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1.
Pestic Biochem Physiol ; 159: 118-126, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31400773

RESUMO

In the plant-insect arms race, plants synthesize toxic compounds to defend against herbivorous insects, whereas insects employ cytochrome P450 monooxygenases (P450s) to detoxify these phytotoxins. As ubiquitous environmental contaminants, heavy metals can be easily absorbed by plants and further accumulated in herbivorous insects through the food chains, resulting in tangible consequences for plant-insect interactions. However, whether heavy metals can influence P450 activities and thereby cause further effects on larval tolerance to phytotoxins remains unknown. In this study, we shown that prior exposure to copper (Cu) enhanced larval tolerance to xanthotoxin in Spodoptera litura, a major polyphagous pest of agriculture. P450 activities were induced in larvae exposed to Cu or xanthotoxin, and a midgut specific expressed P450 gene, CYP6B50 was cross-induced after exposure to these two toxic xenobiotics. Knocking down CYP6B50 by RNA interference (RNAi) rendered the larvae more sensitive to xanthotoxin. As defense against oxidative stress following metal exposure has been demonstrated to affect insecticide resistance, the reactive oxygen species (ROS) generation and antioxidant enzyme activities were assessed. Cu exposure caused the accumulation of hydrogen peroxide (H2O2) and enhanced the activities of superoxide dismutase (SOD) and peroxidase (POD) in larval midgut. In addition, two antioxidant response elements (AREs) were identified from the CYP6B50 promoter, indicating that Cu-induced CYP6B50 expression may be related to the ROS burst. Application of ROS scavenger N-acetylcysteine (NAC) effectively suppressed CYP6B50 expression, inhibited P450 activities and impaired larval tolerance to xanthotoxin that had been induced by Cu. These results indicate that the increase in CYP6B50 expression regulated by Cu-induced H2O2 generation contributed to the enhancement of larval tolerance to xanthotoxin in S. litura. Ingestion of heavy metals from their host plants can inadvertently boost the counter-defense system of herbivorous insects to protect themselves against plant defensive toxins.


Assuntos
Cobre/farmacologia , Peróxido de Hidrogênio/metabolismo , Metoxaleno/farmacologia , Spodoptera/efeitos dos fármacos , Spodoptera/metabolismo , Animais , Elementos de Resposta Antioxidante/genética , Elementos de Resposta Antioxidante/fisiologia , Sistema Enzimático do Citocromo P-450/metabolismo , Peroxidase/genética , Peroxidase/metabolismo , Interferência de RNA , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo
2.
Chem Commun (Camb) ; 55(72): 10792-10795, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31432816

RESUMO

Hypoxia, as an important feature in tumor sites, greatly hinders the performance of photosensitizers, thus affecting the efficacy of photodynamic therapy (PDT). Therefore, designing and preparing new photosensitizer with high photosensitivity under hypoxic condition presents a great challenge that urgently needs to be solved. In this work, a new nano-MOF material using Mn(ii) as the active center can catalytically decompose high concentrations of H2O2 in tumor cells to generate O2, thereby improving the PDT efficacy in hypoxic tumors. The Mn-MOF also produces 1O2 under light irradiation, which finally induces cancer cell apoptosis. This work offers a new strategy for the design and discovery of effective photosensitizers for PDT.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Manganês/farmacologia , Estruturas Metalorgânicas/farmacologia , Fotoquimioterapia , Fármacos Fotossensibilizantes/farmacologia , Hipóxia Tumoral/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Peróxido de Hidrogênio/metabolismo , Manganês/química , Estruturas Metalorgânicas/química , Camundongos , Oxigênio/metabolismo , Tamanho da Partícula , Fármacos Fotossensibilizantes/química , Propriedades de Superfície
3.
Microbiol Res ; 227: 126297, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31421711

RESUMO

Many plant growth promoting rhizobacteria such as Bacillus velezensis GJ11 can produce acetoin to trigger induced systemic resistance (ISR) in plants. For improving acetoin production, the mutant strains were respectively constructed by knockout of the gene of bdh (2,3-butanediol dehydrogenase) and gdh (glycerol dehydrogenase) in GJ11, but only GJ11Δbdh produced a high level of acetoin triggering strong ISR against Pseudomonas syringae infection in plants. GJ11Δbdh could induce H2O2 accumulation in plants by producing a high level of acetoin. H2O2 was necessary for triggering ISR against the pathogen infection because after scavenging H2O2 with ascorbic acid or catalase, the inhibition role to pathogen infection induced by acetoin almost disappeared in plants. Further investigation found the plants treated with GJ11Δbdh in an obvious "priming" state, in which the mild immune response was observed such as a slight increase of H2O2 production, callose deposition, and enzymes activity related with defence response (e.g. POD, PAL and PPO). The plants in "priming" could rapidly respond to the pathogen infection accompanying with a significant increase of H2O2 production, callose deposition, and enzymes activity. Collectively, this study provides new insight into the role of acetoin as a strong elicitor of defense response, and ascribes a new approach to construct the mutant strains with high production of acetoin for triggering stronger ISR against pathogens infection in plants.


