Taurine Chloramine decreases cell viability and cytokine production in blood and spleen lymphocytes from septic rats.

Taurine (Tau) is an abundant amino acid in polymorphonuclear leukocytes that react with hypochlorous acid to form taurine chloramine (TauCl) under inflammatory conditions. We investigated potential interactions between lymphocytes and TauCl in rats submitted to cecal ligation. Animals were divided into sham or CLP groups (24 or 120 h) to isolate lymphocytes from blood and spleen. Lymphocytes were cultured at a concentration of 1×106 cells/mL and activated by concanavalin A. Tau and TauCl were added at 1, 10, and 100 µM. Cells were incubated with MTT to evaluate cell viability and cytokine concentration in the supernatant was determined. TauCl decreased lymphocyte viability and altered the secretion pattern of important inflammatory mediators in non-specific-phenotype manner. The effort to a is elucidate mechanisms of immune cell (dys)function in sepsis is important to better understand the complex regulation of immune system during sepsis development, and further studies are necessary to confirm TauCl as potential target in this context.

Ammonia exposition during gestation induces neonatal oxidative damage in the brain and long-term cognitive alteration in rats.

Ammonia is involved in the pathogenesis of neurological conditions associated with hyperammonemia, including hepatic encephalopathy. Few is known about the effects of gestational exposition to ammonia in the developing brain, and the possible long-term consequences of such exposure. We aimed to evaluate the effects of ammonia exposure during the gestation and the possible long-term cognitive alterations on pups. Eight female rats were divided into two groups: (1) control (saline solution); (2) ammonia (ammonium acetate, 2,5mmol/Kg). Each rat received a single subcutaneous injection during all gestational period. The brains from 1-day-old rats were obtained to the determination of thiobarbituric acid reactive species (TBARS), protein carbonyl and nitrite/nitrate levels. Some animals were followed 30 days after delivery and were subjected to the step-down inhibitory avoidance task. It was observed a significant increase in protein carbonyl, but not TBARS or nitrite/nitrate levels, in pups exposed to ammonia. Rats exposed to ammonia presented long-term cognitive impairment. Gestational exposition to ammonia induces protein oxidative damage in the neonatal rat brain, and long-term cognitive impairment.

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.

CD40-CD40 Ligand Pathway is a Major Component of Acute Neuroinflammation and Contributes to Long-term Cognitive Dysfunction after Sepsis.

Sepsis-associated encephalopathy (SAE) is associated with an increased rate of morbidity and mortality. It is not understood what the exact mechanism is for the brain dysfunction that occurs in septic patients, but brain inflammation and oxidative stress are a possible theory. Such events can occur through the alteration of molecules that perpetuate the inflammatory response. Thus, it is possible to postulate that CD40 may be involved in this process. The aim of this work is to evaluate the role of CD40-CD40L pathway activation in brain dysfunction associated with sepsis in an animal model. Microglia activation induces the upregulation of CD40-CD40L, both in vitro and in vivo. The inhibition of microglia activation decreases levels of CD40-CD40L in the brain and decreases brain inflammation, oxidative damage and blood brain barrier dysfunction. Despite this, anti-CD40 treatment does not improve mortality in this model. However, it is able to improve long-term cognitive impairment in sepsis survivors. In conclusion, there is a major involvement of the CD40-CD40L signaling pathway in long-term brain dysfunction in an animal model of sepsis.

Administration of branched-chain amino acids alters the balance between pro-inflammatory and anti-inflammatory cytokines.

Acute leucine intoxication and neurologic deterioration can develop rapidly at any age as a result of net protein degradation precipitated by infection or psychological stress in patients with maple syrup urine disease (MSUD). Here, we investigated the effects of acute and chronic Hyper-BCAA (H-BCAA) administration on pro- and anti-inflammatory cytokines in the brains of rats. For acute administration, Wistar rats (10 and 30 days) received three injections of BCAA pool (15.8 µL/g at 1-h intervals) or saline, subcutaneously. For chronic administration, Wistar rats (7 days) received of BCAA pool or saline twice a day for 21 days, subcutaneously. Our results showed that acute administration of H-BCAA increased IL-1ß (∼ 78%; p ≤ 0.009) and TNF-α (∼ 155%; p ≤ 0.026) levels in the cerebral cortex but not in the hippocampus of infant rats. Moreover, IL-6 levels were increased in the hippocampus (∼ 135%; p ≤ 0.009) and cerebral cortex (∼ 417%; p ≤ 0.008), whereas IL-10 levels were decreased only in the hippocampus (∼ 42%; p ≤ 0.009). However, repeated administration of H-BCAA decreased IL-1ß (∼ 59%; p ≤ 0.047), IL-6 (∼ 70%; p ≤ 0.009) and IFN-γ (∼ 70%; p ≤ 0.008) levels in the cerebral cortex, whereas the IL-6 (∼ 67%; p ≤ 0.009), IL-10 (∼ 58%; p ≤ 0.01) and IFN-γ (∼ 67%; p ≤ 0.009) levels were decreased in the hippocampus. These findings suggest that a better understanding of the inflammatory response in MSUD patients may be useful to develop therapeutic strategies to modulate the hyperinflammatory/hypoinflammatory axis.

