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1.
Chemosphere ; 238: 124602, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31545211

RESUMO

Polybrominated diphenyl ethers (PBDEs) have been known to exhibit neurotoxicity in rats; however, the underlying mechanism remains unknown and there is no available intervention. In this study, we aimed to investigate the role of oxidative and nitrosative stress in the neurotoxicity in the cerebral cortex and primary neurons in rats following the BDE-153 treatment. Compared to the untreated group, BDE-153 treatment significantly induced the neurotoxic effects in rats, as manifested by the increased lactate dehydrogenase (LDH) activities and cell apoptosis rates, and the decreased neurotrophic factor contents and cholinergic enzyme activities in rats' cerebral cortices and primary neurons. When compared to the untreated group, the oxidative and nitrosative stress had occurred in the cerebral cortex or primary neurons in rats following the BDE-153 treatment, as manifested by the increments in levels of reactive oxygenspecies (ROS), malondialdehyde (MDA), nitric oxide (NO), and neuronal nitric oxide synthase (nNOS) mRNA and protein expressions, along with the decline in levels of superoxide dismutase (SOD) activity, glutathione (GSH) content, and peroxiredoxin I (Prx I) and Prx II mRNA and protein expressions. In addition, the ROS scavenger N-acetyl-l-cysteine (NAC) or NO scavenger NG-Nitro-l-arginine (L-NNA) significantly rescued the LDH leakage and cell survival, reversed the neurotrophin contents and cholinergic enzymes, mainly via regaining balance between oxidation/nitrosation and antioxidation. Overall, our findings suggested that oxidative and nitrosative stresses are involved in the neurotoxicity induced by BDE-153, and that the antioxidation is a potential targeted intervention.


Assuntos
Córtex Cerebral/patologia , Éteres Difenil Halogenados/toxicidade , Síndromes Neurotóxicas/patologia , Estresse Nitrosativo/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Bifenil Polibromatos/toxicidade , Acetilcisteína/farmacologia , Animais , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Córtex Cerebral/efeitos dos fármacos , Glutationa/metabolismo , Éteres Difenil Halogenados/metabolismo , Masculino , Malondialdeído/metabolismo , Fatores de Crescimento Neural/metabolismo , Neurônios/efeitos dos fármacos , Neurotrofina 3/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Estresse Nitrosativo/fisiologia , Estresse Oxidativo/fisiologia , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos
2.
J Basic Microbiol ; 59(8): 834-845, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31210376

RESUMO

A bacterium's ability to thrive in the presence of multiple environmental stressors simultaneously determines its resilience. We showed that activation of the SigB-controlled general stress response by mild environmental or energy stress provided significant cross-protection to subsequent lethal oxidative, disulfide and nitrosative stress in Bacillus subtilis. SigB activation is mediated via the stressosome and RsbP, the main conduits of environmental and energy stress, respectively. Cells exposed to mild environmental stress while lacking the major stressosome components RsbT or RsbRA were highly sensitive to subsequent oxidative stress, whereas rsbRB, rsbRC, rsbRD, and ytvA null mutants showed a spectrum of sensitivity, confirming their redundant roles and suggesting they could modulate the signals generated by environmental or oxidative stress. By contrast, cells encountering stationary phase stress required RsbP but not RsbT to survive subsequent oxidative stress. Interestingly, optimum cross-protection against nitrosative stress caused by sodium nitropruside required SigB but not the known regulators, RsbT and RsbP, suggesting an additional and as yet uncharacterized route of SigB activation independent of the known regulators. Together, these results provide mechanistic information on how B. subtilis promotes enhanced resistance against lethal oxidative stress during mild environmental and energy stress conditions.


Assuntos
Bacillus subtilis/fisiologia , Proteínas de Bactérias/metabolismo , Estresse Oxidativo/fisiologia , Fosfoproteínas Fosfatases/metabolismo , Fator sigma/metabolismo , Transdução de Sinais , Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Proteínas de Bactérias/genética , Deleção de Genes , Viabilidade Microbiana , Estresse Nitrosativo/fisiologia , Fosfoproteínas Fosfatases/genética , Fosfoproteínas/genética , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Fator sigma/genética , Transdução de Sinais/genética
3.
Infect Immun ; 87(9)2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31209149

RESUMO

Enterohemorrhagic Escherichia coli (EHEC) has at least three enzymes, NorV, Hmp, and Hcp, that act independently to lower the toxicity of nitric oxide (NO), a potent antimicrobial molecule. This study aimed to reveal the cooperative roles of these defensive enzymes in EHEC against nitrosative stress. Under anaerobic conditions, combined deletion of all three enzymes significantly increased the NO sensitivity of EHEC determined by the growth at late stationary phase; however, the expression of norV restored the NO resistance of EHEC. On the other hand, the growth of Δhmp mutant EHEC was inhibited after early stationary phase, indicating that NorV and Hmp play a cooperative role in anaerobic growth. Under microaerobic conditions, the growth of Δhmp mutant EHEC was inhibited by NO, indicating that Hmp is the enzyme that protects cells from NO stress under microaerobic conditions. When EHEC cells were exposed to a lower concentration of NO, the NO level in bacterial cells of Δhcp mutant EHEC was higher than those of the other EHEC mutants, suggesting that Hcp is effective at regulating NO levels only at a low concentration. These findings of a low level of NO in bacterial cells with hcp indicate that the NO consumption activity of Hcp was suppressed by Hmp at a low range of NO concentrations. Taken together, these results show that the cooperative effects of NO-metabolizing enzymes are regulated by the range of NO concentrations to which the EHEC cells are exposed.


