Andrea Pereira Rosa

In vivo neuroprotective effect of L-carnitine against oxidative stress in maple syrup urine disease

Maple syrup urine disease (MSUD) is an autosomal recessive inborn error of metabolism caused by deficiency of the activity of the mitochondrial enzyme complex branched-chain α-keto acid dehydrogenase (BCKAD) leading to accumulation of the branched-chain amino acids (BCAA) leucine, isoleucine and valine and their corresponding branched-chain α-keto acids. Affected patients present severe brain dysfunction manifested such as ataxia, seizures, coma, psychomotor delay and mental retardation. The mechanisms of brain damage in this disease remain poorly understood. Recent studies have shown that oxidative stress may be involved in neuropathology of MSUD. L-Carnitine (L-Car) is considered a potential antioxidant through its action against peroxidation as a scavenger of reactive oxygen species and by its stabilizing effect of damage to cell membranes. In this study we evaluate the possible neuroprotective in vivo effects of L-Car against pro-oxidative effects of BCAA in cerebral cortex of rats. L-Car prevented lipoperoxidation, measured by thiobarbituric acid-reactive substances, protein damage, measured by sulfhydryl and protein carbonyl content and alteration on catalase and glutathione peroxidase activity in rat cortex from a chemically-induced model of MSUD. Our data clearly show that L-Car may be an efficient antioxidant, protecting against the oxidative stress promoted by BCAA. If the present results are confirmed in MSUD patients, this could represent an additional therapeutic approach to the patients affected by this disease.

Glutathione metabolism enzymes in brain and liver of hyperphenylalaninemic rats and the effect of lipoic acid treatment

Phenylketonuria (PKU) is a disorder caused by a deficiency in phenylalanine hydroxylase activity, which converts phenylalanine (Phe) to tyrosine, leading to hyperphenylalaninemia (HPA) with accumulation of Phe in tissues of patients. The neuropathophysiology mechanism of disease remains unknown. However, recently the involvement of oxidative stress with decreased glutathione levels in PKU has been reported. Intracellular glutathione (GSH) levels may be maintained by the antioxidant action of lipoic acid (LA). The aim of this study was to evaluate the activity of the enzymes involved in the metabolism and function of GSH, such as glutathione peroxidase (GSH-Px), glucose-6-phosphate dehydrogenase (G6PD), glutathione reductase (GR), glutamate-cysteine ligase (GCL), glutathione-S-transferase (GST) and GSH content in brain and liver of young rats subjected to a chemically induced model of HPA and the effect of LA for a week. In brain, the administration of Phe reduced the activity of the GSH-Px, GR and G6PD and LA prevented these effects totally or partially. GCL activity was increased by HPA and was not affect by LA antioxidant treatment. GST activity did not differ between groups. GSH content was increased by LA and decreased by HPA treatment in brain samples. Considering the liver, all parameters analyzed were increased in studied HPA animals and LA was able to hinder some effects except for the GCL, GST enzymes and GSH content. These results suggested that HPA model alter the metabolism of GSH in rat brain and liver, which may have an important role in the maintenance of GSH function in PKU although liver is not a directly affected organ in this disease. So, an antioxidant therapy with LA may be useful in the treatment of oxidative stress in HPA.

L-carnitine Prevents Oxidative Stress in the Brains of Rats Subjected to a Chemically Induced Chronic Model of MSUD

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.

Neonatal hyperglycemia induces oxidative stress in the rat brain: the role of pentose phosphate pathway enzymes and NADPH oxidase.

Abstract 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.

Diabetic encephalopathy-related depression: experimental evidence that insulin and clonazepam restore antioxidant status in rat brain.

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. Copyright

Enzymatic scavengers in the epididymal fluid: Comparison between pony and miniature breed stallions

The use of stallion semen collected from cauda epididymis for AI has increased due to the new protocols available for cryopreservation. Preserving the genetic material from valuable males that suffer sudden death or other events that prematurely end the stallions reproductive life is an important strategy for Stud breeding management. While protecting spermatozoa from oxidative stress and infectious agents, the epididymis promotes the enhancement of sperm cell morphology and changes in membrane protein profile, increasing its fertility potential. The epididymal fluid must be a balanced redox environment to allow sperm preservation and protein-protein and protein-lipids interactions to quantify. The aim of this study was quantify the enzymatic ROS scavengers in epididymal fluid of pony and miniature breed stallions. Epididymides from 8 pony stallions and 12 miniature breed stallions were dissected and fluid from caput, corpus and cauda epididymis collected. Spermatozoa were separated of epididymal fluid by 2-step centrifugation. The activities of catalase, superoxide dismutase (SOD) and glutathione peroxidase (GPx) were measured and compared between stallion groups and epididymal regions. The three enzymes were present in all epididymal regions tested, with higher activities of catalase and SOD in cauda epididymis in miniature breed stallions (P<0.05). GPx activity was higher in caput epididymis in pony stallions (P<0.05), however with no difference to fluid from cauda epididymis of both breeds. These results show a difference in antioxidant enzymatic scavengers between pony and miniature breed stallions. Also, our data confirm the protective role of cauda epididymis, preserving spermatozoa integrity from oxidative damage. As glutathione peroxidase is involved in several signaling pathways, its constant activity during epididymal transit corroborates the importance of this enzyme for spermatozoa maturation.

Exercício aeróbico agudo restaura a concentração de triptofano em cérebro de ratos com hiperfenilalaninemia

INTRODUCAO: A fenilcetonuria (PKU) e caracterizada pela deficiencia da enzima fenilalanina hidroxilase, causando acumulo de fenilalanina. O diagnostico precoce e a subordinacao a dieta pobre em fenilalanina sao importantes para prevenir os efeitos prejudiciais da hiperfenilalaninemia. Nao aderir estritamente a dieta provoca, entre outros efeitos, um desequilibrio entre os aminoacidos neutros que usam o mesmo transportador da fenilalanina na barreira hematoencefalica, causando, entao, a diminuicao da entrada de triptofano, o precursor de serotonina no cerebro. Esse neurotransmissor tem sido implicado na regulacao dos estados de humor, sendo sua alta producao ligada a fadiga central em individuos submetidos a exercicio prolongado. O exercicio fisico aumenta os niveis de triptofano livre no sangue, o que facilita seu influxo no cerebro, podendo, portanto, ser util nos estados hiperfenilalaninemicos. OBJETIVO: Avaliar se o exercicio aerobico e capaz de normalizar as concentracoes de triptofano no cerebro de ratos com hiperfenilalaninemia. METODOS: Trinta e dois ratos foram separados nos grupos sedentario (Sed) e exercicio (Exe), e cada um deles subdividido em controle (SAL) e hiperfenilalaninemia (PKU). A hiperfenilalaninemia foi induzida pela administracao de alfa-metilfenilalanina e fenilalanina durante tres dias, enquanto os grupos SAL receberam salina. Os grupos Exe realizaram uma sessao de exercicio aerobico com duracao de 60min e velocidade de 12m.min-1. RESULTADOS: A concentracao de triptofano no cerebro nos grupos PKU foi significativamente menor que nos grupos SAL, tanto Sed como Exe, compativel com a condicao hiperfenilalaninemica. O exercicio aumentou a concentracao cerebral de triptofano comparada aos animais sedentarios. O achado mais interessante foi que a concentracao cerebral de triptofano no grupo ExePKU nao foi diferente do SedSAL. CONCLUSAO: Os resultados indicam um importante papel do exercicio aerobico para restaurar a concentracao de triptofano no cerebro em ratos hiperfenilalaninemicos.