Mitochondrial protective effects caused by the administration of mefenamic acid in sepsis.

The pathophysiology of sepsis may involve the activation of the NOD-type receptor containing the pyrin-3 domain (NLPR-3), mitochondrial and oxidative damages. One of the primary essential oxidation products is 8-oxoguanine (8-oxoG), and its accumulation in mitochondrial DNA (mtDNA) induces cell dysfunction and death, leading to the hypothesis that mtDNA integrity is crucial for maintaining neuronal function during sepsis. In sepsis, the modulation of NLRP-3 activation is critical, and mefenamic acid (MFA) is a potent drug that can reduce inflammasome activity, attenuating the acute cerebral inflammatory process. Thus, this study aimed to evaluate the administration of MFA and its implications for the reduction of inflammatory parameters and mitochondrial damage in animals submitted to polymicrobial sepsis. To test our hypothesis, adult male Wistar rats were submitted to the cecal ligation and perforation (CLP) model for sepsis induction and after receiving an injection of MFA (doses of 10, 30, and 50 mg/kg) or sterile saline (1 mL/kg). At 24 h after sepsis induction, the frontal cortex and hippocampus were dissected to analyze the levels of TNF-α, IL-1ß, and IL-18; oxidative damage (thiobarbituric acid reactive substances (TBARS), carbonyl, and DCF-DA (oxidative parameters); protein expression (mitochondrial transcription factor A (TFAM), NLRP-3, 8-oxoG; Bax, Bcl-2 and (ionized calcium-binding adaptor molecule 1 (IBA-1)); and the activity of mitochondrial respiratory chain complexes. It was observed that the septic group in both structures studied showed an increase in proinflammatory cytokines mediated by increased activity in NLRP-3, with more significant oxidative damage and higher production of reactive oxygen species (ROS) by mitochondria. Damage to mtDNA it was also observed with an increase in 8-oxoG levels and lower levels of TFAM and NGF-1. In addition, this group had an increase in pro-apoptotic proteins and IBA-1 positive cells. However, MFA at doses of 30 and 50 mg/kg decreased inflammasome activity, reduced levels of cytokines and oxidative damage, increased bioenergetic efficacy and reduced production of ROS and 8-oxoG, and increased levels of TFAM, NGF-1, Bcl-2, reducing microglial activation. As a result, it is suggested that MFA induces protection in the central nervous system early after the onset of sepsis.

Coadministration of tianeptine alters behavioral parameters and levels of neurotrophins in a chronic model of Maple Syrup Urine disease.

Maple Syrup Urine Disease (MSUD) is caused by the deficiency in the activity of the branched-chain α-ketoacid dehydrogenase complex (BCKDC), resulting in the accumulation of the branched-chain amino acids (BCAA) leucine, isoleucine, and valine, and their respective branched-chain α-keto acids. Patients with MSUD are at high risk of developing chronic neuropsychiatric disorders; however, the pathophysiology of brain damage in these patients remains unclear. We hypothesize that MSUD can cause depressive symptoms in patients. To test our hypothesis, Wistar rats were submitted to the BCAA and tianeptine (antidepressant) administration for 21 days, starting seven days postnatal. Depression-like symptoms were assessed by testing for anhedonia and forced swimming after treatments. After the last test, the brain structures were dissected for the evaluation of neutrophins. We demonstrate that chronic BCAA administration induced depressive-like behavior, increased BDNF levels, and decreased NGF levels, suggesting a relationship between BCAA toxicity and brain damage, as observed in patients with MSUD. However, the administration of tianeptine was effective in preventing behavioral changes and restoring neurotrophins levels.

Exposure to leucine induces oxidative stress in the brain of zebrafish.

