RESUMO
Niemann Pick type C is an inborn error of metabolism (IEM), classified as a lysosomal storage disease (LSD) caused by a dysfunction in NPC transport protein, that leads to intracellular accumulation of non-esterified cholesterol and other lipids. Clinical manifestations are ample, with visceral and neurological symptoms. Miglustat, a molecule that reversibly inhibits glucosylceramide synthase is used as treatment for this disorder. Studies demonstrated the influence of oxidative stress and inflammation in IEM, as well in animal model of NP-C disease. Nonetheless, literature lacks data on patients, so our work aimed to investigate if there is influence of chronic inflammation in the pathophysiology of NP-C disease, and the effect of miglustat, N-acetylcysteine (NAC) and Coenzyme Q10 (CoQ10). We evaluated the plasmatic cytokines in NPC patients at diagnosis and during the treatment with miglustat. Additionally, we performed an in vitro study with antioxidants NAC (1 mM and 2.5 mM) and CoQ10 (5 µM and 10 µM), where we could verify its effect on inflammatory parameters, as well as in cholesterol accumulation. Our results showed that NP-C patients have higher plasmatic levels of pro and anti-inflammatory cytokines (IL-6, IL-8, and IL-10) at diagnosis and the treatment with miglustat was able to restore it. In vitro study showed that treatment with antioxidants in higher concentrations significantly decrease cholesterol accumulation, and NAC at 2.5 mM normalized the level of pro-inflammatory cytokines. Although the mechanism is not completely clear, it can be related to restoration in lipid traffic and decrease in oxidative stress caused by antioxidants.
Assuntos
Doença de Niemann-Pick Tipo C , 1-Desoxinojirimicina/análogos & derivados , Acetilcisteína/farmacologia , Antioxidantes/farmacologia , Colesterol , Citocinas , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Humanos , Inflamação/tratamento farmacológico , Doença de Niemann-Pick Tipo C/tratamento farmacológico , Ubiquinona/análogos & derivadosRESUMO
Sulfite oxidase (SO) deficiency is biochemically characterized by the accumulation of sulfite, thiosulfate and S-sulfocysteine in tissues and biological fluids of the affected patients. The main clinical symptoms include severe neurological dysfunction and brain abnormalities, whose pathophysiology is still unknown. The present study investigated the in vitro effects of sulfite and thiosulfate on mitochondrial homeostasis in rat brain mitochondria. It was verified that sulfite per se, but not thiosulfate, decreased state 3, CCCP-stimulated state and respiratory control ratio in mitochondria respiring with glutamate plus malate. In line with this, we found that sulfite inhibited the activities of glutamate and malate (MDH) dehydrogenases. In addition, sulfite decreased the activity of a commercial solution of MDH, that was prevented by antioxidants and dithiothreitol. Sulfite also induced mitochondrial swelling and reduced mitochondrial membrane potential, Ca(2+) retention capacity, NAD(P)H pool and cytochrome c immunocontent when Ca(2+) was present in the medium. These alterations were prevented by ruthenium red, cyclosporine A (CsA) and ADP, supporting the involvement of mitochondrial permeability transition (MPT) in these effects. We further observed that N-ethylmaleimide prevented the sulfite-elicited swelling and that sulfite decreased free thiol group content in brain mitochondria. These findings indicate that sulfite acts directly on MPT pore containing thiol groups. Finally, we verified that sulfite reduced cell viability in cerebral cortex slices and that this effect was prevented by CsA. Therefore, it may be presumed that disturbance of mitochondrial energy homeostasis and MPT induced by sulfite could be involved in the neuronal damage characteristic of SO deficiency.
