MEDUSA: A Pipeline for Sensitive Taxonomic Classification and Flexible Functional Annotation of Metagenomic Shotgun Sequences.

Metagenomic studies unravel details about the taxonomic composition and the functions performed by microbial communities. As a complete metagenomic analysis requires different tools for different purposes, the selection and setup of these tools remain challenging. Furthermore, the chosen toolset will affect the accuracy, the formatting, and the functional identifiers reported in the results, impacting the results interpretation and the biological answer obtained. Thus, we surveyed state-of-the-art tools available in the literature, created simulated datasets, and performed benchmarks to design a sensitive and flexible metagenomic analysis pipeline. Here we present MEDUSA, an efficient pipeline to conduct comprehensive metagenomic analyses. It performs preprocessing, assembly, alignment, taxonomic classification, and functional annotation on shotgun data, supporting user-built dictionaries to transfer annotations to any functional identifier. MEDUSA includes several tools, as fastp, Bowtie2, DIAMOND, Kaiju, MEGAHIT, and a novel tool implemented in Python to transfer annotations to BLAST/DIAMOND alignment results. These tools are installed via Conda, and the workflow is managed by Snakemake, easing the setup and execution. Compared with MEGAN 6 Community Edition, MEDUSA correctly identifies more species, especially the less abundant, and is more suited for functional analysis using Gene Ontology identifiers.

Influence of spray drying conditions on the properties of avocado powder drink.

Consumers are increasingly looking for foods with attractive taste, easy preparation, quick consumption and nutritious. Avocado is a nutritional fruit that presents a preservation challenge to food industry. The development of powder blends using spray drying of avocado as a powdered drink is an attractive option that generates products with high nutritional value and stable. Two avocado formulations were spray dried using a design of experiments to assess the influence of drying conditions on powder blends. Results showed higher drying temperatures in combination with smaller droplets resulted in higher yields, lower residual moisture, lower water activity, and in smaller, less dense particles with color green. The inclusion of maltodextrin proved to be vital in preserving high contents of protein, ascorbic acid, and phenolic compounds at any drying conditions possibly due to hydrogen bonding stabilization of those compounds. Using a scalable and efficient drying process, avocado high nutritional value was maintained.

A longitudinal study of neurotrophic, oxidative, and inflammatory markers in first-onset depression in midlife women.

Prospective studies have shown during the years preceding and following menopause, also known as "menopause transition", that midlife women are at higher risk for developing first-onset major depressive disorder (MDD). The biological factors associated with risk and resilience in this population are, however, largely unknown. Considering the growing body of evidence suggesting that inflammation, oxidative stress, and brain-derived neurotrophic factor (BDNF) are associated with the pathophysiology of MDD, we investigated serum levels of protein carbonyl, lipid peroxidation (thiobarbituric acid reactive substances-TBARS), thiol group content, BDNF, 3-nitrotyrosine, and heat shock protein 70 (HSP70) in a longitudinal cohort of first-onset MDD. One hundred and forty-eight women from the Harvard Study of Moods and Cycles, a prospective study of midlife women monitored throughout the transition to menopause, were studied. Within- and between-groups analyses of these peripheral markers were conducted in 37 women who developed and 111 women that did not develop MDD during the 3-year follow-up period. In women who developed MDD, HSP70 and 3-nitrotyrosine were elevated at baseline, whereas TBARS were elevated 6 months prior to development of MDD, as compared to those who did not develop MDD. Within-group analyses showed that HSP70, 3-nitrotyrosine, and BDNF decreased over time, whereas protein carbonyl was elevated only at 12 months prior to development of MDD. In women who did not develop MDD, HSP70 and thiol decreased over time. The development of MDD in midlife women may be associated with a systemic cascade of pro-oxidative and pro-inflammatory events including increased HSP70, 3-nitrotyrosine, protein carbonyl, and lipid peroxidation and decreased BDNF.