Maple Syrup Urine Disease (MSUD) is an autosomal recessive inherited disorder caused by a deficiency in the activity of the branched-chain alpha-ketoacid dehydrogenase complex leading to the accumulation of branched-chain amino acids (BCAA) leucine, isoleucine, and valine and their respective branched-chain α-ketoacids and corresponding hydroxy acids. Considering that Danio rerio, known as zebrafish, has been widely used as an experimental model in several research areas because it has favorable characteristics that complement other experimental models, this study aimed to evaluate oxidative stress parameters in zebrafish exposed to high levels of leucine (2 mM and 5 mM), in a model similar of MSUD. Twenty-four hours after exposure, the animals were euthanized, and the brain content dissected for analysis of oxidative stress parameters: thiobarbituric acid reactive substances (TBARS), 2',7'-dichlorofluorescein oxidation assay (DCF); content of sulfhydryl, and superoxide dismutase (SOD) and catalase (CAT) activities. Animals exposed to 2 mM and 5 mM leucine showed an increase in the measurement of TBARS and decreased sulfhydryl content. There were no significant changes in DCF oxidation. In addition, animals exposed to 2 mM and 5 mM leucine were found to have decreased SOD activity and increased CAT activity. Based on these results, exposure of zebrafish to high doses of leucine can act as a promising animal model for MSUD, providing a better understanding of the toxicity profile of leucine exposure and its use in future investigations and strategies related to the pathophysiology of MSUD.

Effects of diet-induced hypercholesterolemia and gold nanoparticles treatment on peripheral tissues.

Cholesterol is a lipid molecule of great biological importance to animal cells. Dysregulation of cholesterol metabolism leads to raised blood total cholesterol levels, a clinical condition called hypercholesterolemia. Evidence has shown that hypercholesterolemia is associated with the development of liver and heart disease. One of the mechanisms underlying heart and liver alterations induced by hypercholesterolemia is oxidative stress. In this regard, in several experimental studies, gold nanoparticles (AuNP) displayed antioxidant properties. We hypothesized that hypercholesterolemia causes redox system imbalance in the liver and cardiac tissues, and AuNP treatment could ameliorate it. Young adult male Swiss mice fed a regular rodent diet or a high cholesterol diet for eight weeks and concomitantly treated with AuNP (2.5 µg/kg) or vehicle by oral gavage. Hypercholesterolemia increased the nitrite concentration and glutathione (GSH) levels and decreased the liver's superoxide dismutase (SOD) activity. Also, hypercholesterolemia significantly enhanced the reactive oxygen species (ROS) and GSH levels in cardiac tissue. Notably, AuNP promoted the redox system homeostasis, increasing the SOD activity in hepatic tissue and reducing ROS levels in cardiac tissue. Overall, our data showed that hypercholesterolemia triggered oxidative stress in mice's liver and heart, which was partially prevented by AuNP treatment.

The metabolic effect of α-ketoisocaproic acid: in vivo and in vitro studies.

Maple syrup urine disease (MSUD) is characterized by a deficiency in the mitochondrial branched-chain α-keto acid dehydrogenase complex activity and, consequently, accumulation of the branched-chain amino acids and their respective branched-chain α-keto acids in fluids and the tissue. MSUD clinical symptoms include neurological alterations. KIC is considered one of the significant neurotoxic metabolites since its increased plasma concentrations are associated with neurological symptoms. We evaluated the effect of KIC intracerebroventricular (ICV) injection in hippocampal mitochondria function in rats. We also investigated the impact of KIC in cells' metabolic activity (using MTT assay) and reactive species (RS) production in HT-22 cells. For this, thirty-day-old male rats were bilaterally ICV injected with KIC or aCSF. Thus, 1 hour after the administration, animals were euthanized, and the hippocampus was harvested for measured the activities of mitochondrial respiratory chain enzymes and RS production. Furthermore, HT-22 cells were incubated with KIC (1-10 mM) in 6, 12, and 24 h. Mitochondrial complexes activities were reduced, and the formation of RS was increased in the hippocampus of rats after KIC administration. Moreover, KIC reduced the cells' metabolic ability to reduce MTT and increased RS production in hippocampal neurons. Impairment in hippocampal mitochondrial function seems to be involved in the neurotoxicity induced by KIC.

Administration of branched-chain amino acids alters epigenetic regulatory enzymes in an animal model of Maple Syrup Urine Disease.