Assuntos
Escherichia coli Êntero-Hemorrágica/enzimologia , Proteínas de Escherichia coli/metabolismo , Óxido Nítrico/metabolismo , Estresse Nitrosativo/fisiologia , Anaerobiose , Escherichia coli Êntero-Hemorrágica/metabolismo , Regulação Bacteriana da Expressão Gênica , Oxirredutases/metabolismo
4.
Nitric Oxide ; 88: 35-44, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-30981896

RESUMO

Vibrio cholerae faces nitrosative stress during successful colonization in intestine. Very little information is available on the nitrosative stress protective mechanisms of V. cholerae. Reports show that NorR regulon control two genes hmpA and nnrS responsible for nitric oxide (NO) detoxification in V. cholerae. In the present study we first time report a novel role of V. cholerae catalases under nitrosative stress. Using zymogram analysis of catalase we showed that KatB and KatG activity were induced within 30 min in V. cholerae in the presence of sodium nitroprusside (SNP), a NO donor compound. Surprisingly, V. cholerae cell survival was found to be decreased under nitrosative stress if catalase activities were blocked by ATz, a catalase inhibitor. Flow cytometry study was conducted to detect reactive oxygen species (ROS) and reactive nitrogen species (RNS) using DHE and DHR123, fluorescent probes respectively. Short exposure of SNP to V. cholerae did not generate ROS but RNS was detectable within 30 min. Total glutathione content was increased in V. cholerae cells under nitrosative stress. Furthermore, Superoxide dismutase (SOD) and Glutathione reductase (GR) activities remained unchanged under nitrosative stress in V. cholerae indicated antioxidant role of NO which could produce peroxynitrite. To investigate the role of catalase induction under nitrosative stress in V. cholerae, we conducted peroxynitrite reductase assay using cell lysates. Interestingly, SNP treated V. cholerae cell lysates showed lowest DHR123 oxidation compared to the control set. The extent of DHR123 oxidation was more in V. cholerae cell lysate when catalases were blocked by ATz.


Assuntos
Proteínas de Bactérias/metabolismo , Catalase/metabolismo , Estresse Nitrosativo/fisiologia , Espécies Reativas de Nitrogênio/fisiologia , Vibrio cholerae/fisiologia , Amitrol (Herbicida)/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/genética , Catalase/antagonistas & inibidores , Catalase/genética , Indução Enzimática , Inibidores Enzimáticos , Doadores de Óxido Nítrico/farmacologia , Nitroprussiato/farmacologia
5.
Neuroscience ; 406: 1-21, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30825584

RESUMO

The global burden of neurodegenerative disorders has increased substantially over the past 2 decades due to rising rates of population aging. Although neurodegenerative disorders differ in their clinical presentation, the underlying pathobiological processes are largely shared. Oxidative stress, among other mechanisms, is strongly implicated in neurodegenerative disorders and aging, and can potentially be targeted by antioxidative agents. Curcumin, a component of turmeric, is a compound that has received considerable attention for its therapeutic properties, and it is considered to be a powerful antioxidant. In this review, we analyzed the evidence for curcumin as an antioxidant in models of neurodegenerative disorders as well as oxido-nitrosative stress. A total of 1451 articles were found from 3 scientific literature databases (PubMed, Scopus, and Web of Science). After all exclusions, a final total of 64 articles were included in this review. The majority of the studies showed that curcumin, or derivatives thereof, were protective against oxidative and/or nitrosative stress in various cellular and animal models. Overall, curcumin protected against lipid and protein oxidation with a reduction in levels of malondialdehyde, and protein carbonyls, thiols and nitrotyrosines. Furthermore, it stimulated the activities of antioxidant enzymes such as superoxide dismutase and glutathione peroxidase. In conclusion, curcumin appears to be a promising compound for phytomedicine. However, due to some concerns about its efficacy, further targeted experiments are needed to identify its exact molecular targets and pathways responsible for its antioxidant effects.


Assuntos
Envelhecimento/efeitos dos fármacos , Antioxidantes/uso terapêutico , Curcumina/uso terapêutico , Doenças Neurodegenerativas/tratamento farmacológico , Estresse Nitrosativo/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Envelhecimento/metabolismo , Animais , Animais Geneticamente Modificados , Antioxidantes/farmacologia , Linhagem Celular , Curcumina/farmacologia , Humanos , Doenças Neurodegenerativas/metabolismo , Estresse Nitrosativo/fisiologia , Estresse Oxidativo/fisiologia
6.
Braz J Microbiol ; 50(2): 501-506, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30864077

RESUMO

Studying biofilm dispersal is important to prevent Listeria monocytogenes persistence in food processing plants and to avoid finished product contamination. Reactive oxygen and nitrogen intermediates (ROI and RNI, respectively) may trigger cell detachment from many bacterial species biofilms, but their roles in L. monocytogenes biofilms have not been fully investigated. This study reports on ROI and RNI quantification in Listeria monocytogenes biofilms formed on stainless steel and glass surfaces; bacterial culture and microscopy combined with fluorescent staining were employed. Nitric oxide (NO) donor and inhibitor putative effects on L. monocytogenes dispersal from biofilms were evaluated, and transcription of genes (prfA, lmo 0990, lmo 0807, and lmo1485) involved in ROI and RNI stress responses were quantified by real-time PCR (qPCR). Microscopy detected the reactive intermediates NO, peroxynitrite, H2O2, and superoxide in L. monocytogenes biofilms. Neither NO donor nor inhibitors interfered in L. monocytogenes growth and gene expression, except for lmo0990, which was downregulated. In conclusion, ROI and RNI did not exert dispersive effects on L. monocytogenes biofilms, indicating that this pathogen has a tight control for protection against oxidative and nitrosative stresses.


