RESUMEN
Phenylketonuria (PKU) is the most common inherited metabolic disorders caused by severe deficiency or absence of phenylalanine hydroxylase activity that converts phenylalanine (Phe) to tyrosine. PKU patients were treated with a Phe restricted diet supplemented with a special formula containing l-carnitine (L-car), well-known antioxidant compound. The lack of treatment can cause neurological and cognitive impairment, as severe mental retardation, neuronal cell loss and synaptic density reduction. Although Phe has been widely demonstrated to be involved in PKU neurotoxicity, the mechanisms responsible for the CNS injury are still not fully known. In this work, we evaluated markers of neurodegeneration, namely BDNF (brain-derived neurotrophic factor), PAI-1 total (Plasminogen activator inhibitor-1 total), Cathepsin D, PDGF AB/BB (platelet-derived growth factor), and NCAM (neuronal adhesion molecule) in plasma of PKU patients at early and late diagnosis and under treatment. We found decreased Phe levels and increased L-car concentrations in PKU patients treated with L-car compared to the other groups, indicating that the proposed treatment was effective. Furthermore, we found increased BDNF levels in the patients under treatment compared to patients at early diagnosis, and a positive correlation between BDNF and L-car and a negative correlation between BDNF and Phe. Our results may indicate that in PKU patients treated with L-car there is an attempt to adjust neuronal plasticity and recover the damage suffered, reflecting a compensatory response to brain injury.
Asunto(s)
Carnitina , Fenilcetonurias , Humanos , Factor Neurotrófico Derivado del Encéfalo , Fenilcetonurias/tratamiento farmacológico , Suplementos Dietéticos , Antioxidantes , Fenilalanina , BecaplerminaRESUMEN
Phenylketonuria (PKU) was the first genetic disease to have an effective therapy, which consists of phenylalanine intake restriction. However, there are patients who do not adhere to treatment and/or are not submitted to neonatal screening. PKU patients present L-carnitine (L-car) deficiency, compound that has demonstrated an antioxidant and anti-inflammatory role in metabolic diseases. This study evaluated the effect caused by exposure time to high Phe levels in PKU patients at early and late diagnosis, through pro- and anti-inflammatory cytokines, as well as the L-car effect in patients under treatment. It was observed that there was a decrease in phenylalanine levels in treated patients compared to patients at diagnosis, and an increase in L-car levels in the patients under treatment. Inverse correlation between Phe versus L-car and nitrate plus nitrite versus L-car in PKU patients was also showed. We found increased proinflammatory cytokines levels: interleukin (IL)-1ß, interferons (IFN)-gamma, IL-2, tumor necrosis factor (TNF)-alpha, IL-8 and IL-6 in the patients at late diagnosis compared to controls, and IL-8 in the patients at early diagnosis and treatment compared to controls. Increased IL-2, TNF-alpha, IL-6 levels in the patients at late diagnosis compared to early diagnosis were shown, and reduced IL-6 levels in the treated patients compared to patients at late diagnosis. Moreover, it verified a negative correlation between IFN-gamma and L-car in treated patients. Otherwise, it was observed that there were increased IL-4 levels in the patients at late diagnosis compared to early diagnosis, and reduction in treated patients compared to late diagnosed patients. In urine, there was an increase in 8-isoprostane levels in the patients at diagnosis compared to controls and a decrease in oxidized guanine species in the treated patients compared to the diagnosed patients. Our results demonstrate for the first time in literature that time exposure to high Phe concentrations generates a proinflammatory status, especially in PKU patients with late diagnosis. A pro-oxidant status was verified in not treated PKU patients. Our results demonstrate the importance of early diagnosis and prompt start of treatment, in addition to the importance of L-car supplementation, which can improve cellular defense against inflammation and oxidative damage in PKU patients.
Asunto(s)
Citocinas , Fenilcetonurias , Recién Nacido , Humanos , Fenilalanina , Diagnóstico Tardío , Interleucina-2 , Interleucina-6 , Interleucina-8 , Carnitina/farmacología , Fenilcetonurias/diagnóstico , Fenilcetonurias/tratamiento farmacológico , Fenilcetonurias/orina , Factor de Necrosis Tumoral alfaRESUMEN
Maple syrup urine disease (MSUD) is a genetic disorder that leads the accumulation of branched-chain amino acids (BCAA) leucine (Leu), isoleucine, valine and metabolites. The symptomatology includes psychomotor delay and mental retardation. MSUD therapy comprises a lifelong protein strict diet with low BCAA levels and is well established that high concentrations of Leu and/or its ketoacid are associated with neurological symptoms. Recently, it was demonstrated that the phenylbutyrate (PBA) have the ability to decrease BCAA concentrations. This work aimed the development of lipid-based nanoparticles loaded with PBA, capable of targeting to the central nervous system in order to verify its action mechanisms on oxidative stress and cell death in brain of rats subjected to a MSUD chronic model. PBA-loaded nanoparticles treatment was effective in significantly decreasing BCAA concentration in plasma and Leu in the cerebral cortex of MSUD animals. Furthermore, PBA modulate the activity of catalase, superoxide dismutase, glutathione peroxidase and glutathione reductase enzymes, as well as preventing the oxidative damage to lipid membranes and proteins. PBA was also able to decrease the glial fibrillary acidic protein concentrations and partially decreased the reactive species production and caspase-3 activity in MSUD rats. Taken together, the data indicate that the PBA-loaded nanoparticles could be an efficient adjuvant in the MSUD therapy, protecting against oxidative brain damage and neuroinflammation.