Assuntos
Encéfalo/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Proteínas de Transporte da Membrana Mitocondrial/efeitos dos fármacos , Compostos de Sulfidrila/química , Sulfitos/farmacologia , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/patologia , Animais , Encéfalo/metabolismo , Proliferação de Células , Citocromos c/metabolismo , Immunoblotting , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias Hepáticas/efeitos dos fármacos , Mitocôndrias Hepáticas/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Poro de Transição de Permeabilidade Mitocondrial , NADP/metabolismo , Consumo de Oxigênio/efeitos dos fármacos , Ratos , Ratos Wistar , Compostos de Sulfidrila/metabolismo , Sulfito Oxidase/deficiência , Sulfito Oxidase/metabolismoRESUMO
The accumulation of octanoic (OA) and decanoic (DA) acids in tissue is the common finding in medium-chain acyl-coenzyme A dehydrogenase deficiency (MCADD), the most frequent defect of fatty acid oxidation. Affected patients present hypoketotic hypoglycemia, rhabdomyolysis, hepatomegaly, seizures and lethargy, which may progress to coma and death. At present, the pathophysiological mechanisms underlying hepatic and skeletal muscle alterations in affected patients are poorly known. Therefore, in the present work, we investigated the in vitro effects of OA and DA, the accumulating metabolites in MCADD, on various bioenergetics and oxidative stress parameters. It was verified that OA and DA decreased complexes I-III, II-III and IV activities in liver and also inhibit complex IV activity in skeletal muscle. In addition, DA decreased complexes II-III activity in skeletal muscle. We also verified that OA and DA increased TBA-RS levels and carbonyl content in both tissues. Finally, DA, but not OA, significantly decreased GSH levels in rat skeletal muscle. Our present data show that the medium-chain fatty acids that accumulate in MCADD impair electron transfer through respiratory chain and elicit oxidative damage in rat liver and skeletal muscle. It may be therefore presumed that these mechanisms are involved in the pathophysiology of the hepatopathy and rhabdomyolysis presented by MCADD-affected patients.
Assuntos
Caprilatos/metabolismo , Decanoatos/metabolismo , Fígado/metabolismo , Músculo Esquelético/metabolismo , Estresse Oxidativo , Acil-CoA Desidrogenase/deficiência , Acil-CoA Desidrogenase/metabolismo , Animais , Caprilatos/farmacologia , Creatina Quinase/metabolismo , Decanoatos/farmacologia , Transporte de Elétrons , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Glutationa/metabolismo , Erros Inatos do Metabolismo Lipídico/metabolismo , Fígado/efeitos dos fármacos , Fígado/enzimologia , Masculino , Mitocôndrias/metabolismo , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/enzimologia , Oxirredução , Carbonilação Proteica , Ratos , Ratos Wistar , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismoRESUMO
Medium-chain fatty acids and acylcarnitines accumulate in medium-chain acyl-CoA dehydrogenase deficiency (MCADD), the most frequent fatty acid oxidation defect clinically characterized by episodic crises with vomiting, seizures and coma. Considering that the pathophysiology of the neurological symptoms observed in MCADD is poorly known and, to our knowledge, there is no report on the involvement of acylcarnitines in the brain damage presented by the affected patients, the objective of the present study was to investigate the in vitro effects of hexanoylcarnitine (HC), octanoylcarnitine, decanoylcarnitine (DC) and cis-4-decenoylcarnitine (cDC) at concentrations varying from 0.01 to 1.0mM on important oxidative stress parameters in cerebral cortex of young rats. HC, DC and cDC significantly induced lipid peroxidation, as determined by increased thiobarbituric acid-reactive substances (TBA-RS) values. In addition, carbonyl formation was significantly augmented and sulfhydryl content diminished by DC, reflecting induction of protein oxidative damage. HC, DC and cDC also decreased glutathione (GSH) levels, the most important brain antioxidant defense. Furthermore, DC-induced elevation of TBA-RS values and decrease of GSH levels were prevented by the free radical scavengers melatonin and α-tocopherol, indicating the involvement of reactive oxygen species in these effects. We also found that l-carnitine itself did not induce lipid and protein oxidative damage, neither reduced the antioxidant defenses. Our present data show that the major medium-chain acylcarnitines accumulating in MCADD elicit oxidative stress in rat brain. It is therefore presumed that these compounds may be involved to a certain extent in the pathogenesis of the neurologic dysfunction of MCADD.
