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1.
J Agric Food Chem ; 67(33): 9265-9276, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31361479

RESUMO

Fungal infections significantly alter the emissions of volatile organic compounds (VOCs) by plants, but the mechanisms for VOCs affecting fungal infections of plants remain largely unknown. Here, we found that infection by Botrytis cinerea upregulated linalool production by strawberries and fumigation with linalool was able to inhibit the infection of fruits by the fungus. Linalool treatment downregulated the expression of rate-limiting enzymes in the ergosterol biosynthesis pathway, and this reduced the ergosterol content in the fungi cell membrane and impaired membrane integrity. Linalool treatment also caused damage to mitochondrial membranes by collapsing mitochondrial membrane potential and also downregulated genes involved in adenosine triphosphate (ATP) production, resulting in a significant decrease in the ATP content. Linalool treatment increased the levels of reactive oxygen species (ROS), in response to which the treated fungal cells produced more of the ROS scavenger pyruvate. RNA-Seq and proteomic analysis data showed that linalool treatment slowed the rates of transcription and translation.


Assuntos
Botrytis/efeitos dos fármacos , Fragaria/metabolismo , Frutas/microbiologia , Monoterpenos/metabolismo , Doenças das Plantas/microbiologia , Compostos Orgânicos Voláteis/metabolismo , Trifosfato de Adenosina/metabolismo , Botrytis/crescimento & desenvolvimento , Fragaria/química , Fragaria/microbiologia , Frutas/química , Frutas/metabolismo , Interações Hospedeiro-Patógeno , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Monoterpenos/farmacologia , Doenças das Plantas/prevenção & controle , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteômica , Espécies Reativas de Oxigênio/metabolismo , Compostos Orgânicos Voláteis/farmacologia
2.
Life Sci ; 231: 116587, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31220526

RESUMO

The endoplasmic reticulum (ER) and mitochondria are two important organelles in cells. Mitochondria-associated membranes (MAMs) are lipid raft-like domains formed in the ER membranes that are in close apposition to mitochondria. They play an important role in signal transmission between these two essential organelles. When cells are exposed to internal or external stressful stimuli, the ER will activate an adaptive response called the ER stress response, which has a significant effect on mitochondrial function. Mitochondrial quality control is an important mechanism to ensure the functional integrity of mitochondria and the effect of ER stress on mitochondrial quality control through MAMs is of great significance. Therefore, in this review, we introduce ER stress and mitochondrial quality control, and discuss how ER stress signals are transmitted to mitochondria through MAMs. We then review the important roles of MAMs in mitochondrial quality control under ER stress.


Assuntos
Estresse do Retículo Endoplasmático/fisiologia , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Animais , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Sinalização do Cálcio , Retículo Endoplasmático/metabolismo , Humanos , Microdomínios da Membrana , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/fisiologia
3.
Med Sci Monit ; 25: 2935-2942, 2019 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-31005958

RESUMO

BACKGROUND Thyroid cancer causes considerable mortality and morbidity across the globe. Owing to the unavailability of biomarkers and the adverse effects of existing drugs, there is an urgent need to develop efficient chemotherapy for the treatment of thyroid cancers. Plants have served as exceptional source of drugs for the treatment of lethal diseases. The purpose of this study was to evaluate the anticancer effects of ferruginol against thyroid cancer cells. MATERIAL AND METHODS We monitored the cell proliferation rate using 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis was detected using 4',6-diamidino-2-phenylindole (DAPI), acridine orange/ethidium bromide (AO/EB), and annexin V/propidium iodide (PI) staining. Reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) levels were examined by fluorescence microscopy. Protein expressed was examined by western blotting. RESULTS We found that ferruginol exerted potent antiproliferative action against thyroid cancer cells, and an IC50 of 12 µM was observed for ferruginol against the MDA-T32 cell line. The toxic effects of ferruginol were less pronounced against normal cells. The anticancer effects of ferruginol were likely due to the induction of apoptosis which was also associated with upregulation of Bax and downregulation of Bcl-2. Ferruginol also caused ROS mediated alterations in the MMP of MDA-T32 cells. In MDA-T32 cells, ferruginol might also block the MAPK and PI3K/AKT signaling pathway, which is believed to be an important therapeutic target of anticancer drugs. CONCLUSIONS In conclusion, in view of the results of this study, it might be suggested that ferruginol might serve as an essential lead molecule for the treatment of thyroid cancer provided further in-depth studies especially studying ferruginol toxicological as well as in vivo studies are needed.