Asunto(s)
Aminoácidos de Cadena Ramificada/sangre , Corteza Cerebral/efectos de los fármacos , Enfermedad de la Orina de Jarabe de Arce/metabolismo , Nanopartículas/administración & dosificación , Estrés Oxidativo/efectos de los fármacos , Fenilbutiratos/administración & dosificación , Animales , Catalasa/metabolismo , Corteza Cerebral/metabolismo , Glutatión Peroxidasa/metabolismo , Enfermedad de la Orina de Jarabe de Arce/sangre , Enfermedad de la Orina de Jarabe de Arce/inducido químicamente , Ratas , Ratas Wistar , Superóxido Dismutasa/metabolismoRESUMEN
Maple syrup urine disease (MSUD) is an inherited deficiency of the branched-chain α-keto dehydrogenase complex, characterized by accumulation of the branched-chain amino acids (BCAAs) and their respective branched chain α-keto-acids (BCKAs), as well as by the presence of alloisoleucine (Allo). Studies have shown that oxidative stress is involved in the pathophysiology of MSUD. In this work, we investigated using the comet assay whether Allo, BCAAs and BCKAs could induce in vitro DNA damage, as well as the influence of l-Carnitine (L-Car) upon DNA damage. We also evaluated urinary 8-hydroxydeoguanosine (8-OHdG) levels, an oxidative DNA damage biomarker, in MSUD patients submitted to a restricted diet supplemented or not with L-Car. All tested concentrations of metabolites (separated or incubated together) induced in vitro DNA damage, and the co-treatment with L-Car reduced these effects. We found that Allo induced the higher DNA damage class and verified a potentiation of DNA damage induced by synergistic action between metabolites. In vivo, it was observed a significant increase in 8-OHdG levels, which was reversed by L-Car. We demonstrated for the first time that oxidative DNA damage is induced not only by BCAAs and BCKAs but also by Allo and we reinforce the protective effect of L-Car.
Asunto(s)
Aminoácidos/administración & dosificación , Carnitina/uso terapéutico , Daño del ADN , Suplementos Dietéticos , Enfermedad de la Orina de Jarabe de Arce , Sustancias Protectoras/uso terapéutico , 8-Hidroxi-2'-Desoxicoguanosina , Aminoácidos/sangre , Aminoácidos/orina , Niño , Preescolar , Ensayo Cometa , Desoxiguanosina/análogos & derivados , Desoxiguanosina/orina , Humanos , Enfermedad de la Orina de Jarabe de Arce/sangre , Enfermedad de la Orina de Jarabe de Arce/dietoterapia , Enfermedad de la Orina de Jarabe de Arce/genética , Enfermedad de la Orina de Jarabe de Arce/orinaRESUMEN
X-linked adrenoleukodystrophy (X-ALD) is an inherited neurometabolic disorder caused by disfunction of the ABCD1 gene, which encodes a peroxisomal protein responsible for the transport of the very long-chain fatty acids from the cytosol into the peroxisome, to undergo ß-oxidation. The mainly accumulated saturated fatty acids are hexacosanoic acid (C26:0) and tetracosanoic acid (C24:0) in tissues and body fluids. This peroxisomal disorder occurs in at least 1 out of 20,000 births. Considering that pathophysiology of this disease is not well characterized yet, and glial cells are widely used in studies of protective mechanisms against neuronal oxidative stress, we investigated oxidative damages and inflammatory effects of vesicles containing lecithin and C26:0, as well as the protection conferred by N-acetyl-L-cysteine (NAC), trolox (TRO), and rosuvastatin (RSV) was assessed. It was verified that glial cells exposed to C26:0 presented oxidative DNA damage (measured by comet assay and endonuclease III repair enzyme), enzymatic oxidative imbalance (high catalase activity), nitrative stress [increased nitric oxide (NO) levels], inflammation [high Interleukin-1beta (IL-1ß) levels], and induced lipid peroxidation (increased isoprostane levels) compared to native glial cells without C26:0 exposure. Furthermore, NAC, TRO, and RSV were capable to mitigate some damages caused by the C26:0 in glial cells. The present work yields experimental evidence that inflammation, oxidative, and nitrative stress may be induced by hexacosanoic acid, the main accumulated metabolite in X-ALD, and that antioxidants might be considered as an adjuvant therapy for this severe neurometabolic disease.
