Melatonin prevents oxidative stress and inhibits reactive gliosis induced by hyperhomocysteinemia in rats

Homocysteine (Hcy), an independent risk factor for atherosclerosis, undergoes auto-oxidation and generates reactive oxygen species, which are thought to be main cause of Hcy neurotoxicity. However, the mechanisms leading to neurodegenerative disorders are poorly understood because studies that have investigated the potential neurotoxicity of hyperhomocysteinemia in vivo are scarce. The purpose of this study was to test whether daily administration of methionine, which induces hyperhomocysteinemia, causes glial hyperactivity, and also to investigate the protective effects of melatonin on the brain tissue against oxidative stress of Hcy in rats. There was a significant development of oxidative stress as indicated by an increase in malondialdehyde + 4-hydroxyalkenals in hippocampus and cortex of hyperhomocysteine mic rats, whereas significant reduction was found in the activity of glutathione peroxidase (GSH-Px). Co-treatment with melatonin inhibited the elevation of lipid peroxidation and significantly increased GSH-Px activity in the brain regions studied. Western blot analysis revealed an increase in glial fibrillary acidic protein (GFAP) contents both in hippocampus and frontal cortex (p < 0.001) of hyperhomocysteinemic rats compared to the controls. Administration of melatonin significantly decreased GFAP contents in hippocampus and cortex (p < 0.05). S100B contents increased only in frontal cortex in hyperhomocysteinemic rats compared to the control (p < 0.01) and was inhibited by melatonin treatment (p < 0.01). The present findings show that Hcy can sensitize glial cells, a mechanism which might contribute to the pathogenesis of neurodegenerative disorders, and further suggest that melatonin can be involved in protecting against the toxicity of Hcy by inhibiting free radical generation and stabilizing glial cell activity.     

Melatonin reduces glial reactivity in the hippocampus, cortex, and cerebellum of streptozotocin-induced diabetic rats

Hyperglycemia plays a critical role in the development and progression of diabetic neuropathy. One of the mechanisms by which hyperglycemia causes neural degeneration is via the increased oxidative stress that accompanies diabetes. Metabolic and oxidative insults often cause rapid changes in glial cells. Key indicators of this response are increased synthesis of glial fibrillary acidic protein (GFAP) and S100B, both astrocytic markers. In the present study, we examined glial reactivity in hippocampus, cortex, and cerebellum of streptozotocin (STZ)-induced diabetic rats by determining the expression of GFAP and S-100B and we evaluated the effect of melatonin on the glial response. Western blot measurement of contents in brain regions after 6 weeks of STZ-induced diabetes indicated significant increases in these constituents compared with those in nondiabetic controls. Administration of melatonin prevented the upregulation of GFAP in all brain regions of diabetic rats. Using GFAP immunohistochemistry, we observed an increase in GFAP immunostaining in the hippocampus of STZ-diabetic rats relative to levels in the control brains. Treatment with melatonin resulted in an obvious reduction of GFAP-immunoreactive astrocytes in hippocampus. Like GFAP, S100B levels also were increased in all three brain areas of diabetic rats, an effect also reduced by melatonin treatment. Finally, the levels of lipid peroxidation products were elevated as a consequence of diabetes, with this change also being prevented by melatonin. These results suggest that diabetes causes increased glial reactivity possibly due to elevated oxidative stress, and administration of melatonin represents an achievable adjunct therapy for preventing gliosis.     

Effect of cerebrolysin on the cerebellum of diabetic rats: An imunohistochemical study

Background

Diabetes mellitus represents one of the disorders in the metabolism that affects all body systems including CNS. Cerebrolysin contains many neurotrophic factors, and many studies reported that it can be used treatment of many neurological disorders.

