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1.
Chem Biol Interact ; 311: 108798, 2019 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-31433962

RESUMO

Natural products are a valuable source of anticancer agents, with many naturally derived compounds currently used in clinical and preclinical treatments. This study aims to investigate the antiproliferative activity and potential mechanism of action of the xanthoquinodin JBIR-99, isolated from fungi Parengyodontium album MEXU 30,054 and identified by single-crystal X-ray crystallography. Cytotoxicity of xanthoquinodin was evaluated in a panel of human cancer cells lines and CCD-112-CoN normal colon cells, using the sulforhodamine B assay. PC-3 prostate cancer cells were used in biochemical assays including cell cycle, mitochondrial transmembrane potential (MTP), reactive oxygen species (ROS) and caspase activity. Expression levels of apoptosis-pathway-related proteins were analyzed by Western blot. The in vivo toxicity of xanthoquinodin was determined using a zebrafish model. Xanthoquinodin showed cytotoxicity in all cancer cell lines but demonstrated relative selective potency against PC-3 cells with an IC50 1.7 µM. In CCD-112-CoN cells, xanthoquinodin was non-cytotoxic at 100 µM. In PC-3 cells, the compound induced loss of MTP, production of ROS, and cell cycle arrest in S phase. The expression and activity of caspase-3 was increased, which correlates with the upregulation of Cyt c, Bax, nuclear factor kappa-B (NF-κB) (p65) and IKKß, and downregulation of poly ADP ribose polymerase (PARP-1) and Bcl-2. Lastly, xanthoquinodin did not cause any visible developmental toxicity in zebrafish at 50 µM. These results demonstrate xanthoquinodin induces apoptosis in PC-3 prostate cancer cells by activation of both intrinsic and extrinsic apoptotic pathways. In addition, the non-toxic effect in vivo indicates that xanthoquinodin could be a useful lead in the development of a novel, anti-cancer agent that is selective for prostate cancer.


Assuntos
Apoptose/efeitos dos fármacos , Ascomicetos/química , Cromonas/farmacologia , Ascomicetos/metabolismo , Linhagem Celular Tumoral , Cromonas/química , Cristalografia por Raios X , Citocromos c/metabolismo , Humanos , Quinase I-kappa B/metabolismo , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Conformação Molecular , Poli(ADP-Ribose) Polimerase-1/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Repressoras/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
2.
BMC Ophthalmol ; 19(1): 168, 2019 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-31375076

RESUMO

BACKGROUND: Age-related macular degeneration (AMD) is the primary cause of blindness and severe vision loss in developed countries and is responsible for 8.7% of blindness globally. Ultraviolet radiation can induce DNA breakdown, produce reactive oxygen species, and has been implicated as a risk factor for AMD. This study investigated the effects of UVA radiation on Human retinal pigment epithelial cell (ARPE-19) growth and protein expression. METHODS: ARPE-19 cells were irradiated with a UVA lamp at different doses (5, 10, 20, 30 and 40 J/cm2) from 10 cm. Cell viability was determined by MTT assay. Visual inspection was first achieved with inverted light microscopy and then the DeadEnd™ Fluorometric TUNEL System was used to observe nuclear DNA fragmentation. Flow cytometry based-Annexin V-FITC/PI double-staining was used to further quantify cellular viability. Mitochondrial membrane potential was assessed with JC-1 staining. 2D electrophoresis maps of exposed cells were compared to nonexposed cells and gel images analyzed with PDQuest 2-D Analysis Software. Spots with greater than a 1.5-fold difference were selected for LC-MS/MS analysis and some confirmed by western blot. We further investigated whether caspase activation, apoptotic-related mitochondrial proteins, and regulators of ER stress sensors were involved in UVA-induced apoptosis. RESULTS: We detected 29 differentially expressed proteins (9 up-regulated and 20 down-regulated) in the exposed cells. Some of these proteins such as CALR, GRP78, NPM, Hsp27, PDI, ATP synthase subunit alpha, PRDX1, and GAPDH are associated with anti-proliferation, induction of apoptosis, and oxidative-stress protection. We also detected altered protein expression levels among caspases (caspase 3 and 9) and in the mitochondrial (cytosolic cytochrome C, AIF, Mcl-1, Bcl-2, Bcl-xl, Bax, Bad, and p-Bad) and ER stress-related (p-PERK, p-eIF2α, ATF4 and CHOP) apoptotic pathways. CONCLUSIONS: UVA irradiation suppressed the proliferation of ARPE-19 cells in a dose-dependent manner, caused quantitative loses in transmembrane potential (ΔΨm), and induced both early and late apoptosis.