Asunto(s)
Acetilcisteína/farmacología , Cromanos/farmacología , Ácidos Grasos/farmacología , Inflamación/patología , Neuroglía/patología , Estrés Nitrosativo , Estrés Oxidativo , Rosuvastatina Cálcica/farmacología , Animales , Antioxidantes/metabolismo , Catalasa/metabolismo , Supervivencia Celular/efectos de los fármacos , Vesículas Citoplasmáticas/metabolismo , Daño del ADN , Interleucina-1beta/metabolismo , Isoprostanos/metabolismo , Neuroglía/metabolismo , Fármacos Neuroprotectores/farmacología , Nitratos/metabolismo , Nitritos/metabolismo , Estrés Nitrosativo/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , RatasRESUMEN
Several studies have examined neonatal diabetes, a rare disease characterized by hyperglycemia and low insulin levels that is usually diagnosed in the first 6 month of life. Recently, the effects of diabetes on the brain have received considerable attention. In addition, hyperglycemia may perturb brain function and might be associated with neuronal death in adult rats. However, few studies have investigated the damaging effects of neonatal hyperglycemia on the rat brain during central nervous system (CNS) development, particularly the mechanisms involved in the disease. Thus, in the present work, we investigated whether neonatal hyperglycemia induced by streptozotocin (STZ) promoted cell death and altered the levels of proteins involved in survival/death pathways in the rat brain. Cell death was assessed using FluoroJade C (FJC) staining and the expression of the p38 mitogen-activated protein kinase (p38), phosphorylated-c-Jun amino-terminal kinase (p-JNK), c-Jun amino-terminal kinase (JNK), protein kinase B (Akt), phosphorylated-protein kinase B (p-Akt), glycogen synthase kinase-3ß (Gsk3ß), B-cell lymphoma 2 (Bcl2) and Bcl2-associated X protein (Bax) protein were measured by Western blotting. The main results of this study showed that the metabolic alterations observed in diabetic rats (hyperglycemia and hypoinsulinemia) increased p38 expression and decreased p-Akt expression, suggesting that cell survival was altered and cell death was induced, which was confirmed by FJC staining. Therefore, the metabolic conditions observed during neonatal hyperglycemia may contribute to the harmful effect of diabetes on the CNS in a crucial phase of postnatal neuronal development.
Asunto(s)
Encéfalo/patología , Muerte Celular/fisiología , Hiperglucemia/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Animales , Encéfalo/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Femenino , Sistema de Señalización de MAP Quinasas/fisiología , Masculino , Neuronas/metabolismo , Fosforilación , Ratas Wistar , Proteína X Asociada a bcl-2/metabolismoRESUMEN
Introduction: Homocysteine (Hcy) tissue accumulation occurs in a metabolic disease characterized biochemically by cystathionine ß-synthase (CBS) deficiency and clinically by mental retardation, vascular problems, and skeletal abnormalities. Previous studies indicate the occurrence of DNA damage secondary to hyperhomocysteinemia and it was observed that DNA damage occurs in leukocytes from CBS-deficient patients. This study aimed to investigate whether an oxidative mechanism could be involved in DNA damage previously found and investigated the in vitro effect of N-acety-L-cysteine (NAC) on DNA damage caused by high Hcy levels. Methods: We evaluated a biomarker of oxidative DNA damage in the urine of CBSdeficient patients, as well as the in vitro effect of NAC on DNA damage caused by high levels of Hcy. Moreover, a biomarker of lipid oxidative damage was also measured in urine of CBS deficient patients. Results: There was an increase in parameters of DNA (8-oxo-7,8-dihydro-2'- deoxyguanosine) and lipid (15-F2t-isoprostanes levels) oxidative damage in CBS-deficient patients when compared to controls. In addition, a significant positive correlation was found between 15-F2t-isoprostanes levels and total Hcy concentrations. Besides, an in vitro protective effect of NAC at concentrations of 1 and 5 mM was observed on DNA damage caused by Hcy 50 µM and 200 µM. Additionally, we showed a decrease in sulfhydryl content in plasma from CBS-deficient patients when compared to controls. Discussion: These results demonstrated that DNA damage occurs by an oxidative mechanism in CBS deficiency together with lipid oxidative damage, highlighting the NAC beneficial action upon DNA oxidative process, contributing with a new treatment perspective of the patients affected by classic homocystinuria.