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.

Cerebral Ketone Body Oxidation Is Facilitated by a High Fat Diet Enriched with Advanced Glycation End Products in Normal and Diabetic Rats.

Diabetes mellitus (DM) causes important modifications in the availability and use of different energy substrates in various organs and tissues. Similarly, dietary manipulations such as high fat diets also affect systemic energy metabolism. However, how the brain adapts to these situations remains unclear. To investigate these issues, control and alloxan-induced type I diabetic rats were fed either a standard or a high fat diet enriched with advanced glycation end products (AGEs) (HAGE diet). The HAGE diet increased their levels of blood ketone bodies, and this effect was exacerbated by DM induction. To determine the effects of diet and/or DM induction on key cerebral bioenergetic parameters, both ketone bodies (ß-hydroxybutyric acid) and lactate oxidation were measured. In parallel, the expression of Monocarboxylate Transporter 1 (MCT1) and 2 (MCT2) isoforms in hippocampal and cortical slices from rats submitted to these diets was assessed. Ketone body oxidation increased while lactate oxidation decreased in hippocampal and cortical slices in both control and diabetic rats fed a HAGE diet. In parallel, the expression of both MCT1 and MCT2 increased only in the cerebral cortex in diabetic rats fed a HAGE diet. These results suggest a shift in the preferential cerebral energy substrate utilization in favor of ketone bodies in animals fed a HAGE diet, an effect that, in DM animals, is accompanied by the enhanced expression of the related transporters.

Dietary n-3 polyunsaturated fatty acids revert renal responses induced by a combination of 2 protocols that increase the amounts of advanced glycation end product in rats.

Renal dysfunction is a severe complication that is caused by diabetes mellitus. Many factors associate the progression of this complication with high levels of proinflammatory and pro-oxidant substances, such as advanced glycation end products (AGEs), which form a heterogeneous group of compounds that can accumulate in tissues such as retinas, joints, and kidneys. The hypothesis of this study is that n-3 polyunsaturated fatty acids (n-3 PUFAs) have a nephroprotective effect on rats after exposing them to a combination of 2 protocols that increase the AGE amounts: a high-fat diet enriched with AGEs and a diabetes rat model. Adult Wistar rats were divided into 6 groups that received the following diets for 4 weeks: (1) control group; 2) HAGE: high AGE fat-containing diet group; (3) HAGE + n-3: high AGE fat-containing diet plus n-3 PUFAs group; (4) diabetic group; (5) Db + HAGE: high AGE fat-containing diet diabetic group; and (6) Db + HAGE + n-3: high AGE fat-containing diet plus n-3 PUFAs diabetic group. Diabetes mellitus was induced by an intraperitoneal injection of alloxan (150 mg kg(-1)). In diabetic and nondiabetic rats, the high HAGE fat-containing diet increased the serum creatinine, tumor necrosis factor-α, thiobarbituric acid reactive substances, and reactive oxygen species levels, as well as the superoxide dismutase/catalase + glutathione peroxidase ratio and the superoxide dismutase 2 and receptor for advanced glycation end products immunocontent of the kidneys. n-3 Polyunsaturated fatty acids attenuated these alterations and influenced the receptor for advanced glycation end products/oxidative stress/tumor necrosis factor-α axis. In summary, this study showed that the extrinsic AGE pathway (HAGE diet) had a greater effect on renal metabolism than the intrinsic AGE pathway (diabetes induction) and that n-3 PUFAs appear to prevent renal dysfunction via antioxidant and anti-inflammatory pathways.

Acute Administration of Branched-Chain Amino Acids Increases the Pro-BDNF/Total-BDNF Ratio in the Rat Brain.