Effect of Acute Administration of l-Tyrosine on Oxidative Stress Parameters in Brain of Young Rats

Tyrosinemia type II, also known as Richner–Hanhart syndrome, is an autosomal recessive inborn error of metabolism caused by a deficiency of hepatic cytosolic 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 studies demonstrated that high concentrations of tyrosine provoke oxidative stress in vitro and in vivo in the cerebral cortex of rats, in the present study we investigate the oxidative stress parameters (enzymatic antioxidant defenses, thiobarbituric acid-reactive substances and protein carbonyl content) in cerebellum, hippocampus and striatum of 30-old-day rats after acute administration of l-tyrosine. Our results demonstrated that the acute administration of l-tyrosine increased the thiobarbituric acid reactive species levels in hippocampus and the carbonyl levels in cerebellum, hippocampus and striatum. In addition, acute administration of l-tyrosine significantly decreased superoxide dismutase activity in cerebellum, hippocampus and striatum, while catalase was increased in striatum. In conclusion, the oxidative stress may contribute, along with other mechanisms, to the neurological dysfunction characteristic of hypertyrosinemia and the administration of antioxidants may be considered as a potential adjuvant therapy for tyrosinemia, especially type II.     

Effects of warfarin and l-carnitine on hemostatic function and oxidative stress in streptozotocin-induced diabetic rats

Diabetes mellitus (DM) is a complex progressive disease characterized by hyperglycemia and a high risk of atherothrombotic disorders affecting the coronary, cerebral, and peripheral arterial trees. Oxidative stress is reported in diabetic patients. We investigated the hemostatic functions and oxidative stress in streptozotocin (STZ)-induced diabetic rats and the effects of warfarin and l-carnitine on those parameters. Forty male Sprague–Dawley rats were divided into four groups: control, DM, and DM received warfarin or l-carnitine. In all rats, blood glucose, insulin, hemoglobin A1c (HbA1c), fibrinogen, factor VII (FVII), plasminogen activator inhibitor-1 (PAI-1), fibrin degradation products (FDP), protein C, antithrombin III (ATIII), malondialdehydes (MDA), and antioxidants (superoxide dismutase, catalase, glutathione peroxidase, glutathione) were measured. Also, prothrombin time (PT), activated partial thromboplastin time (aPTT), coagulation time, and platelet aggregation were evaluated. In diabetic rats, plasma glucose, HbA1c, MDA, fibrinogen, FVII, FDP, PAI-1, and platelet aggregation increased while insulin, PT, aPTT, coagulation time, protein C, ATIII, and antioxidants decreased. Warfarin administration to diabetic rats decreased FVII and FDP and increased PT, aPTT, and coagulation time with no effect on MDA, antioxidants, PAI-1, protein C, ATIII, and platelet aggregation. On the other hand, l-carnitine decreased fibrinogen, FVII, FDP, PAI-1, MDA, and platelet aggregation and increased PT, aPTT, coagulation time, protein C, ATIII, and antioxidants in diabetic rats. Therefore, we concluded that hyperglycemia plays an important role in hypercoagulation state and oxidative stress in STZ-induced DM. While l-carnitine improves oxidative stress and decreases the hypercoagulation state in DM, warfarin normalizes the hypercoagulation state with no effect on oxidative stress.     

Mitochondrial Dysfunction in Brain Cortex Mitochondria of STZ-Diabetic Rats: Effect of l-Arginine

Mitochondrial dysfunction has been implicated in many diseases, including diabetes. It is well known that oxygen free radical species are produced endogenously by mitochondria, and also nitric oxide (NO) by nitric oxide synthases (NOS) associated to mitochondrial membranes, in consequence these organelles constitute main targets for oxidative damage. The aim of this study was to analyze mitochondrial physiology and NO production in brain cortex mitochondria of streptozotocin (STZ) diabetic rats in an early stage of diabetes and the potential effect of l-arginine administration. The diabetic condition was characterized by a clear hyperglycaemic state with loose of body weight after 4 days of STZ injection. This hyperglycaemic state was associated with mitochondrial dysfunction that was evident by an impairment of the respiratory activity, increased production of superoxide anion and a clear mitochondrial depolarization. In addition, the alteration in mitochondrial physiology was associated with a significant decrease in both NO production and nitric oxide synthase type I (NOS I) expression associated to the mitochondrial membranes. An increased level of thiobarbituric acid-reactive substances (TBARS) in brain cortex homogenates from STZ-diabetic rats indicated the presence of lipid peroxidation. l-arginine treatment to diabetic rats did not change blood glucose levels but significantly ameliorated the oxidative stress evidenced by lower TBARS and a lower level of superoxide anion. This effect was paralleled by improvement of mitochondrial respiratory function and a partial mitochondrial repolarization.In addition, the administration of l-arginine to diabetic rats prevented the decrease in NO production and NOSI expression. These results could indicate that exogenously administered l-arginine may have beneficial effects on mitochondrial function, oxidative stress and NO production in brain cortex mitochondria of STZ-diabetic rats.     