Assuntos
Biofilmes/crescimento & desenvolvimento , Listeria monocytogenes/crescimento & desenvolvimento , Estresse Nitrosativo/fisiologia , Estresse Oxidativo/fisiologia , Espécies Reativas de Nitrogênio/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Manipulação de Alimentos/métodos , Listeria monocytogenes/metabolismo , Óxido Nítrico/química , Nitrogênio/química , Oxigênio/química , Transcrição Genética/efeitos dos fármacos , Transcrição Genética/genética
7.
Nitric Oxide ; 86: 1-11, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-30772503

RESUMO

Paracoccidioides brasiliensis is a temperature-dependent dimorphic fungus that cause paracoccidioidomycosis (PCM), the major systemic mycosis in Latin America. The capacity to evade the innate immune response of the host is due to P. brasiliensis ability to respond and to survive the nitrosative stress caused by phagocytic cells. However, the regulation of signal transduction pathways associated to nitrosative stress response are poorly understood. Ras GTPase play an important role in the various cellular events in many fungi. Ras, in its activated form (Ras-GTP), interacts with effector proteins and can initiate a kinase cascade. In this report, we investigated the role of Ras GTPase in P. brasiliensis after in vitro stimulus with nitric oxide (NO). We observed that low concentrations of NO induced cell proliferation in P. brasiliensis, while high concentrations promoted decrease in fungal viability, and both events were reversed in the presence of a NO scavenger. We observed that high levels of NO induced Ras activation and its S-nitrosylation. Additionally, we showed that Ras modulated the expression of antioxidant genes in response to nitrosative stress. We find that the Hog1 MAP kinase contributed to nitrosative stress response in P. brasiliensis in a Ras-dependent manner. Taken together, our data demonstrate the relationship between Ras-GTPase and Hog1 MAPK pathway allowing for the P. brasiliensis adaptation to nitrosative stress.


Assuntos
Proteínas Fúngicas/fisiologia , Sistema de Sinalização das MAP Quinases/fisiologia , Proteínas Quinases Ativadas por Mitógeno/fisiologia , Estresse Nitrosativo/fisiologia , Paracoccidioides/fisiologia , Proteínas ras/fisiologia , Sequência de Aminoácidos , Animais , Morte Celular/fisiologia , Proliferação de Células/fisiologia , Expressão Gênica/fisiologia , Masculino , Camundongos Endogâmicos BALB C , Proteínas Quinases Ativadas por Mitógeno/química , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Óxido Nítrico/química , Óxido Nítrico/farmacologia , Processamento de Proteína Pós-Traducional
8.
Psychopharmacology (Berl) ; 236(10): 2867-2880, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30610349

RESUMO

RATIONALE AND OBJECTIVES: Stress-induced alterations in oxidative and inflammatory parameters have been implicated in the pathophysiology of mood disorders. Based on the antioxidant and anti-inflammatory properties of the selenium-containing compound 3-((4-chlorophenyl)selanyl)-1-methyl-1H-indole (CMI), we assessed its ability to reverse depression-like behavioral alterations, neuroinflammation, and oxidative imbalance induced by acute restraint stress. METHODS: Mice submitted to restraint for 240 min received CMI (1 or 10 mg/kg, orally) 10 min after the end of the stress induction. Behavioral and biochemical tests were carried out after further 30 min. RESULTS: Restraint-induced depression-like behavior in the tail suspension test (TST), splash test, and new object exploration test was reversed by CMI. None of the treatments evoked locomotor alteration. In addition, CMI abrogated restraint-induced increases in plasma levels of corticosterone and in markers of oxidative stress and impaired superoxide dismutase and catalase activity in the prefrontal cortex (PFC) and hippocampus (HC). CMI also blocked stress-induced downregulation of mRNA levels of glucocorticoid receptor and brain-derived neurotrophic factor and upregulation of nuclear factor kappa B, inducible nitric oxide synthase, tumor necrosis alpha, indoelamine-2,3-dioxygenase, and glycogen synthase kinase 3 beta in PFC and HC. CONCLUSIONS: These preclinical results indicate that administration of selenium-containing compounds might help to treat depression associated with inflammation and oxidative stress. Graphical abstract ᅟ.


Assuntos
Depressão/tratamento farmacológico , Indóis/uso terapêutico , Estresse Nitrosativo/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Compostos de Selênio/uso terapêutico , Estresse Psicológico/tratamento farmacológico , Animais , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Depressão/metabolismo , Depressão/psicologia , Indóis/química , Indóis/farmacologia , Inflamação/tratamento farmacológico , Inflamação/psicologia , Masculino , Camundongos , Estresse Nitrosativo/fisiologia , Estresse Oxidativo/fisiologia , Distribuição Aleatória , Restrição Física/métodos , Restrição Física/psicologia , Compostos de Selênio/química , Compostos de Selênio/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Estresse Psicológico/metabolismo , Estresse Psicológico/psicologia
9.
Eur J Ophthalmol ; 29(2): 178-182, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29991295

