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1.
Nat Commun ; 10(1): 3081, 2019 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-31300673

RESUMO

Dimethyl fumarate (DMF) is a first-line-treatment for relapsing-remitting multiple sclerosis (RRMS). The redox master regulator Nrf2, essential for redox balance, is a target of DMF, but its precise therapeutic mechanisms of action remain elusive. Here we show impact of DMF on circulating monocytes and T cells in a prospective longitudinal RRMS patient cohort. DMF increases the level of oxidized isoprostanes in peripheral blood. Other observed changes, including methylome and transcriptome profiles, occur in monocytes prior to T cells. Importantly, monocyte counts and monocytic ROS increase following DMF and distinguish patients with beneficial treatment-response from non-responders. A single nucleotide polymorphism in the ROS-generating NOX3 gene is associated with beneficial DMF treatment-response. Our data implicate monocyte-derived oxidative processes in autoimmune diseases and their treatment, and identify NOX3 genetic variant, monocyte counts and redox state as parameters potentially useful to inform clinical decisions on DMF therapy of RRMS.


Assuntos
Fumarato de Dimetilo/uso terapêutico , Imunossupressores/uso terapêutico , Monócitos/imunologia , Esclerose Múltipla Recidivante-Remitente/tratamento farmacológico , NADPH Oxidases/genética , Adulto , Metilação de DNA/efeitos dos fármacos , Fumarato de Dimetilo/farmacologia , Epigênese Genética/efeitos dos fármacos , Feminino , Humanos , Imunossupressores/farmacologia , Contagem de Leucócitos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Monócitos/metabolismo , Esclerose Múltipla Recidivante-Remitente/sangue , Esclerose Múltipla Recidivante-Remitente/imunologia , NADPH Oxidases/metabolismo , Fator 2 Relacionado a NF-E2/antagonistas & inibidores , Fator 2 Relacionado a NF-E2/metabolismo , Polimorfismo de Nucleotídeo Único , Estudos Prospectivos , Espécies Reativas de Oxigênio/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Resultado do Tratamento
2.
Food Chem ; 293: 285-290, 2019 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-31151613

RESUMO

Exogenous adenosine triphosphate (ATP) treatment at 0, 250, 500, 750, and 1000 µM retarded cap browning in mushrooms by 0, 34, 26, 51 and 32 %, respectively, during storage at 4 °C for 18 days. Triggering signaling H2O2 accumulation arising from elevating NADPH oxidase enzyme activity during 6 days of storage at 4 °C may be pivotal for promoting shikimate dehydrogenase enzyme activity in mushrooms treated with ATP during 18 days of storage at 4 °C. Promoting melatonin accumulation (390 µg kg-1 FW vs. 160 µg kg-1 FW) in mushrooms treated with ATP during cold storage may attribute to signaling H2O2 accumulation. Higher DPPH scavenging capacity (72 % vs. 65 %) in mushrooms treated with ATP may attribute to higher phenols accumulation arising from higher phenylalanine ammonialyase/polyphenol oxidase enzymes activity concomitant with higher alternative oxidase gene expression during 18 days of storage at 4 °C.


Assuntos
Trifosfato de Adenosina/farmacologia , Agaricus/efeitos dos fármacos , Temperatura Baixa , Armazenamento de Alimentos , Reação de Maillard , Trifosfato de Adenosina/administração & dosagem , Agaricus/enzimologia , Agaricus/fisiologia , Oxirredutases do Álcool/metabolismo , Compostos de Bifenilo/química , Relação Dose-Resposta a Droga , Peróxido de Hidrogênio/metabolismo , Melatonina/metabolismo , Proteínas Mitocondriais/genética , NADPH Oxidases/metabolismo , Oxirredutases/genética , Fenóis/metabolismo , Picratos/química , Proteínas de Plantas/genética , Transdução de Sinais
3.
Toxicol Lett ; 313: 108-119, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31251971

RESUMO

Polychlorinated biphenyls (PCBs) are classic persistent organic pollutants (POPs) and are associated with the progression of many cancers, including liver cancer. The present study investigated the effect of 2,3'4,4',5-pentachlorobiphenyl (PCB118) on hepatocellular carcinoma cell proliferation and its underlying mechanisms. The results indicated that PCB118 exposure promotes the proliferation and glycolysis of hepatocellular carcinoma SMMC-7721 cells. Moreover, PCB118 exposure increased the expression level of pyruvate kinase M2 (PKM2) and its nuclear translocation, whereas treatment with PKM2 shRNA suppressed the induction of cell proliferation and glycolysis by PCB118. PCB118 stimulated reactive oxygen species (ROS) production by activating nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Treatment with the antioxidants N-acetyl-L-cysteine (NAC) and superoxide dismutase (SOD) prevented PCB118-induced effects on PKM2, cell proliferation and glycolysis. Furthermore, we found that PCB118 activated NADPH oxidase through the aryl hydrocarbon receptor (AhR) in SMMC-7721 cells. Consistently, treatment with AhR shRNA suppressed PCB118-induced effects on PKM2, cell proliferation and glycolysis. Overall, these results indicated that PCB118 promotes HCC cell proliferation via PKM2-dependent upregulation of glycolysis, which is mediated by AhR/NADPH oxidase-induced ROS production.