Maple Syrup Urine Disease (MSUD) is an autosomal recessive inherited disorder that affects the activity of the branched-chainα-keto acid dehydrogenase complex (BCDK). This deficiency on BCDK complex results in the accumulation of branched-chain amino acids (BCAA) leucine, isoleucine, valine, and their corresponding α-keto acids. Epigenetic changes can negatively affect the metabolism of BCAA. These changes are catalyzed by the epigenetic regulatory enzymes, e.g., DNA methyltransferase (DNMT), histone deacetylases (HDAC), and histone acetyltransferases (HAT). However, the impacts of BCAA administration on the activity of epigenetic regulatory enzymes in the brain of MSUD patients are still unknown. In this study, we aimed to demonstrate the impact of BCAA administration on the activity of DNMT, HDAC, and HAT in the brain structures of infant rats, an animal model of MSUD. For that, we administered a BCAA pool to infant rats for 21 days. We demonstrated that BCAA administration significantly increased the DNMT and HDAC activities in the hippocampus and striatum, but not in the cerebral cortex of MSUD infant rats. A positive correlation was observed between HDAC and DNMT activities in the hippocampus and striatum of animals exposed to BCAA injections. Our results showed that the BCAA administration could modulate epigenetic regulatory enzymes, mainly DNMT and HDAC, in the brains of infant rats. Therefore, we suggest that the increase in the activity of DNMT and HDAC in the hippocampus and striatum could partially explain the neurological impairments presented in animal models of MSUD.

Exposure to a high dose of amoxicillin causes behavioral changes and oxidative stress in young zebrafish.

Autistic spectrum disorder (ASD) is a group of early-onset neurodevelopmental disorders characterized by impaired social and communication skills. Autism is widely described as a behavioral syndrome with multiple etiologies where may exhibit neurobiological, genetic, and psychological deficits. Studies have indicated that long term use of antibiotics can alter the intestinal flora followed by neuroendocrine changes, leading to behavioral changes. Indeed, previous studies demonstrate that a high dose of amoxicillin can change behavioral parameters in murine animal models. The objective was to evaluate behavioral and oxidative stress parameters in zebrafish exposed to a high dose of amoxicillin for 7 days. Young zebrafish were exposed to a daily concentration of amoxicillin (100 mg/L) for 7 days. Subsequently, the behavioral analysis was performed, and the brain content was dissected for the evaluation of oxidative stress parameters. Zebrafish exposed to a high dose of amoxicillin showed locomotor alteration and decreased social interaction behavior. In addition, besides the significant decrease of sulfhydryl content, there was a marked decrease in catalase activity, as well as an increased superoxide dismutase activity in brain tissue. Thus, through the zebrafish model was possible to note a central effect related to the exposition of amoxicillin, the same as observed in murine models. Further, the present data reinforce the relation of the gut-brain-axis and the use of zebrafish as a useful tool to investigate new therapies for autistic traits.

Melatonin ameliorates oxidative stress and DNA damage of rats subjected to a chemically induced chronic model of Maple Syrup Urine Disease.

Maple Syrup Urine Disease (MSUD) is an inborn error of metabolism caused by a deficiency of branched α-ketoacid dehydrogenase complex (BCKDC) activity. Branched-chain amino acids (BCAA) accumulation is, at least in part, responsible for neurological disturbances characteristic of this metabolic disorder. Experimental studies demonstrated that high levels of BCAA induce brain oxidative stress. Considering that many antioxidants are obtained from the diet, the dietary restriction in MSUD patients probably produce deficiency of vitamins and micronutrients involved in antioxidant defenses. Supplementation with synthetic melatonin has been used to prevention and treatment of pathological conditions, including brain diseases. In this study, we aimed at investigating the potential neuroprotective effect of melatonin treatment in a MSUD experimental model. Infant rats (7 day old) received twice daily subcutaneous injections of a BCAA pool (0.21472 g/kg, 190 mmol/L leucine, 59 mmol/L isoleucine and 69 mmol/L valine in saline solution (15.8 µL/g per weight/injection) or saline alone, and supplemented with melatonin (10 mg/kg, intraperitoneal) for 21 days. Oxidative stress parameters, i.e. antioxidant enzyme activity, reactive species production and damage to lipids and proteins, were assessed in the cerebral cortex, hippocampus and striatum at twenty-eight days of age. In addition, the damage to blood cell DNA was evaluated. The chronic administration of BCAA pool in infant rats induced significant oxidative stress (p < 0.05) - such as oxidation of lipids and proteins, imbalance in antioxidant enzymes activities - damages in DNA (p < 0.05) and in brain structures (cerebral cortex, hippocampus and striatum). Notably, melatonin supplementation was able to ameliorate the oxidative (p < 0.05) and antioxidant (p < 0.05) parameters in the brain and blood of the rat model of MSUD. Our results show that melatonin could be a promising therapeutic agent for MSUD.