Assuntos
Degeneração Macular/patologia , Estresse Oxidativo , Proteômica/métodos , Epitélio Pigmentado da Retina/metabolismo , Raios Ultravioleta , Apoptose , Sobrevivência Celular , Células Cultivadas , Cromatografia Líquida , Citocromos c/metabolismo , Humanos , Degeneração Macular/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Epitélio Pigmentado da Retina/patologia , Epitélio Pigmentado da Retina/efeitos da radiação , Transdução de Sinais , Espectrometria de Massas em Tandem
3.
Anticancer Res ; 39(7): 3687-3695, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31262895

RESUMO

BACKGROUND: Neuroblastoma is the main solid extracranial tumor of childhood. The amplification of N-myc oncogene (MYCN) and 1p deletion are the main molecular alterations. These features are what make treatment impossible, especially in high-risk patients with metastases. MATERIALS AND METHODS: Our study investigated the processes undergone by CHP-212 neuroblastoma cells, after being treated with Casiopeínas® (Cas) IIgly, IIIEa, and IIIia for 2, 10, and 24 h. RESULTS: At 2 h, all the treatments Ied to apoptosis [defined by the presence of B-cell lymphoma 2 (BCL2), BCL2-associated X protein, cytochrome c, and caspase-3]. In addition, autophagy with specific molecules beclin-1 and microtubule-associated protein 1A/1B-light chain 3 (LC3)-II/LC3-I (ratio >1). Later at 10 h, autophagy-associated proteins were observed, and at 24 h, only survival proteins nuclear factor kappa-light-chain-enhancer of activated B-cells (NF-κB), and extracellular signal-regulated kinases (ERK)2/ERK1>1 were found. Another relevant finding was the presence of caspase-10 throughout the study, especially in cells treated with CasIIgly and CasIIIEa. CONCLUSION: These relationships indicate a possible mechanism of action of Casiopeínas on neuroblastoma.


Assuntos
Complexos de Coordenação/farmacologia , Neuroblastoma/metabolismo , Fenantrolinas/farmacologia , Autofagia/efeitos dos fármacos , Proteína Beclina-1/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo , Neuroblastoma/tratamento farmacológico , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo
4.
Chem Biol Interact ; 309: 108723, 2019 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-31228469

RESUMO

Ischemic preconditioning and pharmacological preconditioning are common strategies to prevent lethal myocardial injury, especially nutritional preconditioning (NPC). In this study, we investigated the effects of astragaloside IV (Ast), as an NPC agent, on myocardium suffered anoxia/reoxygenation (A/R) injury. Rats received 5 mg/kg Ast daily for 3 weeks by intragastric administration. Then, hearts were harvested and underwent A/R treatment using a Langendorff apparatus. Ast- pretreatment significantly promoted functional recovery of the myocardium, reduced infarct size, and oxidative stress, and decreased the apoptotic index. Similar findings were demonstrated in H9c2 cardiomyocytes that were pretreated with Ast for 24 h. Moreover, Ast-pretreatment significantly upregulated Bcl-2 expression, especially in mitochondria. The effects of Ast treatment against A/R injury were also reflected by increased antioxidant potential, inhibited reactive oxygen species (ROS) burst, increased oxygen consumption rate, maintained mitochondrial membrane potential (MMP), inhibited mitochondrial permeability transition pore (mPTP) opening, and prevented apoptosis. Selective inhibition of Bcl-2 by ABT-737 decreased myocardial injury protection of Ast. Ast-pretreatment resulted in NPC- related effects against A/R, and mitochondria may be the target of a cascade of events elicited by upregulating Bcl-2 expression, promoting translocation of Bcl-2 into mitochondria, maintaining MMP, inhibiting ROS bursts, thereby leading to recovery of mitochondrial respiration, preventing mPTP opening, decreasing cytochrome C release, preventing apoptosis, and ultimately alleviating myocardial injury.


Assuntos
Mitocôndrias/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Saponinas/farmacologia , Triterpenos/farmacologia , Animais , Antioxidantes/química , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Compostos de Bifenilo/farmacologia , Compostos de Bifenilo/uso terapêutico , Caspase 3/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/efeitos dos fármacos , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/metabolismo , Miócitos Cardíacos/citologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Nitrofenóis/farmacologia , Nitrofenóis/uso terapêutico , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Superóxido Dismutase/metabolismo
5.
Yonsei Med J ; 60(6): 509-516, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31124333

RESUMO

PURPOSE: This study was conducted to verify the induction and mechanism of selective apoptosis in G361 melanoma cells using anti-HER2 antibody-conjugated gold nanoparticles (GNP-HER2). MATERIALS AND METHODS: Following GNP-HER2 treatment of G361 cells, cell cycle arrest and apoptosis were measured by WST-1 assay, Hemacolor staining, Hoechst staining, immunofluorescence staining, fluorescence-activated cell sorting analysis, and Western blotting. RESULTS: G361 cells treated with GNP-HER2 showed condensation of nuclei, which is an apoptotic phenomenon, and translocation of apoptosis-inducing factor and cytochrome c from mitochondria into the nucleus and cytoplasm, respectively. Increases in BAX in cells undergoing apoptosis, activation of caspase-3 and -9, and fragmentation of poly (ADP-ribose) polymerase and DNA fragmentation factor 45 (inhibitor of caspase-activated DNase) were observed upon GNP-HER2 treatment. Following GNP-HER2 treatment, an increase of cells in sub-G1 phase, which is a signal of cell apoptosis, was observed. This resulted in the down-regulation of cyclin A, cyclin D1, cyclin E, cdk2, cdk4, and cdc2 and the up-regulation of p21. Thus, GNP-HER2 treatment was confirmed to induce the cessation of cell cycle progression. Also, decreases in phospho-focal adhesion kinase and phospho-human epidermal growth factor receptor, which activate cellular focal adhesion, and decreases in phospho-paxillin, which stimulates the disassembly of filamentous actin, were observed. Reduced cell adhesion and disassembly of the intracellular structure indicated cell deactivation. CONCLUSION: GNP-HER2 can selectively kill G361 melanoma cells without affecting normal cells. The mechanism of G361 cell death upon treatment with GNP-HER2 was apoptosis accompanied by activation of caspases.


