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1.
Hum Mol Genet ; 23(23): 6191-200, 2014 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-24986921

RESUMO

Mitochondrial DNA mutations at MT-ATP6 gene are relatively common in individuals suffering from striatal necrosis syndromes. These patients usually do not show apparent histochemical and/or biochemical signs of oxidative phosphorylation dysfunction. Because of this, MT-ATP6 is not typically analyzed in many other mitochondrial disorders that have not been previously associated to mutations in this gene. To correct this bias, we have performed a screening of the MT-ATP6 gene in a large collection of patients suspected of suffering different mitochondrial DNA (mtDNA) disorders. In three cases, biochemical, molecular-genetics and other analyses in patient tissues and cybrids were also carried out. We found three new pathologic mutations. Two of them in patients showing phenotypes that have not been commonly associated to mutations in the MT-ATP6 gene. These results remark the importance of sequencing the MT-ATP6 gene in patients with striatal necrosis syndromes, but also within other mitochondrial pathologies. This gene should be sequenced at least in all those patients suspected of suffering an mtDNA disorder disclosing normal results for histochemical and biochemical analyses of respiratory chain.


Assuntos
DNA Mitocondrial/genética , ATPases Mitocondriais Próton-Translocadoras/genética , Feminino , Humanos , Doença de Leigh/genética , Masculino , Doenças Mitocondriais/genética , Miopatias Mitocondriais/genética , Mutação , Fenótipo , Retinose Pigmentar/genética
2.
Neuromolecular Med ; 9(4): 285-91, 2007.
Artigo em Inglês | MEDLINE | ID: mdl-17999202

RESUMO

The deafness-dystonia syndrome (DDS) or Mohr-Tranebjaerg syndrome (MTS, MIM 304700) is a rare X-linked recessive neurological disorder resulting from loss-of-function mutations in the nuclear DDP1/TIMM8A gene, involved in the transport and sorting of proteins to the mitochondrial inner membrane. A Mohr-Tranebjaerg patient and his mother were subjected to clinical and molecular studies. Screening of mutations were performed in TIMM8A, TIMM13, and other mitochondrial protein transport genes by conformation sensitive gel electrophoresis (CSGE), followed by direct DNA sequencing of tissue samples from the patient. Mitochondrial DNA of the patient was also sequenced at the genes for COX subunits and some mitochondrial tRNAs. Respiratory chain activities in a muscle biopsy and cultured fibroblasts from the patient were assessed using biochemical methods. mRNA expression of TIMM8A and TIMM13 was determined by RT-PCR in cultured fibroblasts. We identified a new case of Mohr-Tranebjaerg syndrome and report the characteristics of a new pathogenic de novo mutation (c.112C>T, pGln38X) in the TIMM8A gene. Biochemical measures of respiratory chain complex activities in muscle biopsy and fibroblasts did not show a major deficiency or alteration. mRNA expression studies demonstrated increased TIMM8A mRNA levels in cultured fibroblasts from the patient. Phenotypic differences among published cases seem not to be related with the mutation location or type. Our results support the idea that dysfunctions of mitochondrial protein transport, in addition to OXPHOS deficiency, can be the basis of important mitochondrial pathologies.


Assuntos
Proteínas de Membrana Transportadoras/genética , Mutação , Síndromes Orofaciodigitais/genética , Adulto , Surdez/genética , Distonia/genética , Eletroforese , Feminino , Fibroblastos/patologia , Humanos , Masculino , Proteínas de Membrana Transportadoras/isolamento & purificação , Proteínas de Transporte da Membrana Mitocondrial/genética , Proteínas de Transporte da Membrana Mitocondrial/isolamento & purificação , Proteínas do Complexo de Importação de Proteína Precursora Mitocondrial , Linhagem , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Pele/patologia
3.
Med Clin (Barc) ; 126(12): 457-60, 2006 Apr 01.
Artigo em Espanhol | MEDLINE | ID: mdl-16620733