Effects of omega-3 fatty acids supplementation on inflammatory parameters after chronic administration of L-tyrosine.

Tyrosinemia type II is an autosomal recessive inborn error of metabolism caused by hepatic cytosolic tyrosine aminotransferase deficiency. Importantly, this disease is associated with neurological and developmental abnormalities in many patients. Considering that the mechanisms underlying neurological dysfunction in hypertyrosinemic patients are poorly understood, in the present work we investigated the levels of cytokines - tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), IL-6 and IL-10 - in cerebellum, hippocampus, striatum of young rats exposed to chronic administration of L-tyrosine. In addition, we also investigated the impact of the supplementation with Omega-3 fatty acids (n-3 PUFA) on the rodent model of Tyrosinemia. Notably, previous study demonstrated an association between L-tyrosine toxicity and n-3 PUFA deficiency. Our results showed a significant increase in the levels of pro- and anti-inflammatory cytokines in brain structures when animals were administered with L-tyrosine. Cerebral cortex and striatum seem to be more susceptible to the inflammation induced by tyrosine toxicity. Importantly, n-3 PUFA supplementation attenuated the alterations on cytokines levels induced by tyrosine exposure in brain regions of infant rats. In conclusion, the brain inflammation is also an important process related to tyrosine neurotoxicity observed in the experimental model of Tyrosinemia. Finally, n-3 PUFA supplementation could be considered as a potential neuroprotective adjunctive therapy for Tyrosinemias, especially type II.

Evidence of hippocampal astrogliosis and antioxidant imbalance after L-tyrosine chronic administration in rats.

Tyrosinemia type II is a genetic disorder characterized by elevated blood levels of the amino acid tyrosine caused by the deficiency of tyrosine aminotransferase enzyme, resulting in neurologic and developmental difficulties in the patients. Although neurological sequelae are common in Tyrosinemia type II patients, the mechanisms involved are still poorly understood. The oxidative stress appears to be, at least in part, responsible for neurological complication in this inborn error metabolism. We observed that an acute injection of tyrosine in rats caused a massive oxidative stress in different brain structures. The glutathione system and superoxide dismutase enzyme are relevant antioxidant strategies of the cells and tissues, including in the brain. Other important point is the strong relation between oxidative damage and inflammatory events. Herein, we investigated the effects of chronic administration of tyrosine in the hippocampus of young rats, with emphasis in the activity of GSH related enzymes and superoxide dismutase enzyme, and the astrocytosis. We observed that rats exposed to high levels of tyrosine presented an increased content of tyrosine, which was associated with an increment in the activity of glutathione peroxidase and glutathione reductase as well as with a diminished activity of superoxide dismutase. This antioxidant imbalance was accompanied by enhanced glial fibrillary acidic protein immunoreactivity, a marker of astrocytes, in the brain area studied. In conclusion, hippocampus astrogliosis is also a characteristic of brain alteration in Tyrosinemia. In addition, the chronic exposition to high levels of tyrosine is associated with an alteration in the activity of fundamental antioxidant enzymes.

Omega-3 fatty acid supplementation can prevent changes in mitochondrial energy metabolism and oxidative stress caused by chronic administration of L-tyrosine in the brain of rats.