Assuntos
Anticorpos/metabolismo , Apoptose , Ouro/química , Melanoma/patologia , Nanopartículas Metálicas/química , Receptor ErbB-2/metabolismo , Actinas/metabolismo , Fator de Indução de Apoptose/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Forma Celular , Citocromos c/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Adesões Focais/metabolismo , Fase G1 , Humanos , Nanopartículas Metálicas/ultraestrutura
6.
Mol Med Rep ; 19(6): 4852-4862, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059068

RESUMO

Cerebrovascular disease (CVD) is one of the leading causes of mortality worldwide. The role of cytochrome c oxidase subunit 6B1 (COX6B1) in the central nervous system remains unclear. The present study aimed to analyze the role of COX6B1 in rat hippocampal neurons extracted from fetal rats. The subcellular localization of the neuron­specific marker microtubule­associated protein 2 was detected by immunofluorescence assay. Cell viability was assessed using a cell counting kit, and the levels of apoptosis and cytosolic Ca2+ were analyzed by flow cytometry. The expression levels of the molecular factors downstream to COX6B1 were determined using reverse transcription­quantitative polymerase chain reaction and western blotting. Reoxygenation following oxygen­glucose deprivation (OGD) decreased cell viability and the expression levels of COX6B1 in a time­dependent manner, and 60 min of reoxygenation was identified as the optimal time period for establishing an ischemia/reperfusion (I/R) model. Overexpression of COX6B1 was demonstrated to reverse the viability of hippocampal neurons following I/R treatment. Specifically, COX6B1 overexpression decreased the cytosolic concentration of Ca2+ and suppressed neuronal apoptosis, which were increased following I/R treatment. Furthermore, overexpression of COX6B1 increased the protein expression levels of apoptosis regulator BCL­2 and mitochondrial cytochrome c (cyt c), and decreased the protein expression levels of apoptosis regulator BCL2­associated X and cytosolic cyt c in I/R model cells. Collectively, the present study results suggested that COX6B1 overexpression may reverse I/R­induced neuronal damage by increasing the viability of neurons, by decreasing the cytosolic levels of Ca2+ and by suppressing apoptosis. These results may facilitate the development of novel strategies for the prevention and treatment of CVD.


Assuntos
Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/farmacologia , Neurônios/metabolismo , Traumatismo por Reperfusão/prevenção & controle , Lobo Temporal/metabolismo , Animais , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Transtornos Cerebrovasculares/prevenção & controle , Citocromos c/metabolismo , Complexo IV da Cadeia de Transporte de Elétrons/genética , Feminino , Glucose/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Neurônios/patologia , Oxigênio/metabolismo , Substâncias Protetoras/metabolismo , Substâncias Protetoras/farmacologia , Ratos , Ratos Sprague-Dawley , Lobo Temporal/patologia , Proteína bcl-X/metabolismo
7.
Chemosphere ; 230: 182-189, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31103864

RESUMO

Spinosad is one of the most extensively used bio-pesticide in the world. The effects of pesticide in human health are mainly associated with its residue in food or occupational exposure in agricultural production. The lung is the direct target of pesticides exposure, although the study of inhalation damage caused by Spinosad remains unclear. The aim of the present study was to evaluate the cytotoxic effects of the Spinosad in human lung cells. We demonstrated that Spinosad could inhibite the proliferation of human lung epithelial A549 cells, induce the DNA damage and enhance the programmed cell death. Intracellular biochemical assay indicated that DNA double strand breaks, cleaved of PARP, release of cytochrome c, decrease of mitochondrial membrane potential, generation of reactive oxygen species (ROS), activation of caspase-3/9, increase of Bax/Bcl-2 ratio, LC3-II conversion, accumulation of Beclin-1, degradation of p62 and the changes in the phosphorylation of AMPK, mTOR are contributed to the toxic effects of Spinosad in A549 cells. The results showed that the cytotoxicity of Spinosad may be associated with the activity of mitochondrial apoptotic pathways or AMPK/mTOR-mediated autophagy. Meanwhile, the DNA stand breaks caused by the Spinosad suggest it has a potential genotoxic effects on human lung cells. We conclude that Spinosad has a potential risk to human health by inducing the cytotoxic effects.