Maple syrup urine disease (MSUD) is caused by an inborn error in metabolism resulting from a deficiency in the branched-chain α-keto acid dehydrogenase complex activity. This blockage leads to accumulation of the branched-chain amino acids (BCAA) leucine, isoleucine and valine, as well as their corresponding α-keto acids and α-hydroxy acids. High levels of BCAAs are associated with neurological dysfunction and the role of pro- and mature brain-derived neurotrophic factor (BDNF) in the neurological dysfunction of MSUD is still unclear. Thus, in the present study we investigated the effect of an acute BCAA pool administration on BDNF levels and on the pro-BDNF cleavage-related proteins S100A10 and tissue plasminogen activator (tPA) in rat brains. Our results demonstrated that acute Hyper-BCAA (H-BCAA) exposure during the early postnatal period increases pro-BDNF and total-BDNF levels in the hippocampus and striatum. Moreover, tPA levels were significantly decreased, without modifications in the tPA transcript levels in the hippocampus and striatum. On the other hand, the S100A10 mRNA and S100A10 protein levels were not changed in the hippocampus and striatum. In the 30-day-old rats, we observed increased pro-BDNF, total-BDNF and tPA levels only in the striatum, whereas the tPA and S100A10 mRNA expression and the immunocontent of S100A10 were not altered. In conclusion, we demonstrated that acute H-BCAA administration increases the pro-BDNF/total-BDNF ratio and decreases the tPA levels in animals, suggesting that the BCAA effect may depend, at least in part, on changes in BDNF post-translational processing.

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.

Antinociceptive Activity and Redox Profile of the Monoterpenes (+)-Camphene, p-Cymene, and Geranyl Acetate in Experimental Models.

Objective. To evaluate antinocicpetive and redox properties of the monoterpenes (+)-camphene, p-cymene, and geranyl acetate in in vivo and in vitro experimental models. Methods. Evaluation of the in vitro antioxidant activity of (+)-camphene, p-cymene, and geranyl acetate using different free radical-generating systems and evaluation of antinociceptive actions by acetic acid-induced writhing and formalin-induced nociception tests in mice. Results. p-Cymene has the strongest antinociceptive effect, but (+)-camphene and geranyl acetate also present significant activity at high doses (200 mg/kg). (+)-Camphene had the strongest antioxidant effect in vitro at TBARS and TRAP/TAR assays and also had the highest scavenging activities against different free radicals, such as hydroxyl and superoxide radicals. Sodium nitroprussiate-derived NO production was enhanced by (+)-camphene. Geranyl acetate and p-cymene also presented some antioxidant effects, but with a varying profile according the free radical-generating system studied. Conclusion. (+)-Camphene, p-cymene, and geranyl acetate may present pharmacological properties related to inflammation and pain-related processes, being potentially useful for development of new therapeutic strategies, with limited possibilities for p-cymene and geranyl acetate.

Chronic administration of branched-chain amino acids impairs spatial memory and increases brain-derived neurotrophic factor in a rat model.

Maple syrup urine disease (MSUD) is a neurometabolic disorder that leads to the accumulation of branched-chain amino acids (BCAAs) and their α-keto branched-chain by-products. Because the neurotoxic mechanisms of MSUD are poorly understood, this study aimed to evaluate the effects of chronic administration of a BCAA pool (leucine, isoleucine and valine). This study examined the effects of BCAA administration on spatial memory and the levels of brain-derived neurotrophic factor (BNDF). We examined both pro-BDNF and bdnf mRNA expression levels after administration of BCAAs. Furthermore, this study examined whether antioxidant treatment prevented the alterations induced by BCAA administration. Our results demonstrated an increase in BDNF in the hippocampus and cerebral cortex, accompanied by memory impairment in spatial memory tasks. Additionally, chronic administration of BCAAs did not induce a detectable change in pro-BDNF levels. Treatment with N-acetylcysteine and deferoxamine prevented both the memory deficit and the increase in the BDNF levels induced by BCAA administration. In conclusion, these results suggest that when the brain is chronically exposed to high concentrations of BCAA (at millimolar concentrations) an increase in BDNF levels occurs. This increase in BDNF may be related to the impairment of spatial memory. In addition, we demonstrated that antioxidant treatment prevented the negative consequences related to BCAA administration, suggesting that oxidative stress might be involved in the pathophysiological mechanism(s) underlying the brain damage observed in MSUD.

