Anatomopathological characterization of branchial cysts in slaughtered pigs.

Branchial cysts are a congenital anomaly in humans and other animal species. In this study, twenty commercially bred slaughtered pigs ranging from 120 to 150 days of age, sourced from different farms and lots, were found to have cysts in the oropharyngeal region at meat inspection despite the absence of clinical signs. Two cysts were selected for histopathological examination. The first cyst was surrounded by fibrous connective tissue and lined by a simple single cell layer of epithelium. The second cyst comprised a squamous pseudostratified to simple stratified epithelium, accompanied by a mild inflammatory infiltrate. This cyst was also surrounded by fibrous connective tissue and glands. The pathological diagnosis of branchial cysts in slaughtered pigs was established on the basis of their anatomical location and gross and microscopic findings.

Brazilian Red Propolis Presents Promising Anti-H. pylori Activity in In Vitro and In Vivo Assays with the Ability to Modulate the Immune Response.

Helicobacter pylori is a Gram-negative, microaerophilic, curved-rod, flagellated bacterium commonly found in the stomach mucosa and associated with different gastrointestinal diseases. With high levels of prevalence worldwide, it has developed resistance to the antibiotics used in its therapy. Brazilian red propolis has been studied due to its biological properties, and in the literature, it has shown promising antibacterial activities. The aim of this study was to evaluate anti-H. pylori from the crude hydroalcoholic extract of Brazilian red propolis (CHEBRP). For this, in vitro determination of the minimum inhibitory and bactericidal concentration (MIC/MBC) and synergistic activity and in vivo, microbiological, and histopathological analyses using Wistar rats were carried out using CHEBRP against H. pylori strains (ATCC 46523 and clinical isolate). CHEBRP presented MIC/MBC of 50 and 100 µg/mL against H. pylori strains (ATCC 43526 and clinical isolate, respectively) and tetracycline MIC/MBC of 0.74 µg/mL. The association of CHEBRP with tetracycline had an indifferent effect. In the stomach mucosa of rats, all treatments performed significantly decreased the number of H. pylori, and a concentration of 300 mg/kg was able to modulate the inflammatory response in the tissue. Therefore, CHEBRP showed promising anti-H. pylori in in vitro and in vivo assays.

Comparison of immunohistochemical profiles of ovarian germ cells in dysgerminomas of a captive maned wolf and domestic dogs.

We characterized the immunohistochemical expression profiles of dysgerminomas from a 16-y-old maned wolf and 13 domestic dogs using the following biomarkers: Sal-like protein 4 (SALL4), octamer-binding transcription factor 3/4 (OCT3/4), placental alkaline phosphatase (PLAP), c-kit, and vimentin. The maned wolf had nonspecific and long-standing clinical signs of lethargy, anorexia, and weight loss, and was euthanized because of poor prognosis. At autopsy, the left ovary was effaced by a 12 × 8 × 6 cm mass, comprised of anaplastic cells with a mitotic count of 20 mitoses in 10 high power fields. Dysgerminomas from 7 of 13 domestic dogs had nuclear expression of SALL4. Dysgerminomas from the maned wolf and 2 domestic dogs had both nuclear and cytoplasmic expression of SALL4. Cytoplasmic expression of PLAP and OCT3/4 was present in dysgerminomas from the maned wolf and 3 (PLAP) or 4 (OCT3/4) domestic dogs. All dysgerminomas expressed vimentin. Membranous c-kit expression was rare in the dysgerminoma from the maned wolf, and variable in dysgerminomas from 4 domestic dogs. A dysgerminoma from a domestic dog had cytoplasmic expression of c-kit. SALL4 is a useful marker to confirm germ cell origin of dysgerminoma in canids.

In Vitro and In Vivo Models for Studying SARS-CoV-2, the Etiological Agent Responsible for COVID-19 Pandemic.