Memory and Learning Dysfunction Following Copper Toxicity: Biochemical and Immunohistochemical Basis

The prototype disease of Cu toxicity in human is Wilson disease, and cognitive impairment is the presenting symptom of it. There is no study correlating Cu-induced excitotoxicity, apoptosis, and astrocytic reaction with memory dysfunction. We report excitotoxicity, apoptosis, and astrocytic reaction of the hippocampus and frontal cortex with memory dysfunction in rat model of Cu toxicity. Thirty-six rats were divided into group I (control) and group II (100 mg/kgBwt/day CuSO₄ orally). Y-maze was performed for memory and learning at 0, 30, 60, and 90 days. Frontal and hippocampal free Cu concentration, oxidative stress markers [glutathione (GSH), total antioxidant toxicity (TAC), and malondialdehyde (MDA)], and glutamate were measured by atomic absorption spectroscopy, spectrophotometry, and ELISA, respectively. N-methyl-d-aspartate receptors (NMDARs) NR1, NR2A, and NR2B were done by real-time polymerase chain reaction. Immunohistochemistry for caspase-3 and glial fibrillary acidic protein (GFAP) were done and quantified using the ImageJ software. The glutamate level in hippocampus was increased, and NMDAR expression was decreased at 30, 60, and 90 days in group II compared to group I. In the frontal cortex, glutamate was increased at 90 days, but NMDARs were not significantly different in group II compared to group I. Caspase-3 and GFAP expressions were also higher in group II compared to group I, and these changes were more marked in hippocampus than frontal cortex. These changes correlated with respective free tissue Cu, oxidative stress, and Y-maze attention score. Cu toxicity induces apoptosis and astrocytosis of the hippocampus and frontal cortex through direct or glutamate and oxidative stress pathways, and results in impaired memory and learning.     

Transportable and Non-transportable Inhibitors of L-glutamate Uptake Produce Astrocytic Stellation and Increase EAAT2 Cell Surface Expression

Astrocytic excitatory amino acid transporters (EAATs) regulate excitatory transmission and limit excitotoxicity. Evidence for a functional interface between EAATs and glial fibrillary acidic protein (GFAP) relevant to astrocytic morphology led to investigations of actions of transportable (d-Aspartate (d-Asp) and (2S,3S,4R)-2-(carboxycyclopropyl)glycine (l-CCG-III)) and non-transportable (dl-threo-β-benzyloxyaspartate (dl-TBOA)) inhibitors of Glu uptake in murine astrocytes. d-Asp (1 mM), l-CCG-III (0.5 mM) and dl-TBOA (0.5 mM) produced time-dependent (24–72 h) reductions in ³[H]d-Asp uptake (approximately 30–70%) with little or no gliotoxicity. All drugs induced a profound change in phenotype from cobblestone to stellate morphology and image analysis revealed increases in the intensity of GFAP immunolabelling for l-CCG-III and dl-TBOA. Cytochemistry indicated localized changes in F-actin distribution. Cell surface expression of EAAT2, but not EAAT1, was elevated at 72 h. Blockade of Glu uptake by both types of EAAT inhibitor exerts longer-term effects on astrocytic morphology and a compensatory homeostatic rise in EAAT2 abundance.     