RESUMO

AIM:: To quantify the levels of nitric oxide, inducible nitric oxide synthase, and 3-nitrotyrosine in cataractous lenses of smokers and smokers who chewed tobacco in comparison with non-smokers and non-smokers who chewed tobacco. STUDY DESIGN:: A total of 80 cataractous lenses from smokers, non-smokers, smokers with tobacco chewing habit, and non-smokers with tobacco chewing habit were collected from the patients who had enrolled in the Department of Ophthalmology, Mahatma Gandhi Medical College & Research Institute, Puducherry. METHODS:: Levels of nitric oxide, inducible nitric oxide synthase, and 3-nitrotyrosine were quantified using commercially available enzyme-linked immunosorbent assay kits. RESULTS:: The mean concentrations of lens nitric oxide, inducible nitric oxide synthase, and 3-nitrotyrosine are as follows: (a) smokers-112.01, 59.57, and 88.91 µmol/L; (b) smokers who chewed tobacco-175.15, 93.95, and 128.72 µmol/L; (c) non-smokers-76.15, 40.65, and 70.20 µmol/L; and (d) non-smokers who chewed tobacco-96.56, 52.87, and 83.88 µmol/L, respectively. CONCLUSION:: Nitric oxide, inducible nitric oxide synthase, and 3-nitrotyrosine at high levels are the major causative agents for cataractogenesis. The results of this study suggest that smoking and tobacco chewing habit generate nitrosative stress that could enhance the pathogenesis for early cataractogenesis.


Assuntos
Catarata/metabolismo , Cristalino/metabolismo , Estresse Nitrosativo/fisiologia , não Fumantes , Fumantes , Fumar/efeitos adversos , Catarata/diagnóstico , Catarata/etiologia , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Cristalino/patologia , Masculino , Pessoa de Meia-Idade , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismo
10.
Med Mycol ; 57(1): 101-113, 2019 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-29294039

RESUMO

Candida albicans is an important source of device-associated infection because of its capacity for biofilm formation. This yeast has the ability to form biofilms which favors the persistence of the infection. Furthermore, the innate immune response has a critical role in the control of these infections and macrophages (Mø) are vital to this process. An important fungicidal mechanism employed by Mø involves the generation of toxic reactive oxygen species (ROS) and reactive nitrogen intermediates (RNI). The interaction between biofilms and these immune cells, and the contribution of oxidative and nitrosative stress, that is determinant to the course of the infection, remains elusive. The aim of this study was to investigate this interaction. To this purpose, two models of Mø-biofilms contact, early (model 1) and mature (model 2) biofilms, were used; and the production of ROS, RNI and the oxidative stress response (OSR) were evaluated. We found that the presence of Mø decreased the biofilm formation at an early stage and increased the production of ROS and RNI, with activation of ORS (enzymatic and nonenzymatic). On the other hand, the interaction between mature biofilms and Mø resulted in an increasing biofilm formation, with low levels of RNI and ROS production and decrease of OSR. Dynamic interactions between Mø and fungal biofilms were also clearly evident from images obtained by confocal scanning laser microscopy. The prooxidant-antioxidant balance was different depending of C. albicans biofilms stages and likely acts as a signal over their formation in presence of Mø. These results may contribute to a better understanding of the immune-pathogenesis of C. albicans biofilm infections.


Assuntos
Biofilmes/crescimento & desenvolvimento , Candida albicans/fisiologia , Macrófagos/microbiologia , Estresse Oxidativo/fisiologia , Animais , Antifúngicos/farmacologia , Antioxidantes/metabolismo , Biofilmes/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Candida albicans/genética , Farmacorresistência Fúngica/genética , Interações Hospedeiro-Patógeno , Macrófagos/fisiologia , Camundongos , Modelos Biológicos , Mutação , Estresse Nitrosativo/fisiologia , Células RAW 264.7 , Espécies Reativas de Nitrogênio/metabolismo , Espécies Reativas de Oxigênio/metabolismo
11.
FASEB J ; 33(3): 3718-3730, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30521379

RESUMO

Protein tyrosine (Tyr) nitration, the covalent addition of a nitro group (•NO2) to Tyr residues, is emerging as a candidate mechanism of endothelial dysfunction. Previous studies have shown that Tyr nitration is primarily induced by nitrosative stress, a process characterized by the production of reactive nitrogen species, especially peroxynitrite anion (ONOO-), which is considered a secondary product of NO in the presence of superoxide radicals (O2•-). However, the impact of nitrosative stress-induced Tyr nitration on endothelial dysfunction has not been thoroughly elucidated to date. We developed an endothelial dysfunction model, a process called "endothelial-to-mesenchymal transition (EndMT)," and evaluated the production of NO, O2•-, and protein nitration during EndMT. The results showed that TGF-ß1 stimulation induced EndMT and elevated endothelial NO and O2•- production as well as nitration of the catalytic subunit of protein phosphatase (PP)2A. Mass spectrometry analysis showed that Tyr265 was the nitration site in the catalytic subunit of protein phosphatase (PP)2A, and this Tyr nitration increased PP2A activity and disrupted endothelial integrity. To devise an endothelial-targeted anti-PP2Ac nitration strategy, a mimic peptide, tyrosine 265 wild type (Y265WT), conjugated with the cell-penetrating peptide HIV-1 TAT protein (TAT) was synthesized. PP2Ac nitration and PP2A activity were significantly inhibited by pretreatment with TAT-265WT, and the integrity of endothelial cells was maintained. Furthermore, injection of TAT-265WT attenuated renal nitration formation and caused anticapillary rarefaction in a unilateral urethral obstructive nephropathy model. Taken together, these results offer preclinical proof of concept for TAT-265WT as a tractable agent to protect against nitrosative stress-induced endothelial dysfunction in renal microvessels.-Deng,Y., Cai, Y., Liu, L., Lin, X., Lu, P., Guo, Y., Han, M., Xu, G. Blocking Tyr265 nitration of protein phosphatase 2A attenuates nitrosative stress-induced endothelial dysfunction in renal microvessels.