Assuntos
Macrolídeos/toxicidade , Praguicidas/toxicidade , Testes de Toxicidade , Células A549 , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Proteína Beclina-1 , Caspase 3 , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Combinação de Medicamentos , Humanos , Pulmão/metabolismo , Neoplasias Pulmonares/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2 , Espécies Reativas de Oxigênio/metabolismo , Serina-Treonina Quinases TOR
8.
Chem Biol Interact ; 308: 11-19, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31071336

RESUMO

The main aim of this study was to investigate the effects of phloretin loaded chitosan nanoparticles (PhCsNPs) on 7,12-dimethylbenz[a]anthracene (DMBA) induced experimental cancer in hamsters. Oral squamous cell carcinoma (OSCC) was induced in male golden Syrian hamsters by painting with 0.5% DMBA three times a week for 14 weeks. Varying concentration of PhCsNPs (5, 10, and 20 mg/kg b.wt.) was orally administered on alternative days to evaluate the optimum dose. The experiment design was terminated at the end of the 14th week. The development of OSCC was confirmed by histopathological and biochemical analysis (lipid peroxidation, antioxidant profile, and detoxification enzymes) in plasma, erythrocyte, buccal, and liver tissues. Significant increases in oxidation and lipid peroxidation were noticed in DMBA-painted hamsters. Oral administration of PhCsNPs in various doses on alternate days reversed the deleterious effects induced by DMBA. In addition, immunoblot analyses of PhCsNPs treatment enhanced the release of Bcl-2 associated X protein (Bax), cytochrome c, caspase-3, 9 and suppressed the B-cell lymphoma 2 (Bcl-2) expression, which the use of PhCsNPs for mitochondrial-mediated apoptosis. These findings suggest biofabricated PhCsNPs may act as a potent antioxidant and anti-carcinogenic in DMBA induced oral cancer in experimental animals.


Assuntos
Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Quitosana/química , Nanopartículas/química , Floretina/farmacologia , Administração Oral , Animais , Carcinoma de Células Escamosas/induzido quimicamente , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/patologia , Caspase 3/metabolismo , Cricetinae , Sistema Enzimático do Citocromo P-450/metabolismo , Citocromos c/metabolismo , Regulação para Baixo/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Neoplasias Bucais/induzido quimicamente , Neoplasias Bucais/tratamento farmacológico , Neoplasias Bucais/patologia , Floretina/química , Floretina/uso terapêutico , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína X Associada a bcl-2/metabolismo
9.
Eur J Med Chem ; 176: 117-128, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31108261

RESUMO

A series of novel xanthine/NO donor hybrids containing 1,3,8-trisubstituted or 1,8-disubstituted xanthine derivatives were designed and synthesized. The synthesized compounds were tested in a cell viability assay using human mammary gland epithelial cell line (MCF-10A) where all the compounds exhibited no cytotoxic effects and more than 90% cell viability at a concentration of 50 µM. The oxime containing compounds 7a-b and 17-24 were more active as antiproliferative agents than their non-oxime congeners 6a-b and 9-16. Hydroxyimino-phenethyl scaffold compounds 17-24 were more active than the hydroxyimino-ethyl phenyl acetamide 7a-b derivatives. Compounds 18-20 and 22-24 exhibited inhibition of EGFR with IC50 ranging from 0.32 to 2.88 µM. Compounds 18-20 and 22-24 increased the level of active caspase 3 by 4-8 folds, compared to the control cells in Panc-1 cell lines compared to doxorubicin as a reference drug. Compounds 18, 22 and 23 were the most caspase-3 inducers. Compounds 22 and 23 increased the levels of caspase-8 and 9 indicating activation of both intrinsic and extrinsic pathways and showed potent induction of Bax, down-regulation of Bcl-2 protein levels and over-expression of cytochrome c levels in Panc-1 human pancreas cancer cells. Compound 23 exhibited mainly cell cycle arrest at the Pre-G1 and G2/M phases in the cell cycle analysis of Panc-1 cell line. The drug likeness profiles of compounds 18-20 and 22-24 were predicted to have good to excellent drug likeness profiles specially compounds 18-20 and 23. Finally molecular docking study was performed at the EGFR active site to suggest thier possible binding mode. The hydroxyimino-phenethyl scaffold compounds 17-24 represent an interesting starting point to optimize their pharmacokinetics and pharmacodynamics profiles.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Oximas/farmacologia , Xantinas/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/toxicidade , Caspase 3/metabolismo , Caspase 8/metabolismo , Caspase 9/metabolismo , Domínio Catalítico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/química , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Oximas/síntese química , Oximas/química , Oximas/toxicidade , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Relação Estrutura-Atividade , Xantinas/síntese química , Xantinas/química , Xantinas/toxicidade , Proteína X Associada a bcl-2/metabolismo
10.
Mediators Inflamm ; 2019: 4050796, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31065234