Exploring the mechanisms of alcohol-related damage in oral mucosa - is oxidative stress associated with the increase in cell proliferation in rat tongue epithelium?

CONTEXT: Alcohol consumption has been related to a cell proliferation increase in oral epithelium but its mechanism remains unclear. OBJECTIVE: The aim of this study was to investigate whether oxidative stress parameters are implicated in the induction of cell proliferation in rat tongue epithelium after different times of chronic alcohol consumption. MATERIALS AND METHODS: Cell proliferation was assessed in tongue epithelium using AgNOR (argyrophilic proteins related to active nucleolar organizer regions) quantification. Oxidative stress parameters [lipid peroxidation, protein carbonyls, superoxide dismutase activity and catalase (CAT) activity and immunocontent] and Nrf2 immunocontent were quantified in tongue homogenates. RESULTS AND DISCUSSION: Mean AgNOR numbers (mAgNOR) per nucleus was 2.22 ± 0.30 in ventral tongue epithelium after 120 days of alcohol consumption (vs. 1.87 ± 0.18 for control animals and 1.91 ± 0.23 for animals treated with alcohol for 60 days) indicating cell proliferation increase (p < 0.05, ANOVA followed by Tukey post hoc). Interestingly, 60 days of alcohol consumption induced changes in oxidative stress parameters, but no alteration in cell proliferation. Vitamin E co-treatment was conduced in order to evaluate its possible protective effects. The 120 day Tween + vitamin E + alcohol treatment induced an increase in mAgNORs when compared to the Tween + vitamin E treated group (respectively 2.10 ± 0.30 vs. 1.77 ± 0.11, p < 0.05, ANOVA followed by Tukey post hoc), showing that vitamin E co-treatment had no protective effects. In addition, an inverse association was observed between CAT activity and AgNORs quantity (R = -0.32; p < 0.05, Person's correlation) as well as the possible involvement of Nrf2 in alcohol-related damage. CONCLUSIONS: Our findings suggest that the increase in cell proliferation associated with alcohol-related damage has no direct relation with an imbalance in oxidative parameters. In contrast, our results indicate that hydrogen peroxide may be implicated in cellular signaling during proliferation in the oral mucosa.

Similarities in serum oxidative stress markers and inflammatory cytokines in patients with overt schizophrenia at early and late stages of chronicity.

Schizophrenia (SZ) is a debilitating neurodevelopmental disorder that strikes at a critical period of a young person's life. Its pathophysiology could be the result of deregulation of synaptic plasticity, with downstream alterations of inflammatory immune processes regulate by cytokines, impaired antioxidant defense and increased lipid peroxidation. The aim of this study was to examine serum oxidative stress markers and inflammatory cytokines in early and late phases of chronic SZ. Twenty-two patients at early stage (within first 10 years of a psychotic episode), 39 at late stage (minimum 10 years after diagnosis of SZ) and their respective matched controls were included. Each subject had 5 ml blood samples collected by venipuncture to examined thiobarbituric acid-reactive substances (TBARS), total reactive antioxidant potential (TRAP), protein carbonyl content (PCC), Interleukins 6 and 10 (IL-6, IL-10) and tumor necrosis factor alpha (TNF-alpha). TBARS, IL-6 and PCC levels were significantly higher in patients with SZ at early and late stages than in controls. There were no differences for TRAP and TNF-alpha levels in patients with SZ at early and late stages than in controls. IL-10 levels were decreased in patients at late stage and a decrease trend in early stage was found. Results provided evidence consistent with comparable biological markers across chronic SZ. The concept of biochemical staging proposed by others for bipolar disorder is not seen in this cohort of patients with SZ, at least for cytokines and oxidative stress markers. Our findings reinforce the need of assessment of individuals in ultra high risk to develop psychosis and first-episode population.

