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/genéticaRESUMO
BACKGROUND: Mutations causing Leber hereditary optic neuropathy are usually homoplasmic, show incomplete penetrance, and many of the affected positions are not well conserved through evolution. A large percentage of patients harbouring these mutations have no family history of disease. Moreover, the transfer of the mutation in the cybrid model is frequently not accompanied by the transfer of the cellular, biochemical and molecular phenotype. All these features make difficult their classification as the etiologic factors for this disease. We report a patient who exhibits typical clinical features of Leber hereditary optic neuropathy but lacks all three of the most common mitochondrial DNA mutations. METHODS: The diagnosis was made based on clinical studies. The mitochondrial DNA was completely sequenced, and the candidate mutation was analysed in more than 18 000 individuals around the world, its conservation index was estimated in more than 3100 species from protists to mammals, its position was modelled in the crystal structure of a bacteria ortholog subunit, and its functional consequences were studied in a cybrid model. RESULTS: Genetic analysis revealed an m.3472T>C transition in the MT-ND1 gene that changes a phenylalanine to leucine at position 56. Bioinformatics, molecular-genetic analysis and functional studies suggest that this transition is the etiological factor for the disorder. CONCLUSIONS: This mutation expands the spectrum of deleterious changes in mitochondrial DNA-encoded complex I polypeptides associated with this pathology and highlights the difficulties in assigning pathogenicity to new homoplasmic mutations that show incomplete penetrance in sporadic Leber hereditary optic neuropathy patients.
Assuntos
DNA Mitocondrial/genética , Mitocôndrias/genética , NADH Desidrogenase/genética , Atrofia Óptica Hereditária de Leber/genética , Polimorfismo de Nucleotídeo Único , Adulto , Sequência de Bases , Análise Mutacional de DNA , Humanos , Masculino , Dados de Sequência Molecular , Atrofia Óptica Hereditária de Leber/diagnóstico , Fenótipo , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Estrutura Secundária de Proteína , Testes de Campo Visual , Campos VisuaisRESUMO
BACKGROUND: Most patients suffering from Leber hereditary optic neuropathy carry one of the three classic pathologic mutations, but not all individuals with these genetic alterations develop the disease. There are different risk factors that modify the penetrance of these mutations. The remaining patients carry one of a set of very rare genetic variants and, it appears that, some of the risk factors that modify the penetrance of the classical pathologic mutations may also affect the phenotype of these other rare mutations. RESULTS: We describe a large family including 95 maternally related individuals, showing 30 patients with Leber hereditary optic neuropathy. The mutation responsible for the phenotype is a novel transition, m.3734A > G, in the mitochondrial gene encoding the ND1 subunit of respiratory complex I. Molecular-genetic, biochemical and cellular studies corroborate the pathogenicity of this genetic change. CONCLUSIONS: With the study of this family, we confirm that, also for this very rare mutation, sex and age are important factors modifying penetrance. Moreover, this pedigree offers an excellent opportunity to search for other genetic or environmental factors that additionally contribute to modify penetrance.
Assuntos
DNA Mitocondrial , Atrofia Óptica Hereditária de Leber , Humanos , DNA Mitocondrial/genética , Atrofia Óptica Hereditária de Leber/genética , Linhagem , Mutação/genética , FenótipoRESUMO
FK506 is an important immunosuppressive medication. However, it can provoke neurotoxicity, nephrotoxicity, and diabetes as adverse side effects. The decrease in oxygen consumption of rat cells treated with pharmacologically relevant concentrations of FK506, along with other evidences, has insinuated that some of the toxic effects are probably caused by drug-induced mitochondrial dysfunction at the level of gene expression. To confirm this suggestion, we have analyzed cell respiration and mitochondrial protein synthesis in human cell lines treated with FK506. This drug provokes an important decrease in oxygen consumption, accompanied by a slight reduction in the synthesis of mitochondria DNA-encoded proteins. These results are similar to those triggered by rapamycin, another macrolide with immunosuppressive properties, therefore insinuating a common toxic pathway.