Assuntos
Acetoína/metabolismo , Arabidopsis/genética , Bacillus/genética , Bacillus/metabolismo , Resistência à Doença/genética , Imunidade Vegetal/genética , Oxirredutases do Álcool/genética , Arabidopsis/imunologia , Arabidopsis/microbiologia , Ácido Ascórbico/metabolismo , Catalase/metabolismo , Resistência à Doença/fisiologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Técnicas de Inativação de Genes , Genes de Plantas/genética , Peróxido de Hidrogênio/metabolismo , Doenças das Plantas/imunologia , Doenças das Plantas/microbiologia , Imunidade Vegetal/fisiologia , Pseudomonas syringae/patogenicidade , Desidrogenase do Álcool de Açúcar/genética
4.
BMC Plant Biol ; 19(1): 345, 2019 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-31390991

RESUMO

BACKGROUND: Aquaporin (AQP) proteins comprise a group of membrane intrinsic proteins (MIPs) that are responsible for transporting water and other small molecules, which is crucial for plant survival under stress conditions including salt stress. Despite the vital role of AQPs, little is known about them in cucumber (Cucumis sativus L.). RESULTS: In this study, we identified 39 aquaporin-encoding genes in cucumber that were separated by phylogenetic analysis into five sub-families (PIP, TIP, NIP, SIP, and XIP). Their substrate specificity was then assessed based on key amino acid residues such as the aromatic/Arginine (ar/R) selectivity filter, Froger's positions, and specificity-determining positions. The putative cis-regulatory motifs available in the promoter region of each AQP gene were analyzed and results revealed that their promoter regions contain many abiotic related cis-regulatory elements. Furthermore, analysis of previously released RNA-seq data revealed tissue- and treatment-specific expression patterns of cucumber AQP genes (CsAQPs). Three aquaporins (CsTIP1;1, CsPIP2;4, and CsPIP1;2) were the most transcript abundance genes, with CsTIP1;1 showing the highest expression levels among all aquaporins. Subcellular localization analysis in Nicotiana benthamiana epidermal cells revealed the diverse and broad array of sub-cellular localizations of CsAQPs. We then performed RNA-seq to identify the expression pattern of CsAQPs under salt stress and found a general decreased expression level of root CsAQPs. Moreover, qRT-PCR revealed rapid changes in the expression levels of CsAQPs in response to diverse abiotic stresses including salt, polyethylene glycol (PEG)-6000, heat, and chilling stresses. Additionally, transient expression of AQPs in N. benthamiana increased leaf water loss rate, suggesting their potential roles in the regulation of plant water status under stress conditions. CONCLUSIONS: Our results indicated that CsAQPs play important roles in response to salt stress. The genome-wide identification and primary function characterization of cucumber aquaporins provides insight to elucidate the complexity of the AQP gene family and their biological functions in cucumber.


Assuntos
Aquaporinas/fisiologia , Cucumis sativus/genética , Proteínas de Plantas/fisiologia , Aquaporinas/genética , Aquaporinas/metabolismo , Cucumis sativus/metabolismo , Expressão Gênica , Regulação da Expressão Gênica de Plantas , Genoma de Planta , Peróxido de Hidrogênio/metabolismo , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Estresse Fisiológico/genética , Transcriptoma , Água/metabolismo
5.
Plant Mol Biol ; 101(3): 315-323, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31392474

RESUMO

KEY MESSAGE: Pre-treatment of soybean seedlings with 200 µM salicylic acid before fungal inoculation significantly alleviated disease resistance in soybean seedlings against Fusarium solani infection. Sudden death syndrome of soybean is largely caused by Fusarium solani (F. solani). Salicylic acid (SA) has been reported to induce resistance in plants against many pathogens. However, the effect of exogenous SA application on F. solani infection of soybean is less reported. This study investigated the effect of foliar application of SA on soybean seedlings before F. solani infection. Seedlings were sprayed with 200 µM SA and inoculated with F. solani after 24 h of last SA application. After 3 days post-inoculation, seedlings treated with 200 µM SA showed significantly fewer disease symptoms with increased endogenous SA level, SA marker genes expression and antioxidant activities in the SA-treated seedlings more than the untreated control seedlings. Furthermore, the decrease in hydrogen peroxide (H2O2) and malondialdehyde (MDA) levels was observed in the SA-treated plants as compared to the untreated plants. Analysis of the effect of SA application on F. solani showed that the mycelia growth of F. solani was not affected by SA treatment. Further investigation in this study revealed a decreased in F. solani biomass content in the SA treated seedlings. Results from the present study show that pre-treatment of 200 µM SA can induce resistance of soybean seedlings against F. solani infection.