RESUMO

Mitochondrial dysfunction has been established as a common feature of neurodegenerative disorders that contributes to disease pathology by causing impaired cellular energy production. Mitochondrial molecules released into the extracellular space following neuronal damage or death may also play a role in these diseases by acting as signaling molecules called damage-associated molecular patterns (DAMPs). Mitochondrial DAMPs have been shown to initiate proinflammatory immune responses from nonneuronal glial cells, including microglia and astrocytes; thereby, they have the potential to contribute to the chronic neuroinflammation present in these disorders accelerating the degeneration of neurons. In this review, we highlight the mitochondrial DAMPs cytochrome c (CytC), mitochondrial transcription factor A (TFAM), and cardiolipin and explore their potential role in the central nervous system disorders including Alzheimer's disease and Parkinson's disease, which are characterized by neurodegeneration and chronic neuroinflammation.


Assuntos
Inflamação/imunologia , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Doenças Neurodegenerativas/imunologia , Animais , Citocromos c/metabolismo , Proteínas de Ligação a DNA/metabolismo , Humanos , Inflamação/metabolismo , Proteínas Mitocondriais/metabolismo , Doenças Neurodegenerativas/metabolismo , Fatores de Transcrição/metabolismo
11.
Chemosphere ; 226: 463-471, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30951941

RESUMO

Tetrabromobisphenol A (TBBPA) is a commonly used brominated flame retardant, which has a wide range of toxic effects on organisms. This study investigated the cytotoxic effects on human hepatocytes (L02 cells) after treated with 0, 5, 10, 20, and 40 µM of TBBPA. Results showed that TBBPA significantly increased intracellular reactive oxygen species (ROS), malondialdehyde (MDA) and the ratio of oxidized/reduced glutathione (GSSG/GSH) dose-dependently. TBBPA also decreased the cell mitochondrial membrane potential (MMP), caused the release of cytochrome C (Cyt C) to cytoplasm and promoted the expression of caspase-9 and caspase-3, and finally increased the level of apoptosis. The ROS inhibitor N-acetyl-L-cysteine (NAC) relieved the oxidative stress responses, and prevented the decrease of MMP and increase of apoptosis. In addition, TBBPA promoted the expression of antioxidant genes related to Nrf2, such as quinone oxidoreductase 1 (NQO1), catalase (CAT), and heme oxygenase 1 (HO-1). Oxidative stress initiated by TBBPA, activated mitochondrial apoptosis and Nrf2 pathway, and increased the degree of apoptosis in L02 cells.


Assuntos
Apoptose/efeitos dos fármacos , Retardadores de Chama/toxicidade , Mitocôndrias/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Bifenil Polibromatos/toxicidade , Acetilcisteína/farmacologia , Caspase 3/biossíntese , Caspase 9/biossíntese , Catalase/metabolismo , Linhagem Celular , Citocromos c/metabolismo , Glutationa/metabolismo , Heme Oxigenase-1/biossíntese , Heme Oxigenase-1/genética , Hepatócitos/efeitos dos fármacos , Humanos , Malondialdeído/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , NAD(P)H Desidrogenase (Quinona)/biossíntese , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos
12.
Bioelectrochemistry ; 128: 94-99, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30959399

RESUMO

We report on a hybrid bioelectrochemical system that integrates an energy converting part, viz. a glucose/oxygen enzymatic fuel cell, with a charge-storing component, in which the redox features of the immobilized redox protein cytochrome c (cyt c) were utilized. Bilirubin oxidase and pyrroloquinoline quinone-dependent glucose dehydrogenase (PQQ-GDH) were employed as the biocatalysts for dioxygen reduction and glucose oxidation, respectively. A bi-protein PQQ-GDH/cyt c signal chain was created that facilitates electron transfer between the enzyme and the electrode surface. The assembled supercapacitor/biofuel cell hybrid biodevice displays a 15 times higher power density tested in the pulse mode compared to the performance achieved from the continuously operating regime (4.5 and 0.3 µW cm-2, respectively) with an 80% residual activity after 50 charge/discharge pulses. This can be considered as a notable step forward in the field of glucose/oxygen membrane-free, biocompatible hybrid power sources.


Assuntos
Fontes de Energia Bioelétrica , Citocromos c/metabolismo , Enzimas Imobilizadas/metabolismo , Glucose Desidrogenase/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , Técnicas Eletroquímicas/instrumentação , Eletrodos , Transporte de Elétrons , Glucose/metabolismo , Oxirredução
13.
Nat Commun ; 10(1): 1689, 2019 04 11.
Artigo em Inglês | MEDLINE | ID: mdl-30976076

RESUMO

Gasdermin E (GSDME/DFNA5) cleavage by caspase-3 liberates the GSDME-N domain, which mediates pyroptosis by forming pores in the plasma membrane. Here we show that GSDME-N also permeabilizes the mitochondrial membrane, releasing cytochrome c and activating the apoptosome. Cytochrome c release and caspase-3 activation in response to intrinsic and extrinsic apoptotic stimuli are significantly reduced in GSDME-deficient cells comparing with wild type cells. GSDME deficiency also accelerates cell growth in culture and in a mouse model of melanoma. Phosphomimetic mutation of the highly conserved phosphorylatable Thr6 residue of GSDME, inhibits its pore-forming activity, thus uncovering a potential mechanism by which GSDME might be regulated. Like GSDME-N, inflammasome-generated gasdermin D-N (GSDMD-N), can also permeabilize the mitochondria linking inflammasome activation to downstream activation of the apoptosome. Collectively, our results point to a role of gasdermin proteins in targeting the mitochondria to promote cytochrome c release to augment the mitochondrial apoptotic pathway.