Assuntos
Células Endoteliais/metabolismo , Rim/metabolismo , Microvasos/metabolismo , Estresse Nitrosativo/fisiologia , Proteína Fosfatase 2/metabolismo , Tirosina/metabolismo , Doenças Vasculares/metabolismo , Animais , Linhagem Celular , Endotélio/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ácido Peroxinitroso/metabolismo , Espécies Reativas de Nitrogênio/metabolismo , Superóxidos/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
12.
J BUON ; 23(5): 1481-1491, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30570876

RESUMO

PURPOSE: A common feature of malignancies is increased reactive oxygen species (ROS) and reactive nitrogen species (RNS). We analyzed the influence of oxidative and nitrosative stress on the activation of AKT/mTOR signaling pathway in myeloproliferative neoplasms (MPN). METHODS: Oxidative stress-induced gene expression in circulatory CD34+ cells of MPN patients was studied by microarray analysis. Biomarkers of oxidative and nitrosative stress were determined using spectrophotometry in plasma and erythrocyte lysate. The levels of nitrotyrosine, inducible NO synthase (iNOS) and AKT/mTOR/p70S6K phosphorylation were determined by immunocytochemistry and immunoblotting in granulocytes of MPN patients. RESULTS: Antioxidants superoxide dismutase 2 (SOD2) and glutathione peroxidase 1 (GPx1) gene expression were increased in circulatory CD34+ cells, while SOD1 and GPx enzymes were reduced in the erythrocytes of MPN. Plasma malonyl-dialdehyde and protein carbonyl levels were elevated in MPN. The total antioxidant capacity in plasma and erythrocyte catalase (CT) activities was the most prominent in primary myelofibrosis (PMF) with JAK2V617F heterozygosity. The total nitrite/nitrate (NOx) level was augmented in the plasma of PMF patients (p<0.001), while nitrotyrosine and iNOS were generally increased in the granulocytes of MPN patients. Activation of AKT/mTOR signaling was the most significant in PMF (p<0.01), but demonstrated JAK2V617F dependence and consequent p70S6K phosphorylation in the granulocytes of essential thrombocytemia (ET) and polycythemia vera (PV) patients. Hydrogen peroxide stimulated mTOR pathway, iNOS and nitrotyrosine quantities, the last one prevented by the antioxidant n-acetyl-cysteine (NAC) in the granulocytes of MPN. CONCLUSION: Our study showed increased levels of oxidative and nitrosative stress parameters in MPN with JAK2V617F dependence. The ROS enhanced the constitutive activation of AKT/mTOR signaling and nitrosative parameters in MPN.


Assuntos
Transtornos Mieloproliferativos/metabolismo , Estresse Nitrosativo/fisiologia , Estresse Oxidativo/fisiologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Adulto , Idoso , Humanos , Pessoa de Meia-Idade , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/patologia , Transdução de Sinais
13.
Neuroscience ; 393: 273-283, 2018 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-30316912

RESUMO

Chronic psychogenic stress can increase neuronal calcium influx and generate the intracellular accumulation of oxidative (ROS) and nitrosative (RNS) reactive species, disrupting synaptic transmission in the brain. These molecules impair the Na,K-ATPase (NKA) activity, whose malfunction has been related to neuropsychiatric disorders, including anxiety, depression, schizophrenia, and neurodegenerative diseases. In this study, we assessed how 14 days of restraint stress in rats affect NKA activity via oxidative/nitrosative damage in the frontal cortex (FCx), a crucial region for emotional and cognitive control. One day after the last stress session (S14 + 1d), but not immediately after the last stress session (S14), α2,3-NKA activity was significantly reduced in the FCx, without changes in the protein levels. The S14 + 1d animals also showed increased lipid peroxidation, iNOS, and AP-1 activities, as well as TNF-α protein levels, evidencing oxidative stress and neuroinflammation. No cellular death or neurodegeneration was observed in the FCx of S14 + 1d animals. Pharmacological inhibition of iNOS or COX-2 before each stress session prevented lipid peroxidation and the α2,3-NKA activity loss. Our results show that repeated restraint exposure for 14 days decreases the activity of α2,3-NKA in FCx 24 h after the last stress, an effect associated with augmented inflammatory response and oxidative and nitrosative damage and suggest new pathophysiological roles to neuroinflammation in neuropsychiatric diseases.