Ω3-Polyunsaturated fatty acids prevent lipoperoxidation, modulate antioxidant enzymes, and reduce lipid content but do not alter glycogen metabolism in the livers of diabetic rats fed on a high fat thermolyzed diet.

Ω3-Polyunsaturated fatty acids (Ω3-PUFAs) are known to act as hypolipidaemics, but the literature is unclear about the effects that Ω3-PUFAs have on oxidative stress in obese and diabetic patients. In this study, our aim was to investigate the effects of Ω3-PUFAs on oxidative stress, including antioxidant enzyme activity and hepatic lipid and glycogen metabolism in the livers of diabetic and non-diabetic rats fed on a high fat thermolyzed diet. Rats were divided into six groups: (1) the control group (C), (2) the control diabetic group (D), (3) the high fat thermolyzed diet group (HFTD), which were fed a diet that was enriched in fat that was heated for 60 min at 180°C, (4) the high fat thermolyzed diet diabetic group (D + HFTD), (5) the high fat thermolyzed diet + Ω3 polyunsaturated fatty acid group (HFTD + Ω3), and (6) the high fat thermolyzed diet + Ω3 polyunsaturated fatty acid diabetic group (D + HFTD + Ω3). The most important finding of this study was that Ω3-PUFAs are able to reduce triglycerides, non-esterified fatty acid, lipoperoxidation levels, advanced glycation end products, SOD/CAT enzymatic ratio, and CAT immunocontent and increase SOD2 levels in the livers of diabetic rats fed with a HFTD. However, Ω3-PUFAs did not alter the observed levels of protein damage, blood glucose, or glycogen metabolism in the liver. These findings suggest that Ω3-PUFAs may represent an important auxiliary adjuvant in combating some diseases like diabetes mellitus, insulin resistance, and non-alcoholic fatty liver disease.

NFκB inhibitors induce cell death in glioblastomas.

Identification of novel target pathways in glioblastoma (GBM) remains critical due to poor prognosis, inefficient therapies and recurrence associated with these tumors. In this work, we evaluated the role of nuclear-factor-kappa-B (NFκB) in the growth of GBM cells, and the potential of NFκB inhibitors as antiglioma agents. NFκB pathway was found overstimulated in GBM cell lines and in tumor specimens compared to normal astrocytes and healthy brain tissues, respectively. Treatment of a panel of established GBM cell lines (U138MG, U87, U373 and C6) with pharmacological NFκB inhibitors (BAY117082, parthenolide, MG132, curcumin and arsenic trioxide) and NFκB-p65 siRNA markedly decreased the viability of GBMs as compared to inhibitors of other signaling pathways such as MAPKs (ERK, JNK and p38), PKC, EGFR and PI3K/Akt. In addition, NFκB inhibitors presented a low toxicity to normal astrocytes, indicating selectivity to cancerous cells. In GBMs, mitochondrial dysfunction (membrane depolarization, bcl-xL downregulation and cytochrome c release) and arrest in the G2/M phase were observed at the early steps of NFκB inhibitors treatment. These events preceded sub-G1 detection, apoptotic body formation and caspase-3 activation. Also, NFκB was found overstimulated in cisplatin-resistant C6 cells, and treatment of GBMs with NFκB inhibitors overcame cisplatin resistance besides potentiating the effects of the chemotherapeutics, cisplatin and doxorubicin. These findings support NFκB as a potential target to cell death induction in GBMs, and that the NFκB inhibitors may be considered for in vivo testing on animal models and possibly on GBM therapy.

Vitamin A supplementation induces oxidative stress and decreases the immunocontent of catalase and superoxide dismutase in rat lungs.