Assuntos
Carcinógenos Ambientais/toxicidade , Carcinoma Hepatocelular/enzimologia , Proteínas de Transporte/metabolismo , Proliferação de Células/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Neoplasias Hepáticas/enzimologia , Proteínas de Membrana/metabolismo , Bifenilos Policlorados/toxicidade , Hormônios Tireóideos/metabolismo , Transporte Ativo do Núcleo Celular , Fatores de Transcrição Hélice-Alça-Hélice Básicos/agonistas , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Proteínas de Transporte/genética , Linhagem Celular Tumoral , Humanos , Neoplasias Hepáticas/patologia , Proteínas de Membrana/genética , NADPH Oxidases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores de Hidrocarboneto Arílico/agonistas , Receptores de Hidrocarboneto Arílico/genética , Receptores de Hidrocarboneto Arílico/metabolismo , Transdução de Sinais/efeitos dos fármacos , Hormônios Tireóideos/genética
4.
Mol Plant Microbe Interact ; 32(11): 1508-1516, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31230563

RESUMO

During interactions, both plants and pathogens produce reactive oxygen species (ROS). Plants generate ROS for defense induction, while pathogens synthesize ROS for growth, sporulation, and virulence. NADPH oxidase (NOX) complex in the plasma membrane represents a main protein complex for ROS production in pathogens. Although NOX plays a crucial role in pathogenicity of pathogens, the underlying molecular mechanisms of NOX, especially the proteins regulated by NOX, remain largely unknown. Here, we applied an iodoacetyl tandem mass tag-based redox proteomic assay to investigate the protein redox dynamics in deletion mutant of bcnoxR, which encodes a regulatory subunit of NOX in the fungal pathogen Botrytis cinerea. In total, 214 unique peptidyl cysteine (Cys) thiols from 168 proteins were identified and quantified in both the wild type and ∆bcnoxR mutant. The Cys thiols in the ∆bcnoxR mutant were generally more oxidized than those in the wild type, suggesting that BcNoxR is essential for maintaining the equilibrium of the redox state in B. cinerea. Site-specific thiol oxidation analysis indicated that 142 peptides containing the oxidized thiols changed abundance significantly in the ∆bcnoxR mutant. Proteins containing these differential peptides are classified into various functional categories. Functional analysis revealed that one of these proteins, 6-phosphate dehydrogenase, played roles in oxidative stress response and pathogenesis of B. cinerea. These results provide insight into the potential target proteins and the ROS signal transduction pathway regulated by NOX.


Assuntos
Botrytis , Homeostase , NADPH Oxidases , Botrytis/enzimologia , Botrytis/genética , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Homeostase/genética , Mutação , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Oxirredução , Proteômica , Espécies Reativas de Oxigênio , Transdução de Sinais
5.
Vet Microbiol ; 233: 93-101, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31176418

RESUMO

Actinobacillus pleuropneumoniae (APP) and porcine circovirus type 2 (PCV2) are both important pathogens of the porcine respiratory disease complex (PRDC), which results in significant worldwide economic losses. Recently, PCV2 and APP coinfection has been described in the worldwide pork industry, and represents an extremely complex situation in veterinary medicine. However, the mechanism of their coinfection has not been investigated. In this study, we found that PCV2 promoted APP adhesion to and invasion of porcine alveolar macrophages (PAMs) during coinfection. Additionally, PCV2 suppressed reactive oxygen species (ROS) production by inhibiting cytomembrane NADPH oxidase activity, which was beneficial for APP survival in PAMs in vitro. During coinfection, PCV2 weakened the inflammatory response and macrophage antigen presentation by decreasing TNF-α, IFN-γ and IL-4 expression, and reduced clearance of the invading bacteria. The host-cell experimental results were verified in a mouse model. The findings provide a deeper and novel understanding of porcine coinfection, and will be extremely helpful for the design of strategies for PRDC control.