Assuntos
Diterpenos de Abietano/farmacologia , Neoplasias da Glândula Tireoide/tratamento farmacológico , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , China , Diterpenos/metabolismo , Diterpenos/farmacologia , Diterpenos de Abietano/metabolismo , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Membranas Mitocondriais/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinases/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Extratos Vegetais/farmacologia , Proteínas Proto-Oncogênicas c-akt/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos
4.
Biomed Pharmacother ; 111: 859-872, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30841465

RESUMO

This study investigated the effects of post-treatment with kolaviron on a 2-Vessel Occlusion (2-VO) model of cerebral ischemia/reperfusion (I/R) injury in rats to ascertain its level of efficacy as a potential therapeutic agent for stroke. Male Wistar rats submitted to 30 min of bilateral common carotid artery occlusion and 24 h of reperfusion were treated with kolaviron (25-100 mg/kg) or 20 mg/kg quercetin immediately after reperfusion and 2 h post reperfusion. At the end of the period of reperfusion, animals were scored for motor and cognitive deficits. Brain relative weight and water content were determined. Cortices, striata and hippocampi were dissected and processed for estimation of markers of oxidative stress, inflammation, neurotransmitter dysregulation and excitotoxicity. In addition, assessment of hippocampal mitochondrial integrity and histopathological examination of the cortical, striatal and hippocampal regions were carried out. There was reversal of 2-VO ischemia/reperfusion (I/R) induced motor and cognitive deficits by kolaviron post-treatment. Post-treatment with kolaviron also attenuated I/R-induced oxidative stress, neuroinflammatory events, excitotoxicity as well as mitochondrial dysfunction in brain tissues. Histopathological findings showed amelioration of I/R-induced neuronal cell damage by kolaviron post-treatment. The results revealed the multi-target neurotherapeutic activity of kolaviron and suggest that it is a promising candidate for drug development against stroke.


Assuntos
Isquemia Encefálica/tratamento farmacológico , Transporte de Elétrons/efeitos dos fármacos , Flavonoides/farmacologia , Mitocôndrias/efeitos dos fármacos , Neurotransmissores/metabolismo , Oxirredução/efeitos dos fármacos , Traumatismo por Reperfusão/tratamento farmacológico , Animais , Isquemia Encefálica/metabolismo , Modelos Animais de Doenças , Masculino , Mitocôndrias/metabolismo , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Fármacos Neuroprotetores/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Wistar , Traumatismo por Reperfusão/metabolismo , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/metabolismo
5.
Nat Commun ; 10(1): 688, 2019 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-30737374

RESUMO

Corticosteroids are host-directed drugs with proven beneficial effect on survival of tuberculosis (TB) patients, but their precise mechanisms of action in this disease remain largely unknown. Here we show that corticosteroids such as dexamethasone inhibit necrotic cell death of cells infected with Mycobacterium tuberculosis (Mtb) by facilitating mitogen-activated protein kinase phosphatase 1 (MKP-1)-dependent dephosphorylation of p38 MAPK. Characterization of infected mixed lineage kinase domain-like (MLKL) and tumor necrosis factor receptor 1 (TNFR1) knockout cells show that the underlying mechanism is independent from TNFα-signaling and necroptosis. Our results link corticosteroid function and p38 MAPK inhibition to abrogation of necrotic cell death mediated by mitochondrial membrane permeability transition, and open new avenues for research on novel host-directed therapies (HDT).


Assuntos
Corticosteroides/farmacologia , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/metabolismo , Dexametasona/farmacologia , Humanos , Fosforilação/efeitos dos fármacos , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
6.
Lipids Health Dis ; 18(1): 53, 2019 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-30764880

RESUMO

BACKGROUND: Supplemented fatty acids can incorporate into cardiolipin (CL) and affect its remodeling. The change in CL species may alter the mitochondrial membrane composition, potentially disturbing the mitochondrial structure and function during inflammation. METHOD: To investigate the effect of the unsaturation of fatty acids on CL, we supplemented macrophage-like RAW264.7 cells with 18-carbon unsaturated fatty acids including oleic acid (OA, 18:1), linoleic acid (LA, 18:2), α-linolenic acid (ALA, 18:3), γ-linolenic acid (GLA, 18:3), and stearidonic acid (SDA, 18:4). Mitochondrial changes in CL were measured through mass spectrometry. RESULT: Our data indicated that OA(18:1) was the most efficient fatty acid that incorporated into CL, forming symmetrical CL without fatty acid elongation and desaturation. In addition, LA(18:2) and ALA(18:3) were further elongated before incorporation, significantly increasing the number of double bonds and the chain length of CL. GLA and SDA were not optimal substrates for remodeling enzymes. The findings of RT-qPCR experiments revealed that none of these changes in CL occurred through the regulation of CL remodeling- or synthesis-related genes. The fatty acid desaturase and transportation genes-Fads2 and Cpt1a, respectively-were differentially regulated by the supplementation of five unsaturated 18-carbon fatty acids. CONCLUSIONS: The process of fatty acid incorporation to CL was regulated by the fatty acid desaturation and transportation into mitochondria in macrophage. The double bonds of fatty acids significantly affect the incorporation process and preference. Intact OA(18:1) was incorporated to CL; LA(18:2) and ALA(18:3) were desaturated and elongated to long chain fatty acid before the incorporation; GLA(18:3) and SDA(18:4) were unfavorable for the CL incorporation.