Assuntos
Imunossupressores/efeitos adversos , Mitocôndrias/efeitos dos fármacos , Biossíntese de Proteínas/efeitos dos fármacos , Tacrolimo/efeitos adversos , Animais , Diabetes Mellitus/induzido quimicamente , Diabetes Mellitus/metabolismo , Diabetes Mellitus/patologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Imunossupressores/uso terapêutico , Proteínas Mitocondriais/efeitos dos fármacos , Consumo de Oxigênio/efeitos dos fármacos , Ratos , Tacrolimo/uso terapêuticoRESUMO
Multiple sclerosis is likely caused by a complex interaction of multiple genes and environmental factors. The contribution of mitochondrial DNA genetic backgrounds has been frequently reported. To evaluate the effect of mitochondrial DNA haplogroups in the same genetic and environmental circumstances, we have built human transmitochondrial cell lines and simulated the effect of axon demyelination, one of the hallmarks of multiple sclerosis pathology, by altering the ionic gradients through the plasmalemma and increasing ATP consumption. In this model, mitochondrial biogenesis is observed. This process is larger in Uk cybrids, which mirrors their lower oxidative phosphorylation capacity in basal conditions. This model replicates a process occurring in both patients suffering from multiple sclerosis and several animal models of axon demyelination. Therefore, it can be used to analyze the contribution of various mitochondrial DNA genotypes to multiple sclerosis. In this sense, a longer or stronger energy stress, such as that associated with demyelinated axons in multiple sclerosis, could make Uk individuals more susceptible to this pathology. Finally, pharmacologic compounds targeted to mitochondrial biogenesis could be a potential therapy for multiple sclerosis.
Assuntos
Axônios/patologia , DNA Mitocondrial , Mitocôndrias/patologia , Esclerose Múltipla/patologia , Bainha de Mielina/patologia , Axônios/metabolismo , Linhagem Celular , Sobrevivência Celular/fisiologia , Haplótipos , Humanos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Esclerose Múltipla/genética , Esclerose Múltipla/metabolismo , Bainha de Mielina/genética , Bainha de Mielina/metabolismo , Consumo de Oxigênio/fisiologiaRESUMO
BACKGROUND: Leigh syndrome (LS) is a serious genetic disease that can be caused by mutations in dozens of different genes. METHODS: Clinical study of a deafness pedigree in which some members developed LS. Cellular, biochemical and molecular genetic analyses of patients' tissues and cybrid cell lines were performed. RESULTS: mitochondrial DNA (mtDNA) m.1555A>G/MT-RNR1 and m.9541T>C/MT-CO3 mutations were found. The first one is a well-known pathologic mutation. However, the second one does not appear to contribute to the high hearing loss penetrance and LS phenotype observed in this family. CONCLUSION: The m.1555A>G pathological mutation, accompanied with an unknown nuclear DNA (nDNA) factor, could be the cause of the phenotypic manifestations in this pedigree.
Assuntos
Doença de Leigh/genética , Mutação , RNA Mitocondrial/genética , Adulto , Pré-Escolar , Feminino , Humanos , Lactente , Doença de Leigh/patologia , Masculino , Linhagem , Fenótipo , RNA Ribossômico , Adulto JovemRESUMO
The oxidative phosphorylation system is important for adipocyte differentiation. Therefore, xenobiotics inhibitors of the oxidative phosphorylation system could affect adipocyte differentiation and adipokine secretion. As adipokines impact the overall health status, these xenobiotics may have wide effects on human health. Some of these xenobiotics are widely used therapeutic drugs, such as ribosomal antibiotics. Because of its similarity to the bacterial one, mitochondrial translation system is an off-target for these compounds. To study the influence of the ribosomal antibiotic linezolid on adipokine production, we analyzed its effects on adipocyte secretome. Linezolid, at therapeutic concentrations, modifies the levels of apolipoprotein E and several adipokines and proteins related with the extracellular matrix. This antibiotic also alters the global methylation status of human adipose tissue-derived stem cells and, therefore, its effects are not limited to the exposure period. Besides their consequences on other tissues, xenobiotics acting on the adipocyte oxidative phosphorylation system alter apolipoprotein E and adipokine production, secondarily contributing to their systemic effects.
Assuntos
Adipócitos/efeitos dos fármacos , Adipocinas/metabolismo , Linezolida/farmacologia , Fosforilação Oxidativa , Inibidores da Síntese de Proteínas/farmacologia , Adipócitos/metabolismo , Células Cultivadas , Exocitose , HumanosRESUMO
BACKGROUND: The oxidative phosphorylation system (OXPHOS) includes nuclear chromosome (nDNA)- and mitochondrial DNA (mtDNA)-encoded polypeptides. Many rare OXPHOS disorders, such as striatal necrosis syndromes, are caused by genetic mutations. Despite important advances in sequencing procedures, causative mutations remain undetected in some patients. It is possible that etiologic factors, such as environmental toxins, are the cause of these cases. Indeed, the inhibition of a particular enzyme by a poison could imitate the biochemical effects of pathological mutations in that enzyme. Moreover, environmental factors can modify the penetrance or expressivity of pathological mutations. OBJECTIVES: We studied the interaction between mitochondrially encoded ATP synthase 6 (p.MT-ATP6) subunit and an environmental exposure that may contribute phenotypic differences between healthy individuals and patients suffering from striatal necrosis syndromes or other mitochondriopathies. METHODS: We analyzed the effects of the ATP synthase inhibitor tributyltin chloride (TBTC), a widely distributed environmental factor that contaminates human food and water, on transmitochondrial cell lines with or without an ATP synthase mutation that causes striatal necrosis syndrome. Doses were selected based on TBTC concentrations previously reported in human whole blood samples. RESULTS: TBTC modified the phenotypic effects caused by a pathological mtDNA mutation. Interestingly, wild-type cells treated with this xenobiotic showed similar bioenergetics when compared with the untreated mutated cells. CONCLUSIONS: In addition to the known genetic causes, our findings suggest that environmental exposure to TBTC might contribute to the etiology of striatal necrosis syndromes. CITATION: López-Gallardo E, Llobet L, Emperador S, Montoya J, Ruiz-Pesini E. 2016. Effects of tributyltin chloride on cybrids with or without an ATP synthase pathologic mutation. Environ Health Perspect 124:1399-1405; http://dx.doi.org/10.1289/EHP182.