The emergence and rapid worldwide spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has prompted the scientific community to rapidly develop in vitro and in vivo models that could be applied in COVID-19 research. In vitro models include two-dimensional (2D) cultures of immortalized cell lines or primary cells and three-dimensional (3D) cultures derived from lung, alveoli, bronchi, and other organs. Although cell-based systems are economic and allow strict control of experimental variables, they do not always resemble physiological conditions. Thus, several in vivo models are being developed, including different strains of mice, hamsters, ferrets, dogs, cats, and non-human primates. In this review, we summarize the main models of SARS-CoV-2 infection developed so far and discuss their advantages, drawbacks and main uses.

Acute necrotic hepatotoxicity caused by Lantana camara L. ingestion in dairy cattle

ABSTRACT: This study describes an outbreak of acute necrotic hepatopathy associated with spontaneous poisoning by Lantana camara L. in dairy cattle. A herd of 15 cows and heifers was introduced into a native pasture with limited food supply, and, sixteen days later, eight animals had spontaneous nasal hemorrhage, fever, lethargy, jaundice, and dry, dark stools with mucus and blood. The clinical course varied from two to five days. In the pasture where the cattle were kept, abundant adult specimens of L. camara L. with evident signs of consumption were observed. In total, seven cattle died and necropsy was performed in three of them. All animals had moderate jaundice, hemorrhage in the subcutaneous tissue and on the surface of different organs. The liver was slightly enlarged, with orange discoloration and enhanced lobular pattern. Histologically, multifocal areas of coagulative necrosis of hepatocytes in the centrilobular area, occasionally extending to the midzonal area, were observed, as well as marked hepatocellular degeneration and prominent cholestasis. The current study suggests that L. camara L. poisoning should be considered a differential diagnosis of acute and necrotic hepatotoxicity in cattle, despite the absence of photosensitization.

RESUMO: Esse estudo descreve um surto de hepatopatia necrótica aguda associada a intoxicação espontânea por Lantana camara L. em bovinos leiteiros. Um lote de 15 vacas e novilhas foram introduzidas em um piquete com campo nativo, com escassa oferta de alimento. Após dezesseis dias, oito animais manifestaram epistaxe, febre, apatia, icterícia, fezes ressecadas e escuras com muco e sangue. A evolução do quadro clínico variou de dois a cinco dias. No piquete em que os bovinos estavam alojados havia grande quantidade de L. camara L. com sinais evidentes de consumo. No total, sete bovinos morreram, e destes, o exame post mortem foi realizado em três. Os bovinos exibiam moderada icterícia, hemorragias no tecido subcutâneo e na superfície de diferentes órgãos. O fígado estava discretamente aumentado, com coloração alaranjada e padrão lobular evidente. As lesões histológicas consistiam em acentuada necrose de coagulação de hepatócitos em região centrolobular, por vezes se estendendo a região mediozonal, além de acentuada degeneração dos hepatócitos e evidente Colestase. O presente trabalho alerta para que intoxicação por L. camara L. seja levada em consideração nos diagnósticos diferenciais de hepatotoxicidade necrótica aguda em bovinos, mesmo sem indícios de fotossensibilização.

Acute necrotic hepatotoxicity caused by Lantana camara L. ingestion in dairy cattle / Hepatotoxicidade necrótica aguda por ingestão de Lantana camara L. em bovinos leiteiros

This study describes an outbreak of acute necrotic hepatopathy associated with spontaneous poisoning by Lantana camara L. in dairy cattle. A herd of 15 cows and heifers was introduced into a native pasture with limited food supply, and, sixteen days later, eight animals had spontaneous nasal hemorrhage, fever, lethargy, jaundice, and dry, dark stools with mucus and blood. The clinical course varied from two to five days. In the pasture where the cattle were kept, abundant adult specimens of L. camara L. with evident signs of consumption were observed. In total, seven cattle died and necropsy was performed in three of them. All animals had moderate jaundice, hemorrhage in the subcutaneous tissue and on the surface of different organs. The liver was slightly enlarged, with orange discoloration and enhanced lobular pattern. Histologically, multifocal areas of coagulative necrosis of hepatocytes in the centrilobular area, occasionally extending to the midzonal area, were observed, as well as marked hepatocellular degeneration and prominent cholestasis. The current study suggests that L. camara L. poisoning should be considered a differential diagnosis of acute and necrotic hepatotoxicity in cattle, despite the absence of photosensitization.(AU)