Cerebral Oedema, Blood-Brain Barrier Breakdown and the Decrease in Na(+),K(+)-ATPase Activity in the Cerebral Cortex and Hippocampus are Prevented by Dexamethasone in an Animal Model of Maple Syrup Urine Disease.

Maple syrup urine disease (MSUD) is a rare metabolic disorder associated with acute and chronic brain dysfunction. This condition has been shown to lead to macroscopic cerebral alterations that are visible on imaging studies. Cerebral oedema is widely considered to be detrimental for MSUD patients; however, the mechanisms involved are still poorly understood. Therefore, we investigated whether acute administration of branched-chain amino acids (BCAA) causes cerebral oedema, modifies the Na(+),K(+)-ATPase activity, affects the permeability of the blood-brain barrier (BBB) and alters the levels of cytokines in the hippocampus and cerebral cortex of 10-day-old rats. Additionally, we investigated the influence of concomitant administration of dexamethasone on the alterations caused by BCAA. Our results showed that the animals submitted to the model of MSUD exhibited an increase in the brain water content, both in the cerebral cortex and in the hippocampus. By investigating the mechanism of cerebral oedema, we discovered an association between H-BCAA and the Na(+),K(+)-ATPase activity and the permeability of the BBB to small molecules. Moreover, the H-BCAA administration increases Il-1ß, IL-6 and TNF-α levels in the hippocampus and cerebral cortex, whereas IL-10 levels were decreased in the hippocampus. Interestingly, we showed that the administration of dexamethasone successfully reduced cerebral oedema, preventing the inhibition of Na(+),K(+)-ATPase activity, BBB breakdown and the increase in the cytokines levels. In conclusion, these findings suggest that dexamethasone can improve the acute cerebral oedema and brain injury associated with high levels of BCAA, either through a direct effect on brain capillary Na(+),K(+)-ATPase or through a generalized effect on the permeability of the BBB to all compounds.

Evaluation of the role of the cannabidiol system in an animal model of ischemia/reperfusion kidney injury.

OBJECTIVE: This work aimed to investigate the effects of the administration of cannabidiol in a kidney ischemia/reperfusion animal model. METHODS: Kidney injury was induced by 45 minutes of renal ischemia followed by reperfusion. Cannabidiol (5mg/kg) was administered immediately after reperfusion. RESULTS: Ischemia/reperfusion increased the IL-1 and TNF levels, and these levels were attenuated by cannabidiol treatment. Additionally, cannabidiol was able to decrease lipid and protein oxidative damage, but not the nitrite/nitrate levels. Kidney injury after ischemia/reperfusion seemed to be independent of the cannabidiol receptor 1 and cannabidiol receptor 2 (CB1 and CB2) expression levels, as there was no significant increase in these receptors after reperfusion. CONCLUSION: The cannabidiol treatment had a protective effect against inflammation and oxidative damage in the kidney ischemia/reperfusion model. These effects seemed to be independent of CB1/CB2 receptor activation.

Coadministration of branched-chain amino acids and lipopolysaccharide causes matrix metalloproteinase activation and blood-brain barrier breakdown.

Maple syrup urine disease (MSUD) is an inborn error of metabolism caused by a severe deficiency in the activity of the branched-chain α-keto acid dehydrogenase complex, leading to accumulation of the branched-chain amino acids (BCAA) leucine, isoleucine, and valine. Infections have a significant role in precipitating acute metabolic decompensation in patients with MSUD; however, the mechanisms underlying the neurotoxicity in this disorder are poorly understood. In this study, we subjected rats to the coadministration of lipopolysaccharide (LPS), which is a major component of gram-negative bacteria cell walls, and high concentrations of BCAA (H-BCAA) to determine their effects on the permeability of the blood-brain barrier (BBB) and on the levels of matrix metalloproteinases (MMP-2 and MMP-9). Our results demonstrated that the coadministration of H-BCAA and LPS causes breakdown of the BBB and increases the levels of MMP-2 and MMP-9 in the hippocampus of these rats. On the other hand, examination of the cerebral cortex of the 10- and 30-day-old rats revealed a significant difference in Evan's Blue content after coadministration of H-BCAA and LPS, as MMP-9 levels only increased in the cerebral cortex of the 10-day-old rats. In conclusion, these results suggest that the inflammatory process associated with high levels of BCAA causes BBB breakdown. Thus, we suggest that BBB breakdown is relevant to the perpetuation of brain inflammation and may be related to the brain dysfunction observed in MSUD patients.

Il1-ß involvement in cognitive impairment after sepsis.