Assuntos
Cardiolipinas/biossíntese , Ácidos Graxos Ômega-3/farmacologia , Ácido Linoleico/farmacologia , Membranas Mitocondriais/efeitos dos fármacos , Ácido Oleico/farmacologia , Ácido alfa-Linoleico/farmacologia , Ácido gama-Linolênico/farmacologia , Animais , Transporte Biológico , Carnitina O-Palmitoiltransferase/genética , Carnitina O-Palmitoiltransferase/metabolismo , Ácidos Graxos Dessaturases/genética , Ácidos Graxos Dessaturases/metabolismo , Ácidos Graxos Ômega-3/química , Ácidos Graxos Ômega-3/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Ácido Linoleico/química , Ácido Linoleico/metabolismo , Camundongos , Mitocôndrias/química , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Membranas Mitocondriais/química , Membranas Mitocondriais/metabolismo , Ácido Oleico/química , Ácido Oleico/metabolismo , Células RAW 264.7 , Relação Estrutura-Atividade , Ácido alfa-Linoleico/química , Ácido alfa-Linoleico/metabolismo , Ácido gama-Linolênico/química , Ácido gama-Linolênico/metabolismo
7.
Mol Cell ; 73(5): 1028-1043.e5, 2019 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-30733118

RESUMO

Mutations in PTEN-induced kinase 1 (PINK1) can cause recessive early-onset Parkinson's disease (PD). Import arrest results in PINK1 kinase activation specifically on damaged mitochondria, triggering Parkin-mediated mitophagy. Here, we show that PINK1 import is less dependent on Tim23 than on mitochondrial membrane potential (ΔΨm). We identified a negatively charged amino acid cluster motif that is evolutionarily conserved just C-terminal to the PINK1 transmembrane. PINK1 that fails to accumulate at the outer mitochondrial membrane, either by mutagenesis of this negatively charged motif or by deletion of Tom7, is imported into depolarized mitochondria and cleaved by the OMA1 protease. Some PD patient mutations also are defective in import arrest and are rescued by the suppression of OMA1, providing a new potential druggable target for PD. These results suggest that ΔΨm loss-dependent PINK1 import arrest does not result solely from Tim23 inactivation but also through an actively regulated "tug of war" between Tom7 and OMA1.


Assuntos
Proteínas de Membrana/metabolismo , Metaloendopeptidases/metabolismo , Mitocôndrias/enzimologia , Membranas Mitocondriais/enzimologia , Proteínas Mitocondriais/metabolismo , Doença de Parkinson/enzimologia , Proteínas Quinases/metabolismo , Motivos de Aminoácidos , Antiparkinsonianos/farmacologia , Transporte Biológico , Desenho de Drogas , Ativação Enzimática , Células HeLa , Humanos , Potencial da Membrana Mitocondrial , Proteínas de Membrana/genética , Metaloendopeptidases/genética , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Proteínas de Transporte da Membrana Mitocondrial/genética , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Membranas Mitocondriais/efeitos dos fármacos , Proteínas Mitocondriais/genética , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/genética , Domínios e Motivos de Interação entre Proteínas , Proteínas Quinases/genética , Proteólise , Transdução de Sinais , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
8.
Clinics (Sao Paulo) ; 73: e113, 2018 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-29972436

RESUMO

OBJECTIVES: The objective of the present study was to evaluate the protective effect of pre-conditioning treatment with laser light on hepatic injury in rats submitted to partial ischemia using mitochondrial function and liver fatty acid binding protein as markers. METHODS: Rats were divided into four groups (n=5): 1) Control, 2) Control + Laser, 3) Partial Ischemia and 4) Partial Ischemia + Laser. Ischemia was induced by clamping the hepatic pedicle of the left and middle lobes of the liver for 60 minutes. Laser light at 660 nm was applied to the liver immediately prior to the induction of ischemia at 22.5 J/cm2, with 30 seconds of illumination at five individual points. The animals were sacrificed after 30 minutes of reperfusion. Blood and liver tissues were collected for analysis of mitochondrial function, determination of malondialdehyde and analysis of fatty acid binding protein expression by Western blot. RESULTS: Mitochondrial function decreased in the Partial Ischemia group, especially during adenosine diphosphate-activated respiration (state 3), and the expression of fatty acid binding protein was also reduced. The application of laser light prevented bioenergetic changes and restored the expression of fatty acid binding protein. CONCLUSION: Prophylactic application of laser light to the livers of rats submitted to partial ischemia was found to have a protective effect in the liver, with normalization of both mitochondrial function and fatty acid binding protein tissue expression.


Assuntos
Proteínas de Ligação a Ácido Graxo/metabolismo , Precondicionamento Isquêmico/métodos , Fígado/irrigação sanguínea , Fígado/efeitos da radiação , Terapia com Luz de Baixa Intensidade/métodos , Traumatismo por Reperfusão/prevenção & controle , Alanina Transaminase/sangue , Animais , Aspartato Aminotransferases/sangue , Western Blotting , Proteínas de Ligação a Ácido Graxo/análise , Fígado/metabolismo , Malondialdeído/análise , Malondialdeído/efeitos da radiação , Membranas Mitocondriais/efeitos dos fármacos , Dilatação Mitocondrial/efeitos da radiação , Ratos Wistar , Reprodutibilidade dos Testes
9.
Int J Mol Sci ; 19(7)2018 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-30011897