Assuntos
Resistência à Doença/efeitos dos fármacos , Fusarium/patogenicidade , Doenças das Plantas/microbiologia , Ácido Salicílico/farmacologia , Soja/microbiologia , Regulação da Expressão Gênica de Plantas , Peróxido de Hidrogênio/metabolismo , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/microbiologia , Plântula/efeitos dos fármacos , Plântula/microbiologia , Soja/efeitos dos fármacos
6.
Exp Parasitol ; 205: 107748, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31442453

RESUMO

Trypanosoma cruzi (the causative agent of Chagas disease) presents a complex life cycle that involves adaptations in vertebrate and invertebrate hosts. As a protozoan parasite of hematophagous insects and mammalian hosts, T. cruzi is exposed to reactive oxygen species (ROS). To investigate the functionality of T. cruzi tartrate-resistant acid phosphatase type 5 (TcACP5), we cloned, superexpressed and purified the enzyme. Purified TcACP5 exhibited a Vmax and apparent Km for pNPP hydrolysis of 7.7 ±â€¯0.2 nmol pNP × µg-1 × h-1 and 169.3 ±â€¯22.6 µM, respectively. The pH dependence was characterized by sharp maximal activity at pH 5.0, and inhibition assays demonstrated its sensitivity to acid phosphatase inhibitors. Similar activities were obtained with saturating concentrations of P-Ser and P-Thr as substrates. The enzyme metabolizes hydrogen peroxide (H2O2) in vitro, and parasites superexpressing this enzyme were more resistant to oxidative stress promoted by H2O2. Taken together, these results suggest that TcACP5 plays a central role in phosphoryl transfer and redox reactions.


Assuntos
Peróxido de Hidrogênio/farmacologia , Estresse Oxidativo/fisiologia , Fosfatase Ácida Resistente a Tartarato/metabolismo , Trypanosoma cruzi/enzimologia , Sequência de Aminoácidos , Imunofluorescência , Regulação Enzimológica da Expressão Gênica , Peróxido de Hidrogênio/metabolismo , Concentração de Íons de Hidrogênio , Microscopia Confocal , Oxirredução , Especificidade por Substrato , Fosfatase Ácida Resistente a Tartarato/antagonistas & inibidores , Fosfatase Ácida Resistente a Tartarato/química , Transfecção , Trypanosoma cruzi/efeitos dos fármacos
7.
Ecotoxicol Environ Saf ; 182: 109421, 2019 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-31301592

RESUMO

The environmental contamination of soil by metal oxide nanomaterials is a growing global concern because of their potential toxicity. We investigated the effects of Mg doped ZnO (Mg-nZnO) nanoparticles on a model soil microorganism Bacillus subtilis. Mg-nZnO exhibited only a moderate toxic effect on B. subtilis vegetative cells but was able to prevent biofilm formation and destroy already formed biofilms. Similarly, Mg-nZnO (≤1 mg/mL) was moderately toxic towards Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, Salmonella enterica, Saccharomyces cerevisiae and murine macrophages. Engineered Mg-nZnO produced H2O2 and O2•- radicals in solutions of various salt and organic molecule compositions. A quantitative proteomic analysis of B. subtilis membrane proteins showed that Mg-nZnO increased the expression of proteins involved in detoxification of ROS, translation and biofilm formation. Overall, our results suggest that Mg-nZnO released into the environment may hinder the spreading, colonization and biofilm formation by B. subtilis but also induce a mechanism of bacterial adaptation.


Assuntos
Bacillus subtilis/efeitos dos fármacos , Nanopartículas/toxicidade , Poluentes do Solo/toxicidade , Óxido de Zinco/toxicidade , Animais , Biofilmes , Escherichia coli/efeitos dos fármacos , Peróxido de Hidrogênio/metabolismo , Camundongos , Óxidos/metabolismo , Proteômica , Solo , Microbiologia do Solo , Staphylococcus aureus
8.
Biomater Sci ; 7(9): 3898-3905, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31317137

RESUMO

To target a response to a high oxidative stress environment of inflammatory or tumor sites, various reactive oxygen species (ROS) sensitive polymers have been developed as drug delivery systems. In this study, a novel oxidation sensitive copolymer, phenylboronic acid pinacol ester-functionalized methoxyl poly(ethylene glycol)-block-poly(phthalic anhydride-alter-glycidyl propargyl ether) (mPEG-b-P(PA-alt-GPBAe)), was designed and synthesized by ring-opening alternating copolymerization (ROAP) and click reaction. The copolymers could self-assemble into micelles in aqueous solution with an average size of 20.3 ± 9.3 nm, and are able to load hydrophobic anticancer drug (doxorubicin, DOX) with a high encapsulation efficiency of 75.2%. Interestingly, the encapsulated drug showed accelerated release in the trigger of H2O2, or at low pH values. The copolymers have low cytotoxicity indicated by the 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay towards 4T1 cells, which showed cell viabilities of more than 80% with treatment of our copolymers at concentrations up to 0.5 mg mL-1. The effective uptake of the drug-loaded micelles by 4T1 cells was investigated by confocal laser scanning microscopy (CLSM) and flow cytometry (FCM) analysis. Finally, compared with free DOX, the DOX-loaded nanoparticles exhibited a better antitumor effect and had lower systemic toxicity in 4T1 tumor-bearing mice. Therefore, this new kind of copolymer acting as a stimuli-responsive nanocarrier should represent a promising therapeutic platform for cancer therapy.