Esse estudo descreve um surto de hepatopatia necrótica aguda associada a intoxicação espontânea por Lantana camara L. em bovinos leiteiros. Um lote de 15 vacas e novilhas foram introduzidas em um piquete com campo nativo, com escassa oferta de alimento. Após dezesseis dias, oito animais manifestaram epistaxe, febre, apatia, icterícia, fezes ressecadas e escuras com muco e sangue. A evolução do quadro clínico variou de dois a cinco dias. No piquete em que os bovinos estavam alojados havia grande quantidade de L. camara L. com sinais evidentes de consumo. No total, sete bovinos morreram, e destes, o exame post mortem foi realizado em três. Os bovinos exibiam moderada icterícia, hemorragias no tecido subcutâneo e na superfície de diferentes órgãos. O fígado estava discretamente aumentado, com coloração alaranjada e padrão lobular evidente. As lesões histológicas consistiam em acentuada necrose de coagulação de hepatócitos em região centrolobular, por vezes se estendendo a região mediozonal, além de acentuada degeneração dos hepatócitos e evidente Colestase. O presente trabalho alerta para que intoxicação por L. camara L. seja levada em consideração nos diagnósticos diferenciais de hepatotoxicidade necrótica aguda em bovinos, mesmo sem indícios de fotossensibilização.(AU)

Hematological findings and factors associated with feline leukemia virus (FeLV) and feline immunodeficiency virus (FIV) positivity in cats from southern Brazil / Achados hematológicos e fatores associados a positividade pelo vírus da leucemia felina (felv) e vírus da imunodeficiência felina em gatos do sul do Brasil

Using a retrospective study, 493 cats tested for FeLV and FIV were selected for analysis of the association between hematologic findings and positivity at immunoassay test. Individual and hematologic variables were assessed considering the influence of results using univariate and multivariate logistic regression analysis. Out 153 of the 493 cats were positive for FeLV (31%), 50 were positive for FIV (10.1%) and 22 were positive for both FIV and FeLV (4.4%). Multivariate analysis detected significant associations between FeLV infection and age below 1 year (p=0.01), age from 1 to 10 years (p=0.03), and crossbreed (p=0.04). Male cats were more likely to be FIV-positive (p=0.002). Regarding hematological changes, FeLV-positive cats have higher odds to anemia, leukopenia and lymphopenia than FeLV-negative cats. FIV-positive cats are more likely to have anemia than negative. Identification of associated factors related to animal status and correlation of hematological disorders with infection by retroviruses in cats could be useful for detecting these retroviral diseases in cats.(AU)

Através de um estudo retrospectivo, 493 gatos testados para FeLV e FIV foram selecionados para análise da associação entre as alterações hematológicas e a positividade no teste imunoenzimático. Variáveis individuais e hematológicas foram consideradas para verificar a influência dos resultados utilizando análise de regressão logística univariada e multivariada. Um total de 153 de 493 gatos avaliados foram positivos para o FeLV (31%), 50 foram positivos para o FIV (10,1%) e 22 foram positivos para FIV e FeLV (4,4%). Análise multivariada detectou uma associação significativa entre a infecção pelo FeLV e a idade abaixo de 1 ano (P=0,01), idade entre 1 a 10 anos (P=0,03) e raça mista (P=0,04). Gatos machos foram mais predispostos a serem positivos para FIV (P=0,002). Com base nas alterações hematológicas, gatos positivos para o FeLV tem maior odds para apresentar anemia, leucopenia e linfopenia que os negativos. Gatos positivos para FIV possuem maiores chances de apresentarem anemia que os gatos negativos. A identificação dos fatores associados à infecção relacionados ao perfil do animal e a correlação com os distúrbios hematológicos com a infecção, pode ser útil para detecção das doenças retrovirais em gatos.(AU)

Inhibition of brain energy metabolism by the branched-chain amino acids accumulating in maple syrup urine disease.

