética , Acidose Láctica/etiologia , Acidose Láctica/patologia , Adulto , Sequência de Bases , Criança , Pré-Escolar , Análise Mutacional de DNA/métodos , DNA Mitocondrial/química , DNA Mitocondrial/genética , Deficiências do Desenvolvimento/etiologia , Deficiências do Desenvolvimento/patologia , Complexo I de Transporte de Elétrons/metabolismo , Evolução Fatal , Feminino , Humanos , Lactente , Recém-Nascido , Encefalomiopatias Mitocondriais/complicações , Encefalomiopatias Mitocondriais/enzimologia , Fosforilação Oxidativa , Transtornos Psicomotores/etiologia , Transtornos Psicomotores/patologiaRESUMO
Mitochondrial changes are frequently encountered in sporadic inclusion-body myositis (s-IBM). Cytochrome c oxidase (COX)-deficient muscle fibers and large-scale mitochondrial DNA (mtDNA) deletions are more frequent in s-IBM than in age-matched controls. COX deficient muscle fibers are due to clonal expansion of mtDNA deletions and point mutations in segments of muscle fibers. Such segments range from 75 microm to more than 1,000 microm in length. Clonal expansion of the 4977 bp "common deletion" is a frequent cause of COX deficient muscle fiber segments, but many other deletions also occur. The deletion breakpoints cluster in a few regions that are similar to what is found in human mtDNA deletions in general. Analysis in s-IBM patients of three nuclear genes associated with multiple mtDNA deletions, POLG1, ANT1 and C10orf2, failed to demonstrate any mutations. In s-IBM patients with high number of COX-deficient fibers, the impaired mitochondrial function probably contribute to muscle weakness and wasting. Treatment that has positive effects in mitochondrial myopathies may be tried also in s-IBM.
Assuntos
Mitocôndrias Musculares/patologia , Miosite de Corpos de Inclusão/patologia , Envelhecimento/genética , Sequência de Bases , DNA Mitocondrial/genética , Complexo IV da Cadeia de Transporte de Elétrons/análise , Humanos , Mitocôndrias Musculares/enzimologia , Mitocôndrias Musculares/fisiologia , Miopatias Mitocondriais/genética , Dados de Sequência Molecular , Fibras Musculares Esqueléticas/enzimologia , Fibras Musculares Esqueléticas/patologia , Mutação , Miosite de Corpos de Inclusão/metabolismo , Fosforilação Oxidativa , RNA de Transferência/genética , Deleção de SequênciaRESUMO
In this study we have analyzed the mtDNA encoded ATPase 6 and 8 genes ( MTATP6 and MTATP8) in two children with Leigh syndrome (LS) and reduced Mg (2+) ATPase activity in muscle mitochondria. In patient 1, with a mild and reversible phenotype, mutational analysis revealed a heteroplasmic T --> C mutation at nt position 9185 (T9185C) in the MTATP6. The mutation resulted in substitution of a highly conserved leucine to proline at codon 220. The proportion of the mutation was > 97 % in the patient's blood and muscle and 85 % in blood of his asymptomatic mother. Patient 2, with severe clinical phenotype and death at 2 years of age, exhibited a novel heteroplasmic T9191C missense mutation in the MTATP6, which converted a highly conserved leucine to a proline at position 222 of the polypeptide. The proportion of the mutation was 90 % in fibroblasts and 94 % muscle tissue. This mutation was absent in the patient's parents and sister suggesting that the mutation was de novo. Our findings expand the spectrum of mutations causing LS and emphasize the role of MTATP6 gene mutations in pathogenesis of LS.
Assuntos
Doença de Leigh/genética , ATPases Mitocondriais Próton-Translocadoras/genética , Mutação , Sequência de Aminoácidos/fisiologia , Criança , Pré-Escolar , Análise Mutacional de DNA/métodos , DNA Mitocondrial/genética , Humanos , Doença de Leigh/fisiopatologia , RespiraçãoRESUMO
Inclusion body myositis (IBM) is a sporadic progressive myopathy, which is morphologically characterized by inflammatory cell infiltrates and rimmed vacuoles in muscle fibers. Mitochondrial changes are regularly present with ragged-red fibers showing deficiency of cytochrome c oxidase. In these muscle fiber segments, there is accumulation of mitochondria with mitochondrial DNA (mtDNA) deletions. There are different deletions in different muscle fibers. In this study, we have sequenced for the first time the multiple mtDNA deletions in muscle from four patients with IBM. The deletion breakpoints were sequenced from cloned polymerase chain reaction (PCR)-amplified mtDNA fragments. The sequencing was performed directly from the bacterial colonies used for cloning. Of 122 analyzed clones, 33 different deletions were identified. The majority of these have not previously been described. There was a marked predominance of deletion breakpoints in certain regions of mtDNA. These predominant breakpoint regions are similar to those described in other conditions with multiple deletions, such as autosomal dominant progressive external ophthalmoplegia (adPEO) and normal aging, but different from those described in diseases due to single deletions such as Kearns-Sayre syndrome and sporadic PEO. These findings indicate that common factors are involved in the development of multiple mtDNA deletions in IBM, adPEO, and aging.