Assuntos
Actinobacillus pleuropneumoniae/fisiologia , Circovirus/fisiologia , Coinfecção/veterinária , Macrófagos Alveolares/microbiologia , Macrófagos Alveolares/virologia , Espécies Reativas de Oxigênio/metabolismo , Infecções por Actinobacillus/imunologia , Infecções por Actinobacillus/veterinária , Animais , Anticorpos Antivirais/imunologia , Apresentação do Antígeno , Aderência Bacteriana , Infecções por Circoviridae/imunologia , Infecções por Circoviridae/veterinária , Citocinas/genética , Citocinas/imunologia , Feminino , Inflamação , Masculino , Camundongos , Camundongos Endogâmicos ICR , Viabilidade Microbiana , NADPH Oxidases/metabolismo , Suínos
6.
J Dairy Sci ; 102(8): 7421-7434, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31178179

RESUMO

It is generally accepted that intracellular killing of microorganisms by production of reactive oxygen species (ROS) in the phagosome of the neutrophil is an important arm of innate defense. High-producing dairy cows are prone to periparturient metabolic and infectious diseases. Both myeloperoxidase (MPO) activity and ROS production decrease the day of parturition. Several studies have demonstrated changes in the expression of genes involved in, for example, metabolism and defense in the circulating neutrophil during peripartum. In this study, we wanted to further characterize the periparturient neutrophil in terms of its oxidative killing capacity by analyzing the oxidative burst at 3 levels. First, the ROS phenotype was evaluated using chemiluminescence. The cows (sampled within 24 h after parturition and at 135 d in milk) showed a significantly slower production of ROS at parturition. Both primiparous (n = 13) and multiparous (n = 12) cows were included in this study, but parity did not affect the kinetics of ROS production. Second, the expression of 11 genes involved in ROS production was measured in the same cows: cytochrome b-245 α and ß chain (CYBA, CYBB; coding for membrane-bound constituents of NADPH oxidase); neutrophil cytosolic factors 1, 2, and 4 (NCF1, NCF2, and NCF4); Rac family small GTPase 1 and 2 (RAC1 and RAC2; coding for regulatory proteins of NADPH oxidase); superoxide dismutase 2 (SOD2); catalase (CAT); myeloperoxidase (MPO; coding for enzymes involved in metabolizing downstream ROS); and spleen-associated tyrosine kinase (SYK; involved in signaling). During peripartum, a shift in expression in the oxidative killing pathway was observed, characterized by a downregulation of MPO and a simultaneous upregulation of the genes coding for NADPH oxidase. Third, as total DNA methylation is known to change during pregnancy, we investigated whether the observed differences were due to different methylation patterns. Promotor regions initiate transcription of particular genes; therefore, we analyzed the methylation status in annotated CpG islands of MPO and SOD2, 2 genes with a significant difference in expression between both lactation stages. The differences in methylation of these CpG islands were nonsignificant. High-throughput techniques may be necessary to obtain more detailed information on the total DNA methylation dynamics in bovine neutrophils and increase our understanding of how gene expression is controlled in neutrophils.


Assuntos
Bovinos/genética , Ilhas de CpG , Metilação de DNA , Regulação Enzimológica da Expressão Gênica , Neutrófilos/metabolismo , Peroxidase/genética , Superóxido Dismutase/genética , Animais , Feminino , Lactação , Leite/metabolismo , NADPH Oxidases/metabolismo , Paridade , Período Periparto , Peroxidase/metabolismo , Gravidez , Espécies Reativas de Oxigênio/metabolismo , Explosão Respiratória , Superóxido Dismutase/metabolismo
7.
Phytother Res ; 33(8): 2044-2055, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31209937

RESUMO

Mulberry leaf was reported that it has antidiabetic activity, although the mechanisms underlying the function have not been fully elucidated. In the present study, the results of network pharmacology suggested that mulberry leaves could regulate key biological process in development of diabetes, and the process implicates multiple signaling pathways, such as JAK-STAT, MAPK, VEGF, PPAR, and Wnt. Then, the research in vitro indicated that mulberry leaves remarkably ameliorated high glucose-induced epithelial to mesenchymal transition, which was characterized with significant reduction of intracellular reactive oxygen species (ROS) levels as well as downregulation of NADPH oxidase subunits NOX1, NOX2, and NOX4, and it was found to be connected with the ERK1/2 signaling pathway in human tubular epithelial cells (HK-2). Moreover, the results of bioinformatics and the dual luciferase report showed that ZEB1 might be a target gene of miR-302a; decreased miR-302a and increased ZEB1 expressions could significantly promote epithelial to mesenchymal transition. However, mulberry leaves could reverse these modulations. Our results demonstrated that network pharmacology could provide a guidance role for traditional Chinese medicine research, and mulberry leaves could be of benefit in preventing high glucose-induced EMT in HK-2 cells, which proved that it was related to the upregulation of miR-302a by targeting ZEB1 and the inhibition of NADPH oxidase/ROS/ERK1/2 pathway.