Assuntos
Antineoplásicos/administração & dosagem , Ácidos Borônicos/química , Doxorrubicina/administração & dosagem , Nanocápsulas/química , Polímeros/química , Animais , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Sobrevivência Celular/efeitos dos fármacos , Liberação Controlada de Fármacos , Ésteres/química , Humanos , Peróxido de Hidrogênio/metabolismo , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Camundongos , Micelas , Oxirredução , Polietilenoglicóis/química
9.
Anticancer Res ; 39(7): 3443-3451, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31262868

RESUMO

BACKGROUND/AIM: This study aimed to investigate aclarubicin (ACR)-induced oxidative DNA damage and apoptosis. MATERIALS AND METHODS: ACR-induced apoptosis was analyzed using HL-60 leukemia cells and HP100 cells, hydrogen peroxide (H2O2)-resistant cells derived from HL-60 cells. ACR-induced DNA damage was analyzed using plasmid DNA. RESULTS: HL-60 cells were more sensitive to ACR than HP100 cells. In HP100 cells, DNA ladder formation and caspase-3/7 activity induced by ACR were suppressed or delayed in comparison to those in HL-60 cells. ACR-induced DNA damage occurred in the presence of Cu(II), and scavenger experiments showed that the reactive species causing DNA damage appeared to be generated from H2O2 and Cu(I). Moreover, we detected intracellular Cu(I) induced by ACR in HL-60 cells, using CopperGREEN™, a fluorescent probe for detection of Cu(I) ion specifically. CONCLUSION: ACR-induced DNA damage and apoptosis can be accounted for by the involvement of H2O2 and Cu(I).


Assuntos
Aclarubicina/efeitos adversos , Antibióticos Antineoplásicos/efeitos adversos , Apoptose/efeitos dos fármacos , Cobre/farmacologia , Dano ao DNA , Peróxido de Hidrogênio/metabolismo , Linhagem Celular Tumoral , Humanos , Neoplasias/metabolismo
10.
Chem Pharm Bull (Tokyo) ; 67(7): 717-720, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31257327

RESUMO

This study demonstrates the relation between the redox properties and cytotoxicity of anthraquinone derivatives with a hydroxyl and methoxy group. The redox behavior of the anthraquinone derivatives was initially observed via cyclic voltammetry and their characteristics were investigated using molecular orbital calculations. The cytotoxicity of the anthraquinone derivatives was then evaluated using human leukemia HL-60 and H2O2 resistant HP100 cells, and its correlation with the redox properties of these compounds was investigated. Therefore, it was suggested that the anthraquinone derivatives express cytotoxicity through H2O2 production, and that generation of the oxidized radical form influences their cytotoxicity.


Assuntos
Antraciclinas/química , Antraquinonas/química , Antineoplásicos/química , Antraciclinas/farmacologia , Antraquinonas/farmacologia , Antineoplásicos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Técnicas Eletroquímicas , Células HL-60 , Humanos , Peróxido de Hidrogênio/metabolismo , Oxirredução , Teoria Quântica
11.
Arch Insect Biochem Physiol ; 102(1): e21595, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31276240

RESUMO

Honey bees Apis mellifera L. are one of the most studied insect species due to their economic importance. The interest in studying honey bees chiefly stems from the recent rapid decrease in their world population, which has become a problem of food security. Nevertheless, there are no systemic studies on the properties of the mitochondria of honey bee flight muscles. We conducted a research of the mitochondria of the flight muscles of A. mellifera L. The influence of various organic substrates on mitochondrial respiration in the presence or absence of adenosine diphosphate (ADP) was investigated. We demonstrated that pyruvate is the optimal substrate for the coupled respiration. A combination of pyruvate and glutamate is required for the maximal respiration rate. We also show that succinate oxidation does not support the oxidative phosphorylation and the generation of membrane potential. We also studied the production of reactive oxygen species by isolated mitochondria. The greatest production of H2 O2 (as a percentage of the rate of oxygen consumed) in the absence of ADP was observed during the respiration supported by α-glycerophosphate, malate, and a combination of malate with another NAD-linked substrate. We showed that honey bee flight muscle mitochondria are unable to uptake Ca2+ -ions. We also show that bee mitochondria are able to oxidize the respiration substrates effectively at the temperature of 50°Ð¡ compared to Bombus terrestris mitochondria, which were more adapted to lower temperatures.