Assuntos
Lobo Frontal/metabolismo , Estresse Nitrosativo/fisiologia , Estresse Oxidativo/fisiologia , Restrição Física , ATPase Trocadora de Sódio-Potássio/metabolismo , Animais , Ciclo-Oxigenase 2/metabolismo , Depressão/metabolismo , Masculino , Ratos Wistar , Restrição Física/métodos
14.
Nat Commun ; 9(1): 3505, 2018 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-30158522

RESUMO

Argininosuccinate lyase (ASL) belongs to the hepatic urea cycle detoxifying ammonia, and the citrulline-nitric oxide (NO) cycle producing NO. ASL-deficient patients present argininosuccinic aciduria characterised by hyperammonaemia, multiorgan disease and neurocognitive impairment despite treatment aiming to normalise ammonaemia without considering NO imbalance. Here we show that cerebral disease in argininosuccinic aciduria involves neuronal oxidative/nitrosative stress independent of hyperammonaemia. Intravenous injection of AAV8 vector into adult or neonatal ASL-deficient mice demonstrates long-term correction of the hepatic urea cycle and the cerebral citrulline-NO cycle, respectively. Cerebral disease persists if ammonaemia only is normalised but is dramatically reduced after correction of both ammonaemia and neuronal ASL activity. This correlates with behavioural improvement and reduced cortical cell death. Thus, neuronal oxidative/nitrosative stress is a distinct pathophysiological mechanism from hyperammonaemia. Disease amelioration by simultaneous brain and liver gene transfer with one vector, to treat both metabolic pathways, provides new hope for hepatocerebral metabolic diseases.


Assuntos
Argininossuccinato Liase/metabolismo , Acidúria Argininossuccínica/metabolismo , Acidúria Argininossuccínica/terapia , Animais , Argininossuccinato Liase/genética , Acidúria Argininossuccínica/genética , Encefalopatias/genética , Encefalopatias/metabolismo , Encefalopatias/terapia , Citrulina/metabolismo , Terapia Genética , Hiperamonemia/genética , Hiperamonemia/metabolismo , Hiperamonemia/terapia , Fígado/citologia , Camundongos , Neurônios/metabolismo , Óxido Nítrico/metabolismo , Estresse Nitrosativo/genética , Estresse Nitrosativo/fisiologia
15.
Pulm Pharmacol Ther ; 51: 32-40, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29964173

RESUMO

Pulmonary fibrosis (PF) is a lethal end stage of interstitial lung disease with increasing prevalence. The disease burden of PF has seen a sharp surge in the past two decades owing to entry of heavy amount of particulate matter due to industrialization and urbanization. In this work, we developed an oropharyngeal aspiration model of silica (1.5 mg/mice) induced pulmonary fibrosis as a homogeneous, reproducible, simple and alternative strategy in Swiss albino mice. Various BALF (protein, albumin, cell count), biochemical parameters (MDA, GSH, hydroxyproline), cytokines (IL-1ß, IL-6, TNF-α and TGF-ß1), histological (H&E and PSR staining) and protein expression (N-cadherin, vimentin, α-SMA, CTGF, collagen-1) studies were conducted to validate the model. Oropharyngeal administration of silica in Swiss mice produced significantly changes in lung morphology with statistically higher lung weights compared to normal control animals. The silica treated mice showed profoundly elevated BALF soluble and cytological parameters and enhanced oxidative and nitrosative stress in lungs. The levels of hydroxyproline were increased by 2.6 fold in the silica treated mice. The expression of pro-inflammatory cytokines were profoundly increased in silica treated mice. The histology and PSR staining indicated increased inflammatory infiltration and staggering fibrosis in silica treated group. In addition, the expression of EMT markers (N-cadherin, vimentin, α-SMA and CTGF) were significantly increased indicating their role in silica induced pulmonary fibrosis. Our work clearly demonstrates the superiority of stress free oropharyngeal instillation of silica with dose reduction over the conventional invasive and non-homogeneous intratracheal route.


Assuntos
Modelos Animais de Doenças , Doenças Profissionais/fisiopatologia , Fibrose Pulmonar/fisiopatologia , Dióxido de Silício/toxicidade , Administração Oral , Animais , Líquido da Lavagem Broncoalveolar , Citocinas/metabolismo , Hidroxiprolina/metabolismo , Masculino , Camundongos , Estresse Nitrosativo/fisiologia , Estresse Oxidativo/fisiologia , Reprodutibilidade dos Testes , Dióxido de Silício/administração & dosagem
16.
Nitric Oxide ; 78: 95-102, 2018 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-29885902

RESUMO

AIM: Inducible nitric oxide synthase (iNOS) is a key regulator of the innate immune system. The aim of the current study was to explore whether innate immune-mediated iNOS and reactive nitrogen species acutely perturb acinar cell physiology and calcium homeostasis of exocrine salivary tissues. METHODS: Innate immunity in the submandibular gland of C57BL/6 mice was locally activated via intraductal retrograde infusion of polyinosinic:polycytidylic acid (poly (I:C). Expressions of iNOS and the activity of the reactive nitrogen species peroxynitrite, were evaluated by immunohistochemistry. Mice were pre-treated with the selective iNOS inhibitor aminoguanidine in order to substantiate the injurious effect of the nitrosative signal on the key calcium regulator sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA2b) and calcium signalling. RESULTS: Challenging salivary gland innate immunity with poly (I:C) prompted upregulated expression of iNOS and the generation of peroxynitrite. Inhibition of iNOS/peroxynitrite revealed the role played by upregulated nitrosative signalling in: dysregulated expression of SERCA2b, perturbed calcium homeostasis and loss of saliva secretion. CONCLUSION: iNOS mediates disruption of exocrine calcium signalling causing secretory dysfunction following activation of innate immunity in a novel salivary gland injury model.