Assuntos
Nefropatias Diabéticas/tratamento farmacológico , Células Epiteliais/metabolismo , Nefropatias/tratamento farmacológico , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Morus/química , NADPH Oxidases/metabolismo , Folhas de Planta/química , Espécies Reativas de Oxigênio/metabolismo , Transição Epitelial-Mesenquimal , Humanos , Nefropatias/patologia
8.
Mol Med Rep ; 19(6): 5115-5122, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059037

RESUMO

3,3'­Diindolylmethane (DIM) is a naturally derived indole compound found in the Brassica family of vegetables. DIM has several beneficial effects, including anti­cancer, anti­inflammatory and anti­angiogenic functions. However, the effects of DIM on acute kidney injury (AKI) stimulated by lipopolysaccharide (LPS) are poorly studied. In this present study, male BALB/c mouse models of AKI were established using intraperitoneal injections of 10 mg/kg LPS. DIM (40 mg/kg) was administered intraperitoneally 24 and 2 h before LPS exposure. The results indicated that DIM significantly mitigated histopathological changes in the kidneys and improved the levels of blood urea nitrogen and serum creatinine. DIM also suppressed the LPS­induced production of reactive oxygen species and cell apoptosis. Furthermore, DIM treatment significantly decreased the expression of NADPH oxidase 2 (NOX2) and NOX4 in LPS­treated mice. Therefore, DIM may exert its renoprotective actions by inhibiting NOX­mediated oxidative stress and the apoptosis of renal tubular epithelial cells.


Assuntos
Lesão Renal Aguda/prevenção & controle , Apoptose/efeitos dos fármacos , Indóis/farmacologia , Lipopolissacarídeos/toxicidade , NADPH Oxidases/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Lesão Renal Aguda/etiologia , Lesão Renal Aguda/patologia , Animais , Nitrogênio da Ureia Sanguínea , Creatinina/sangue , Regulação para Baixo/efeitos dos fármacos , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Glutationa/metabolismo , Rim/patologia , Túbulos Renais/citologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , NADPH Oxidases/genética , Espécies Reativas de Oxigênio/metabolismo
9.
Int J Mol Sci ; 20(9)2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-31067654

RESUMO

As a gaseous biological signaling molecule, nitric oxide (NO) regulates many physiological processes in plants. Over the last decades, this low molecular weight compound has been identified as a key signaling molecule to regulate plant stress responses, and also plays an important role in plant development. However, elucidation of the molecular mechanisms for NO in leaf development has so far been limited due to a lack of mutant resources. Here, we employed the NO-deficient mutant nia1nia2 to examine the role of NO in leaf development. We have found that nia1nia2 mutant plants displayed very different leaf phenotypes as compared to wild type Col-0. Further studies have shown that reactive oxygen species (ROS) levels are higher in nia1nia2 mutant plants. Interestingly, ROS-related enzymes ascorbate peroxidase (APX), catalases (CAT), and peroxidases (POD) have shown decreases in their activities. Our transcriptome data have revealed that the ROS synthesis gene RBOHD was enhanced in nia1nia2 mutants and the photosynthesis-related pathway was impaired, which suggests that NO is required for chloroplast development and leaf development. Together, these results imply that NO plays a significant role in plant leaf development by regulating ROS homeostasis.


Assuntos
Arabidopsis/metabolismo , Homeostase , Óxido Nítrico/metabolismo , Folhas de Planta/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Nitrato Redutase/genética , Nitrato Redutase/metabolismo , Fotossíntese , Folhas de Planta/crescimento & desenvolvimento
10.
Life Sci ; 228: 121-127, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-31039364

RESUMO

AIMS: B1- and B2-kinin receptors play a major role in several cardiovascular diseases. Therefore, we aimed to evaluate cardiac functional consequences of B1- and B2-kinin receptors ablation, focusing on the cardiac ROS and NO generation. MAIN METHODS: Cardiac contractility, ROS, and NO generation, and protein expression were evaluated in male wild-type (WT), B1- (B1-/-) and B2-kinin (B2-/-) knockout mice. KEY FINDINGS: Impaired contractility in B1-/- and B2-/- hearts was associated with oxidative stress through upregulation of NADPH oxidase p22phox subunit. B1-/- and B2-/- hearts presented higher NO and peroxynitrite levels than WT. Despite decreased sarcoplasmic reticulum Ca2+ ATPase pump (SERCA2) expression, nitration at tyrosine residues of SERCA2 was markedly higher in B1-/- and B2-/- hearts. SIGNIFICANCE: B1- and B2-kinin receptors govern ROS generation, while disruption of B1- and B2-kinin receptors leads to impaired cardiac dysfunction through excessive tyrosine nitration on the SERCA2 structure.


Assuntos
Cardiopatias/genética , Coração/fisiopatologia , Receptor B1 da Bradicinina/genética , Receptor B2 da Bradicinina/genética , Animais , Deleção de Genes , Cardiopatias/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Contração Miocárdica , NADPH Oxidases/metabolismo , Óxido Nítrico/metabolismo , Oxirredução , Espécies Reativas de Oxigênio/metabolismo , Receptor B1 da Bradicinina/metabolismo , Receptor B2 da Bradicinina/metabolismo
11.
J Agric Food Chem ; 67(28): 7968-7976, 2019 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-31062982