RESUMO

Platinum(II) drugs are activated intracellularly by aquation of the leaving groups and then bind to DNA, forming DNA adducts capable to activate various signal-transduction pathways. Mostly explored in recent years are Pt(IV) complexes which allow the presence of two additional ligands in the axial positions suitable for the attachment of other cancer-targeting ligands. Here we have extended this strategy by coordinating in the axial positions of kiteplatin ([PtCl2(cis-1,4-DACH)], DACH = Diaminocyclohexane) and its CBDCA (1,1-cyclobutanedicarboxylate) analogue the antioxidant α-Lipoic acid (ALA), an inhibitor of the mitochondrial pyruvate dehydrogenase kinase (PDK). The new compounds (cis,trans,cis-[Pt(CBDCA)(ALA)2(cis-1,4-DACH)], 2, and cis,trans,cis-[PtCl2(ALA)2(cis-1,4-DACH)], 3), after intracellular reduction, release the precursor Pt(II) species and two molecules of ALA. The Pt residue is able to target DNA, while ALA could act on mitochondria as activator of the pyruvate dehydrogenase complex, thus suppressing anaerobic glycolysis. Compounds 2 and 3 were tested in vitro on a panel of five human cancer cell lines and compared to cisplatin, oxaliplatin, and kiteplatin. They proved to be much more effective than the reference compounds, with complex 3 most effective in 3D spheroid tumor cultures. Notably, treatment of human A431 carcinoma cells with 2 and 3 did not determine increase of cellular ROS (usually correlated to inhibition of mitochondrial PDK) and did not induce a significant depolarization of the mitochondrial membrane or alteration of other morphological mitochondrial parameters.


Assuntos
Mitocôndrias/metabolismo , Compostos Organoplatínicos/metabolismo , Platina/metabolismo , Pró-Fármacos/metabolismo , Ácido Tióctico/metabolismo , Antineoplásicos/química , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cisplatino/química , Cisplatino/metabolismo , Cisplatino/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Células MCF-7 , Mitocôndrias/efeitos dos fármacos , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Estrutura Molecular , Compostos Organoplatínicos/química , Compostos Organoplatínicos/farmacologia , Oxaliplatina , Platina/química , Platina/farmacologia , Pró-Fármacos/química , Pró-Fármacos/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Ácido Tióctico/química , Ácido Tióctico/farmacologia
10.
Biomed Pharmacother ; 105: 1062-1071, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30021342

RESUMO

Seventeen endophytic fungi were isolated from various tissues of Cassia fistula and the ethyl acetate extracts obtained from 21-day cultures of all the endophytic fungal isolates were initially screened for their cytotoxicity against HeLa (human cervical carcinoma) cells using MTT assay. Of these, Penicillium sclerotiorum extract (PSE), significantly affected the viability of HeLa cells in a dose-dependent manner. The extract of P. Sclerotiorum was further analyzed by GC-MS, which showed three compounds, hexadecanoic acid, oleic acid and benzoic acid to be the major active principles in the extracts.The extract was further tested for invitro cytotoxicity against five cancer cell lines. Of the cell lines tested, HeLa cells showed maximum sensitivity followed by A549, while A431 and U251 were moderately sensitive and MCF-7 was insensitive to the treatment. In addition, normal human embryonic kidney cells, HEK293 remained insensitive to the treatment. Furthermore, the mechanism of cytotoxic activity exhibited by PSE was investigated by evaluating cell cycle progression and apoptotic induction in HeLa cells. Cell cycle analysis revealed that the PSE arrested cells at S and G2/M phase of the cell cycle in a dose-dependent manner. Annexin V- Propidium iodide double staining showed that, the extract potentiates apoptosis rather than necrosis in cells. This was supported by the down regulation in the proapoptotic protein BCL2 and up regulation of BAX (BCL2 Associated X), tumor suppressor protein, p53 and Apaf-1 [Apoptotic Peptidase Activating Factor 1]. Loss of mitochondrial membrane potential and a distinct DNA fragmentation pattern observed following the treatment, suggest that the PSE treatment leads to activation of mitochondrial pathway of apoptosis. Further, the extract also exhibited both antioxidant and anti-angiogenic properties. These results indicate that endophytic fungi isolated from medicinal plants may serve as potential sources of the anti-cancerous compounds.


Assuntos
Apoptose/efeitos dos fármacos , Cassia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Endófitos/efeitos dos fármacos , Membranas Mitocondriais/efeitos dos fármacos , Penicillium , Neoplasias do Colo do Útero/metabolismo , Células A549 , Apoptose/fisiologia , Pontos de Checagem do Ciclo Celular/fisiologia , Relação Dose-Resposta a Droga , Endófitos/fisiologia , Feminino , Células HEK293 , Células HeLa , Humanos , Células MCF-7 , Membranas Mitocondriais/fisiologia , Penicillium/isolamento & purificação , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Neoplasias do Colo do Útero/tratamento farmacológico
11.
Chem Biol Interact ; 291: 192-201, 2018 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-29935161