Effect of novel synthetic evodiamine analogue on sexual behavior in male rats

Currently phosphodiestrase5 (PDE5) inhibitors are the first-line treatment for erectile dysfunction. Drugs such as sildenafil and tadalafil are available as PDE5 inhibitors which are potent and reversible but lack selectivity with side effects such as headache, facial flushing, dyspepsia, and visual disturbances. We herein report for the first time novel condensed thienopyrimidines as evodiamine analogue and their effect on sexual behavior in male rats hitherto unreported. Novel synthetic evodiamine significantly showed improvement in male rat copulatory behavior. The test compound MKAC9 could be of promising importance in the treatment of sexual disorders like desire disorder or erectile dysfunction.

Activation of AMP-activated protein kinase is required for berberine-induced reduction of atherosclerosis in mice: the role of uncoupling protein 2

Aims

Berberine, a botanical alkaloid purified from Coptidis rhizoma, is reported to activate the AMP-activated protein kinase (AMPK). Whether AMPK is required for the protective effects of berberine in cardiovascular diseases remains unknown. This study was designed to determine whether AMPK is required for berberine-induced reduction of oxidative stress and atherosclerosis in vivo.

Methods

ApoE (ApoE^-/-) mice and ApoE^-/-/AMPK alpha 2^-/- mice that were fed Western diets were treated with berberine for 8 weeks. Atherosclerotic aortic lesions, expression of uncoupling protein 2 (UCP2), and markers of oxidative stress were evaluated in isolated aortas.

Results

In ApoE^-/- mice, chronic administration of berberine significantly reduced aortic lesions, markedly reduced oxidative stress and expression of adhesion molecules in aorta, and significantly increased UCP2 levels. In contrast, in ApoE^-/-/AMPK alpha 2^-/- mice, berberine had little effect on those endpoints. In cultured human umbilical vein endothelial cells (HUVECs), berberine significantly increased UCP2 mRNA and protein expression in an AMPK-dependent manner. Transfection of HUVECs with nuclear respiratory factor 1 (NRF1)-specific siRNA attenuated berberine-induced expression of UCP2, whereas transfection with control siRNA did not. Finally, berberine promoted mitochondrial biogenesis that contributed to up-regulation of UCP2 expression.

Conclusion

We conclude that berberine reduces oxidative stress and vascular inflammation, and suppresses atherogenesis via a mechanism that includes stimulation of AMPK-dependent UCP2 expression.     

A Novel and Efficient Gene Transfer Strategy Reduces Glial Reactivity and Improves Neuronal Survival and Axonal Growth In Vitro

Background

The lack of axonal regeneration in the central nervous system is attributed among other factors to the formation of a glial scar. This cellular structure is mainly composed of reactive astrocytes that overexpress two intermediate filament proteins, the glial fibrillary acidic protein (GFAP) and vimentin. Indeed, in vitro, astrocytes lacking GFAP or both GFAP and vimentin were shown to be the substrate for increased neuronal plasticity. Moreover, double knockout mice lacking both GFAP and vimentin presented lower levels of glial reactivity in vivo, significant axonal regrowth and improved functional recovery in comparison with wild-type mice after spinal cord hemisection. From these results, our objective was to develop a novel therapeutic strategy for axonal regeneration, based on the targeted suppression of astroglial reactivity and scarring by lentiviral-mediated RNA-interference (RNAi).

Methods and Findings

In this study, we constructed two lentiviral vectors, Lv-shGFAP and Lv-shVIM, which allow efficient and stable RNAi-mediated silencing of endogenous GFAP or vimentin in vitro. In cultured cortical and spinal reactive astrocytes, the use of these vectors resulted in a specific, stable and highly significant decrease in the corresponding protein levels. In a second model — scratched primary cultured astrocytes — Lv-shGFAP, alone or associated with Lv-shVIM, decreased astrocytic reactivity and glial scarring. Finally, in a heterotopic coculture model, cortical neurons displayed higher survival rates and increased neurite growth when cultured with astrocytes in which GFAP and vimentin had been invalidated by lentiviral-mediated RNAi.

Conclusions

Lentiviral-mediated knockdown of GFAP and vimentin in astrocytes show that GFAP is a key target for modulating reactive gliosis and monitoring neuron/glia interactions. Thus, manipulation of reactive astrocytes with the Lv-shGFAP vector constitutes a promising therapeutic strategy for increasing glial permissiveness and permitting axonal regeneration after central nervous system lesions.     