Assuntos
Inflamassomos/metabolismo , Melanoma Experimental/patologia , Mitocôndrias/fisiologia , Piroptose/fisiologia , Receptores Estrogênicos/metabolismo , Neoplasias Cutâneas/patologia , Animais , Caspase 3/metabolismo , Citocromos c/metabolismo , Fibroblastos , Técnicas de Inativação de Genes , Células HEK293 , Células HeLa , Humanos , Macrófagos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Membranas Mitocondriais/metabolismo , Mutação , Fosforilação/fisiologia , Cultura Primária de Células , Domínios Proteicos/genética , Receptores Estrogênicos/genética , Treonina/metabolismo
14.
Mar Drugs ; 17(4)2019 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-31013932

RESUMO

Phloroglucinol (PG) is a component of phlorotannins, which are abundant in marine brown alga species. Recent studies have shown that PG is beneficial in protecting cells from oxidative stress. In this study, we evaluated the protective efficacy of PG in HaCaT human skin keratinocytes stimulated with oxidative stress (hydrogen peroxide, H2O2). The results showed that PG significantly inhibited the H2O2-induced growth inhibition in HaCaT cells, which was associated with increased expression of heme oxygenase-1 (HO-1) by the activation of nuclear factor erythroid 2-related factor-2 (Nrf2). PG remarkably reversed H2O2-induced excessive ROS production, DNA damage, and apoptosis. Additionally, H2O2-induced mitochondrial dysfunction was related to a decrease in ATP levels, and in the presence of PG, these changes were significantly impaired. Furthermore, the increases of cytosolic release of cytochrome c and ratio of Bax to Bcl-2, and the activation of caspase-9 and caspase-3 by the H2O2 were markedly abolished under the condition of PG pretreatment. However, the inhibition of HO-1 function using zinc protoporphyrin, a HO-1 inhibitor, markedly attenuated these protective effects of PG against H2O2. Overall, our results suggest that PG is able to protect HaCaT keratinocytes against oxidative stress-induced DNA damage and apoptosis through activating the Nrf2/HO-1 signaling pathway.


Assuntos
Apoptose/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Heme Oxigenase-1/metabolismo , Queratinócitos/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/metabolismo , Floroglucinol/farmacologia , Substâncias Protetoras/farmacologia , Trifosfato de Adenosina/metabolismo , Antioxidantes/farmacologia , Caspase 3/metabolismo , Linhagem Celular , Citocromos c/metabolismo , Humanos , Peróxido de Hidrogênio/farmacologia , Queratinócitos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos
15.
Artif Cells Nanomed Biotechnol ; 47(1): 1248-1255, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30945565

RESUMO

Mitochondrial dysfunction is a major contributory factor for myocardial ischemia-reperfusion (I/R) injury. It has been reported that Pink1-Parkin-mediated mitochondrial autophagy could effectively remove damaged mitochondria and excess ROS to ensure the stability of intracellular mitochondria. The present study was designed to evaluate whether the polymerized porcine haemoglobin (pPolyHb), a novel type of haemoglobin oxygen carrier, has an effect on I/R injury via regulating the Pink1-Parkin mediated mitochondrial autophagy pathway in myocardial H9C2 cells. The results revealed that pPolyHb could effectively reduce apoptosis and improve the survival rates of H9C2 cells. In addition, Pink1 and Parkin levels gradually decreased with pPolyHb reoxygenation. The inhibition of mitochondrial autophagy through mitochondrial-division inhibitor-1(mdivi-1) resulted in a decrease in anti-apoptotic protein Bcl-2 and an increase in pro-apoptotic protein Bax and CytC. In conclusion, pPolyHb has a protective effect on myocardial ischemia-reperfusion injury by regulating moderate mitochondrial autophagy.