Assuntos
Abelhas/metabolismo , Mitocôndrias Musculares/metabolismo , Animais , Cálcio/metabolismo , Respiração Celular , Feminino , Voo Animal , Peróxido de Hidrogênio/metabolismo , Masculino , Potenciais da Membrana , Camundongos , Músculos/metabolismo , Temperatura Ambiente
12.
Chem Biol Interact ; 310: 108739, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31288001

RESUMO

Phenol red (PR) is the standard pH indicator in various cell and tissue culture media, as it provides a quick check for the health of the culture. PR has also been used in multiple protocols to detect cellular hydrogen peroxide as well as peroxidase activity from human peroxidase enzymes. The majority of promyelocytic leukemia cell lines (e.g. HL-60 cells) express myeloperoxidase (MPO), which may react with PR, especially as the latter is present in cell culture media at sufficient concentrations (~15 µM) to partake in redox reactions. Moreover, phenolic molecules are often efficient donor substrates for peroxidase enzymes. In this study, we hypothesized that MPO metabolism of PR via MPO-expressing HL-60 cells could result in PR metabolite(s) that could modulate cell viability. We used purified human MPO for UV-visible spectrophotometry, electron paramagnetic resonance (EPR) and LC-MS analyses to investigate PR peroxidation. 2-chloro-5,5-dimethyl-1,3-cyclohexanedione (monochloro-dimedone, MCD) was used to assess the effect of PR on MPO-catalyzed chlorination activity, and we assessed PR uptake by HL-60 cells using LC-MS analysis. Lastly, we investigated the impact of PR metabolism by intracellular MPO on cell viability (ATP, using CellTiter-Glo®), cytotoxicity (using trypan blue), and on reduced and oxidized glutathione (using GSH/GSSG-Glo™). Our results demonstrate that PR undergoes oxidative halogenation via MPO, resulting in its UV-vis spectral changes due to the formation of mono- and di-halogenated products. Moreover, a significant increase in MPO-catalyzed chlorination of MCD and an increase in glutathionyl radical detection (using EPR) were observed in the presence of PR. Our in-vitro studies revealed that PR is readily taken up by HL-60 cells and its metabolism by intracellular MPO leads to a significant decrease in cellular glutathione as well as a significant increase in glutathione disulphide formation. In spite of the latter, PR had no considerable effect on HL-60 cell viability. These results provide evidence that while no overt decrease in cell viability may be observed, PR does impart redox activity, which investigators should be wary of in experimental protocols.


Assuntos
Protocolos Clínicos/normas , Concentração de Íons de Hidrogênio , Peroxidase/metabolismo , Fenolsulfonaftaleína/farmacologia , Células HL-60 , Halogenação , Humanos , Peróxido de Hidrogênio/metabolismo , Leucemia Promielocítica Aguda/enzimologia , Leucemia Promielocítica Aguda/metabolismo , Leucemia Promielocítica Aguda/patologia , Oxirredução , Fenolsulfonaftaleína/química , Fenolsulfonaftaleína/metabolismo , Fenolsulfonaftaleína/farmacocinética , Espectrofotometria
13.
Parasit Vectors ; 12(1): 339, 2019 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-31292008

RESUMO

BACKGROUND: The primary cause of parasitic gastroenteritis in small ruminants in temperate regions is the brown stomach worm, Teladorsagia circumcincta. Host immunity to this parasite is slow to develop, consistent with the ability of T. circumcincta to suppress the host immune response. Previous studies have shown that infective fourth-stage T. circumcincta larvae produce excretory-secretory products that are able to modulate the host immune response. The objective of this study was to identify immune modulatory excretory-secretory proteins from populations of fourth-stage T. circumcincta larvae present in two different host-niches: those associated with the gastric glands (mucosal-dwelling larvae) and those either loosely associated with the mucosa or free-living in the lumen (lumen-dwelling larvae). RESULTS: In this study excretory-secretory proteins from mucosal-dwelling and lumen-dwelling T. circumcincta fourth stage larvae were analysed using comparative 2-dimensional gel electrophoresis. A total of 17 proteins were identified as differentially expressed, with 14 proteins unique to, or enriched in, the excretory-secretory proteins of mucosal-dwelling larvae. One of the identified proteins, unique to mucosal-dwelling larvae, was a putative peroxiredoxin (T. circumcincta peroxiredoxin 1, Tci-Prx1). Peroxiredoxin orthologs from the trematode parasites Schistosoma mansoni and Fasciola hepatica have previously been shown to alternatively activate macrophages and play a key role in promoting parasite induced Th2 type immunity. Here we demonstrate that Tci-Prx1 is expressed in all infective T. circumcincta life-stages and, when produced as a recombinant protein, has peroxidase activity, whereby hydrogen peroxide (H2O2) is reduced and detoxified. Furthermore, we use an in vitro macrophage stimulation assay to demonstrate that, unlike peroxiredoxins from trematode parasites Schistosoma mansoni and Fasciola hepatica, Tci-Prx1 is unable to alternatively activate murine macrophage cells. CONCLUSIONS: In this study, we identified differences in the excretory-secretory proteome of mucosal-dwelling and lumen-dwelling infective fourth-stage T. circumcincta larvae, and demonstrated the utility of this comparative proteomic approach to identify excretory-secretory proteins of potential importance for parasite survival and/or host immune modulation.