RESUMO

Brominated flame retardants are used in consumer goods to increase product resistance to fire and/or high temperatures. Polybrominated diphenyl ethers (PBDEs) are the most commonly employed class of brominated flame retardants because they are inexpensive and can effectively prevent flame from spreading. PBDEs are persistent, can bioaccumulate, are transported over long distances, and display toxicity. However, their toxic mechanisms of action have not been well established. Because mitochondria are recognized as the main energy-producing cell organelle and play a vital role in cellular function maintenance, here we apply mitochondria as an experimental model to evaluate the toxic effects of the PBDE congener BDE-153 (Hexa-BDE) at concentrations ranging from 0.1 to 25 µM. We also assess BDE-153 cytotoxicity to HepG2 cells in order to elucidate its mechanisms of toxicity. Exposure to BDE-153 affects isolated mitochondria: this congener can interact with the mitochondrial membrane, to dissipate the membrane potential and to induce significant ATP depletion. Furthermore, BDE-153 can diminish MTT reduction and cell proliferation and can interfere in cell cycle, as evaluated in cell cultures. These cytotoxic effects are related to mitochondrial dysfunction due to mitochondrial membrane potential dissipation and reactive oxygen species accumulation. These effects result in apoptotic cell death, as demonstrated by phosphatidylserine maintenance on the cell membrane external surface, nuclear condensation and fragmentation, and presence of pro-apoptotic factors such as cytochrome c and Apoptosis-inducing Factor (AIF) plus caspase 3 activation in the cytosol. Together, our results show PBDEs can induce cytotoxicity, reinforcing the idea that these compounds pose a risk to the exposed population.


Assuntos
Apoptose/efeitos dos fármacos , Fígado/patologia , Mitocôndrias Hepáticas/patologia , Bifenil Polibromatos/toxicidade , Trifosfato de Adenosina/metabolismo , Animais , Cálcio/metabolismo , Células Hep G2 , Humanos , Fígado/efeitos dos fármacos , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias Hepáticas/efeitos dos fármacos , Mitocôndrias Hepáticas/metabolismo , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Dilatação Mitocondrial/efeitos dos fármacos , Fosforilação Oxidativa/efeitos dos fármacos , Fosfatidilserinas/metabolismo , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo
12.
Int J Oncol ; 53(3): 1257-1268, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29956777

RESUMO

Aged garlic extract (AGE) has been shown to possess therapeutic properties in cancer; however its mechanisms of action are unclear. In this study, we demonstrate by MTT assay that AGE exerts an anti-proliferative effect on a panel of both sensitive and multidrug-resistant (MDR) human cancer cell lines and enhances the effects of hyperthermia (42˚C) on M14 melanoma cells. The evaluation of the mitochondrial activity in whole cancer cells treated with AGE, performed by cytofluorimetric analysis in the presence of the lipophilic cationic fluorochrome JC-1, revealed the occurrence of dose-dependent mitochondrial membrane depolarization. Membrane potential was measured by the TPP+ selective electrode. In order to shed light on its mechanisms of action, the effects of AGE on isolated rat liver mitochondria were also examined. In this regard, AGE induced a mitochondrial membrane hyperpolarization of approximately 15 mV through a mechanism that was similar to that observed with the ionophores, nigericin or salinomycin, by activating an exchange between endogenous K+ with exogenous H+. The prolonged incubation of the mitochondria with AGE induced depolarization and matrix swelling, indicative of mitochondrial permeability transition induction that, however, occurs through a different mechanism from the well-known one. In particular, the transition pore opening induced by AGE was due to the rearrangement of the mitochondrial membranes following the increased activity of the K+/H+ exchanger. On the whole, the findings of this study indicate that AGE exerts cytotoxic effects on cancer cells by altering mitochondrial permeability. In particular, AGE in the mitochondria activates K+/H+ exchanger, causes oxidative stress and induces mitochondrial permeability transition (MPT).


Assuntos
Antioxidantes/farmacologia , Alho/química , Membranas Mitocondriais/efeitos dos fármacos , Neoplasias/terapia , Extratos Vegetais/farmacologia , Animais , Antioxidantes/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Terapia Combinada/métodos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Humanos , Hipertermia Induzida/métodos , Ionóforos/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Mitocôndrias Hepáticas/efeitos dos fármacos , Mitocôndrias Hepáticas/metabolismo , Membranas Mitocondriais/metabolismo , Neoplasias/patologia , Estresse Oxidativo/efeitos dos fármacos , Permeabilidade/efeitos dos fármacos , Extratos Vegetais/uso terapêutico , Antiportadores de Potássio-Hidrogênio/metabolismo , Ratos , Ratos Wistar
13.
PLoS One ; 13(5): e0196805, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29723246

RESUMO

Blasts from different patients with acute myeloid leukemia (AML) vary in the agent(s) to which they are most responsive. With a myriad of novel agents to evaluate, there is a lack of predictive biomarkers to precisely assign targeted therapies to individual patients. Primary AML cells often survive poorly in vitro, thus confounding conventional cytotoxicity assays. The purpose of this work was to assess the potential of two same-day functional predictive assays in AML cell lines to predict long-term response to chemotherapy. (i) Ribosomal protein S6 (rpS6) is a downstream substrate of PI3K/akt/mTOR/ kinase and MAPK kinase pathways and its dephosphorylation is also triggered by DNA double strand breaks. Phospho-rpS6 is reliably measurable by flow cytometry and thus has the potential to function as a biomarker of responsiveness to several therapeutic agents. (ii) A cell's propensity for apoptosis can be interrogated via a functional assay termed "Dynamic BH3 Profiling" in which mitochondrial outer membrane permeabilization in drug-treated cells can be driven by pro-apoptotic BH3 domain peptides such as PUMA-BH3. The extent to which a particular cell is primed for apoptosis by the drug can be determined by measuring the amount of cytochrome C released on addition of BH3 peptide. We demonstrate that phospho-rpS6 expression and PUMA-BH3 peptide-induced cytochrome C release after 4 hours both predict long term chemoresponsiveness to tyrosine kinase inhibitors and DNA double strand break inducers in AML cell lines. We also describe changes in expression levels of the prosurvival BCL-2 family member Mcl-1 and the pro-apoptotic protein BIM after short term drug culture.