RESUMO

BACKGROUND AND OBJECTIVE: Our purpose was to report the neurological manifestations and molecular-genetic analysis of mitochondrial DNA associated with chronic progressive external ophthalmoplegia (CPEO) and raged red fibers (RRFs). PATIENTS AND METHOD: Two patients, a male and a female (32 and 28 year-old, respectively), were studied due to progressive palpebral ptosis associated with RRFs in muscle biopsy. Both patients were subjected to neurological, histochemical and enzymatic analysis of muscular biopsy, analysis of cerebro-spinal fluid, and molecular analysis of mitochondrial DNA. RESULTS: Symptoms started at ages 24 and 17 years. Initial symptoms were palpebral ptosis, progressive limitation of vertical and horizontal gaze, fatigue and exercise intolerance, and weakness of proximal muscles. Brain MRIs were normal in both patients. Both patients had deletions of muscle mitochondrial DNA with similar size (5,425 and 5,112 base pairs) and location. CONCLUSIONS: CPEO with RRFs is usually associated with huge deletions in mitochondrial DNA. Fatigue and proximal muscle weakness can be found during the follow-up.


Assuntos
DNA Mitocondrial/análise , Oftalmoplegia Externa Progressiva Crônica/genética , Adulto , Brasil , Feminino , Humanos , Masculino , Oftalmoplegia Externa Progressiva Crônica/patologia
4.
FEBS Lett ; 579(30): 6909-13, 2005 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-16337195

RESUMO

We have studied the production of reactive oxygen species (ROS) in transmitochondrial cells, harboring homoplasmic levels of the T14487C mtDNA mutation in the ND6 gene of mitochondrial DNA (mtDNA). Previous work has shown that this mutation causes complex I deficiency. Here, we show that this mutation causes an overproduction of ROS leading to an increase in the oxidation of lipids and mtDNA without modification of antioxidant enzyme activities. We suggest that mutations in mtDNA affecting complex I activity may result in oxidative cellular damage, and reinforce the possible role of ROS-mediated mechanisms participating in some mtDNA-related disorders.


Assuntos
DNA Mitocondrial/genética , Mitocôndrias/metabolismo , Mutação , NADH Desidrogenase/genética , Espécies Reativas de Oxigênio/metabolismo , Humanos , Lipídeos/fisiologia , Mitocôndrias/genética , NADH Desidrogenase/química , NADH Desidrogenase/metabolismo , Oxirredução , Estresse Oxidativo/genética , Subunidades Proteicas/química , Subunidades Proteicas/genética
5.
Hum Gene Ther ; 14(5): 463-72, 2003 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-12691611

RESUMO

Herpes simplex virus thymidine kinase (HSV-tk) converts ganciclovir (GCV) into an active compound, which can be incorporated into DNA molecules and terminate DNA synthesis. Gene transfer of HSV-tk followed by GCV administration has been used with success to treat experimental cancer and this strategy has entered into clinical trials. Although it is thought that the cytotoxic effect occurs mainly in tumoral dividing cells, where mitotic activity favors integration of the genotoxic compound into nuclear DNA, there are concerns of potential damage to normal nondividing cells. In the present work we have explored the mechanisms of HSV-tk/GCV toxicity and in particular whether this therapy may cause lesions of mitochondrial DNA (mtDNA) and mitochondrial dysfunction. We found that the administration of GCV to rats injected with adenovirus encoding HSV-tk induced hepatocellular damage characterized by the presence of apoptotic bodies, ballooning of hepatocytes, and severe hepatic steatosis with mitochondria enlargement and cristae dissolution at the ultrastructural level. Remarkably, Southern blot analysis showed substantial reduction in the amount of mtDNA in the liver. Using radiolabeled GCV we could demonstrate incorporation of this compound into both nuclear and mtDNA in HSV-tk-transduced rat hepatocytic cell line MCA-RH7777 and subsequent alteration of mitochondrial function. Our observations confirm that GCV can damage both nuclear and mtDNA in cells transduced with HSV-tk and that this effect could be responsible for severe mitochondrial dysfunction and toxicity in normal nondividing cells. These data are relevant for the design of clinical trials using adenoviral vectors encoding HSV-tk.