Assuntos
Proteínas de Helminto/metabolismo , Peroxirredoxinas/metabolismo , Trichostrongyloidea/metabolismo , Animais , Eletroforese em Gel Bidimensional , Proteínas de Helminto/genética , Interações Hospedeiro-Parasita , Peróxido de Hidrogênio/metabolismo , Enteropatias Parasitárias , Larva/imunologia , Larva/metabolismo , Camundongos , Membrana Mucosa/parasitologia , Peroxirredoxinas/genética , Proteoma/análise , Proteômica , Ovinos/parasitologia , Doenças dos Ovinos/parasitologia , Trichostrongyloidea/imunologia
14.
Cancer Sci ; 110(9): 2856-2866, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31314163

RESUMO

4-Hydroxynonenal (HNE) is an important product of plasma membrane lipid peroxidation, which is a cause of cell and tissue injury. Mitochondrial DNA (mtDNA)-depleted ρ0 cells were established using human cervical cancer and oral squamous cell carcinoma cell lines. We investigated the effect of reactive oxygen species in ρ0 cells, especially the mechanism of hydrogen peroxide (H2 O2 )-mediated cell death. These cell were subjected to high oxidative stress and, compared with their parental cells, showed greater sensitivity to H2 O2 and high lipid peroxidation. Upregulation of HNE in the plasma membrane was observed prior to the increase in intracellular H2 O2 . The amount of oxidized lipid present changed H2 O2 permeability and administration of oxidized lipid led to further cell death after treatment with H2 O2 . Expression levels of lipoxygenase ALOX genes (ie ALOX5, ALOX12, and ALOX15) were upregulated in ρ0 cells, as were expression levels of ALOX12 and ALOX15 proteins. ALOX5 protein was mainly distributed in the nucleus, while ALOX12 and ALOX15 proteins were distributed in the nucleus and the cytoplasm. Although expression of COX2 gene was upregulated, its protein expression did not increase. ALOX (especially ALOX15) may be involved in the sensitivity of cancer cells to treatment. These data offer promise for the development of novel anticancer agents by altering the oxidation state of the plasma membrane. Our results showed that lipid peroxidation status is important for H2 O2 sensitivity and that ALOX15 is involved in lipid peroxidation status.


Assuntos
Apoptose/efeitos dos fármacos , Permeabilidade da Membrana Celular/genética , DNA Mitocondrial/genética , Peróxido de Hidrogênio/administração & dosagem , Peroxidação de Lipídeos/genética , Neoplasias/patologia , Aldeídos/metabolismo , Araquidonato 15-Lipoxigenase/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Resistencia a Medicamentos Antineoplásicos , Complexo de Proteínas da Cadeia de Transporte de Elétrons/genética , Complexo de Proteínas da Cadeia de Transporte de Elétrons/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo , Peróxido de Hidrogênio/farmacocinética , Mitocôndrias/genética , Mitocôndrias/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Estresse Oxidativo/efeitos dos fármacos , Éteres Fosfolipídicos/administração & dosagem , Regulação para Cima
15.
BMC Evol Biol ; 19(1): 146, 2019 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-31324143

RESUMO

BACKGROUND: Antioxidative enzymes contribute to a parasite's ability to counteract the host's intracellular killing mechanisms. The facultative intracellular oyster parasite, Perkinsus marinus, a sister taxon to dinoflagellates and apicomplexans, is responsible for mortalities of oysters along the Atlantic coast of North America. Parasite trophozoites enter molluscan hemocytes by subverting the phagocytic response while inhibiting the typical respiratory burst. Because P. marinus lacks catalase, the mechanism(s) by which the parasite evade the toxic effects of hydrogen peroxide had remained unclear. We previously found that P. marinus displays an ascorbate-dependent peroxidase (APX) activity typical of photosynthetic eukaryotes. Like other alveolates, the evolutionary history of P. marinus includes multiple endosymbiotic events. The discovery of APX in P. marinus raised the questions: From which ancestral lineage is this APX derived, and what role does it play in the parasite's life history? RESULTS: Purification of P. marinus cytosolic APX activity identified a 32 kDa protein. Amplification of parasite cDNA with oligonucleotides corresponding to peptides of the purified protein revealed two putative APX-encoding genes, designated PmAPX1 and PmAPX2. The predicted proteins are 93% identical, and PmAPX2 carries a 30 amino acid N-terminal extension relative to PmAPX1. The P. marinus APX proteins are similar to predicted APX proteins of dinoflagellates, and they more closely resemble chloroplastic than cytosolic APX enzymes of plants. Immunofluorescence for PmAPX1 and PmAPX2 shows that PmAPX1 is cytoplasmic, while PmAPX2 is localized to the periphery of the central vacuole. Three-dimensional modeling of the predicted proteins shows pronounced differences in surface charge of PmAPX1 and PmAPX2 in the vicinity of the aperture that provides access to the heme and active site. CONCLUSIONS: PmAPX1 and PmAPX2 phylogenetic analysis suggests that they are derived from a plant ancestor. Plant ancestry is further supported by the presence of ascorbate synthesis genes in the P. marinus genome that are similar to those in plants. The localizations and 3D structures of the two APX isoforms suggest that APX fulfills multiple functions in P. marinus within two compartments. The possible role of APX in free-living and parasitic stages of the life history of P. marinus is discussed.