Assuntos
Sinalização do Cálcio/fisiologia , Óxido Nítrico Sintase Tipo II/metabolismo , Estresse Nitrosativo/fisiologia , Doenças da Glândula Submandibular/fisiopatologia , Células Acinares/fisiologia , Animais , Cálcio/metabolismo , Inibidores Enzimáticos/farmacologia , Feminino , Guanidinas/farmacologia , Imunidade Inata/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Ácido Peroxinitroso/metabolismo , Poli I-C , Saliva/metabolismo , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Glândula Submandibular/efeitos dos fármacos , Glândula Submandibular/patologia , Doenças da Glândula Submandibular/induzido quimicamente , Doenças da Glândula Submandibular/imunologia , Tirosina/análogos & derivados , Tirosina/metabolismo , Regulação para Cima/efeitos dos fármacos
17.
Asian J Androl ; 20(6): 600-607, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29956685

RESUMO

Peroxynitrite is a highly reactive nitrogen species and a potent inducer of apoptosis and necrosis in somatic cells. Peroxynitrite-induced nitrosative stress has emerged as a major cause of impaired sperm function; however, its ability to trigger cell death has not been described in human spermatozoa. The objective here was to characterize biochemical and morphological features of cell death induced by peroxynitrite-mediated nitrosative stress in human spermatozoa. For this, spermatozoa were incubated with and without (untreated control) 3-morpholinosydnonimine (SIN-1), in order to generate peroxynitrite. Sperm viability, mitochondrial permeability transition (MPT), externalization of phosphatidylserine, DNA oxidation and fragmentation, caspase activation, tyrosine nitration, and sperm ultrastructure were analyzed. The results showed that at 24 h of incubation with SIN-1, the sperm viability was significantly reduced compared to untreated control (P < 0.001). Furthermore, the MPT was induced (P < 0.01) and increment in DNA oxidation (P < 0.01), DNA fragmentation (P < 0.01), tyrosine nitration (P < 0.0001) and ultrastructural damage were observed when compared to untreated control. Caspase activation was not evidenced, and although phosphatidylserine externalization increased compared to untreated control (P < 0.001), this process was observed in <10% of the cells and the gradual loss of viability was not characterized by an important increase in this parameter. In conclusion, peroxynitrite-mediated nitrosative stress induces the regulated variant of cell death known as MPT-driven necrosis in human spermatozoa. This study provides a new insight into the pathophysiology of nitrosative stress in human spermatozoa and opens up a new focus for developing specific therapeutic strategies to better preserve sperm viability or to avoid cell death.


Assuntos
Morte Celular , Mitocôndrias/patologia , Estresse Nitrosativo/fisiologia , Espermatozoides/patologia , Adulto , Caspases/metabolismo , Ativação Enzimática , Humanos , Masculino , Mitocôndrias/metabolismo , Necrose , Permeabilidade , Ácido Peroxinitroso/farmacologia , Fosfatidilserinas/metabolismo , Espermatozoides/metabolismo , Espermatozoides/ultraestrutura
18.
Arch Endocrinol Metab ; 62(3): 309-318, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29791650

RESUMO

OBJECTIVE: The aim of this study is to evaluate the influence of the body mass index (BMI) and the metabolic syndrome (MetS) parameters on oxidative and nitrosative stress in overweight and obese subjects. SUBJECTS AND METHODS: Individuals were divided into three groups: the control group (G1, n = 131) with a BMI between 20 and 24.9 kg/m2, the overweight group (G2, n = 120) with a BMI between 25 and 29.9 kg/m2 and the obese group (G3, n = 79) with a BMI ≥ 30 kg/m2. RESULTS: G3 presented higher advanced oxidation protein products (AOPPs) in relation to G1 and G2 (p = 0.001 and p = 0.011, respectively) whereas G2 and G3 had lower levels of nitric oxide (NO) (p = 0.009 and p = 0.048, respectively) compared to G1. Adjusted for the presence of MetS to evaluate its influence, the levels of AOPPs did not differ between the groups, whereas NO remained significantly lower. Data adjusted by the BMI showed that subjects with higher triacylglycerol levels had higher AOPPs (p = 0.001) and decreased total radical-trapping antioxidant parameter/uric Acid (p = 0.036). Subjects with lower high-density lipoprotein (HDL) levels and patients with higher blood pressure showed increased AOPPs (p = 0.001 and p = 0.034, respectively) and lower NO levels (p = 0.017 and p = 0.043, respectively). Subjects who presented insulin resistance had higher AOPPs (p = 0.024). CONCLUSIONS: Nitrosative stress was related to BMI, and protein oxidation and nitrosative stress were related to metabolic changes and hypertension. MetS components were essential participants in oxidative and nitrosative stress in overweight and obese subjects.


Assuntos
Produtos da Oxidação Avançada de Proteínas/metabolismo , Síndrome Metabólica/metabolismo , Estresse Nitrosativo/fisiologia , Obesidade/metabolismo , Adolescente , Adulto , Idoso , Estudos de Casos e Controles , Feminino , Humanos , Masculino , Síndrome Metabólica/fisiopatologia , Pessoa de Meia-Idade , Obesidade/fisiopatologia , Sobrepeso/metabolismo , Sobrepeso/fisiopatologia , Adulto Jovem
19.
J Sex Med ; 15(5): 654-661, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29606625