In the present work we investigated the in vitro effect of the branched-chain amino acids (BCAA) accumulating in maple syrup urine disease (MSUD) on some parameters of energy metabolism in cerebral cortex of rats. 14CO2 production from [1-14C]acetate, [1-5-14C]citrate and [U-14C]glucose, as well as glucose uptake by the brain were evaluated by incubating cortical prisms from 30-day-old rats in the absence (controls) or presence of leucine (Leu), valine (Val) or isoleucine (Ile). All amino acids significantly reduced 14CO2 production by around 20-55%, in contrast to glucose utilization, which was significantly increased by up to 90%. Furthermore, Leu significantly inhibited the activity of the respiratory chain complex IV, whereas Val and Ile markedly inhibited complexes II-III, III and IV by up to 40%. We also observed that trolox (alpha-tocopherol) and creatine totally prevented the inhibitory effects provoked by the BCAA on the respiratory chain complex activities, suggesting that free radicals were involved in these effects. The results indicate that the major metabolites accumulating in MSUD disturb brain aerobic metabolism by compromising the citric acid cycle and the electron flow through the respiratory chain. We presume that these findings may be of relevance to the understanding of the pathophysiology of the neurological dysfunction of MSUD patients.

Evidence that 3-hydroxy-3-methylglutaric acid promotes lipid and protein oxidative damage and reduces the nonenzymatic antioxidant defenses in rat cerebral cortex.

In the present work we investigated the in vitro effect of 3-hydroxy-3-methylglutarate (HMG) that accumulates in 3-hydroxy-3-methylglutaryl-CoA lyase deficiency (HMGLD) on important parameters of oxidative stress in rat cerebral cortex. It was observed that HMG induced lipid peroxidation by significantly increasing chemiluminescence and levels of thiobarbituric acid-reactive substances (TBA-RS). This effect was prevented by the antioxidants alpha-tocopherol, melatonin, N-acetylcysteine, and superoxide dismutase plus catalase, suggesting that free radicals were involved in the lipid oxidative damage. On the other hand, HMG did not change TBA-RS levels in intact or disrupted mitochondrial preparations, indicating that generation of oxidants by this organic acid was dependent on cytosolic mechanisms. HMG also induced protein oxidative damage in cortical supernatants, which was reflected by increased carbonyl content and sulfhydryl oxidation. Furthermore, HMG significantly reduced the nonenzymatic antioxidant defenses total-radical trapping antioxidant potential, total antioxidant reactivity, and reduced glutathione (GSH) levels in rat cerebral cortex. HMG-induced GSH reduction was totally blocked by melatonin pretreatment. We also verified that the decrease of GSH levels provoked by HMG in cortical supernatants was not due to a direct oxidative effect of this organic acid, because exposition of commercial GSH and purified membrane protein-bound thiol groups to HMG in the absence of cortical supernatants did not decrease the reduced sulfhydryl groups. Finally, the activities of the main antioxidant enzymes were not altered by HMG exposure. Our data indicate that oxidative stress elicited in vitro by HMG may possibly contribute at least in part to the pathophysiology of the brain injury in HMGLD.

Evidence that glutaric acid reduces glutamate uptake by cerebral cortex of infant rats.