Asunto(s)
Humanos , Femenino , Niño , Adolescente , Adulto , Adulto Joven , Acetilcisteína/farmacología , Daño del ADN , Estrés Oxidativo , Cistationina/metabolismo , Desoxiguanosina/orina , Homocistinuria/genética , Antioxidantes/farmacología , Biomarcadores/orina , Estudios de Casos y Controles , Creatinina/orina , Ensayo Cometa , Cistationina/biosíntesis , Cistationina/sangre , Isoprostanos/análisis , Desoxiguanosina/análogos & derivados , Homocisteína/sangre , Homocistinuria/sangreRESUMEN
Lysosomal Storage Disorders (LSD) comprise a heterogeneous group of >50 genetic disorders caused by mutations in genes that encode lysosomal enzymes, transport proteins or other gene products essential for a functional lysosomal system. As a result, abnormal accumulation of substrates within the lysosome leads to a progressive cellular impairment and dysfunction of numerous organs and systems. The exact mechanisms underlying the pathophysiology of LSD remain obscure. Previous studies proposed a relationship between oxidative stress and the pathogenesis of several inborn errors of metabolism, including LSD. Considering these points, in this paper it was reviewed oxidative stress and emerging antioxidant therapy in LSD, emphasizing studies with biological samples from patients affected by this group of conditions. These studies allow presuming that metabolites accumulated in LSD cause an increase of lysosomes' number and size, which may induce excessive production of reactive species and/or deplete the tissue antioxidant capacity, leading to damage in biomolecules. In vitro and in vivo evidence showed that cell oxidative process occurs in LSD and probably contributes to the pathophysiology of these disorders. In this context, it is possible to suggest that, in the future, antioxidants could come to be used as adjuvant therapy for LSD patients.
Asunto(s)
Enfermedades por Almacenamiento Lisosomal/fisiopatología , Estrés Oxidativo , Antioxidantes/uso terapéutico , Biomarcadores/análisis , Humanos , Enfermedades por Almacenamiento Lisosomal/diagnóstico , Enfermedades por Almacenamiento Lisosomal/tratamiento farmacológico , Enfermedades por Almacenamiento Lisosomal/metabolismo , Oxidación-Reducción/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiologíaRESUMEN
Mucopolysaccharidosis type II (MPS II) is a lysosomal storage disease caused by a deficient activity of iduronate-2-sulfatase, leading to abnormal accumulation of glycosaminoglycans (GAG). The main treatment for MPS II is enzyme replacement therapy (ERT). Previous studies described potential benefits of six months of ERT against oxidative stress in patients. Thus, the aim of this study was to investigate oxidative, nitrative and inflammatory biomarkers in MPS II patients submitted to long term ERT. It were analyzed urine and blood samples from patients on ERT (mean time: 5.2years) and healthy controls. Patients presented increased levels of lipid peroxidation, assessed by urinary 15-F2t-isoprostane and plasmatic thiobarbituric acid-reactive substances. Concerning to protein damage, urinary di-tyrosine (di-Tyr) was increased in patients; however, sulfhydryl and carbonyl groups in plasma were not altered. It were also verified increased levels of urinary nitrate+nitrite and plasmatic nitric oxide (NO) in MPS II patients. Pro-inflammatory cytokines IL-1ß and TNF-α were increased in treated patients. GAG levels were correlated to di-Tyr and nitrate+nitrite. Furthermore, IL-1ß was positively correlated with TNF-α and NO. Contrastingly, we did not observed alterations in erythrocyte superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase activities, in reduced glutathione content and in the plasmatic antioxidant capacity. Although some parameters were still altered in MPS II patients, these results may suggest a protective role of long-term ERT against oxidative stress, especially upon oxidative damage to protein and enzymatic and non-enzymatic defenses. Moreover, the redox imbalance observed in treated patients seems to be GAG- and pro-inflammatory cytokine-related.
Asunto(s)
Citocinas/metabolismo , Terapia de Reemplazo Enzimático , Glicosaminoglicanos/metabolismo , Mucopolisacaridosis II/tratamiento farmacológico , Mucopolisacaridosis II/metabolismo , Adolescente , Adulto , Estudios de Casos y Controles , Niño , Humanos , Iduronato Sulfatasa/uso terapéutico , Interleucina-1beta/metabolismo , Masculino , Mucopolisacaridosis II/inmunología , Estrés Nitrosativo/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Factor de Necrosis Tumoral alfa/metabolismo , Adulto JovenRESUMEN
Maple syrup urine disease (MSUD), or branched-chain α-keto aciduria, is an inherited disorder that is caused by a deficiency in branched-chain α-keto acid dehydrogenase complex (BCKAD) activity. Blockade of this pathway leads to the accumulation of the branched-chain amino acids (BCAAs), leucine, isoleucine, and valine, and their respective ketoacids in tissues. The main clinical symptoms presented by MSUD patients include ketoacidosis, hypoglycemia, opisthotonos, poor feeding, apnea, ataxia, convulsions, coma, psychomotor delay, and mental retardation. Although increasing evidence indicates that oxidative stress is involved in the pathophysiology of this disease, the mechanisms of the brain damage caused by this disorder remain poorly understood. In the present study, we investigated the effect of BCAAs on some oxidative stress parameters and evaluated the efficacy of L-carnitine (L-car), an efficient antioxidant that may be involved in the reduction of oxidative damage observed in some inherited neurometabolic diseases, against these possible pro-oxidant effects of a chronic MSUD model in the cerebral cortex and cerebellum of rats. Our results showed that chronic BCAA administration was able to promote both lipid and protein oxidation, impair brain antioxidant defenses, and increase reactive species production, particularly in the cerebral cortex, and that L-car was able to prevent these effects. Taken together, the present data indicate that chronic BCAA administration significantly increased oxidative damage in the brains of rats subjected to a chronic model of MSUD and that L-car may be an efficient antioxidant in this disorder.