Lungs require an adequate supply of vitamin A for normal embryonic development, postnatal maturation, and maintenance and repair during adult life. However, recent intervention studies revealed that supplementation with retinoids resulted in a higher incidence of lung cancer, although the mechanisms underlying this effect are still unknown. Here, the authors studied the effect of vitamin A supplementation on oxidative stress parameters in lungs of Wistar rats. Vitamin A supplementation either at therapeutic (1000 and 2500 IU/kg) or excessive (4500 and 9000 IU/kg) doses for 28 days induced lipid peroxidation, protein carbonylation, and oxidation of protein thiol groups, as well as change in catalase (EC 1.11.1.6; CAT) and superoxide dismutase (EC 1.15.1.1, SOD) activities and immunocontents. These results altogether suggest that vitamin A supplementation causes significant changes in redox balance the free radical status in lungs, which are frequently associated to severe lung dysfunction.

Antioxidant and antigenotoxic activities of purple grape juice--organic and conventional--in adult rats.

Oxidative damage to biomolecules occurs by the accumulation of molecular damage due to free radicals and/or a diminution of antioxidant protection. The aim of this study was to evaluate the protection of organic and conventional purple grape juices in brain, liver, and plasma from adult Wistar rats (7 months old) against the oxidative damage provoked by carbon tetrachloride (CCl(4)). Adult rats were divided into three groups (control, conventional purple grape juice, and organic purple grape juice). Half of the rats received CCl(4), and the other half received the vehicle (vegetable oil). The chemical analytical determination showed that the highest levels of total phenolic, resveratrol, and catechins were seen in organic purple grape juices. Considering the treatment groups, it was observed that in all tissues (brain structures and liver) and plasma, CCl(4) treatment increased the lipid peroxidation (LP) levels. Both grape juices were capable to reduce LP levels in cerebral cortex and hippocampus; however, in the striatum and substantia nigra only the organic grape juice reduced LP level. CCl(4) caused an increase in catalase activity in cerebral cortex, hippocampus, and substantia nigra and in superoxide dismutase activity in substantia nigra. This increase was reduced by both juices in substantia nigra and hippocampus structures (P < .05). In the alkaline version of the comet assay performed on whole blood, it was observed that CCl(4) was capable of inducing mainly DNA damage class 4 and 3 frequencies, which was significantly reduced in groups that received both purple grape juices. This implies that both grape juices have an important antigenotoxic activity.

The NF kappa B-mediated control of RS and JNK signaling in vitamin A-treated cells: duration of JNK-AP-1 pathway activation may determine cell death or proliferation.

Nuclear factor kappa B (NFkappaB) has emerged as a crucial regulator of cell survival, playing important functions in cellular resistance to oxidants and chemotherapeutic agents. Recent studies showed that NFkappaB mediates cell survival through suppression of the accumulation of reactive species (RS) and a control of sustained activation of the Jun-N-terminal kinase (JNK) cascade. This work was undertaken in order to evaluate the role of NFkappaB in modulating the pro-oxidant effects of supplementation with vitamin A (retinol, ROH) in Sertoli cells, a major ROH physiological target. In this work, we reported that ROH treatment increases mitochondrial RS formation leading to a redox-dependent activation of NFkappaB. NFkappaB activation played a pivotal role in counteract RS accumulation in ROH-treated cells, since NFkappaB inhibition with DNA decoy oligonucleotides or pharmacological inhibitors (BAY-117082) potentiated ROH-induced RS accumulation and oxidative damage. In the presence of NFkappaB inhibition, ROH-induced oxidative stress promoted a prolonged activation of the JNK-activator protein 1 (AP-1) pathway and induced significant decreases in cell viability. Inhibition of JNK-AP-1 with decoy oligonucleotides to AP-1 or JNK inhibitor SP600125 prevented the decreases in cell viability. Antioxidants blocked the persistent JNK-AP-1 activation, cell oxidative damage, and the decreases in cell viability induced by NFkappaB inhibition. Finally, our data point superoxide dismutase (SOD)2 as a potential antioxidant factor involved in NFkappaB protective effects against ROH-induced oxidative stress. Taken together, data presented here show that NFkappaB mediates cellular resistance to the pro-oxidant effects of vitamin A by inhibiting RS accumulation and the persistent and redox-dependent activation of JNK-AP-1 cascade.