Assuntos
Córtex Cerebral/fisiopatologia , Homeostase/fisiologia , Erros Inatos do Metabolismo Lipídico/patologia , Acil-CoA Desidrogenase/deficiência , Animais , Carnitina/análogos & derivados , Carnitina/toxicidade , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Glutationa/metabolismo , Homeostase/efeitos dos fármacos , Homeostase/ética , Erros Inatos do Metabolismo Lipídico/induzido quimicamente , Erros Inatos do Metabolismo Lipídico/prevenção & controle , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Melatonina/administração & dosagem , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Carbonilação Proteica/efeitos dos fármacos , Ratos , Ratos Wistar , Estatísticas não Paramétricas , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , alfa-Tocoferol/administração & dosagemRESUMO
Accumulation of long-chain 3-hydroxy fatty acids is the biochemical hallmark of long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) and mitochondrial trifunctional protein (MTP) deficiencies. These disorders are clinically characterized by neurological symptoms, such as convulsions and lethargy, as well as by cardiomyopathy and muscle weakness. In the present work we investigated the in vitro effect of 3-hydroxydodecanoic (3HDA), 3-hydroxytetradecanoic (3HTA) and 3-hydroxypalmitic (3HPA) acids, which accumulate in these disorders, on important oxidative stress parameters in cerebral cortex of young rats in the hope to clarify the mechanisms leading to the brain damage found in patients affected by these disorders. It was first verified that these compounds significantly induced lipid peroxidation, as determined by increased thiobarbituric acid-reactive substances levels. In addition, carbonyl formation was significantly increased and sulfhydryl content decreased by 3HTA and 3HPA, which indicates that these fatty acids elicit protein oxidative damage. 3HTA and 3HPA also diminished the reduced glutathione (GSH) levels, without affecting nitrate and nitrite production. Finally, we observed that the addition of the antioxidants and free radical scavengers trolox and deferoxamine (DFO) was able to partially prevent lipid oxidative damage, whereas DFO fully prevented the reduction on GSH levels induced by 3HTA. Our present data showing that 3HDA, 3HTA and 3HPA elicit oxidative stress in rat brain indicate that oxidative damage may represent an important pathomechanism involved in the neurologic symptoms manifested by patients affected by LCHAD and MTP deficiencies.
Assuntos
3-Hidroxiacil-CoA Desidrogenases/deficiência , Encefalopatias Metabólicas/metabolismo , Encéfalo/metabolismo , Transtornos do Metabolismo dos Lipídeos/metabolismo , Complexos Multienzimáticos/deficiência , Ácidos Mirísticos/toxicidade , Estresse Oxidativo/fisiologia , Ácidos Palmíticos/toxicidade , Animais , Encéfalo/efeitos dos fármacos , Encefalopatias Metabólicas/induzido quimicamente , Ácidos Decanoicos/metabolismo , Ácidos Decanoicos/toxicidade , Ácidos Graxos/metabolismo , Ácidos Graxos/toxicidade , Transtornos do Metabolismo dos Lipídeos/induzido quimicamente , 3-Hidroxiacil-CoA Desidrogenase de Cadeia Longa , Masculino , Proteína Mitocondrial Trifuncional , Ácidos Mirísticos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ácidos Palmíticos/metabolismo , Ratos , Ratos WistarRESUMO
Medium-chain acyl-CoA dehydrogenase deficiency (MCADD) is the most frequent disorder of fatty acid oxidation with a similar prevalence to that of phenylketonuria. Affected patients present tissue accumulation of the medium-chain fatty acids octanoate (OA), decanoate (DA) and cis-4-decenoate. Clinical presentation is characterized by neurological symptoms, such as convulsions and lethargy that may develop into coma and sudden death. The aim of the present work was to investigate the in vitro effect of OA and DA, the metabolites that predominantly accumulate in MCADD, on oxidative stress parameters in rat cerebral cortex homogenates. It was first verified that both DA and OA significantly increased chemiluminescence and thiobarbituric acid-reactive species levels (lipoperoxidation) and decreased the non-enzymatic antioxidant defenses, measured by the decreased total antioxidant capacity. DA also enhanced carbonyl content and oxidation of sulfhydryl groups (protein damage) and decreased reduced glutathione (GSH) levels. We also verified that DA-induced GSH decrease and sulfhydryl oxidation were not observed when cytosolic preparations (membrane-free supernatants) were used, suggesting a mitochondrial mechanism for these actions. Our present data show that the medium-chain fatty acids DA and OA that most accumulate in MCADD cause oxidative stress in rat brain. It is therefore presumed that this pathomechanism may be involved in the pathophysiology of the neurologic symptoms manifested by patients affected by MCADD.