Deficiency of hepatic enzyme tyrosine aminotransferase characterizes the innate error of autosomal recessive disease Tyrosinemia Type II. Patients may develop neurological and developmental difficulties due to high levels of the amino acid tyrosine in the body. Mechanisms underlying the neurological dysfunction in patients are poorly known. Importantly, Tyrosinemia patients have deficient Omega-3 fatty acids (n-3 PUFA). Here, we investigated the possible neuroprotective effect of the treatment with n-3 PUFA in the alterations caused by chronic administration of L-tyrosine on important parameters of energetic metabolism and oxidative stress in the hippocampus, striatum and cerebral cortex of developing rats. Chronic administration of L-tyrosine causes a decrease in the citrate synthase (CS) activity in the hippocampus and cerebral cortex, as well as in the succinate dehydrogenase (SDH) and isocitrate dehydrogenase (IDH) activities, and an increase in the α-ketoglutarate dehydrogenase activity in the hippocampus. Moreover, in the striatum, L-tyrosine administration caused a decrease in the activities of CS, SDH, creatine kinase, and complexes I, II-III and IV of the mitochondrial respiratory chain. We also observed that the high levels of L-tyrosine are related to oxidative stress in the brain. Notably, supplementation of n-3 PUFA prevented the majority of the modifications caused by the chronic administration of L-tyrosine in the cerebral enzyme activities, as well as ameliorated the oxidative stress in the brain regions of rats. These results indicate a possible neuroprotective and antioxidant role for n-3 PUFA and may represent a new therapeutic approach and potential adjuvant therapy to Tyrosinemia Type II individuals.

N-acetylcysteine effects on a murine model of chronic critical limb ischemia.

During chronic limb ischemia, oxidative damage and inflammation are described. Besides oxidative damage, the decrease of tissue oxygen levels is followed by several adaptive responses. The purpose of this study was to determine whether supplementation with N-acetylcysteine (NAC) is effective in an animal model of chronic limb ischemia. Chronic limb ischemia was induced and animals were treated once a day for 30 consecutive days with NAC (30mg/kg). After this time clinical scores were recorded and soleus muscle was isolated and lactate levels, oxidative damage and inflammatory parameters were determined. In addition, several mechanisms associated with hypoxia adaptation were measured (vascular endothelial growth factor - VEGF and hypoxia inducible factor - HIF levels, ex vivo oxygen consumption, markers of autophagy/mitophagy, and mitochondrial biogenesis). The adaptation to chronic ischemia in this model included an increase in muscle VEGF and HIF levels, and NAC was able to decrease VEGF, but not HIF levels. In addition, ex vivo oxygen consumption under hypoxia was increased in muscle from ischemic animals, and NAC was able to decrease this parameter. This effect was not mediated by a direct effect of NAC on oxygen consumption. Ischemia was followed by a significant increase in muscle myeloperoxidase activity, as well as interleukin-6 and thiobarbituric acid reactive substances species levels. Supplementation with NAC was able to attenuate inflammatory and oxidative damage parameters, and improve clinical scores. In conclusion, NAC treatment decreases oxidative damage and inflammation, and modulates oxygen consumption under hypoxic conditions in a model of chronic limb ischemia.

Role of antioxidant treatment on DNA and lipid damage in the brain of rats subjected to a chemically induced chronic model of tyrosinemia type II.