Assuntos
DNA Mitocondrial/genética , Miosite de Corpos de Inclusão/genética , Deleção de Sequência , Adulto , Idoso , Feminino , Humanos , Masculino , Reação em Cadeia da PolimeraseRESUMO
Sporadic progressive external ophthalmoplegia and Kearns-Sayre syndrome are usually associated with single large-scale mitochondrial DNA deletions in muscle. In progressive external ophthalmoplegia with autosomal dominant inheritance, multiple mitochondrial DNA deletions have been reported. We studied several members of a Swedish family with autosomal dominant progressive external ophthalmoplegia and multiple mitochondrial DNA deletions by polymerase chain reaction analysis of single muscle fibers and by in situ hybridization, combined with enzyme histochemical analysis. Muscle fiber segments with deficiency of cytochrome c oxidase, which is partially encoded by mitochondrial DNA, had accumulated mitochondrial DNA with deletions and showed reduced levels of wild-type mitochondrial DNA. The deletions varied between individual muscle fibers. There was one predominant deletion in each cytochrome c oxidase-deficient muscle fiber segment. Sequencing of the deletion breakpoints showed that most but not all of the deletions were flanked by direct repeats. Young, clinically affected individuals of this family without limb muscle symptoms did not show mitochondrial DNA deletions or cytochrome c oxidase-deficient muscle fibers. Our results indicate that a nuclear factor predisposes to the development of somatic multiple mitochondrial DNA deletions. Mitochondrial DNA with multiple different deletions shows clonal expansion, which leads to mitochondrial myopathy with ragged-red fibers and muscle weakness.
Assuntos
DNA Mitocondrial/genética , Músculo Esquelético/patologia , Oftalmoplegia Externa Progressiva Crônica/genética , Deleção de Sequência , Adolescente , Adulto , Sequência de Bases , Southern Blotting , Clonagem Molecular , Deficiência de Citocromo-c Oxidase , Sondas de DNA , DNA Complementar , DNA Mitocondrial/biossíntese , DNA Mitocondrial/isolamento & purificação , Feminino , Genes Dominantes , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Fibras Musculares Esqueléticas/enzimologia , Fibras Musculares Esqueléticas/patologia , Músculo Esquelético/metabolismo , Oftalmoplegia Externa Progressiva Crônica/patologia , Linhagem , Reação em Cadeia da Polimerase , SuéciaRESUMO
Inclusion body myositis (IBM) is a chronic inflammatory myopathy. The muscle histology is characterized by infiltration of T cells, which invade and apparently destroy muscle fibres. This study was performed to investigate whether predominant clones of muscle-infiltrating T cells are identical in different muscles and whether they persist over time in IBM. By reverse transcriptase-polymerase chain reaction, 25 T-cell receptor (TCR) variable beta (Vbeta) chain families and the complementarity-determining region 3 (CDR3) of the TCR were analysed in two different muscle biopsies of four patients with IBM. In two of the patients, the muscle biopsies were obtained from different muscles at one time point, whereas in two patients, the second biopsy was obtained 9 years after the first biopsy. T cells expressing predominant Vbeta families were analysed for clonality by fragment length analysis of the CDR3. Predominant Vbeta families were analysed by DNA sequencing to identify identical clones. Immunohistochemical staining of Vbeta families was performed to study the distribution of T cells expressing identified predominant Vbeta families. The muscle-infiltrating lymphocytes showed restricted expression of TCR Vbeta families. DNA sequencing proved that clonally expanded T cells were identical in different muscles and persisted 9 years after the first biopsy. Immunohistochemical analysis with Vbeta family-specific antibodies demonstrated the endomysial localization of these T cells in inflammatory cell infiltrates. Our results show that in IBM there is clonal restriction of TCR expression in muscle-infiltrating lymphocytes. Identical T-cell clones predominate in different muscles, and these clones persist for many years. These results indicate an important, continuous, antigen-driven inflammatory reaction in IBM.