Assuntos
Autofagia/efeitos dos fármacos , Hemoglobinas/farmacologia , Mitocôndrias/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/patologia , Miócitos Cardíacos/efeitos dos fármacos , Proteínas Quinases/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Citoproteção/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Hemoglobinas/química , Humanos , L-Lactato Desidrogenase/metabolismo , Mitocôndrias/patologia , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Oxigênio/metabolismo , Multimerização Proteica , Estrutura Quaternária de Proteína , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Suínos , Proteína X Associada a bcl-2/metabolismo
16.
Neurochem Res ; 44(7): 1653-1664, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30949935

RESUMO

Neuroinflammation has been acknowledged as a primary factor contributing to the pathogenesis of neurodegenerative disease. However, the molecular mechanism underlying inflammation stress-mediated neuronal dysfunction is not fully understood. The aim of our study was to explore the influence of mammalian STE20-like kinase 1 (Mst1) in neuroinflammation using TNFα and CATH.a cells in vitro. The results of our study demonstrated that the expression of Mst1 was dose-dependently increased after TNFα treatment. Interestingly, knockdown of Mst1 using siRNA transfection significantly repressed TNFα-induced neuronal death. We also found that TNFα treatment was associated with mitochondrial stress, including mitochondrial ROS overloading, mitochondrial permeability transition pore (mPTP) opening, mitochondrial membrane potential reduction, and mitochondrial pro-apoptotic factor release. Interestingly, loss of Mst1 attenuated TNFα-triggered mitochondrial stress and sustained mitochondrial function in CATH.a cells. We found that Mst1 modulated mitochondrial homeostasis and cell viability via the JNK pathway in a TNFα-induced inflammatory environment. Inhibition of the JNK pathway abolished TNFα-mediated CATH.a cell death and mitochondrial malfunction, similar to the results obtained via silencing of Mst1. Taken together, our results indicate that inflammation-mediated neuronal dysfunction is implicated in Mst1 upregulation, which promotes mitochondrial stress and neuronal death by activating the JNK pathway. Accordingly, our study identifies the Mst1-JNK-mitochondria axis as a novel signaling pathway involved in neuroinflammation.


Assuntos
Inflamação/fisiopatologia , Sistema de Sinalização das MAP Quinases/genética , Mitocôndrias/metabolismo , Estresse Oxidativo/fisiologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Antracenos/farmacologia , Apoptose/fisiologia , Linhagem Celular Tumoral , Citocromos c/metabolismo , Relação Dose-Resposta a Droga , Técnicas de Silenciamento de Genes , Inflamação/induzido quimicamente , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Doenças Neurodegenerativas/induzido quimicamente , Doenças Neurodegenerativas/fisiopatologia , Estresse Oxidativo/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Fator de Necrose Tumoral alfa
17.
Carbohydr Polym ; 215: 99-107, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-30981376

RESUMO

Suaeda salsa, is an annual herbaceous plant that contains various bio-functional macromolecules. Herein, an acidic polysaccharide from Suaeda salsa, denoted as SSP2-2, with a molecular weight of 53.8 kDa was isolated, which composed of mannose, rhamnose, glucuronic acid, galacturonic acid, galactose and xylose in a molar ratio of 0.6: 8.0: 1.0: 83.6: 5.0: 7.2. An MTT assay showed that SSP2-2 induced apoptosis of MCF-7 cells in a dose-dependent manner in vitro. Morphological analysis and flow cytometry experiments indicated that SSP2-2 promotes MCF-7 cells death via apoptosis, while JC-1 staining results revealed that mitochondrial membrane potential was reduced in a dose-dependent manner. The data from the western blot showed an increase in the levels of Bax, cytochrome C (Cyto-c), caspase-3 and caspase-9 and a decrease in the level of Bcl-2 further demonstrated that SSP2-2 could induce apoptosis via a mitochondrial pathway. These results suggest that SSP2-2 can potentially be used as an antitumor agent.


Assuntos
Antineoplásicos , Apoptose/efeitos dos fármacos , Chenopodiaceae/química , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Polissacarídeos , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Caspase 3/metabolismo , Caspase 9/metabolismo , Citocromos c/metabolismo , Humanos , Células MCF-7 , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Polissacarídeos/química , Polissacarídeos/isolamento & purificação , Polissacarídeos/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína X Associada a bcl-2/metabolismo
18.
Pestic Biochem Physiol ; 156: 63-71, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31027582

RESUMO

Phenazine-1-carboxylic acid (PCA), a secondary metabolite produced by Pseudomonas spp., exhibits a high inhibitory effect in Xanthomonas oryzae pv. oryzae (Xoo), but less inhibitory effect in Xanthomonas oryzae pv. oryzicola (Xoc), and almost no inhibitory effect in Xanthomonas campestris pv. campestris (Xcc). In our previous study, reactive oxygen species (ROS) scavenging system was reported to be involved in PCA tolerance in Xanthomonas spp. However, the PCA tolerance mechanism of Xanthomonas spp. is unclear. In the current study, we constructed a Tn5-based transposon mutant library in Xcc and four highly PCA-sensitive insertion mutants were obtained. TAIL-PCR further confirmed that the Tn5 transposon was inserted in the cytochrome c maturation (CCM) system (XC_1893, XC_1897) of these mutants. Disruption of the CCM system significantly decreased the growth, motility and tolerance of Xcc to PCA and other phenazines, such as phenazine and 1-OH-phenazine. The CCM system is responsible for the covalent attachment of the apocytochrome and heme. Disruption of the transmembrane thioredox protein (Dsb) pathway (XC_0531), an essential process for the formation of mature apocytochrome, also exhibited a decreased tolerance to PCA, suggesting that the defect of cytochrome c caused decreased tolerance of Xcc to PCA. Meanwhile, disruption of the CCM system or Dsb pathway interfered with the functions of cytochrome c proteins, causing an increased sensitivity to H2O2. Collectively, we concluded that the CCM system and Dsb pathway, regulate the tolerance of Xcc to phenazines by influencing the functions of cytochrome c. Therefore, these results provide important references for revealing the action mechanism of PCA in Xanthomonas spp.