The role of excitotoxicity in the cerebral damage of glutaryl-CoA dehydrogenase deficiency (GDD) is under intense debate. We therefore investigated the in vitro effect of glutaric (GA) and 3-hydroxyglutaric (3-OHGA) acids, which accumulate in GDD, on [(3)H]glutamate uptake by slices and synaptosomal preparations from cerebral cortex and striatum of rats aged 7, 15 and 30 days. Glutamate uptake was significantly decreased by high concentrations of GA in cortical slices of 7-day-old rats, but not in cerebral cortex from 15- and 30-day-old rats and in striatum from all studied ages. Furthermore, this effect was not due to cellular death and was prevented by N-acetylcysteine preadministration, suggesting the involvement of oxidative damage. In contrast, glutamate uptake by brain slices was not affected by 3-OHGA exposure. Immunoblot analysis revealed that GLAST transporters were more abundant in the cerebral cortex compared to the striatum of 7-day-old rats. Moreover, the simultaneous addition of GA and dihydrokainate (DHK), a specific inhibitor of GLT1, resulted in a significantly higher inhibition of [(3)H]glutamate uptake by cortical slices of 7-day-old rats than that induced by the sole presence of DHK. We also observed that both GA and 3-OHGA exposure did not alter the incorporation of glutamate into synaptosomal preparations from cerebral cortex and striatum of rats aged 7, 15 and 30 days. Finally, GA in vivo administration did not alter glutamate uptake into cortical slices from 7-day-old rats. Our findings may explain at least in part why cortical neurons are more vulnerable to damage at birth as evidenced by the frontotemporal cortical atrophy observed in newborns affected by GDD.

Age and brain structural related effects of glutaric and 3-hydroxyglutaric acids on glutamate binding to plasma membranes during rat brain development.

(1) In the present study we determined the effects of glutaric (GA, 0.01-1 mM) and 3-hydroxyglutaric (3-OHGA, 1.0-100 microM) acids, the major metabolites accumulating in glutaric acidemia type I (GA I), on Na(+)-independent and Na(+)-dependent [(3)H]glutamate binding to synaptic plasma membranes from cerebral cortex and striatum of rats aged 7, 15 and 60 days. (2) GA selectively inhibited Na(+)-independent [(3)H]glutamate binding (binding to receptors) in cerebral cortex and striatum of rats aged 7 and 15 days, but not aged 60 days. In contrast, GA did not alter Na(+)-dependent glutamate binding (binding to transporters) to synaptic membranes from brain structures of rats at all studied ages. Furthermore, experiments using the glutamatergic antagonist CNQX indicated that GA probably binds to non-NMDA receptors. In addition, GA markedly inhibited [(3)H]kainate binding to synaptic plasma membranes in cerebral cortex of 15-day-old rats, indicating that this effect was probably directed towards kainate receptors. On the other hand, experiments performed with 3-OHGA revealed that this organic acid did not change Na(+)-independent [(3)H]glutamate binding to synaptic membranes from cerebral cortex and striatum of rats from all ages, but inhibited Na(+)-dependent [(3)H]glutamate binding to membranes in striatum of 7-day-old rats, but not in striatum of 15- and 60-day-old rats and in cerebral cortex of rats from all studied ages. We also provided some evidence that 3-OHGA competes with the glutamate transporter inhibitor L-trans-pyrrolidine-2,4-dicarboxylate, suggesting a possible interaction of 3-OHGA with glutamate transporters on synaptic membranes. (3) These results indicate that glutamate binding to receptors and transporters can be inhibited by GA and 3-OHGA in cerebral cortex and striatum in a developmentally regulated manner. It is postulated that a disturbance of glutamatergic neurotransmission caused by the major metabolites accumulating in GA I at early development may possibly explain, at least in part, the window of vulnerability of striatum and cerebral cortex to injury in patients affected by this disorder.

Evidence that 3-hydroxyglutaric acid interacts with NMDA receptors in synaptic plasma membranes from cerebral cortex of young rats.