Asunto(s)
Encéfalo/patología , Carnitina/farmacología , Enfermedad de la Orina de Jarabe de Arce/inducido químicamente , Enfermedad de la Orina de Jarabe de Arce/patología , Estrés Oxidativo/efectos de los fármacos , Aminoácidos de Cadena Ramificada/farmacología , Animales , Catalasa/metabolismo , Modelos Animales de Enfermedad , Glutatión/metabolismo , Modelos Biológicos , Carbonilación Proteica/efectos de los fármacos , Ratas Wistar , Superóxido Dismutasa/metabolismo , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismoRESUMEN
Oxidative stress has been proposed as an important pathophysiologic feature of various inborn errors of metabolism, including phenylketonuria (PKU). Considering that there are few studies relating oxidative stress and inflammation directly in PKU disease, the aim of this study was to evaluate and correlate oxidative damage to biomolecules, antioxidant defenses, pro-inflammatory cytokines, phenylalanine (Phe) and its metabolites (phenyllactic acid--PLA and phenylacetic acid--PAA) levels in urine and plasma from patients with PKU under dietary treatment. We observed a marked increase of isoprostanes, which is a lipid peroxidation biomarker, in urine from these treated patients. Next, we demonstrated that protein oxidative damage, measured by di-tyrosine formation, was significantly increased in urine from PKU treated patients and that decreased urinary antioxidant capacity was also observed. Our findings concerning to the inflammatory cytokines interleukin-6 and interleukin-1ß, both significantly increased in these patients, provide evidence that the pro-inflammatory state occurs. Besides, interleukin-1ß was positively correlated with isoprostanes. We observed a negative correlation between interleukin-6 and interleukin-10, an anti-inflammatory cytokine. Di-tyrosine was positively correlated with Phe, which indicates oxidative damage to proteins, as well as with PAA. These findings may suggest that the protein damage may be induced by Phe and its metabolite PAA in PKU. Our results indicate that pro-oxidant and pro-inflammatory states occur and are, in part, correlated and protein oxidation seems to be induced by Phe and PPA in PKU patients.
Asunto(s)
Biomarcadores/orina , Citocinas/sangre , Estrés Oxidativo , Fenilcetonurias/sangre , Fenilcetonurias/orina , Adolescente , Niño , Creatina Quinasa/sangre , Dinoprost/análogos & derivados , Dinoprost/orina , Femenino , Humanos , Peroxidación de Lípido , Masculino , Fenilalanina , Especies Reactivas de Oxígeno , Superóxido Dismutasa/orina , Sustancias Reactivas al Ácido Tiobarbitúrico , Tirosina , Adulto JovenRESUMEN
Maple Syrup Urine Disease (MSUD) is a metabolic disorder caused by a severe deficiency of the branched-chain α-keto acid dehydrogenase complex activity which leads to the accumulation of branched-chain amino acids (BCAA) leucine (Leu), isoleucine and valine and their respective α-keto-acids in body fluids. The main symptomatology presented by MSUD patients includes ketoacidosis, failure to thrive, poor feeding, apnea, ataxia, seizures, coma, psychomotor delay and mental retardation, but, the neurological pathophysiologic mechanisms are poorly understood. The treatment consists of a low protein diet and a semi-synthetic formula restricted in BCAA and supplemented with essential amino acids. It was verified that MSUD patients present L-carnitine (L-car) deficiency and this compound has demonstrated an antioxidant and anti-inflammatory role in metabolic diseases. Since there are no studies in the literature reporting the inflammatory profile of MSUD patients and the L-car role on the inflammatory response in this disorder, the present study evaluates the effect of L-car supplementation on plasma inflammatory cytokines interleukin-1ß (IL-1ß), interleukin-6 (IL-6), interferon-gamma (INF-É£), and a correlation with malondialdehyde (MDA), as a marker of oxidative damage, and with free L-car plasma levels in treated MSUD patients. Significant increases of IL-1ß, IL-6, and INF-É£ were observed before the treatment with L-car. Moreover, there is a negative correlation between all cytokines tested and L-car concentrations and a positive correlation among the MDA content and IL-1ß and IL-6 values. Our data show that L-car supplementation can improve cellular defense against inflammation and oxidative stress in MSUD patients and may represent an additional therapeutic approach to the patients affected by this disease.