Tyrosine levels are abnormally elevated in tissues and body fluids of patients with inborn errors of tyrosine metabolism. Tyrosinemia type II, which is caused by tyrosine aminotransferase deficiency, provokes eyes, skin, and central nervous system disturbances in affected patients. However, the mechanisms of brain damage are still poorly known. Considering that studies have demonstrated that oxidative stress may contribute, along with other mechanisms, to the neurological dysfunction characteristic of hypertyrosinemia, in the present study we investigated the effects of antioxidant treatment (NAC and DFX) on DNA damage and oxidative stress markers induced by chronic administration of L-tyrosine in cerebral cortex, hippocampus, and striatum of rats. The results showed elevated levels of DNA migration, and thus DNA damage, after chronic administration of L-tyrosine in all the analyzed brain areas, and that the antioxidant treatment was able to prevent DNA damage in cerebral cortex and hippocampus. However, the co-administration of NAC plus DFX did not prevent the DNA damage in the striatum. Moreover, we found a significant increase in thiobarbituric acid-reactive substances (TBA-RS) and DCFH oxidation in cerebral cortex, as well as an increase in nitrate/nitrite levels in the hippocampus and striatum. Additionally, the antioxidant treatment was able to prevent the increase in TBA-RS levels and in nitrate/nitrite levels, but not the DCFH oxidation. In conclusion, our findings suggest that reactive oxygen and nitrogen species and oxidative stress can play a role in DNA damage in this disorder. Moreover, NAC/DFX supplementation to tyrosinemia type II patients may represent a new therapeutic approach and a possible adjuvant to the current treatment of this disease.

Acute and long-term effects of intracerebroventricular administration of α-ketoisocaproic acid on oxidative stress parameters and cognitive and noncognitive behaviors.

Maple Syrup Urine Disease (MSUD) is biochemically characterized by elevated levels of leucine, isoleucine and valine, as well as their corresponding transaminated branched-chain α-keto acids in tissue and biological fluids. Neurological symptoms and cerebral abnormalities, whose mechanisms are still unknown, are typical of this metabolic disorder. In the present study, we evaluated the early effects (1 h after injection) and long-term effects (15 days after injection) of a single intracerebroventricular administration of α-ketoisocaproic acid (KIC) on oxidative stress parameters and cognitive and noncognitive behaviors. Our results showed that KIC induced early and long-term effects; we found an increase in TBARS levels, protein carbonyl content and DNA damage in the hippocampus, striatum and cerebral cortex both one hour and 15 days after KIC administration. Moreover, SOD activity increased in the hippocampus and striatum one hour after injection, whereas after 15 days, SOD activity decreased only in the striatum. On the other hand, KIC significantly decreased CAT activity in the striatum one hour after injection, but 15 days after KIC administration, we found a decrease in CAT activity in the hippocampus and striatum. Finally, we showed that long-term cognitive deficits follow the oxidative damage; KIC induced impaired habituation memory and long-term memory impairment. From the biochemical and behavioral findings, it we presume that KIC provokes oxidative damage, and the persistence of brain oxidative stress is associated with long-term memory impairment and prepulse inhibition.

Omega-3 fatty acids and mood stabilizers alter behavioral and oxidative stress parameters in animals subjected to fenproporex administration.

Studies have shown that oxidative stress is involved in the pathophysiology of bipolar disorder (BD). It is suggested that omega-3 (ω3) fatty acids are fundamental to maintaining the functional integrity of the central nervous system. The animal model used in this study displayed fenproporex-induced hyperactivity, a symptom similar to manic BD. Our results showed that the administration of fenproporex, in the prevent treatment protocol, increased lipid peroxidation in the prefrontal cortex (143%), hippocampus (58%) and striatum (181%), and ω3 fatty acids alone prevented this change in the prefrontal cortex and hippocampus, whereas the co-administration of ω3 fatty acids with VPA prevented the lipoperoxidation in all analyzed brain areas, and the co-administration of ω3 fatty acids with Li prevented this increase only in the prefrontal cortex and striatum. Moreover, superoxide dismutase (SOD) activity was decreased in the striatum (54%) in the prevention treatment, and the administration of ω3 fatty acids alone or in combination with Li and VPA partially prevented this inhibition. On the other hand, in the reversal treatment protocol, the administration of fenproporex increased carbonyl content in the prefrontal cortex (25%), hippocampus (114%) and striatum (91%), and in prefrontal coxter the administration of ω3 fatty acids alone or in combination with Li and VPA reversed this change, whereas in the hippocampus and striatum only ω3 fatty acids alone or in combination with VPA reversed this effect. Additionally, the administration of fenproporex resulted in a marked increase of TBARS in the hippocampus and striatum, and ω3 fatty acids alone or in combination with Li and VPA reversed this change. Finally, fenproporex administration decreased SOD activity in the prefrontal cortex (85%), hippocampus (52%) and striatum (76%), and the ω3 fatty acids in combination with VPA reversed this change in the prefrontal cortex and striatum, while the co-administration of ω3 fatty acids with Li reversed this inhibition in the hippocampus and striatum. In conclusion, our results support other studies showing the importance of ω3 fatty acids in the brain and the potential for these fatty acids to aid in the treatment of BD.