Assuntos
DNA Mitocondrial/análise , Ganciclovir/toxicidade , Herpesvirus Humano 1/genética , Falência Hepática/etiologia , Mitocôndrias/fisiologia , Timidina Quinase/genética , Adenoviridae/genética , Alanina Transaminase/sangue , Alanina Transaminase/química , Animais , Antígenos Glicosídicos Associados a Tumores/análise , Aspartato Aminotransferases/sangue , Aspartato Aminotransferases/química , Complexo IV da Cadeia de Transporte de Elétrons/análise , Vetores Genéticos , Células HeLa , Herpesvirus Humano 1/enzimologia , Humanos , Ácido Láctico/biossíntese , Falência Hepática/genética , Falência Hepática/patologia , Masculino , Mitocôndrias/efeitos dos fármacos , RNA/análise , RNA Mitocondrial , Ratos , Ratos Wistar , Células Tumorais Cultivadas
6.
Springerplus ; 1(1): 63, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23487597

RESUMO

Cardiac healing, which follows myocardial infarction, is a complex process guided by intricate interactions among different components. Some resident cell populations with a potential role in cardiac healing have already been described in cardiac tissues. These non-cardiomyocyte cell subsets, globally described as cardiac pluripotent/progenitor cells (CPCs), are able to differentiate into all three major cardiac cell lineages (endothelial, smooth muscle and cardiomyocyte cells) in experimental settings. Nevertheless, physiological cardiac healing results in a fibrous scar, which remains to be fully modelled experimentally. Since a role for complement anaphylatoxins (C3a and C5a) has been described in several regeneration/repair processes, we examined the effects that C3a and C5a exert on a defined population of CPCs. We found that C3a and C5a are able to enhance CPC migration and proliferation. In vitro studies showed that this effect is linked to activation of telomerase mRNA and partial preservation of telomere length, in an NFκB-dependent manner. In addition, anaphylatoxin signalling modulates the CPC phenotype, increasing myofibroblast differentiation and reducing endothelial and cardiac gene expression. These findings may denote that C3a and C5a are able to maintain/increase the cardiac stem cell pool within the heart, whilst simultaneously facilitating and modulating resident cell differentiation. We found that this modulation was directed towards scar forming cells, which increased fibroblast/myofibroblast generation and suggests that both these anaphylatoxins could play a relevant role in the damage-coupled activation of resident cells, and regulation of the cardiac healing process after injury.

7.
Ann Neurol ; 59(2): 394-8, 2006 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-16365882

RESUMO

OBJECTIVE: Our aim was to describe a child with an incomplete form of Kearns-Sayre syndrome who presented profound cerebrospinal fluid (CSF) folate deficiency and his response to folinic acid supplementation METHODS: CSF 5-methyltetrahydrofolate was analyzed by HPLC with fluorescence detection and mitochondrial DNA deletions by southern blot hybridization. RESULTS: Cranial magnetic resonance imaging showed a leukoencephalopathy. Profound CSF 5-methyltetrahydrofolate deficiency was observed with normal blood folate values and decreased CSF/serum folate ratio, suggesting a transport defect across the blood-brain barrier. Folinic acid treatment was established, and after 1 year clinical response to folinic supplementation was remarkable, with almost normal white matter image. INTERPRETATION: The clinical response after folinic therapy highlights the need for the study of cerebral folate deficiency in patients with mitochondrial disorders and white matter lesions.


Assuntos
DNA Mitocondrial/genética , Deficiência de Ácido Fólico/genética , Deleção de Genes , Síndrome de Kearns-Sayre/genética , Química Encefálica/genética , Criança , Análise Mutacional de DNA/métodos , Ácido Fólico/sangue , Ácido Fólico/líquido cefalorraquidiano , Deficiência de Ácido Fólico/complicações , Deficiência de Ácido Fólico/patologia , Humanos , Síndrome de Kearns-Sayre/líquido cefalorraquidiano , Síndrome de Kearns-Sayre/etiologia , Síndrome de Kearns-Sayre/patologia , Imageamento por Ressonância Magnética/métodos , Masculino , Músculos/metabolismo
8.
Mol Genet Metab ; 89(3): 283-5, 2006 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-16603396