Retinoic acid induces apoptosis by a non-classical mechanism of ERK1/2 activation.

Even though RA is involved in differentiation and apoptosis of normal and cancer cells, being sometimes used as adjuvant in chemotherapy, its mechanisms of action involve multiple overlapping pathways that still remain unclear. Recent studies point out that RA exerts rapid and non-genomic effects, which are independent of RAR/RXR-mediated gene transcription. In this work, we reported that RA treatment for 24 h decreases cell viability, induces apoptosis dependent on caspase-3 activation, and activates the transcription factor AP-1 in cultured Sertoli cells. Moreover, RA induced a rapid and non-classical stimulation of ERK1/2. ERK1/2 activation was mediated by MEK1/2, and the protein synthesis inhibitor cycloheximide did not alter the pattern of RA-induced ERK1/2 phosphorylation. Pharmacological inhibition of MEK1/2-ERK1/2 pathway with UO126 blocked caspase-3 activation, decreased AP-1 binding to DNA and inhibited apoptosis. Overall, our data suggest that a rapid and non-genomic effect of RA upon MEK1/2-ERK1/2 pathway leads to caspase-3 activation and caspase-3-dependent apoptosis in cultured Sertoli cells. The non-canonical RA signaling presented in this work evokes new perspectives of RA action, which may play an important role in mediating early biological effects of RA modulating cell death in normal and tumor cells.

Retinol and retinoic acid modulate catalase activity in Sertoli cells by distinct and gene expression-independent mechanisms.

Vitamin A (retinol) exerts a major role in several biological functions. However, it was observed that retinol induces oxidative stress on different cellular types. Catalase (EC 1.11.1.6; CAT) is a hydrogen peroxide metabolizing enzyme, and its activity and expression is widely used as an index to measure oxidative stress and perturbations in the cellular redox state. The aim of this study was to investigate the effects of retinol and its major biologically active metabolite, all-trans retinoic acid (RA), on CAT regulation. For this purpose, cultured Sertoli cells (a physiological target of vitamin A) were treated with retinol or RA. Retinol (7 microM, 14 microM) and RA (100 nM, 1 microM) enhanced intracellular reactive species production and increased CAT activity after 24 h of treatment. Retinol increased CAT immunocontent but did not alter CAT mRNA expression, while the increase in CAT activity by RA was not related to alterations in immunocontent or mRNA expression. In vitro incubation of purified CAT with retinol or RA did not alter enzyme activity.

Protective effects of purple grape juice on carbon tetrachloride-induced oxidative stress in brains of adult Wistar rats.

The antioxidant properties of purple grape juice, organic and conventional, in brain tissues are not well known. In this study our objective was to evaluate the antioxidant activity in substantia nigra and striatum of rats chronically treated with organic or conventional purple grape juice and to correlate the results obtained with the polyphenol content (total polyphenolic content, resveratrol, and anthocyanins [malvidin, delphinidin, peonidin, and cyanidin]). We observed that CCl(4) damage decreased significantly in the grape juice-treated groups when compared with the control group. In the grape juice-treated groups we further observed a decrease of lipid (thiobarbituric acid-reactive substances assay) and protein (carbonyl) peroxidation, as well as a significant antioxidant protection through the increase of enzyme activity. Antioxidant activities were significantly correlated with polyphenol content. These findings demonstrated that both grape juices have potent antioxidant properties and these activities could be at least attributed to the high phenolic content present in these juices.