Diabetic nephropathy in type 2 diabetes: MPO T-764C genotype is associated with oxidative stress

Background

Oxidative stress is a single mechanism relating all major pathways responsible for diabetic damage and plays an important role in diabetes development, progression and related vascular complications. To investigate the impact of oxidative stress related gene polymorphisms on development of diabetic nephropathy (DN), we tested 7 polymorphic variants that could hypothetically affect the ability of the antioxidant defense system and thus accelerate oxidative stress.

Methodology

197 Slovenian (Caucasian) type 2 diabetic (T2D) patients, age 34?C83, classified into two groups according to the presence of DN, were tested for SOD2 Val16Ala (rs4880), p22 phox C242T (rs4673), CAT C-262T (rs1001179), MPO T-764C (rs2243828), GSTP1 Ile105Val (rs1695), GSTT1 and GSTM1 deletion polymorphisms using PCR, RFLP and qPCR. Oxidative stress was assessed through serum 8-hydroxy-2-deoxyguanosine (8-OHdG) level. Results were analyzed using ANOVA, Chi-square test and multivariate logistic regression.

Results and Conclusions

Despite the commonly recognized link between oxidative stress and diabetes and its complications we found no association between the selected polymorphisms and DN. However, we confirmed an association between oxidative stress level and MPO T-764C genotype, which was tested in relation to DN for the first time.     

Advances in establishment and analysis of three-dimensional tumor spheroid-based functional assays for target validation and drug evaluation

Background

Glutamate and ??-aminobutyric acid (GABA) transporters play important roles in balancing excitatory and inhibitory signals in the brain. Increasing evidence suggest that they may act concertedly to regulate extracellular levels of the neurotransmitters.

Results

Here we present evidence that glutamate uptake-induced release of GABA from astrocytes has a direct impact on the excitability of pyramidal neurons in the hippocampus. We demonstrate that GABA, synthesized from the polyamine putrescine, is released from astrocytes by the reverse action of glial GABA transporter (GAT) subtypes GAT-2 or GAT-3. GABA release can be prevented by blocking glutamate uptake with the non-transportable inhibitor DHK, confirming that it is the glutamate transporter activity that triggers the reversal of GABA transporters, conceivably by elevating the intracellular Na⁺ concentration in astrocytes. The released GABA significantly contributes to the tonic inhibition of neurons in a network activity-dependent manner. Blockade of the Glu/GABA exchange mechanism increases the duration of seizure-like events in the low-[Mg²⁺] in vitro model of epilepsy. Under in vivo conditions the increased GABA release modulates the power of gamma range oscillation in the CA1 region, suggesting that the Glu/GABA exchange mechanism is also functioning in the intact hippocampus under physiological conditions.

Conclusions

The results suggest the existence of a novel molecular mechanism by which astrocytes transform glutamatergic excitation into GABAergic inhibition providing an adjustable, in situ negative feedback on the excitability of neurons.     

Microglial p38α MAPK is critical for LPS-induced neuron degeneration, through a mechanism involving TNFα

Background

The identification of factors that compromise neurogenesis is aimed at improving stem cell-based approaches in the field of regenerative medicine. Interferon gamma (IFNγ) is a main pro-inflammatory cytokine and up-regulated during several neurological diseases. IFNγ is generally thought to beneficially enhance neurogenesis from fetal or adult neural stem/precursor cells (NSPCs).

Results

We now provide direct evidence to the contrary that IFNγ induces a dysfunctional stage in a substantial portion of NSPC-derived progeny in vitro characterized by simultaneous expression of glial fibrillary acid protein (GFAP) and neuronal markers, an abnormal gene expression and a functional phenotype neither typical for neurons nor for mature astrocytes. Dysfunctional development of NSPCs under the influence of IFNγ was finally demonstrated by applying the microelectrode array technology. IFNγ exposure of NSPCs during an initial 7-day proliferation period prevented the subsequent adequate differentiation and formation of functional neuronal networks.