Assuntos
Antioxidantes/metabolismo , Ascorbato Peroxidases/metabolismo , Catalase/metabolismo , Parasitos/enzimologia , Fotossíntese , Sequência de Aminoácidos , Animais , Ascorbato Peroxidases/química , Ascorbato Peroxidases/genética , Ascorbato Peroxidases/isolamento & purificação , Peróxido de Hidrogênio/metabolismo , Cinética , Modelos Moleculares , Parasitos/genética , Filogenia , Homologia Estrutural de Proteína , Frações Subcelulares/metabolismo
16.
Chemosphere ; 233: 905-912, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31340418

RESUMO

We investigated the interconnected roles of reactive oxygen species (ROS) generated upon seed exposure to glyphosate and/or gibberellic acid (GA3), and the possible interaction between the herbicide and the plant hormone during germination of sorghum seeds. GA3 decreased antioxidant enzyme activity in embryos, and the over accumulation of hydrogen peroxide (H2O2) in 1000 mM GA3-treated seeds resulted in the lowest germinability among treatments. The deleterious effects of glyphosate on germination rate, in contrast, were not related to H2O2 accumulation, but to its interference with the mitochondrial electron transport chain. However, interactions among glyphosate, GA3 and H2O2 during seed germination were observed. Similar to paclobutrazol, glyphosate appears to interfere with the de novo synthesis of gibberellin, which modulates seed germination through oxidative metabolism. Seeds experiencing increased oxidative status due to GA3 (100 mM) or H2O2 (50 mM) applications had the effects of glyphosate on germination rate reversed. Since decreased ATP synthesis is a secondary effect of glyphosate, increased H2O2 concentrations in embryos must facilitate germination by decreasing the energy required by ATP-demanding metabolism. Our results showed that glyphosate affect seed germination of sorghum, and that the herbicide interacts with oxidative and gibberellin metabolisms.


Assuntos
Germinação/efeitos dos fármacos , Giberelinas/metabolismo , Glicina/análogos & derivados , Herbicidas/farmacologia , Peróxido de Hidrogênio/metabolismo , Reguladores de Crescimento de Planta/metabolismo , Sorghum/metabolismo , Antioxidantes/metabolismo , Grão Comestível/metabolismo , Transporte de Elétrons/efeitos dos fármacos , Glicina/farmacologia , Sementes/efeitos dos fármacos
17.
Food Chem ; 298: 125093, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31260960

RESUMO

The purpose of this study was to investigate the impact of ozonation process on the level of oxidative stress markers in raspberries stored at room temperature. Raspberry fruit was ozonated with an ozone concentration of 8-10 ppm for 30 min, every 12 h, for 72 h of storage at room temperature. Research showed that ozonated raspberries were characterized by higher activity of superoxide dismutase, ascorbate peroxidase and phenylalanine ammonia-lyase. In turn, the ability to generate superoxide anion radical and hydrogen peroxide by ozone-treated fruit was significantly lower than in the control sample due to higher activities of ROS detoxification systems.


Assuntos
Armazenamento de Alimentos/métodos , Frutas/metabolismo , Estresse Oxidativo , Ozônio/química , Rubus/metabolismo , Antioxidantes/química , Antioxidantes/metabolismo , Ascorbato Peroxidases/metabolismo , Biomarcadores/química , Biomarcadores/metabolismo , Frutas/química , Peróxido de Hidrogênio/química , Peróxido de Hidrogênio/metabolismo , Fenilalanina Amônia-Liase/metabolismo , Espécies Reativas de Oxigênio/química , Espécies Reativas de Oxigênio/metabolismo , Rubus/química , Superóxido Dismutase/metabolismo , Superóxidos/química , Superóxidos/metabolismo , Temperatura Ambiente
18.
Food Chem ; 299: 125116, 2019 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-31295637

RESUMO

The effects of exogenous melatonin treatment on the enzymatic browning and nutritional quality of fresh-cut pear fruit were investigated. Fresh-cut fruit soaked with 0, 0.05, 0.1 and 0.5 mM melatonin were stored at 4 °C. Our results showed that 0.1 mM melatonin treatment was optimal for reducing the surface browning and maintaining the titratable acidity of the fresh-cut fruit, which significantly decreased MDA and H2O2 contents and the growth of microorganism, enhanced total phenolic content and antioxidant capacity, and delayed the reduction of ascorbic acid. Furthermore, melatonin treatment at 0.1 mM decreased the expression of genes involving in enzymatic browning pathway including POD, PPO1, PPO5 and LOX1, and reduced PPO activity. Moreover, this treatment increased the expression of PAL and CHS, and enhanced PAL and CHS activities. These results showed that melatonin treatment might be a promising strategy to alleviate browning and improve the nutritional quality of fresh-cut pear fruit.