RESUMO

Curcumin exhibits efficient antimicrobial activity; nevertheless, its effect on the postharvest decay of fruit has not been examined. Here, effects of curcumin on the fruit gray mold of kiwifruit infected by Botrytis cinerea were analyzed. Results demonstrated that curcumin induced reactive oxygen species (ROS) production and triggered apoptosis in B. cinerea hyphae. Use of N-acetylcysteine, a ROS scavenger, partially ameliorated the inhibition of curcumin on B. cinerea. The NADPH oxidase inhibitor, diphenyleneiodonium chlorine, abrogated the ROS production induced by curcumin, suggesting that curcumin induces oxidative stress in B. cinerea via a NADPH-oxidase-dependent mechanism. Disease severity of gray mold in curcumin-treated kiwifruit was significantly reduced. The malondialdehyde content decreased while the antioxidant enzyme activity increased in kiwifruit with the application of increasing concentrations of curcumin. Collectively, these results indicate that curcumin can be used to control gray mold and elevate antioxidant activity in kiwifruit.


Assuntos
Actinidia/microbiologia , Botrytis/efeitos dos fármacos , Curcumina/farmacologia , Fungicidas Industriais/farmacologia , Doenças das Plantas/microbiologia , Botrytis/metabolismo , Frutas/microbiologia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo
12.
Cell Physiol Biochem ; 52(6): 1398-1411, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31075190

RESUMO

BACKGROUND/AIMS: Visfatin is known to act as a mediator in several metabolic disorders, such as obesity, diabetes, and cardiovascular diseases. This study aimed to investigate the effect of visfatin on the adhesion of THP-1 monocytes to human vascular endothelial cells and the underlying mechanism. METHODS: Monocytes adhesion to endothelial cells was determined by using fluorescence-labeled monocytes. ICAM-1 and VCAM-1 expression in endothelial cells were measured by western blotting. Production of reactive oxygen species (ROS) was measured by using a fluorescent dye. The amounts of nuclear factor-kappa B (NF-κB) and phosphorylation of inhibitory factor of NF-κB (IκB) were determined by using western blot analysis. The translocation of NF-κB from the cytoplasm to the nucleus was determined by using immunofluorescence. RESULTS: Here we showed that visfatin significantly caused the upregulation of intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in endothelial cells, as well as enhanced monocyte adhesion to endothelial cells. Moreover, we found that inhibition of PI3K, Akt, and p38 MAPK activation significantly prevented visfatin-enhanced expression of ICAM-1 and VCAM-1 and monocyte adhesion to endothelial cells. Visfatin enhanced ROS production and IKK/NF-кB activation and then led to upregulation of ICAM-1 and VCAM-1 and enhanced monocyte adhesion to endothelial cells. These effects were also p38/PI3K/Akt-dependent. CONCLUSION: These results demonstrated that visfatin promoted monocyte-endothelial cell adhesion by increasing ICAM-1 and VCAM-1 expression via the activation of p38/PI3K/Akt signaling and downstream ROS production and IKK/NF-кB activation.


Assuntos
Adesão Celular/efeitos dos fármacos , Nicotinamida Fosforribosiltransferase/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Humanos , Proteínas I-kappa B/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Monócitos/citologia , NADPH Oxidases/metabolismo , NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/antagonistas & inibidores , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Molécula 1 de Adesão de Célula Vascular/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
13.
Cell Biol Int ; 43(6): 678-694, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30977575

RESUMO

We sought to determine the mechanism by which angiotensin II (ANGII) stimulates NADPH oxidase-mediated superoxide (O2 .- ) production in bovine pulmonary artery smooth muscle cells (BPASMCs). ANGII-induced increase in phospholipase D (PLD) and NADPH oxidase activities were inhibited upon pretreatment of the cells with chemical and genetic inhibitors of PLD2, but not PLD1. Immunoblot study revealed that ANGII treatment of the cells markedly increases protein kinase C-α (PKC-α), -δ, -ε, and -ζ levels in the cell membrane. Pretreatment of the cells with chemical and genetic inhibitors of PKC-ζ, but not PKC-α, -δ, and -ε, attenuated ANGII-induced increase in NADPH oxidase activity without a discernible change in PLD activity. Transfection of the cells with p47phox small interfering RNA inhibited ANGII-induced increase in NADPH oxidase activity without a significant change in PLD activity. Pretreatment of the cells with the chemical and genetic inhibitors of PLD2 and PKC-ζ inhibited ANGII-induced p47phox phosphorylation and subsequently translocation from cytosol to the cell membrane, and also inhibited its association with p22phox (a component of membrane-associated NADPH oxidase). Overall, PLD-PKCζ-p47phox signaling axis plays a crucial role in ANGII-induced increase in NADPH oxidase-mediated O2 .- production in the cells.