Neurological symptoms are common in patients with glutaric acidemia type I (GA-I). Although the pathophysiology of this disorder is not yet fully established, 3-hydroxyglutaric acid (3-HGA), which accumulates in affected patients, has recently been demonstrated to be excitotoxic to embryonic chick and neonatal rat neurons probably via NMDA glutamate receptors. In the present study, we investigated the in vitro effects of 3-HGA on the [(3)H]glutamate and [(3)H]MK-801 (dizocilpine) binding to rat synaptic plasma membranes from cerebral cortex of young rats in order to elucidate the interactions of 3-HGA with glutamate receptors and its possible contribution to the in vitro excitotoxic properties of 3-HGA. 3-HGA (10-100 microM) significantly decreased Na(+)-dependent (up to 62%) and Na(+)-independent (up to 30%) [(3)H]glutamate binding to synaptic membranes, reflecting a possible competition between glutamate and 3-HGA for the glutamate transporter and receptor sites, respectively. Since a decrease in Na(+)-independent glutamate binding might represent an interaction of 3-HGA with glutamate receptors, we next investigated whether 3-HGA interacts with NMDA receptors by adding NMDA alone or combined with 3-HGA and measuring Na(+)-independent [(3)H]glutamate binding to synaptic membranes (binding to receptors). We verified that 3-HGA and NMDA, at 10 and 100 microM concentrations, decreased glutamate binding by up to 20 and 45%, respectively, and that the simultaneous addition of both substances did not provoke an additive effect, implying that they bind to NMDA receptors at the same site. Furthermore, the binding of the NMDA-channel blocker [(3)H ]MK-801 was significantly increased (approximately 32-40%) by 10 and 100 microM 3-HGA, implying that 3-HGA was able to open the NMDA channel allowing MK-801 binding, which is a characteristic of NMDA agonists. On the other hand, glutamate had a much higher stimulatory effect on this binding (180% increase), reflecting its strong NMDA agonist property. Furthermore, the simultaneous addition of 3-HGA and glutamate provoked an additive stimulatory effect on [(3)H]MK-801 binding to the NMDA receptor. These data indicate that, relatively to glutamate, 3-HGA is a weak agonist of NMDA receptors. Finally, we demonstrated that 3-HGA provoked a significant increase of extracellular calcium uptake by cerebral cortex slices, strengthening therefore, the view that 3-HGA activates NMDA receptors. The present study therefore, demonstrates at the molecular level that 3-HGA modulates glutamatergic neurotransmission and may explain previous findings relating the neurotoxic actions of this organic acid with excitotoxicity.

Inhibition of creatine kinase activity from rat cerebral cortex by D-2-hydroxyglutaric acid in vitro.

D-2-Hydroxyglutaric acid (DGA) is the biochemical hallmark of patients affected by the neurometabolic disorder known as D-2-hydroxyglutaric aciduria (DHGA). Although this disease is predominantly characterized by severe neurological findings, the underlying mechanisms of brain injury are virtually unknown. In the present study, we investigated the effect of DGA on total, cytosolic, and mitochondrial creatine kinase (CK) activities from cerebral cortex of 30-day-old Wistar rats. Total CK activity (tCK) was measured in whole cell homogenates, whereas cytosolic and mitochondrial activities were measured in the cytosolic and mitochondrial preparations from cerebral cortex. We verified that CK activities were significantly inhibited by DGA (11-34% inhibition) at concentrations as low as 0.25 mM, being the mitochondrial fraction the most affected activity. Kinetic studies revealed that the inhibitory effect of DGA was non-competitive in relation to phosphocreatine. We also observed that this inhibition was fully prevented by pre-incubation of the homogenates with reduced glutathione, suggesting that the inhibitory effect of DGA on tCK activity is possibly mediated by oxidation of essential thiol groups of the enzyme. Considering the importance of CK activity for brain metabolism homeostasis, our results suggest that inhibition of this enzyme by increased levels of DGA may be related to the neurodegeneration of patients affected by DHGA.

Inhibition of brain energy metabolism by the alpha-keto acids accumulating in maple syrup urine disease.