Assuntos
Frutas/efeitos dos fármacos , Melatonina/farmacologia , Valor Nutritivo , Pyrus/efeitos dos fármacos , Antioxidantes/análise , Antioxidantes/metabolismo , Ácido Ascórbico/análise , Ácido Ascórbico/metabolismo , Armazenamento de Alimentos , Frutas/química , Frutas/metabolismo , Frutas/microbiologia , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Peróxido de Hidrogênio/análise , Peróxido de Hidrogênio/metabolismo , Malondialdeído/análise , Malondialdeído/metabolismo , Fenóis/análise , Pyrus/química , Pyrus/genética , Pyrus/metabolismo
19.
Ann Hematol ; 98(9): 2045-2052, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31243572

RESUMO

Thalassemia has a high prevalence in Thailand. Oxidative damage to erythroid cells is known to be one of the major etiologies in thalassemia pathophysiology. Oxidative stress status of thalassemia is potentiated by the heme, nonheme iron, and free iron resulting from imbalanced globin synthesis. In addition, levels of antioxidant proteins are reduced in α-thalassemia and ß-thalassemia erythrocytes. However, the primary molecular mechanism for this phenotype remains unknown. Our study showed a high expression of miR-144 in ß- and α-thalassemia. An increased miR-144 expression leads to decreased expression of nuclear factor erythroid 2-related factor 2 (NRF2) target, especially in α-thalassemia. In α-thalassemia, miR-144 and NRF2 target are associated with glutathione level and anemia severity. To study the effect of miR-144 expression, the gain-loss of miR-144 expression was performed by miR inhibitor and mimic transfection in the erythroblastic cell line. This study reveals that miR-144 expression was upregulated, whereas NRF2 expression and glutathione levels were decreased in comparison with the untreated condition after miR mimic transfection, while the reduction of miR-144 expression contributed to the increased NRF2 expression and glutathione level compared with the untreated condition after miR inhibitor transfection. Moreover, miR-144 overexpression leads to significantly increased sensitivity to oxidative stress at indicated concentrations of hydrogen peroxide (H2O2) and rescued by miR-144 inhibitor. Taken together, our findings suggest that dysregulation of miR-144 may play a role in the reduced ability of erythrocyte to deal with oxidative stress and increased RBC hemolysis susceptibility especially in thalassemia.


Assuntos
Eritrócitos/metabolismo , MicroRNAs/biossíntese , Fator 2 Relacionado a NF-E2/biossíntese , Estresse Oxidativo , Regulação para Cima , Talassemia alfa/metabolismo , Talassemia beta/metabolismo , Eritrócitos/patologia , Feminino , Glutationa/biossíntese , Glutationa/genética , Hemólise , Humanos , Peróxido de Hidrogênio/metabolismo , Células K562 , Masculino , MicroRNAs/genética , Fator 2 Relacionado a NF-E2/genética , Talassemia alfa/genética , Talassemia alfa/patologia , Talassemia beta/genética , Talassemia beta/patologia
20.
Microbiol Res ; 223-225: 99-109, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31178057

RESUMO

Streptococcus suis has received increasing attention for its involvement in severe infections in pigs and humans; however, their pathogenesis remains unclear. ClpX and ClpP, two subunits of the ATP-dependent caseinolytic protease Clp, play key roles in bacterial adaptation to various environmental stresses. In this study, a virulent S. suis serotype 2 strain, ZY05719, was employed to construct clpX and clpP deletion mutants (ΔclpX and ΔclpP, respectively) and their complementation strains. Both ΔclpX and ΔclpP displayed significantly reduced adaptability compared with the wild-type strain, evident through several altered phenotypes: formation of long cell chains, tendency to aggregate in culture, and reduced growth under acidic pH and H2O2-induced oxidative stress. ClpP and ClpX were required for the optimal growth during heat and cold stress, respectively. An in vitro experiment on RAW264.7 macrophage cells showed significantly increased sensitivity of ΔclpX and ΔclpP to phagocytosis compared with the wild-type strain. Mouse infection assays verified the deletion of clpX and clpP led to not only fewer clinical symptoms and lower mortality but also to a marked attenuation in bacterial colonization. These virulence-related phenotypes were restored by genetic complementation. Furthermore, the deletion of clpX or clpP caused a significant decrease in the expression of sodA, tpx, and apuA compared with the wild-type strain, suggesting that these genes may be regulated by ClpX and ClpP as downstream response factors to facilitate the bacterial tolerance against various environmental stresses. Taken together, these results suggest that ClpX and ClpP play important roles in stress tolerance for achieving the full virulence of S. suis serotype 2 during infection.


Assuntos
ATPases Associadas a Diversas Atividades Celulares/metabolismo , Proteínas de Bactérias/metabolismo , Endopeptidase Clp/metabolismo , Chaperonas Moleculares/metabolismo , Streptococcus suis/metabolismo , Streptococcus suis/patogenicidade , Animais , Proteínas de Bactérias/genética , Biofilmes , Resposta ao Choque Frio , Endopeptidase Clp/genética , Deleção de Genes , Regulação Bacteriana da Expressão Gênica , Resposta ao Choque Térmico , Peróxido de Hidrogênio/metabolismo , Concentração de Íons de Hidrogênio , Camundongos , Chaperonas Moleculares/genética , Pressão Osmótica , Estresse Oxidativo , Fagocitose , Células RAW 264.7 , Infecções Estreptocócicas/microbiologia , Streptococcus suis/genética , Streptococcus suis/crescimento & desenvolvimento , Transcriptoma , Virulência/genética , Fatores de Virulência/genética , Fatores de Virulência/fisiologia
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