Assuntos
Angiotensina II/farmacologia , NADPH Oxidases/metabolismo , Fosfolipase D/metabolismo , Angiotensina II/metabolismo , Angiotensina II/fisiologia , Animais , Bovinos , Técnicas de Cultura de Células/métodos , Membrana Celular/metabolismo , Miócitos de Músculo Liso/metabolismo , NADPH Oxidases/fisiologia , Oxirredução , Fosfolipase D/antagonistas & inibidores , Fosfoproteínas/metabolismo , Fosforilação , Proteína Quinase C/metabolismo , Proteína Quinase C-alfa/metabolismo , Artéria Pulmonar/metabolismo , Transdução de Sinais/efeitos dos fármacos , Superóxidos/metabolismo
14.
Eur J Pharmacol ; 853: 381-387, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-31009636

RESUMO

Diabetic retinopathy is a common complication of diabetes that affects the retina due to a sustained high blood sugar level. Recent studies have demonstrated that high glucose-driven oxidative stress plays an important role in the microvascular complications of retina in diabetes. Oxidative stress occurs due to the excess of reactive oxygen species, which causes oxidative damage to retina, leading to the leak of tiny blood vessels, or acts as signaling molecules to trigger neovascularization, resulting in new fragile vessels. NADPH oxidase (NOX) is a key enzymatic source of reactive oxygen species in the retina, and it is involved in the early as well as the advanced stage of diabetic retinopathy. To date, at least 7 NOX isoforms, including NOX1 to NOX5, dual oxidase1 and dual oxidase 2, have been identified. It has been shown that NOX isoforms exert different roles in the pathogenesis of diabetic retinopathy. Intervention of NOX by its inhibitors or modulators shows beneficial effect on improving the retinal functions in the models of diabetic retinopathy in vivo or in vitro. Thereby, NOX might be a potential target for the therapy of diabetic retinopathy. The present review focuses on the role of NOX, particularly the NOX isoforms, in promoting the development of diabetic retinopathy. In addition, NOX isoforms as potential targets for therapy of diabetic retinopathy are also discussed.


Assuntos
Retinopatia Diabética/tratamento farmacológico , Retinopatia Diabética/enzimologia , Terapia de Alvo Molecular/métodos , NADPH Oxidases/metabolismo , Animais , Humanos
15.
Oxid Med Cell Longev ; 2019: 1050476, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31007831

RESUMO

The progression of Alzheimer's dementia is associated with neurovasculature impairment, which includes inflammation, microthromboses, and reduced cerebral blood flow. Here, we investigate the effects of ß amyloid peptides on the function of platelets, the cells driving haemostasis. Amyloid peptide ß1-42 (Aß1-42), Aß1-40, and Aß25-35 were tested in static adhesion experiments, and it was found that platelets preferentially adhere to Aß1-42 compared to other Aß peptides. In addition, significant platelet spreading was observed over Aß1-42, while Aß1-40, Aß25-35, and the scAß1-42 control did not seem to induce any platelet spreading, which suggested that only Aß1-42 activates platelet signalling in our experimental conditions. Aß1-42 also induced significant platelet adhesion and thrombus formation in whole blood under venous flow condition, while other Aß peptides did not. The molecular mechanism of Aß1-42 was investigated by flow cytometry, which revealed that this peptide induces a significant activation of integrin αIIbß3, but does not induce platelet degranulation (as measured by P-selectin membrane translocation). Finally, Aß1-42 treatment of human platelets led to detectable levels of protein kinase C (PKC) activation and tyrosine phosphorylation, which are hallmarks of platelet signalling. Interestingly, the NADPH oxidase (NOX) inhibitor VAS2870 completely abolished Aß1-42-dependent platelet adhesion in static conditions, thrombus formation in physiological flow conditions, integrin αIIbß3 activation, and tyrosine- and PKC-dependent platelet signalling. In summary, this study highlights the importance of NOXs in the activation of platelets in response to amyloid peptide ß1-42. The molecular mechanisms described in this manuscript may play an important role in the neurovascular impairment observed in Alzheimer's patients.


Assuntos
Peptídeos beta-Amiloides/toxicidade , NADPH Oxidases/metabolismo , Fragmentos de Peptídeos/toxicidade , Adesividade Plaquetária/efeitos dos fármacos , Trombose/patologia , Benzoxazóis/farmacologia , Humanos , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Triazóis/farmacologia
16.
Int J Mol Sci ; 20(7)2019 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-30987055

RESUMO

BACKGROUND: Previous studies demonstrated that calcium/calmodulin (Ca2+/CaM) activates nicotinamide adenine dinucleotide phosphate oxidases (NOX). In endothelial cells, the elevation of intracellular Ca2+ level consists of two components: Ca2+ mobilization from the endoplasmic reticulum (ER) and the subsequent store-operated Ca2+ entry. However, little is known about which component of Ca2+ increase is required to activate NOX in endothelial cells. Here, we investigated the mechanism that regulates NOX-derived reactive oxygen species (ROS) production via a Ca2+/CaM-dependent pathway. METHODS: We measured ROS production using a fluorescent indicator in endothelial cells and performed phosphorylation assays. RESULTS: Bradykinin (BK) increased NOX-derived cytosolic ROS. When cells were exposed to BK with either a nominal Ca2+-free or 1 mM of extracellular Ca2+ concentration modified Tyrode's solution, no difference in BK-induced ROS production was observed; however, chelating of cytosolic Ca2+ by BAPTA/AM or the depletion of ER Ca2+ contents by thapsigargin eliminated BK-induced ROS production. BK-induced ROS production was inhibited by a CaM inhibitor; however, a Ca2+/CaM-dependent protein kinase II (CaMKII) inhibitor did not affect BK-induced ROS production. Furthermore, BK stimulation did not increase phosphorylation of NOX2, NOX4, and NOX5. CONCLUSIONS: BK-induced NOX-derived ROS production was mediated via a Ca2+/CaM-dependent pathway; however, it was independent from NOX phosphorylation. This was strictly regulated by ER Ca2+ contents.