Asunto(s)
Carnitina/uso terapéutico , Suplementos Dietéticos , Mediadores de Inflamación/sangre , Enfermedad de la Orina de Jarabe de Arce/sangre , Enfermedad de la Orina de Jarabe de Arce/tratamiento farmacológico , Niño , Preescolar , Femenino , Humanos , Inflamación/sangre , Inflamación/tratamiento farmacológico , MasculinoRESUMEN
Maple syrup urine disease (MSUD) is an inherited disorder caused by severe deficient activity of the branched-chain α-keto acid dehydrogenase complex involved in the degradation pathway of branched-chain amino acids (BCAAs) and their α-ketoacid derivatives. MSUD patients generally present ketoacidosis, poor feeding, ataxia, coma, psychomotor delay, mental retardation and brain abnormalites. Treatment consists of dietary restriction of the BCAA (low protein intake) supplemented by a BCAA-free amino acid mixture. Although the mechanisms of brain damage in MSUD are poorly known, previous studies have shown that oxidative stress may be involved in the neuropathology of this disorder. In this regard, it was recently reported that MSUD patients have deficiency of l-carnitine (l-car), a compound with antioxidant properties that is used as adjuvant therapy in various inborn errors of metabolism. In this work, we investigated DNA damage determined by the alkaline comet assay in peripheral whole blood leukocytes of MSUD patients submitted to a BCAA-restricted diet supplemented or not with l-car. We observed a significant increase of DNA damage index (DI) in leukocytes from MSUD patients under BCAA-restricted diet as compared to controls and that l-car supplementation significantly decreased DNA DI levels. It was also found a positive correlation between DI and MDA content, a marker of lipid peroxidation, and an inverse correlation between DI and l-car levels. Taken together, our present results suggest a role for reactive species and the involvement of oxidative stress in DNA damage in this disorder. Since l-car reduced DNA damage, it is presumed that dietary supplementation of this compound may serve as an adjuvant therapeutic strategy for MSUD patients in addition to other therapies.
Asunto(s)
Carnitina/administración & dosificación , Daño del ADN , Leucocitos/metabolismo , Enfermedad de la Orina de Jarabe de Arce/tratamiento farmacológico , Enfermedad de la Orina de Jarabe de Arce/metabolismo , Estrés Oxidativo/efectos de los fármacos , Complejo Vitamínico B/administración & dosificación , Niño , Preescolar , Femenino , Humanos , Leucocitos/patología , Masculino , Enfermedad de la Orina de Jarabe de Arce/genética , Enfermedad de la Orina de Jarabe de Arce/patologíaRESUMEN
Cystathionine-ß-synthase (CBS) deficiency is the main cause of homocystinuria. Homocysteine (Hcy), methionine, and other metabolites of Hcy accumulate in the body of affected patients. Despite the fact that thromboembolism represents the major cause of morbidity in CBS-deficient patients, the mechanisms of cardiovascular alterations found in homocystinuria remain unclear. In this work, we evaluated the lipid and inflammatory profile, oxidative protein damage, and the activities of the enzymes paraoxonase (PON1) and butyrylcholinesterase (BuChE) in plasma of CBS-deficient patients at diagnosis and during the treatment (protein-restricted diet supplemented with pyridoxine, folic acid, betaine, and vitamin B12). We also investigated the effect of folic acid and vitamin B12 on these parameters. We found a significant decrease in HDL cholesterol and apolipoprotein A1 (ApoA-1) levels, as well as in PON1 activity in both untreated and treated CBS-deficient patients when compared to controls. BuChE activity and IL-6 levels were significantly increased in not treated patients. Furthermore, significant positive correlations between PON1 activity and sulphydryl groups and between IL-6 levels and carbonyl content were verified. Moreover, vitamin B12 was positively correlated with PON1 and ApoA-1 levels, while folic acid was inversely correlated with total Hcy concentration, demonstrating the importance of this treatment. Our results also demonstrated that CBS-deficient patients presented important alterations in biochemical parameters, possibly caused by the metabolites of Hcy, as well as by oxidative stress, and that the adequate adherence to the treatment is essential to revert or prevent these alterations.
Asunto(s)
Arildialquilfosfatasa/sangre , Butirilcolinesterasa/sangre , Homocistinuria/sangre , Lípidos/sangre , Oxidantes/sangre , Adolescente , Adulto , Estudios de Casos y Controles , Niño , Preescolar , Cistationina betasintasa/deficiencia , Cistationina betasintasa/genética , Femenino , Ácido Fólico/sangre , Ácido Fólico/fisiología , Homocistinuria/genética , Humanos , Masculino , Estrés Oxidativo/fisiología , Vitamina B 12/sangre , Vitamina B 12/fisiología , Adulto JovenRESUMEN
Toxic metabolites accumulation and oxidative stress have been associated to the pathophysiology of X-linked adrenoleukodystrophy (X-ALD), an inborn error of peroxisome metabolism. Parameters of oxidative damage to proteins and lipids in X-ALD patients were already described in literature; however, DNA injuries were not studied yet. Considering that, the aims were to investigate DNA damage by comet assay in heterozygotes and symptomatic X-ALD patients, to look for associations between DNA damage and lipid peroxidation as measured by urinary 15-F2t-isoprostane; and to evaluate the in vitro effect of N-acetyl-l-cysteine (NAC), trolox (TRO) and rosuvastatin (RSV) on DNA damage in leukocytes from symptomatic patients. Symptomatic patients presented higher DNA damage levels than those found in heterozygotes and controls; heterozygotes and controls showed similar results. In order to investigate the in vitro antioxidant effect on DNA damage, whole blood cells from symptomatic patients were incubated with NAC (1 and 2.5mM), TRO (25 and 75 µM) and RSV (0.5, 2 and 5 µM) before DNA damage analysis. NAC, TRO and RSV, at all tested concentrations, were all capable to reduce DNA damage in symptomatic X-ALD patients until control levels. Finally, DNA damage correlated with urinary isoprostanes and plasmatic levels of TBA-RS and DCFH-DA, allowing to hypothesize that DNA damage might be induced by lipid peroxidation in symptomatic patients. The present work yields experimental evidence that NAC, TRO and RSV reduce the in vitro DNA injury in symptomatic X-ALD patients, what may suggest that the administration of these antioxidants might be considered as an adjuvant therapy for X-ALD.