RESUMO

The mechanisms underlying the appearance of lipomas in patients bearing mutations in the tRNA(Lys) gene of mitochondrial DNA are unknown. We investigated changes in gene expression patterns in lipomas from three patients bearing A8344G or G8363A tRNA(Lys) gene mutations. Uncoupling protein-1 mRNA was detected in the lipomas, in contrast with undetectable expression in normal adipose tissue. However, expression of other markers of brown fat, such as PGC-1alpha, was unaltered. PPARgamma and retinoblastoma gene expression was down regulated in the lipomas, but C/EBPalpha mRNA was not affected. The expression of Pref-1 was dramatically down regulated. Thus, lipomatosis due to tRNA(Lys) mutations is associated with a pattern of altered expression of master regulators of adipogenesis consistent with enhanced proliferation but maintenance of adipocyte features, and with a distorted pattern of brown versus white adipocyte differentiation.


Assuntos
Adipogenia/genética , DNA Mitocondrial/genética , Regulação Neoplásica da Expressão Gênica , Genes Reguladores , Lipoma/genética , Mutação Puntual/genética , RNA de Transferência de Lisina/genética , Feminino , Humanos , Pessoa de Meia-Idade , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
9.
Pediatr Nephrol ; 19(7): 790-3, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15133725

RESUMO

We report the molecular findings in a child presenting with sideroblastic anemia and proximal tubulopathy. Analysis of mitochondrial DNA (mtDNA) from fibroblasts showed the presence of a 3.3-kb single deletion in 50% of the genomes. This mutation is, unlike other previously reported deletions in tubulopathy patients, not flanked by direct repeat sequences but by palindrome sequences at the deletion breakpoints, suggesting an unusual mechanism for production of deletion. These findings further expand our knowledge of the syndrome of anemia and tubulopathy due to single deletions of mtDNA.


Assuntos
DNA Mitocondrial/genética , Síndrome de Fanconi/genética , Deleção de Sequência , Sequência de Bases , Humanos , Lactente , Masculino
10.
Pediatr Res ; 56(1): 55-9, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15128915

RESUMO

Peripheral neuropathy has been identified in children with mitochondrial encephalomyopathies but not as a main clinical landmark. Here we report the clinical, electrophysiologic, biochemical, and genetic findings in a family who harbors the G8363A mutation in the tRNALys gene of mitochondrial DNA. Affected individuals presented with peripheral neuropathy and ataxia as the main clinical sign. Additional involvement included muscle weakness and multiple lipomatosis. Other common clinical characteristics associated with the G8363A mutation, such as cardiomyopathy and myoclonus epilepsy, were not observed. These findings suggest that a mitochondrial disease should be considered in the differential diagnosis of children with heredoataxic syndrome and peripheral neuropathy of unknown origin.


Assuntos
Ataxia/genética , DNA Mitocondrial/genética , Doenças Mitocondriais/genética , Doenças do Sistema Nervoso Periférico/genética , Mutação Puntual , Adolescente , Adulto , Ataxia/patologia , Biópsia , Criança , Feminino , Humanos , Masculino , Doenças Mitocondriais/patologia , Músculo Esquelético/patologia , Linhagem , Doenças do Sistema Nervoso Periférico/patologia , Polimorfismo de Fragmento de Restrição
11.
Ann Neurol ; 54(4): 527-30, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-14520668

RESUMO

We report the molecular findings in two independent patients presenting with progressive generalized dystonia and bilateral striatal necrosis in whom we have identified a mutation (T14487C) in the mitochondrial ND6 gene. The mutation is heteroplasmic in all samples analyzed, and it fulfills all accepted criteria of pathogenicity. Transmitochondrial cell lines harboring 100% mutant mitochondrial DNA showed a marked decrease in the activity of complex I of the respiratory chain supporting the pathogenic role of T14487C.


Assuntos
Corpo Estriado/patologia , DNA Mitocondrial/genética , Distonia/genética , Mutação Puntual , Adolescente , Análise Mutacional de DNA , Distonia/sangue , Humanos , Masculino , Dados de Sequência Molecular , Necrose , Consumo de Oxigênio/genética , Fatores de Tempo , Transfecção , Células Tumorais Cultivadas
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