Assuntos
Proteínas de Bactérias/metabolismo , Citocromos c/metabolismo , Fenazinas/farmacologia , Xanthomonas campestris/efeitos dos fármacos , Xanthomonas campestris/metabolismo , Mutação/genética , Espécies Reativas de Oxigênio/metabolismo
19.
Med Sci Monit ; 25: 1976-1983, 2019 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-30877718

RESUMO

BACKGROUND Nasopharyngeal carcinoma results in high patient morbidity and mortality, due to early metastasis, and toxicity due to chemotherapy. Mukonal is plant-derived carbazole alkaloid that has been used in traditional Chinese medicine to treat several types of cancer. This study aimed to investigate the effects of mukonal on cell proliferation, apoptosis, autophagy, and the mitochondrial membrane potential of nasopharyngeal carcinoma cells in vitro. MATERIAL AND METHODS CNE1 human nasopharyngeal carcinoma cells and NP69 normal nasopharyngeal epithelial cells were cultured with and without treatment with increasing doses of mukonal. Cell viability was determined by the MTT assay. Fluorescence microscopy was used to detect reactive oxygen species (ROS), mitochondrial membrane potential, and the release of cytochrome C. Flow cytometry was used to examine changes in the cell cycle, electron microscopy examined cell autophagy, and Western blot was performed to measure levels of proteins associated with autophagy and apoptosis. RESULTS Mukonal had an antiproliferative effect on CNE1 cells, with an IC50 of 9 µM and there were effects of toxicity on normal NP69 cells. Mukonal triggered ROS-mediated changes in mitochondrial membrane potential which was also accompanied by the discharge of cytochrome C in the CNE1 cells. Mukonal activated autophagy and apoptosis in CNE1 cells, which was also associated with upregulation of the autophagy-related proteins, LC3 II and beclin-1, as well as apoptosis-associated proteins, Bax, cleaved caspase-3 and -9. Mukonal treatment also resulted in CNE1 cells cycle arrest at G2/M. CONCLUSIONS Mukonal inhibited the growth of human CNE1 nasopharyngeal carcinoma cells in vitro.


Assuntos
Carbazóis/farmacologia , Carcinoma Nasofaríngeo/tratamento farmacológico , Extratos Vegetais/uso terapêutico , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Proteína Beclina-1/metabolismo , Carcinoma/tratamento farmacológico , Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , China , Citocromos c/metabolismo , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Carcinoma Nasofaríngeo/patologia , Espécies Reativas de Oxigênio/metabolismo
20.
Chemistry ; 25(37): 8760-8768, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-30908783

RESUMO

We report the first electrochemical study of a lanthanoid-dependent methanol dehydrogenase (Eu-MDH) from the acidophilic verrucomicrobial methanotroph Methylacidiphilum fumariolicum SolV with its own physiological cytochrome cGJ electron acceptor. Eu-MDH harbours a redox active 2,7,9-tricarboxypyrroloquinoline quinone (PQQ) cofactor which is non-covalently bound but coordinates trivalent lanthanoid elements including Eu3+ . Eu-MDH and the cytochrome were co-adsorbed with the biopolymer chitosan and cast onto a mercaptoundecanol (MU) monolayer modified Au working electrode. Cyclic voltammetry of cytochrome cGJ reveals a well-defined quasi-reversible FeIII/II redox couple at +255 mV vs. NHE at pH 7.5 and this response is pH independent. The reversible one-electron response of the cytochrome cGJ transforms into a sigmoidal catalytic wave in the presence of Eu-MDH and its substrates (methanol or formaldehyde). The catalytic current was pH-dependent and pH 7.3 was found to be optimal. Kinetic parameters (pH dependence, activation energy) obtained by electrochemistry show the same trends as those obtained from an artificial phenazine ethosulfate/dichlorophenol indophenol assay.


Assuntos
Oxirredutases do Álcool/metabolismo , Citocromos c/química , Európio/química , Oxirredutases do Álcool/química , Biocatálise , Domínio Catalítico , Citocromos c/metabolismo , Técnicas Eletroquímicas , Eletrodos , Cinética , Metanol/química , Metanol/metabolismo , Oxirredução , Cofator PQQ/química , Cofator PQQ/metabolismo , Espectrofotometria , Especificidade por Substrato , Temperatura Ambiente , Verrucomicrobia/enzimologia
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