Antioxidants reverse the changes in energy metabolism of rat brain after chronic administration of L.-tyrosine.

Tyrosinemia type II is a rare autosomal recessive disease caused by deficiency of hepatic tyrosine aminotransferase and is associated with neurologic and development difficulties in numerous patients. Considering that the mechanisms underlying the neurological dysfunction in hypertyrosinemic patients are poorly known and that high concentrations of tyrosine provoke mitochondrial dysfunction and oxidative stress, in the present study we investigated the in vivo influence of antioxidants (N-acetylcysteine, NAC; and deferoxamine, DFX) administration on the inhibitory effects on parameters of energy metabolism in cerebral cortex, hippocampus and striatum of rats, provoked by chronic administration of L.-tyrosine. Our results showed that chronic administration of L.-tyrosine results in a marked decrease in the activity of citrate synthase in all the analyzed structures and succinate dehydrogenase activities in hippocampus and striatum, and that antioxidants administration can prevent this inhibition in hippocampus and striatum. Moreover, chronic administration of L.-tyrosine inhibited the activity of complex I, II-III and IV in the striatum, which can be prevented by antioxidant treatment. However, the co-administration of NAC plus DFX could not prevent the inhibition of creatine kinase activity in the striatum. In conclusion, the present study demonstrates that the administration of antioxidants NAC and DFX attenuates the L.-tyrosine effects on enzymes of the Krebs cycle and the mitochondrial respiratory chain, suggesting that impairment of energy metabolism can be involved with oxidative stress. These results also indicate a possible neuroprotective role for NAC and DFX as a potential adjuvant therapy to the patients with Tyrosinemia type II.

Apoptotic signaling pathways induced by acute administration of branched-chain amino acids in an animal model of maple syrup urine disease.

Maple Syrup Urine Disease (MSUD) is an inborn error of metabolism caused by a deficiency of the branched-chain α-keto acid dehydrogenase complex activity. This blockage leads to accumulation of the branched-chain amino acids leucine, isoleucine and valine, as well as their corresponding α-keto acids and α-hydroxy acids. The affected patients present severe neurological symptoms, such as coma and seizures, as well as edema and cerebral atrophy. Considering that the mechanisms of the neurological symptoms presented by MSUD patients are still poorly understood, in this study, protein levels of apoptotic factors are measured, such as Bcl-2, Bcl-xL, Bax, caspase-3 and -8 in hippocampus and cerebral cortex of rats submitted to acute administration of branched-chain amino acids during their development. The results in this study demonstrated that BCAA acute exposure during the early postnatal period did not significantly change Bcl-2, Bcl-xL, Bax and caspase-8 protein levels. However, the Bax/Bcl-2 ratio and procaspase-3 protein levels were decreased in hippocampus. On the other hand, acute administration of BCAA in 30-day-old rats increase in Bax/Bcl-2 ratio followed by an increased caspase-3 activity in cerebral cortex, whereas BCAA induces apoptosis in hippocampus through activation and cleavage of caspase-3 and -8 without changing the Bax/Bcl-2 ratio. In conclusion, the results suggest that apoptosis could be of pivotal importance in the developmental neurotoxic effects of BCAA. In addition, the current studies also suggest that multiple mechanisms may be involved in BCAA-induced apoptosis in the cerebral cortex and hippocampus.

Omega-3 fatty acid supplementation decreases DNA damage in brain of rats subjected to a chemically induced chronic model of Tyrosinemia type II.