Conclusions

Our results show that immunocytochemical analyses of NSPC-derived progeny are not necessarily indicating the correct cellular phenotype specifically under inflammatory conditions and that simultaneous expression of neuronal and glial markers rather point to cellular dysregulation. We hypothesize that inhibiting the impact of IFNγ on NSPCs during neurological diseases might contribute to effective neurogenesis and regeneration.     

Aluminum affects glial system and behavior of rats

Aluminum (Al) has been associated with neuronal dysfunction. These neuronal changes may involve glial alterations. We intend to evaluate the consequence of Al on the glial system and the behavior of rats exposed chronically to 0.3% of aluminum chloride in drinking water during 4 months in adulthood (A) or since intra-uterine age (IU); animals from this latter group were sacrificed at four months of age. Our data show an intense glial fibrillary acidic protein (GFAP)-immunoreactivity with a high density of astrocytes in both treated groups compared with controls. However, in IU rats, astrocytes display prominent glial cell bodies and processes. A and IU rat groups perform a significantly reduced locomotor activity. However, using the dark/light box test, the IU rats prefer to spend more time in the enlightened compartment compared to other groups. Behavioral and glial changes caused by Al exposure bring support for the role of Al in brain dysfunction involving glial alterations.     

l-Tyrosine Induces DNA Damage in Brain and Blood of Rats

Mutations in the tyrosine aminotransferase gene have been identified to cause tyrosinemia type II which is inherited in an autosomal recessive manner. Studies have demonstrated that an excessive production of ROS can lead to reactions with macromolecules, such as DNA, lipids, and proteins. Considering that the l-tyrosine may promote oxidative stress, the main objective of this study was to investigate the in vivo effects of l-tyrosine on DNA damage determined by the alkaline comet assay, in brain and blood of rats. In our acute protocol, Wistar rats (30 days old) were killed 1 h after a single intraperitoneal l-tyrosine injection (500 mg/kg) or saline. For chronic administration, the animals received two subcutaneous injections of l-tyrosine (500 mg/kg, 12-h intervals) or saline administered for 24 days starting at postnatal day (PD) 7 (last injection at PD 31), 12 h after the last injection, the animals were killed by decapitation. We observed that acute administration of l-tyrosine increased DNA damage frequency and damage index in cerebral cortex and blood when compared to control group. Moreover, we observed that chronic administration of l-tyrosine increased DNA damage frequency and damage index in hippocampus, striatum, cerebral cortex and blood when compared to control group. In conclusion, the present work demonstrated that DNA damage can be encountered in brain from animal models of hypertyrosinemia, DNA alterations may represent a further means to explain neurological dysfunction in this inherited metabolic disorder and to reinforce the role of oxidative stress in the pathophysiology of tyrosinemia type II.     

Differential cellular FGF-2 upregulation in the rat facial nucleus following axotomy, functional electrical stimulation and corticosterone: a possible therapeutic target to Bell's palsy

Background

The etiology of Bell's palsy can vary but anterograde axonal degeneration may delay spontaneous functional recovery leading the necessity of therapeutic interventions. Corticotherapy and/or complementary rehabilitation interventions have been employed. Thus the natural history of the disease reports to a neurotrophic resistance of adult facial motoneurons leading a favorable evolution however the related molecular mechanisms that might be therapeutically addressed in the resistant cases are not known. Fibroblast growth factor-2 (FGF-2) pathway signaling is a potential candidate for therapeutic development because its role on wound repair and autocrine/paracrine trophic mechanisms in the lesioned nervous system.

Methods

Adult rats received unilateral facial nerve crush, transection with amputation of nerve branches, or sham operation. Other group of unlesioned rats received a daily functional electrical stimulation in the levator labii superioris muscle (1 mA, 30 Hz, square wave) or systemic corticosterone (10 mgkg^-1). Animals were sacrificed seven days later.