Assuntos
Cálcio/metabolismo , Calmodulina/metabolismo , Retículo Endoplasmático/metabolismo , Células Endoteliais/metabolismo , NADPH Oxidases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Bradicinina/farmacologia , Citosol/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Suínos
17.
BMC Complement Altern Med ; 19(1): 88, 2019 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-31023287

RESUMO

BACKGROUND: Stevia rebaudiana Bertoni has various pharmacological actions, which includes antidiabetic, antioxidant, anti-inflammatory activities. The superoxide and consequently NADPH oxidase (Nox) are relevant targets involved in biological effects of Stevia. The presence of NADPH-containing superoxide-producing lipoprotein (suprol) in Stevia leaves has not yet been tested. The mechanism of producing superoxide radicals (O2-) by suprol was determined in vitro, which is associated with the electron transfer from NADPH in the composition of suprol by traces of transition metal ions (Fe3+ or Cu2+) to molecular oxygen, turning it into O2-. It is expected that the therapeutic efficacy of Stevia leaves is caused by specific activity of superoxide-producing lipoprotein fraction. METHODS: For the first time, from the dry leaves of Stevia the NADPH-containing superoxide-producing lipoprotein was isolated and purified. The specific content of suprol (milligrams in 1 g of Stevia leaves- mg/g) was determined after desalination of suprol and lyophilization. RESULTS: According to the method provided, the specific content of the isolated suprol from Stevia's leaves was up to 4.5 ± 0.2 mg / g (yields up to 68.5 ± 4.7%, p < 0.05, n = 6). Nox forms a stable complex with suprol. The optical absorption spectrum of the Nox-suprol complex represents the overlapping suprol and Nox spectra, with a certain background increase and characteristic features of optical absorption for Nox. Due to O2- producing capacity suprol-Nox complex discolors KMnO4 solutions, Coomassie brilliant blue, restores nitrotetrazolium blue to formazan and oxidizes epinephrine to adrenochrome. The oxidation activity of adrenaline is 50.3 ± 5.1 U / mg / ml (p < 0.05, n = 6). CONCLUSION: Superoxide-producing lipoprotein fraction-Nox complex from Stevia leaves (membranes) can modulate redox regulated signaling pathways and may play a positive role in type-2 diabetes by means of adrenaline oxidation mechanism.


Assuntos
Lipoproteínas , NADP , Proteínas de Plantas , Stevia/química , Superóxidos , Lipoproteínas/química , Lipoproteínas/metabolismo , NADP/química , NADP/metabolismo , NADPH Oxidases/química , NADPH Oxidases/metabolismo , Oxirredução , Extratos Vegetais/química , Extratos Vegetais/metabolismo , Folhas de Planta/química , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Superóxidos/química , Superóxidos/metabolismo
18.
Oxid Med Cell Longev ; 2019: 1897316, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31019650

RESUMO

Background: Cellular stress is involved in ischemia/reperfusion- (I/R-) induced acute kidney injury (AKI). This study is aimed at investigating the effects of pretreatment with cholecalciferol on renal oxidative stress and endoplasmic reticulum (ER) stress during I/R-induced AKI. Methods: I/R-induced AKI was established by cross-clamping renal pedicles for 90 minutes and then reperfusion. In the Chol + I/R group, mice were orally administered with three doses of cholecalciferol (25 µg/kg) at 1, 24, and 48 h before ischemia. Renal cellular stress and kidney injury were measured at different time points after reperfusion. Results: I/R-induced AKI was alleviated in mice pretreated with cholecalciferol. In addition, I/R-induced renal cell apoptosis, as determined by TUNEL, was suppressed by cholecalciferol. Additional experiment showed that I/R-induced upregulation of renal GRP78 and CHOP was inhibited by cholecalciferol. I/R-induced renal IRE1α and eIF2α phosphorylation was attenuated by cholecalciferol. Moreover, I/R-induced renal GSH depletion, lipid peroxidation, and protein nitration were blocked in mice pretreated with cholecalciferol. I/R-induced upregulation of renal NADPH oxidases, such as p47phox, gp91phox, and nox4, was inhibited by cholecalciferol. I/R-induced upregulation of heme oxygenase- (HO-) 1, gshpx and gshrd, was attenuated in mice pretreated with cholecalciferol. Conclusions: Pretreatment with cholecalciferol protects against I/R-induced AKI partially through suppressing renal cellular stress response.