Sepsis is defined as the host's reaction to infection and characterised by a systemic inflammatory response with important clinical implications. Central nervous system dysfunction secondary to sepsis is associated with local generation of pro- and anti-inflammatory cytokines, impaired cerebral microcirculation, an imbalance of neurotransmitters, apoptosis and cognitive impairment. It's known that the IL-1ß is one of the first cytokines to be altered. Thus, the objective of this study was to evaluate the role of IL-1ß in cognitive parameters in brain tissue through the use of an IL-1ß (IL-1ra) receptor antagonist up to 10 days and to assess blood-brain barrier permeability, cytokine levels, oxidative parameters and energetic metabolism up to 24 h, after sepsis induction. To this aim, we used sham-operated Wistar rats or submitted to the cecal ligation and perforation (CLP) procedure. Immediately after, the animals received one dose of 10 µg of IL-1ra. After 24 h, the rats were killed and were evaluated for biochemical parameters in the pre-frontal cortex, hippocampus and striatum. After 10 days, the animals were submitted to the habituation to the open field and step-down inhibitory avoidance task. We observed that the use of IL-1ra reverted the increase of blood-brain barrier permeability in the pre-frontal cortex, hippocampus and striatum; the increase of IL-1ß, IL1-6 and TNF-α levels in the pre-frontal cortex and striatum; the decrease of complex I activity in the pre-frontal, hippocampus and striatum; the increase of oxidative parameters in pre-frontal cortex, hippocampus and striatum; and cognitive impairment. In conclusion, the results observed in this study reinforce the role of acute brain inflammatory response, in particular, the IL1ß response, in the cognitive impairment associated with sepsis.

Correlation of acute phase inflammatory and oxidative markers with long-term cognitive impairment in sepsis survivors rats.

Oxidative damage and inflammation occur early in the brain after sepsis and are resolved when long-term cognitive impairment occurs. There is no information of a direct relation between acute levels of brain inflammation and oxidative damage and long-term cognitive deficits. We hypothesized that higher levels of early oxidative damage and inflammation are followed by long-term cognitive deficits, and this is related to a decrease in the levels of brain-derived neurotropic factor (BDNF). Wistar rats were subjected to sham operation or cecal ligation and perforation and the cerebrospinal fluid (CSF) was obtained 6 and 24 h after the determination of thiobarbituric acid-reactive species, interleukin 1 (IL-1), IL-10, and tumor necrosis factor α (TNF-α). Animals were followed until 30 days after surgery and were subjected to the step-down inhibitory avoidance (IA) task, and the hippocampus levels of BDNF were determined. At 6 h, higher CSF levels of thiobarbituric acid-reactive species and TNF-α were observed in septic animals that had a better performance in the IA task and presented higher BDNF levels in the hippocampus. At 24 h, higher CSF levels of IL-1ß and TNF-α were observed in septic animals that had a worse performance in the IA task, and this was associated with lower BDNF levels. The persistence of brain inflammation during the acute phase of sepsis is associated with long-term hippocampus levels of BDNF and memory impairment in sepsis survivors.

Avaliação do papel do sistema canabidiol em um modelo de lesão renal por isquemia/reperfusão em animais / Evaluation of the role of the cannabidiol system in an animal model of ischemia/reperfusion kidney injury

RESUMO Objetivo: Investigar os efeitos da administração de canabidiol em um modelo de isquemia/reperfusão renal em animais. Métodos: Foi induzida uma lesão renal, por meio de 45 minutos de isquemia renal seguida por reperfusão. Administrou-se canabidiol (5mg/kg) imediatamente após a reperfusão. Resultados: A isquemia/reperfusão aumentou os níveis de interleucina 1 e fator de necrose tumoral, o que foi atenuado pelo tratamento com canabidiol. Além disso, o canabidiol foi capaz de diminuir o dano oxidativo de lipídios e proteínas, mas não os níveis de nitrito/nitrato. A lesão renal após isquemia/reperfusão pareceu ser independente da expressão dos receptores canabidiol-1 e canabidiol-2, já que não houve aumento significante desses receptores após a reperfusão. Conclusão: O tratamento com canabidiol teve um efeito protetor contra a inflamação e o dano oxidativo em um modelo de isquemia/reperfusão renal. Esses efeitos parecem não ocorrer via ativação dos receptores canabidiol-1/canabidiol-2.

ABSTRACT Objective: This work aimed to investigate the effects of the administration of cannabidiol in a kidney ischemia/reperfusion animal model. Methods: Kidney injury was induced by 45 minutes of renal ischemia followed by reperfusion. Cannabidiol (5mg/kg) was administered immediately after reperfusion. Results: Ischemia/reperfusion increased the IL-1 and TNF levels, and these levels were attenuated by cannabidiol treatment. Additionally, cannabidiol was able to decrease lipid and protein oxidative damage, but not the nitrite/nitrate levels. Kidney injury after ischemia/reperfusion seemed to be independent of the cannabidiol receptor 1 and cannabidiol receptor 2 (CB1 and CB2) expression levels, as there was no significant increase in these receptors after reperfusion. Conclusion: The cannabidiol treatment had a protective effect against inflammation and oxidative damage in the kidney ischemia/reperfusion model. These effects seemed to be independent of CB1/CB2 receptor activation.