Asunto(s)
Adrenoleucodistrofia/sangre , Antioxidantes/uso terapéutico , Daño del ADN/efectos de los fármacos , Leucocitos/patología , Adulto , Relación Dosis-Respuesta a Droga , Ácidos Grasos/metabolismo , Femenino , Humanos , Leucocitos/efectos de los fármacos , Peroxidación de Lípido/efectos de los fármacos , Masculino , Estrés Oxidativo , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismoRESUMEN
Maple syrup urine disease (MSUD) is a disorder of branched-chain amino acids (BCAA). The defect in the branched-chain α-keto acid dehydrogenase complex activity leads to an accumulation of these compounds and their corresponding α-keto-acids and α-hydroxy-acids. Studies have shown that oxidative stress may be involved in neuropathology of MSUD. L-carnitine (L-car), which has demonstrated an important role as antioxidant by reducing and scavenging free radicals formation and by enhancing the activity of antioxidant enzymes, have been used in the treatment of some metabolic rare disorders. This study evaluated the oxidative stress parameters, di-tyrosine, isoprostanes and antioxidant capacity, in urine of MSUD patients under protein-restricted diet supplemented or not with L-car capsules at a dose of 50 mg kg(-1) day(-1). It was also determined urinary α-keto isocaproic acid levels as well as blood free L-car concentrations in blood. It was found a deficiency of carnitine in patients before the L-car supplementation. Significant increases of di-tyrosine and isoprostanes, as well as reduced antioxidant capacity, were observed before the treatment with L-car. The L-car supplementation induced beneficial effects on these parameters reducing the di-tyrosine and isoprostanes levels and increasing the antioxidant capacity. It was also showed a significant increase in urinary of α-ketoisocaproic acid after 2 months of L-car treatment, compared to control group. In conclusion, our results suggest that L-car may have beneficial effects in the treatment of MSUD by preventing oxidative damage to the cells and that urine can be used to monitorize oxidative damage in patients affected by this disease.
Asunto(s)
Biomarcadores/orina , Suplementos Dietéticos , Enfermedad de la Orina de Jarabe de Arce/orina , Aminoácidos/orina , Análisis de Varianza , Antioxidantes/metabolismo , Niño , Preescolar , Dinoprost/análogos & derivados , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Isoprostanos/orina , Cetoácidos/orina , Masculino , Enfermedad de la Orina de Jarabe de Arce/dietoterapia , Espectrometría de Masas en Tándem , Tirosina/orinaRESUMEN
Recently, the consequences of diabetes on the central nervous system (CNS) have received great attention. However, the mechanisms by which hyperglycemia affects the central nervous system remain poorly understood. In addition, recent studies have shown that hyperglycemia induces oxidative damage in the adult rat brain. In this regard, no study has assessed oxidative stress as a possible mechanism that affects the brain normal function in neonatal hyperglycemic rats. Thus, the present study aimed to investigate whether neonatal hyperglycemia elicits oxidative stress in the brain of neonate rats subjected to a streptozotocin-induced neonatal hyperglycemia model (5-day-old rats). The activities of glucose-6-phosphate-dehydrogenase (G6PD), 6-phosphogluconate-dehydrogenase (6-PGD), NADPH oxidase (Nox), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSHPx), the production of superoxide anion, the thiobarbituric acid-reactive substances (TBA-RS), and the protein carbonyl content were measured. Neonatal hyperglycemic rats presented increased activities of G6PD, 6PGD, and Nox, which altogether may be responsible for the enhanced production of superoxide radical anion that was observed. The enhanced antioxidant enzyme activities (SOD, CAT, and GSHPx) that were observed in neonatal hyperglycemic rats, which may be caused by a rebound effect of oxidative stress, were not able to hinder the observed lipid peroxidation (TBA-RS) and protein damage in the brain. Consequently, these results suggest that oxidative stress could represent a mechanism that explains the harmful effects of neonatal hyperglycemia on the CNS.