RESUMO

BACKGROUND: Excess reactive oxygen species and reactive nitrogen species are implicated in male infertility and impaired spermatogenesis. AIM: To investigate the effect of excess reactive nitrogen species and nitrosative stress on testicular function and the hypothalamic-pituitary-gonadal axis using the S-nitrosoglutathione reductase-null (Gsnor-/-) mouse model. METHODS: Testis size, pup number, and epididymal sperm concentration and motility of Gsnor-/- mice were compared with those of age-matched wild-type (WT) mice. Reproductive hormones testosterone (T), luteinizing hormone (LH), and follicle-stimulating hormone were compared in Gsnor-/- and WT mice. Immunofluorescence for Gsnor-/- and WT testis was performed for 3ß-hydroxysteroid dehydrogenase and luteinizing hormone receptor (LHR) and compared. Human chorionic gonadotropin and gonadotropin-releasing hormone stimulation tests were performed to assess and compare testicular and pituitary functions of Gsnor-/- and WT mice. OUTCOMES: Evaluation of fertility and reproductive hormones in Gsnor-/- vs WT mice. Response of Gsnor-/- and WT mice to human chorionic gonadotropin and gonadotropin-releasing hormone to evaluate LH and T production. RESULTS: Gsnor-/- mice had smaller litters (4.2 vs 8.0 pups per litter; P < .01), smaller testes (0.08 vs 0.09 g; P < .01), and decreased epididymal sperm concentration (69 vs 98 × 106; P < .05) and motility (39% vs 65%; P < .05) compared with WT mice. Serum T (44.8 vs 292.2 ng/dL; P < .05) and LH (0.03 vs 0.74 ng/mL; P = .04) were lower in Gsnor-/- than in WT mice despite similar follicle-stimulating hormone levels (63.98 vs 77.93 ng/mL; P = .20). Immunofluorescence of Gsnor-/- and WT testes showed similar staining of 3ß-hydroxysteroid dehydrogenase and LHR. Human chorionic gonadotropin stimulation of Gsnor-/- mice increased serum T (>1,680 vs >1,680 ng/dL) and gonadotropin-releasing hormone stimulation increased serum LH (6.3 vs 8.9 ng/mL; P = .20) similar to WT mice. CLINICAL TRANSLATION: These findings provide novel insight to a possible mechanism of secondary hypogonadism from increased reactive nitrogen species and excess nitrosative stress. STRENGTHS AND LIMITATIONS: Limitations of this study are its small samples and variability in hormone levels. CONCLUSION: Deficiency of S-nitrosoglutathione reductase results in secondary hypogonadism, suggesting that excess nitrosative stress can affect LH production from the pituitary gland. Masterson TA, Arora H, Kulandavelu S, et al. S-Nitrosoglutathione Reductase (GSNOR) Deficiency Results in Secondary Hypogonadism. J Sex Med 2018;15:654-661.


Assuntos
Aldeído Oxirredutases/deficiência , Hipogonadismo/etiologia , Hipogonadismo/patologia , 17-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Gonadotropina Coriônica/metabolismo , Hormônio Foliculoestimulante/metabolismo , Hormônio Liberador de Gonadotropina/metabolismo , Hormônio Luteinizante/metabolismo , Masculino , Camundongos , Estresse Nitrosativo/fisiologia , Contagem de Espermatozoides , Testículo/patologia , Testosterona/metabolismo
20.
mBio ; 9(2)2018 03 27.
Artigo em Inglês | MEDLINE | ID: mdl-29588408

RESUMO

In all eukaryotic kingdoms, mitogen-activated protein kinases (MAPKs) play critical roles in cellular responses to environmental cues. These MAPKs are activated by phosphorylation at highly conserved threonine and tyrosine residues in response to specific inputs, leading to their accumulation in the nucleus and the activation of their downstream targets. A specific MAP kinase can regulate different downstream targets depending on the nature of the input signal, thereby raising a key question: what defines the stress-specific outputs of MAP kinases? We find that the Hog1 MAPK contributes to nitrosative-stress resistance in Candida albicans even though it displays minimal stress-induced phosphorylation under these conditions. We show that Hog1 becomes oxidized in response to nitrosative stress, accumulates in the nucleus, and regulates the nitrosative stress-induced transcriptome. Mutation of specific cysteine residues revealed that C156 and C161 function together to promote stress resistance, Hog1-mediated nitrosative-stress-induced gene expression, resistance to phagocytic killing, and C. albicans virulence. We propose that the oxidation of Hog1, rather than its phosphorylation, contributes to the nitrosative-stress-specific responses of this MAP kinase.IMPORTANCE Mitogen-activated protein kinases play key roles in the responses of eukaryotic cells to extracellular signals and are critical for environmental-stress resistance. The widely accepted paradigm is that MAP kinases are activated by phosphorylation, which then triggers their nuclear accumulation and the activation of target proteins and genes that promote cellular adaptation. Our data suggest that alternative forms of posttranslational modification can modulate MAP kinase functionality in Candida albicans We demonstrate that Hog1 is not significantly phosphorylated in response to nitrosative stress, yet it displays nuclear accumulation and contributes to the global transcriptional response to this stress, as well as promoting nitrosative-stress resistance. Instead, nitrosative stress triggers changes in the redox status of Hog1. We also show that specific Hog1 cysteine residues influence its activation of stress genes. Therefore, alternative posttranslational modifications appear to regulate the stress-specific outputs of MAP kinases.


Assuntos
Proteínas Quinases Ativadas por Mitógeno/metabolismo , Estresse Nitrosativo/fisiologia , Candida albicans/metabolismo , Regulação Fúngica da Expressão Gênica/genética , Regulação Fúngica da Expressão Gênica/fisiologia , Proteínas Quinases Ativadas por Mitógeno/genética , Estresse Nitrosativo/genética , Oxirredução , Fosforilação/genética , Fosforilação/fisiologia
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