Assuntos
Lesão Renal Aguda/etiologia , Lesão Renal Aguda/patologia , Colecalciferol/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Rim/patologia , Estresse Oxidativo/efeitos dos fármacos , Traumatismo por Reperfusão/complicações , Animais , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Glutationa/metabolismo , Rim/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Camundongos Endogâmicos ICR , NADPH Oxidases/metabolismo , Receptores de Calcitriol/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos
19.
Virology ; 531: 269-279, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30974383

RESUMO

The study evaluated the effects of nucleoprotein viral and the infectious virus in SHK-1 cells. The results show a strong respiratory burst activation and the induction of p47phox, SOD, GLURED, and apoptotic genes. Additionally, the cells alter the profile of SUMOylated proteins by the effect of transfection and infection experiments. In silico analyses show a set of structural motifs in NP susceptible of post-translational modification by the SUMO protein. Interestingly, the inhibition of the NADPH oxidase complex blocked the production of reactive oxygen species and the high level of cellular ROS due to the nucleoprotein and the ISAv. At the same time, the blocking of the p38MAPK signaling pathway and the use of Aristotelia chilensis, decreased viral progeny production. These results suggest that the NP triggers a strong production of ROS and modifying the post-translational profile mediated by SUMO-2/3, a phenomenon that favors the production of new virions.


Assuntos
Doenças dos Peixes/metabolismo , Proteínas de Peixes/metabolismo , Isavirus/metabolismo , NADPH Oxidases/metabolismo , Nucleoproteínas/metabolismo , Infecções por Orthomyxoviridae/veterinária , Estresse Oxidativo , Proteínas Virais/metabolismo , Animais , Doenças dos Peixes/genética , Doenças dos Peixes/virologia , Proteínas de Peixes/genética , Interações Hospedeiro-Patógeno , Isavirus/genética , NADPH Oxidases/genética , Nucleoproteínas/genética , Infecções por Orthomyxoviridae/genética , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/virologia , Espécies Reativas de Oxigênio/metabolismo , Explosão Respiratória , Salmão , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/genética , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Sumoilação , Proteínas Virais/genética , Vírion/genética , Vírion/metabolismo
20.
Med Sci Monit ; 25: 2132-2140, 2019 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-30901320

RESUMO

BACKGROUND Endothelial injury is the main mechanism of atherosclerosis, and is caused by oxidized low-density lipoprotein (ox-LDL). Astragaloside IV (AS-IV) is the primary active ingredient of the Chinese herb Huangqi, and exhibits antioxidant and anti-inflammatory properties in cardiovascular diseases. This study investigated the protective effect of AS-IV in human umbilical vein endothelial cells (HUVECs). MATERIAL AND METHODS HUVEC cells were induced with ox-LDL to establish an in vitro atherosclerosis model. Then HUVECs were pretreated for 1 h with AS-IV at different concentrations (10, 20, and 50 µM) and then exposed to ox-LDL (100 µg/mL) for 48 h. The cell viability, lactate dehydrogenase (LDH) release, apoptosis, migration, intracellular reactive oxygen species (ROS), and NADPH oxidase activity of HUVECs were measured. qRT-PCR was performed to measure the mRNA expressions of Nrf2, HO-1, TNFalpha, and IL-6. Enzyme-linked immunosorbent assay (ELISA) was performed to measure the supernatant contents of TNFalpha and IL-6. RESULTS Exposure of HUVECs to ox-LDL reduced cell viability and migration, induced apoptosis, and increased intracellular ROS production and NADPH oxidase. Pretreatment with AS-IV (10, 20, and 50 µM) significantly enhanced the cell viability and migration, suppressed LDH release, apoptosis, ROS production, and NADPH oxidase in HUVECs, in a concentration-dependent manner. The AS-IV (50 µM) alone did not show significant differences from control. AS-IV increased mRNA expressions of Nrf2 and HO-1 and decreased mRNA expressions of TNFalpha and IL-6 in the ox-LDL-HUEVC cells. Furthermore, AS-IV reduced supernatant contents of TNFalpha and IL-6. CONCLUSIONS Astragaloside IV prevents ox-LDL-induced endothelial cell injury by reducing apoptosis, oxidative stress, and inflammatory response.


Assuntos
Células Endoteliais/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Saponinas/farmacologia , Triterpenos/farmacologia , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Endoteliais/fisiologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Inflamação/metabolismo , L-Lactato Desidrogenase/análise , Lipoproteínas LDL/metabolismo , NADP/análise , NADP/efeitos dos fármacos , NADPH Oxidases/metabolismo , Substâncias Protetoras/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Saponinas/metabolismo , Triterpenos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
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