Assuntos
Apoptose/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Leucemia Mieloide Aguda/metabolismo , Proteínas de Neoplasias/metabolismo , Processamento de Proteína Pós-Traducional , Proteína S6 Ribossômica/metabolismo , Transdução de Sinais/efeitos dos fármacos , Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose/farmacologia , Proteína 11 Semelhante a Bcl-2/biossíntese , Proteína 11 Semelhante a Bcl-2/genética , Biomarcadores , Linhagem Celular Tumoral , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Humanos , Leucemia Mieloide Aguda/patologia , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Proteína de Sequência 1 de Leucemia de Células Mieloides/biossíntese , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Proteínas de Neoplasias/antagonistas & inibidores , Fragmentos de Peptídeos/farmacologia , Permeabilidade/efeitos dos fármacos , Fosforilação , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas/farmacologia , Proteínas Quinases S6 Ribossômicas/metabolismo
14.
Mol Med Rep ; 18(2): 1710-1717, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29845198

RESUMO

The mitochondrial pyruvate carrier (MPC) complex, located on the inner mitochondrial membrane, transports pyruvate to the mitochondrial matrix for oxidative phosphorylation. Previous studies have shown that the MPC complex is a key regulator of glycolysis in tumor cells. The present study evaluated the role of the MPC under hypoxic conditions in human umbilical vein endothelial cells, which rely on glycolysis for energy generation. It was indicated that hypoxia led to an increase in lactate secretion and a decrease in MPC1 and MPC2 levels, which were upregulated following re­oxygenation. In addition, the knockdown of MPC1 or treatment with the MPC inhibitor UK5099 increased the levels of glycolytic enzymes, HK2, PFKFB3, and LDHA, promoting glycolysis and lactate secretion. Taken together, the present data revealed that hypoxia can induce lactate secretion and glycolytic efflux by downregulating MPC levels.


Assuntos
Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Proteínas de Transporte da Membrana Mitocondrial/genética , Oxigênio/farmacologia , Acrilatos/farmacologia , Hipóxia Celular , Glicólise/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Lactato Desidrogenases/genética , Lactato Desidrogenases/metabolismo , Ácido Láctico/biossíntese , Mitocôndrias/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/antagonistas & inibidores , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Fosfofrutoquinase-2/genética , Fosfofrutoquinase-2/metabolismo , Ácido Pirúvico/metabolismo
15.
Biomed Pharmacother ; 102: 823-832, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29605770

RESUMO

Doxorubicin (Dox) is a widely administered chemotherapeutic drug and incidences of cardiotoxicity associated with its administration have been of general concern. Extensive research proposes several mechanisms as a cause of Dox induced cardiotoxicity. However, none of these studies have been able to suggest a find one, cure all antidote for the same. To this end, several studies involving plant based compounds or natural products have gained acclaim for their ability to address at least one factor contributing to drug induced pathogenesis. We had previously reported that p-coumaric (pCA) has a protective effect on Dox induced oxidative stress in rat-derived cardiomyoblasts. In this study we investigated the effects of pCA on the regulation of Nrf-2, mitochondrial viability, autophagy and apoptosis in Doxorubicin treated H9c2 cardiomyocytes. ROS induced mitochondrial stress, changes in mitochondrial membrane potential, loss of membrane integrity; nuclear damage as single/double stranded breaks, autophagy and the effects of pre and co-treatment of pCA on Nrf-2 mediated signaling was evaluated by various approaches. The effect of pCA on drug uptake was evaluated through confocal Raman Spectroscopy. We find that nuclear translocation of Nrf-2 is prominently marked by protein-specific antibody conjugated fluorophore in Dox treated cells especially. Cell survival is mediated to a certain extent by the expression of the anti-apoptotic BCl2 in pCA treated cells. However, mRNA levels of autophagy related (Atg) genes suggest that autophagy plays a decisive role in deciding cellular fate. Caspase-3 activation is also observed in pCA treated cells which suggest an alternative function of caspase-3 in pCA mediated cell survival. Expression of antioxidant enzymes confirm the oxidative stress induced by Dox treatment in cells and the modulation of cell redox homeostasis through treatment with pCA.