Asunto(s)
Encéfalo/enzimología , Encéfalo/patología , Hiperglucemia/patología , NADPH Oxidasas/metabolismo , Estrés Oxidativo , Vía de Pentosa Fosfato , Animales , Animales Recién Nacidos , Catalasa/metabolismo , Glutatión Peroxidasa/metabolismo , Carbonilación Proteica , Ratas Wistar , Superóxidos/metabolismo , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismoRESUMEN
There is increasing evidence suggesting that oxidative stress plays an important role in the development of many chronic and degenerative conditions such as diabetic encephalopathy and depression. Considering that diabetic rats and mice present higher depressive-like behaviour when submitted to the forced swimming test and that treatment with insulin and/or clonazepam is able to reverse the behavioural changes of the diabetic rats, the present work investigated the antioxidant status, specifically total antioxidant reactivity and antioxidant potential of insulin and clonazepam, as well as the effect of this drugs upon protein oxidative damage and reactive species formation in cortex, hippocampus and striatum from diabetic rats submitted to forced swimming test. It was verified that longer immobility time in diabetic rats and insulin plus clonazepam treatment reversed this depressive-like behaviour. Moreover, data obtained in this study allowed to demonstrate through different parameters such as protein carbonyl content, 2'7'-dichlorofluorescein oxidation, catalase, superoxide dismutase, glutathione peroxidase assay, total radical-trapping antioxidant potential and total antioxidant reactivity that there is oxidative stress in cortex, hippocampus and striatum from diabetic rats under depressive-like behaviour and highlight the insulin and/or clonazepam effect in these different brain areas, restoring antioxidant status and protein damage.
Asunto(s)
Anticonvulsivantes/uso terapéutico , Encefalopatías/complicaciones , Clonazepam/uso terapéutico , Depresión/tratamiento farmacológico , Diabetes Mellitus Experimental/complicaciones , Hipoglucemiantes/uso terapéutico , Insulina/uso terapéutico , Animales , Antioxidantes/metabolismo , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Cuerpo Estriado/metabolismo , Cuerpo Estriado/patología , Depresión/etiología , Hipocampo/metabolismo , Hipocampo/patología , Masculino , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Maple syrup urine disease (MSUD) is an inherited aminoacidopathy caused by a deficiency in branched-chain α-keto acid dehydrogenase complex activity that leads to the accumulation of the branched-chain amino acids (BCAAs) leucine (Leu), isoleucine, and valine and their respective α-keto-acids, α-ketoisocaproic acid (KIC), α keto-ß-methylvaleric acid, and α-ketoisovaleric acid. The major clinical features presented by MSUD patients include ketoacidosis, failure to thrive, poor feeding, apnea, ataxia, seizures, coma, psychomotor delay, and mental retardation; however, the pathophysiology of this disease is poorly understood. MSUD treatment consists of a low protein diet supplemented with a mixture containing micronutrients and essential amino acids but excluding BCAAs. Studies have shown that oxidative stress may be involved in the neuropathology of MSUD, with the existence of lipid and protein oxidative damage in affected patients. In recent years, studies have demonstrated the antioxidant role of L-carnitine (L-Car), which plays a central function in cellular energy metabolism and for which MSUD patients have a deficiency. In this work, we investigated the in vitro effect of Leu and KIC in the presence or absence of L-Car on DNA damage in peripheral whole blood leukocytes using the alkaline comet assay with silver staining and visual scoring. Leu and KIC resulted in a DNA damage index that was significantly higher than that of the control group, and L-Car was able to significantly prevent this damage, mainly that due to KIC.
Asunto(s)
Carnitina/farmacología , Daño del ADN/efectos de los fármacos , Cetoácidos/metabolismo , Leucina/metabolismo , Enfermedad de la Orina de Jarabe de Arce/metabolismo , Complejo Vitamínico B/farmacología , Ensayo Cometa , Metabolismo Energético/efectos de los fármacos , Humanos , Leucocitos/metabolismo , Enfermedad de la Orina de Jarabe de Arce/patología , Estrés OxidativoRESUMEN
The objective of this study was to test whether macromolecule oxidative damage and altered enzymatic antioxidative defenses occur in patients with medium-chain acyl coenzyme A dehydrogenase (MCAD) deficiency. We performed a cross-sectional observational study of in vivo parameters of lipid and protein oxidative damage and antioxidant defenses in asymptomatic, nonstressed, MCAD-deficient patients and healthy controls. Patients were subdivided into three groups based on therapy: patients without prescribed supplementation, patients with carnitine supplementation, and patients with carnitine plus riboflavin supplementation. Compared with healthy controls, nonsupplemented MCAD-deficient patients and patients receiving carnitine supplementation displayed decreased plasma sulfhydryl content (indicating protein oxidative damage). Increased erythrocyte superoxide dismutase (SOD) activity in patients receiving carnitine supplementation probably reflects a compensatory mechanism for scavenging reactive species formation. The combination of carnitine plus riboflavin was not associated with oxidative damage. These are the first indications that MCAD-deficient patients experience protein oxidative damage and that combined supplementation of carnitine and riboflavin may prevent these biochemical alterations. Results suggest involvement of free radicals in the pathophysiology of MCAD deficiency. The underlying mechanisms behind the increased SOD activity upon carnitine supplementation need to be determined. Further studies are necessary to determine the clinical relevance of oxidative stress, including the possibility of antioxidant therapy.