Tyrosinemia type II is an inborn error of metabolism caused by a mutation in a gene encoding the enzyme tyrosine aminotransferase leading to an accumulation of tyrosine in the body, and is associated with neurologic and development difficulties in numerous patients. Because the accumulation of tyrosine promotes oxidative stress and DNA damage, the main aim of this study was to investigate the possible antioxidant and neuroprotective effects of omega-3 treatment in a chemically-induced model of Tyrosinemia type II in hippocampus, striatum and cerebral cortex of rats. Our results showed chronic administration of L-tyrosine increased the frequency and the index of DNA damage, as well as the 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels in the hippocampus, striatum and cerebral cortex. Moreover, omega-3 fatty acid treatment totally prevented increased DNA damage in the striatum and hippocampus, and partially prevented in the cerebral cortex, whereas the increase in 8-OHdG levels was totally prevented by omega-3 fatty acid treatment in hippocampus, striatum and cerebral cortex. In conclusion, the present study demonstrated that the main accumulating metabolite in Tyrosinemia type II induce DNA damage in hippocampus, striatum and cerebral cortex, possibly mediated by free radical production, and the supplementation with omega-3 fatty acids was able to prevent this damage, suggesting that could be involved in the prevention of oxidative damage to DNA in this disease. Thus, omega-3 fatty acids supplementation to Tyrosinemia type II patients may represent a new therapeutic approach and a possible adjuvant to the curren t treatment of this disease.

Activity of Krebs cycle enzymes in mdx mice.

INTRODUCTION: Duchenne muscular dystrophy (DMD) is a degenerative disease of skeletal, respiratory, and cardiac muscles caused by defects in the dystrophin gene. More recently, brain involvement has been verified. Mitochondrial dysfunction and oxidative stress may underlie the pathophysiology of DMD. In this study we evaluate Krebs cycle enzymes activity in the cerebral cortex, diaphragm, and quadriceps muscles of mdx mice. METHODS: Cortex, diaphragm, and quadriceps tissues from male dystrophic mdx and control mice were used. RESULTS: We observed increased malate dehydrogenase activity in the cortex; increased malate dehydrogenase and succinate dehydrogenase activities in the diaphragm; and increased citrate synthase, isocitrate dehydrogenase, and malate dehydrogenase activities in the quadriceps of mdx mice. CONCLUSION: This study showed increased activity of Krebs cycle enzymes in cortex, quadriceps, and diaphragm in mdx mice.

Intracerebroventricular administration of α-ketoisocaproic acid decreases brain-derived neurotrophic factor and nerve growth factor levels in brain of young rats.

Maple syrup urine disease (MSUD) is an inherited aminoacidopathy resulting from dysfunction of the branched-chain keto acid dehydrogenase complex, leading to accumulation of the branched-chain amino acids (BCAA) leucine, isoleucine and valine as well as their corresponding transaminated branched-chain α-ketoacids. This disorder is clinically characterized by ketoacidosis, seizures, coma, psychomotor delay and mental retardation whose pathophysiology is not completely understood. Recent studies have shown that oxidative stress may be involved in neuropathology of MSUD. However, the effect of accumulating α-ketoacids in MSUD on neurotrophic factors has not been investigated. Thus, the objective of the present study was to evaluate the effects of acute intracerebroventricular administration of α-ketoisocaproic acid (KIC) on brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) levels in the brains of young male rats. Ours results showed that intracerebroventricular administration of KIC decreased BDNF levels in hippocampus, striatum and cerebral cortex, without induce a detectable change in pro-BDNF levels. Moreover, NGF levels in the hippocampus were reduced after intracerebroventricular administration of KIC. In conclusion, these data suggest that the effects of KIC on demyelination and memory processes may be mediated by reduced trophic support of BDNF and NGF. Moreover, lower levels of BDNF and NGF are consistent with the hypothesis that a deficit in this neurotrophic factor may contribute to the structural and functional alterations of brain underlying the psychopathology of MSUD, supporting the hypothesis of a neurodegenerative process in MSUD.