Assuntos
Autofagia , Cardiotoxicidade/tratamento farmacológico , Cardiotoxicidade/patologia , Ácidos Cumáricos/farmacologia , Citoproteção/efeitos dos fármacos , Doxorrubicina/efeitos adversos , Fator 2 Relacionado a NF-E2/metabolismo , Animais , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/genética , Autofagia/efeitos dos fármacos , Autofagia/genética , Cardiotoxicidade/genética , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Citoproteção/genética , Dano ao DNA/genética , Regulação da Expressão Gênica/efeitos dos fármacos , L-Lactato Desidrogenase/metabolismo , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Ratos , Análise Espectral Raman , Coloração e Rotulagem
16.
Part Fibre Toxicol ; 15(1): 16, 2018 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-29650039

RESUMO

BACKGROUND: The rapid increase in carbon black poses threats to human health. We evaluated the effect of CB (Printex 90) on the osteogenesis of bone-marrow-derived mesenchymal stem cells (MSCs). Mitochondria play an important role in the osteogenesis of MSCs and are potential targets of nanomaterials, so we studied the role of mitochondria in the CB Printex 90-induced effects on osteogenesis. RESULTS: Low doses of Printex 90 (3 ng/mL and 30 ng/mL) that did not cause deleterious effects on MSCs' viability significantly inhibited osteogenesis of MSCs. Printex 90 caused down-regulation of osteoblastic markers, reduced activity of alkaline phosphatase (ALP), and poor mineralization of osteogenically induced MSCs. Cellular ATP production was decreased, mitochondrial respiration was impaired with reduced expression of ATPase, and the mitochondrial membrane was depolarized. The quantity and quality of mitochondria are tightly controlled by mitochondrial biogenesis, mitochondrial dynamics and mitophagy. The transcriptional co-activator and transcription factors for mitochondrial biogenesis, PGC-1α, Nrf1 and TFAM, were suppressed by Printex 90 treatment, suggesting that decreased biogenesis was caused by Printex 90 treatment during osteogenesis. Mitochondrial fusion and fission were significantly inhibited by Printex 90 treatment. PINK1 accumulated in Printex 90-treated cells, and more Parkin was recruited to mitochondria, indicating that mitophagy increased to remove the damaged mitochondria. CONCLUSIONS: This is the first report of the inhibitory effects of CB on the osteogenesis of MSCs and the involvement of mitochondria in CB Printex 90-induced suppression of MSC osteogenesis.


Assuntos
Células-Tronco Mesenquimais/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Degradação Mitocondrial/efeitos dos fármacos , Dinâmica Mitocondrial/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Fuligem/toxicidade , Fosfatase Alcalina/metabolismo , Relação Dose-Resposta a Droga , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Fatores de Transcrição/metabolismo
17.
Biochim Biophys Acta Bioenerg ; 1859(9): 789-796, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29630891

RESUMO

Enzymes in the respiratory chain are increasingly seen as potential targets against multi-drug resistance of human pathogens and cancerous cells. However, a detailed understanding of the mechanism and specificity determinants of known inhibitors is still lacking. Oligomycin, for example, has been known to be an inhibitor of the membrane motor of the mitochondrial ATP synthase for over five decades, and yet little is known about its mode of action at the molecular level. In a recent breakthrough, a crystal structure of the S. cerevisiae c-subunit ring with bound oligomycin revealed the inhibitor docked on the outer face of the proton-binding sites, deep into the transmembrane region. However, the structure of the complex was obtained in an organic solvent rather than detergent or a lipid bilayer, and therefore it has been unclear whether this mode of recognition is physiologically relevant. Here, we use molecular dynamics simulations to address this question and gain insights into the mechanism of oligomycin inhibition. Our findings lead us to propose that oligomycin naturally partitions into the lipid/water interface, and that in this environment the inhibitor can indeed bind to any of the c-ring proton-carrying sites that are exposed to the membrane, thereby becoming an integral component of the proton-coordinating network. As the c-ring rotates within the membrane, driven either by downhill proton permeation or ATP hydrolysis, one of the protonated, oligomycin-bound sites eventually reaches the subunit-a interface and halts the rotary mechanism of the enzyme.


Assuntos
Trifosfato de Adenosina/metabolismo , Inibidores Enzimáticos/metabolismo , Membranas Mitocondriais/metabolismo , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Oligomicinas/metabolismo , Saccharomyces cerevisiae/enzimologia , Sítios de Ligação , Inibidores Enzimáticos/química , Membranas Mitocondriais/efeitos dos fármacos , ATPases Mitocondriais Próton-Translocadoras/antagonistas & inibidores , ATPases Mitocondriais Próton-Translocadoras/química , Simulação de Dinâmica Molecular , Oligomicinas/química , Conformação Proteica
18.
Dalton Trans ; 47(16): 5714-5724, 2018 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-29632937