Assuntos
Exposição Ambiental , Poluentes Ambientais/toxicidade , ATPases Mitocondriais Próton-Translocadoras/genética , Mutação , Compostos de Trialquitina/toxicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Necrose/genéticaRESUMO
Besides the advance in scientific knowledge and the production of different compounds, cell culture can now be used to obtain cells for regenerative medicine. To avoid microbial contamination, antibiotics were usually incorporated into culture media. However, these compounds affect cell biochemistry and may modify the differentiation potential of cultured cells. To check this possibility, we grew human adipose tissue-derived stem cells and differentiated them to adipocyte with or without antibiotics commonly used in these culture protocols, such as a penicillin-streptomycin-amphotericin mix or gentamicin. We show that these antibiotics affect cell differentiation. Therefore, antibiotics should not be used in cell culture because aseptic techniques make these compounds unnecessary.
Assuntos
Antibacterianos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Meios de Cultura/química , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Tecido Adiposo/citologia , Biomarcadores/metabolismo , Células Cultivadas , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/enzimologia , Espécies Reativas de Oxigênio/metabolismo , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacosRESUMO
Adipogenesis is accompanied by differentiation of adipose tissue-derived stem cells to adipocytes. As part of this differentiation, biogenesis of the oxidative phosphorylation system occurs. Many chemical compounds used in medicine, agriculture or other human activities affect oxidative phosphorylation function. Therefore, these xenobiotics could alter adipogenesis. We have analyzed the effects on adipocyte differentiation of some xenobiotics that act on the oxidative phosphorylation system. The tested concentrations have been previously reported in human blood. Our results show that pharmaceutical drugs that decrease mitochondrial DNA replication, such as nucleoside reverse transcriptase inhibitors, or inhibitors of mitochondrial protein synthesis, such as ribosomal antibiotics, diminish adipocyte differentiation and leptin secretion. By contrast, the environmental chemical pollutant tributyltin chloride, which inhibits the ATP synthase of the oxidative phosphorylation system, can promote adipocyte differentiation and leptin secretion, leading to obesity and metabolic syndrome as postulated by the obesogen hypothesis.
Assuntos
Adipócitos/efeitos dos fármacos , Adipogenia/efeitos dos fármacos , Tecido Adiposo/citologia , Fosforilação Oxidativa/efeitos dos fármacos , Células-Tronco/efeitos dos fármacos , Xenobióticos/farmacologia , Adipócitos/metabolismo , Forma Celular/efeitos dos fármacos , Células Cultivadas , Replicação do DNA/efeitos dos fármacos , DNA Mitocondrial/biossíntese , DNA Mitocondrial/efeitos dos fármacos , Humanos , Leptina/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Fenótipo , Células-Tronco/metabolismo , Fatores de Tempo , Compostos de Trialquitina/toxicidade , Xenobióticos/sangue , Xenobióticos/toxicidadeRESUMO
BACKGROUND AND AIMS: Tacrolimus (Tac) is an immunosuppressive drug widely used to avoid organ rejection. New-onset diabetes after transplantation (NODAT) is a major complication among transplanted patients who receive Tac. The increased risk for NODAT could be partly mediated by the effect of Tac on mitochondria from pancreatic beta-cells. Common and rare mitochondrial DNA variants have been linked to the risk of diabetes. Our aim was to determine whether mtDNA polymorphisms/haplogroups were associated with NODAT in Tac-treated kidney transplanted. METHODS: Seven polymorphisms that define the common European haplogroups were determined in 115 NODAT and 197 no-NODAT patients. RESULTS: Haplogroup H was significantly more frequent in the NODAT group (50% vs. 35%; p=0.01, OR=1.82). There was no difference between patients without and with (n=106) D2M prior to the transplant. CONCLUSIONS: Mitochondrial haplogroup H was associated with the risk for NODAT among Tac-treated transplanted patients. The reported differences between the mtDNA variants could explain the increased NODAT-risk among H-patients.