Results

Crush and transection lesions promoted no changes in the number of neurons but increased the neurofilament in the neuronal neuropil of axotomized facial nuclei. Axotomy also elevated the number of GFAP astrocytes (143% after crush; 277% after transection) and nuclear FGF-2 (57% after transection) in astrocytes (confirmed by two-color immunoperoxidase) in the ipsilateral facial nucleus. Image analysis reveled that a seven days functional electrical stimulation or corticosterone led to elevations of FGF-2 in the cytoplasm of neurons and in the nucleus of reactive astrocytes, respectively, without astrocytic reaction.

Conclusion

FGF-2 may exert paracrine/autocrine trophic actions in the facial nucleus and may be relevant as a therapeutic target to Bell's palsy.     

Role of gold in a complex cascade reaction involving two electrocyclization steps

Quantum chemical computations (B3LYP/LACVP**) were applied to assess the impact of Au(I) complexation on activation barriers for sequential electrocyclization reactions (one a 1,2-dihydroazete ring-opening and another a pentadienyl cation ring-closure) proposed to occur during a complex reaction cascade that converts alkynes and imines to cyclopentenimines.

Effects of isoquinoline alkaloid berberine on lipid peroxidation,antioxidant defense system,and liver damage induced by lead acetate in rats

Objectives: Liver is considered a target organ affected by lead toxicity. Oxidative stress is among the mechanisms involved in liver damage. Here we investigated the effects of the natural alkaloid berberine on oxidative stress and hepatotoxicity induced by lead in rats.

Methods: Animals received an aqueous solution of lead acetate (500?mg Pb/l in the drinking water) and/or daily oral gavage of berberine (50?mg/kg) for 8 weeks. Rats were then weighed and used for the biochemical, molecular, and histological evaluations.

Results: Lead-induced oxidative stress, shown by increasing lipid peroxidation along with a concomitant decrease in hepatic levels of thiol groups, total antioxidant capacity, the activities of superoxide dismutase, catalase, glutathione peroxidase, and glutathione-S-transferase, and reduced versus oxidized glutathione ratio. Berberine corrected all the disturbances in oxidative stress markers induced by lead administration. Berberine also prevented the elevated levels of enzymes (alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase) and the decrease in body weight and albumin. The protective effects of berberine were comparable with silymarin. Furthermore, berberine attenuated liver damage, shown by decreased necrosis and inflammatory cell infiltration.

Discussion: Berberine represents a potential therapeutic option against lead-induced hepatotoxicity through inhibiting lipid peroxidation and enhancing antioxidant defenses.

Conclusion: Berberine exerted protective effects on lead-induced oxidative stress and hepatotoxicity in rats.     

Long-Term Consumption of High-Fat Diet in Rats: Effects on Microglial and Astrocytic Morphology and Neuronal Nitric Oxide Synthase Expression

Obesity in humans is associated with cognitive decline and elevated risk of neurodegenerative diseases of old age. Variations of high-fat diet are often used to model these effects in animal studies. However, we previously reported improvements in markers of memory and learning, as well as larger hippocampi and higher metabolite concentrations in Wistar rats fed high-fat, high-carbohydrate diet (HFCD, 60 % energy from fat, 28 % from carbohydrates) for 1 year; this diet leads to mild ketonemia (Setkowicz et al. in PLoS One 10:e0139987, 2015). In the present study, we follow up on this cohort to assess glial morphology and expression of markers related to gliosis. Twenty-five male Wistar rats were kept on HFCD and twenty-five on normal chow. At 12 months of age, the animals were sacrificed and processed for immunohistochemical staining for astrocytic (glial fibrillary acidic protein), microglial (Iba1), and neuronal (neuronal nitric oxide synthetase, nNOS) markers in the hippocampus. We have found changes in immunopositive area fraction and cellular complexity, as studied by a simplified Sholl procedure. To our knowledge, this study is the first to apply this methodology to the study of glial cells in HFCD animals. GFAP and Iba1 immunoreactive area fraction in the hippocampi of HFCD-fed rats were decreased, while the mean number of intersections (an indirect measure of cell complexity) was decreased in GFAP-positive astrocytes, but not in Iba1-expressing microglia. At the same time, nNOS expression was lowered after HFCD in both the cortex and the hippocampus.