RESUMO

The [Os(η6-pcym)(dpa)(VP)]PF6 (1-VP) complex contains the histone deacetylase (HDAC) inhibitor valproate (2-propylpentanoate; VP) as a monodentate O-donor ligand and shows ca. 3-fold higher in vitro cytotoxicity against A2780 human ovarian carcinoma cells than its chlorido analogue [Os(η6-pcym)(dpa)Cl]PF6 (1-Cl); pcym = 1-methyl-4-(propan-2-yl)benzene (p-cymene), dpa = 2,2'-dipyridylamine. The complex 1-VP showed promising selectivity towards the A2780 ovarian carcinoma cell line (IC50 = 20.9 µM) over normal human hepatocytes (IC50 > 200.0 µM). Moreover, the complex 1-VP was found to be inactive against MCF-7 (breast adenocarcinoma), PANC-1 (pancreatic adenocarcinoma) and HT-29 (colon carcinoma) up to a concentration of 100 µM. Detailed flow cytometry studies indicated that treatment of A2780 cells with complex 1-VP led to induction of apoptosis, production of reactive oxygen species (ROS) and superoxide (SO) anion radicals, as well as mitochondrial membrane potential depletion and cell cycle perturbations. The microscopic assessment (standard hematoxylin/eosin staining) revealed signs of morphological changes associated with the progression of apoptosis in A2780 cells treated with the IC50 concentration of the complex 1-VP. Consistent with the intracellular production of ROS and SO, the complex 1-VP induced hydroxyl radical formation, as proved by EPR spin trapping experiments. This case study suggests that replacement of the chlorido ligand of half-sandwich Os(ii) complexes by a releasable monodentate biologically active ligand (e.g., VP used in this study) is an effective strategy for the development of novel non-platinum cytotoxic agents.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Complexos de Coordenação/química , Complexos de Coordenação/farmacologia , Osmio/química , 2,2'-Dipiridil/análogos & derivados , 2,2'-Dipiridil/química , Antineoplásicos/síntese química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/síntese química , Humanos , Concentração Inibidora 50 , Membranas Mitocondriais/efeitos dos fármacos , Monoterpenos/química , Espécies Reativas de Oxigênio/metabolismo , Superóxidos/metabolismo , Ácido Valproico/análogos & derivados , Ácido Valproico/química
19.
Mar Drugs ; 16(4)2018 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-29584673

RESUMO

Acquired drug resistance constitutes an enormous hurdle in cancer treatment, and the search for effective compounds against resistant cancer is still advancing. Marine organisms are a promising natural resource for the discovery and development of anticancer agents. In this study, we examined whether gliotoxin (GTX), a secondary metabolite isolated from marine-derived Aspergillus fumigatus, inhibits the growth of adriamycin (ADR)-resistant non-small-cell lung cancer (NSCLC) cell lines A549/ADR. We investigated the effects of GTX on A549/ADR cell viability with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and the induction of apoptosis in A549/ADR cells treated with GTX via fluorescence-activated cell sorting analysis, Hoechst staining, annexin V/propidium iodide staining, tetraethylbenzimidazolylcarbocyanine iodide (JC-1) staining, and western blotting. We found that GTX induced apoptosis in A549/ADR cells through the mitochondria-dependent pathway by disrupting mitochondrial membrane potential and activating p53, thereby increasing the expression levels of p21, p53 upregulated modulator of apoptosis (PUMA), Bax, cleaved poly (ADP-ribose) polymerase (PARP), and cleaved caspase-9. More importantly, we discovered that GTX works in conjunction with ADR to exert combinational effects on A549/ADR cells. In conclusion, our results suggest that GTX may have promising effects on ADR-resistant NSCLC cells by inducing mitochondria-dependent apoptosis and through the combined effects of sequential treatment with ADR.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Doxorrubicina/farmacologia , Gliotoxina/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Células A549 , Antineoplásicos/administração & dosagem , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/administração & dosagem , Resistencia a Medicamentos Antineoplásicos , Gliotoxina/administração & dosagem , Humanos , Membranas Mitocondriais/efeitos dos fármacos
20.
Pharmazie ; 73(2): 87-91, 2018 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-29442010

RESUMO

Ganoderma lucidum extracts have shown antiepileptic effects in in vivo and in vitro studies. In this work, primary hippocampal neurons cultured in magnesium-free medium were used to study the neuroprotective effects of ganoderic acid A and B (GA-A and GA-B) on superoxide dismutase (SOD) activity and mitochondrial membrane potential, to improve our understanding of their antiepileptic effect. The activity of SOD was determined by the xanthine oxidase assay, the variations of mitochondrial membrane potential and cell apoptosis were measured by JC-1 fluorescent staining and flow cytometry. It was found that the SOD activity and mitochondrial membrane potential (118.84 U/mg protein and 244.08 Δψm) of the epileptic hippocampal neurons were significantly lower than control values (135.95 U/mg protein and 409.81 Δψm), associated with an increase of cell apoptosis (31.88% vs. 8.84%). These circumstances can be improved by treatment of GA-A/GA-B (for SOD, 127.15±3.82 / 120.52±4.30 U/mg protein; for membrane potential (Δψm), 372.35 / 347.28; and for cell apoptosis (%), 14.93 / 20.52). Results indicated that GA-A significantly improved SOD activity, while both GA-A/GA-B tranquillized the mitochondrial membrane potential of hippocampal neurons, and thereby protected these neurons by inhibiting apoptosis.


Assuntos
Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Ácidos Heptanoicos/farmacologia , Hipocampo/citologia , Lanosterol/análogos & derivados , Membranas Mitocondriais/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Animais , Meios de Cultura , Sinergismo Farmacológico , Hipocampo/efeitos dos fármacos , Lanosterol/farmacologia , Magnésio , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Cultura Primária de Células , Ratos , Ratos Wistar , Superóxido Dismutase/metabolismo
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