Neurological dysfunction is a common finding in patients with maple syrup urine disease (MSUD). However, the mechanisms underlying the neuropathology of brain damage in this disorder are poorly known. In the present study, we investigated the effect of the in vitro effect of the branched chain alpha-keto acids (BCKA) accumulating in MSUD on some parameters of energy metabolism in cerebral cortex of rats. [14CO(2)] production from [14C] acetate, glucose uptake and lactate release from glucose were evaluated by incubating cortical prisms from 30-day-old rats in Krebs-Ringer bicarbonate buffer, pH 7.4, in the absence (controls) or presence of 1-5 mM of alpha-ketoisocaproic acid (KIC), alpha-keto-beta-methylvaleric acid (KMV) or alpha-ketoisovaleric acid (KIV). All keto acids significantly reduced 14CO(2) production by around 40%, in contrast to lactate release and glucose utilization, which were significantly increased by the metabolites by around 42% in cortical prisms. Furthermore, the activity of the respiratory chain complex I-III was significantly inhibited by 60%, whereas the other activities of the electron transport chain, namely complexes II, II-III, III and IV, as well as succinate dehydrogenase were not affected by the keto acids. The results indicate that the major metabolites accumulating in MSUD compromise brain energy metabolism by blocking the respiratory chain. We presume that these findings may be of relevance to the understanding of the pathophysiology of the neurological dysfunction of MSUD patients.

Inhibition of mitochondrial creatine kinase activity by D-2-hydroxyglutaric acid in cerebellum of young rats.

D-2-Hydroxyglutaric aciduria (DHGA) is a neurometabolic disorder biochemically characterized by tissue accumulation and excretion of high amounts of D-2-hydroxyglutaric acid (DGA). Although the affected patients have predominantly severe neurological findings, the underlying mechanisms of brain injury are virtually unknown. In previous studies we have demonstrated that DGA, at concentrations as low as 0.25 mM, significantly decreased creatine kinase activity and other parameters of energy metabolism in cerebral cortex of young rats. In the present study, we investigated the effect of DGA (0.25-5 mM) on total creatine kinase (tCK) activity, as well as on CK activity in cytosolic (Cy-CK) and mitochondrial (Mi-CK) preparations from cerebellum of 30-day-old Wistar rats in order to test whether the inhibitory effect of DGA on CK was tissue specific. We verified that tCK (22% inhibition) and Mi-CK (40% inhibition) activities were moderately inhibited by DGA at concentrations of 2.5 mM and higher, in contrast to Cy-CK, which was not affected by the acid. Kinetic studies revealed that the inhibitory effect of DGA was noncompetitive in relation to phosphocreatine. We also observed that this inhibition was fully prevented by preincubation of the homogenates with reduced glutathione, suggesting that the inhibition of CK activity by DGA is possibly mediated by modification of essential thiol groups of the enzyme. Our present results therefore demonstrate a relatively weak inhibitory effect of DGA on cerebellum Mi-CK activity, as compared to that provoked in cerebral cortex, and may possibly be related to the neuropathology of DHGA, characterized by cerebral cortex abnormalities.

Evidence that antioxidants prevent the inhibition of Na+,K(+)-ATPase activity induced by octanoic acid in rat cerebral cortex in vitro.

The objective of the present study was to investigate the in vitro effects of octanoic acid, which accumulates in medium-chain acyl-CoA dehydrogenase (MCAD) deficiency and in Reye syndrome, on key enzyme activities of energy metabolism in the cerebral cortex of young rats. The activities of the respiratory chain complexes I-IV, creatine kinase, and Na+,K(+)-ATPase were evaluated. Octanoic acid did not alter the electron transport chain and creatine kinase activities, but, in contrast, significantly inhibited Na+,K(+)-ATPase activity both in synaptic plasma membranes and in homogenates prepared from cerebral cortex. Furthermore, decanoic acid, which is also increased in MCAD deficiency, and oleic acid strongly reduced Na+,K(+)-ATPase activity, whereas palmitic acid had no effect. We also examined the effects of incubating glutathione and trolox (alpha-tocopherol) alone or with octanoic acid on Na+,K(+)-ATPase activity. Tested compounds did not affect Na+,K(+)-ATPase activity by itself, but prevented the inhibitory effect of octanoic acid. These results suggest that inhibition of Na+,K(+)-ATPase activity by octanoic acid is possibly mediated by oxidation of essential groups of the enzyme. Considering that Na+,K(+)-ATPase is critical for normal brain function, it is feasible that the significant inhibition of this enzyme activity by octanoate and also by decanoate may be related to the neurological dysfunction found in patients affected by MCAD deficiency and Reye syndrome.