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1.
Int J Mol Sci ; 22(17)2021 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-34502052

RESUMO

The role of reactive oxygen species (ROS) in ABA-induced increase in hydraulic conductivity was hypothesized to be dependent on an increase in aquaporin water channel (AQP) abundance. Single ABA application or its combination with ROS manipulators (ROS scavenger ascorbic acid and NADPH oxidase inhibitor diphenyleneiodonium chloride (DPI)) were studied on detached roots of barley plants. We measured the osmotically driven flow rate of xylem sap and calculated root hydraulic conductivity. In parallel, immunolocalization of ABA and HvPIP2;2 AQPs was performed with corresponding specific antibodies. ABA treatment increased the flow rate of xylem, root hydraulic conductivity and immunostaining for ABA and HvPIP2;2, while the addition of antioxidants prevented the effects of this hormone. The obtained results confirmed the involvement of ROS in ABA effect on hydraulic conductivity, in particular, the importance of H2O2 production by ABA-treated plants for the effect of this hormone on AQP abundance.


Assuntos
Ácido Abscísico/farmacologia , Aquaporinas/metabolismo , Osmose , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Antioxidantes/farmacologia , Ácido Ascórbico/farmacologia , Inibidores Enzimáticos/farmacologia , Hordeum/efeitos dos fármacos , Hordeum/metabolismo , NADPH Oxidases/antagonistas & inibidores , NADPH Oxidases/metabolismo , Oniocompostos/farmacologia , Raízes de Plantas/efeitos dos fármacos , Xilema/efeitos dos fármacos , Xilema/metabolismo
2.
Exp Cell Res ; 406(1): 112719, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34273405

RESUMO

Hepatic ischemia/reperfusion injury (IRI) is an adverse effect for liver transplantation which is characterized by immune response mediated inflammation. Recent studies report that neutrophil extracellular traps (NETs) are implicated in hepatic IRI. The aim of this study was to explore the mechanism of action of tetramethylpyrazine (TMP), the main chemical composition of Ligusticum chuanxiong in treatment of ischemic related diseases. Data showed that hepatic IRI increases the leak of alanine aminotransferase (ALT) and aspartate transaminase (AST), and stimulates formation of NETs. Extracellular DNA/NETs assay, hematoxylin-eosin (HE) staining, immunofluorescence assay, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) and Western blot assay, showed that TMP significantly reduces formation of NETs and alleviates hepatic IRI. Moreover, TMP and Diphenyleneiodonium (DPI) suppressed ROS production in neutrophils. In addition, analysis showed that activation of NADPH oxidase plays a role in formation of NETs triggered by hepatic IRI. Notably, TMP inhibited formation of NETs though inhibition of NADPH oxidase. Additionally, Combination treatment using TMP and DPI was more effective compared with monotherapy of either of the two drugs. These findings show that combination therapy using TMP and DPI is a promising method for treatment hepatic IRI.


Assuntos
Antioxidantes/farmacologia , Armadilhas Extracelulares/efeitos dos fármacos , Transplante de Fígado/reabilitação , Oniocompostos/farmacologia , Pirazinas/farmacologia , Traumatismo por Reperfusão/tratamento farmacológico , Alanina Transaminase/metabolismo , Animais , Aspartato Aminotransferases/metabolismo , Ácidos Nucleicos Livres/antagonistas & inibidores , Ácidos Nucleicos Livres/sangue , Modelos Animais de Doenças , Sinergismo Farmacológico , Quimioterapia Combinada/métodos , Armadilhas Extracelulares/metabolismo , Marcação In Situ das Extremidades Cortadas , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/cirurgia , Transplante de Fígado/métodos , Masculino , NADPH Oxidases/antagonistas & inibidores , NADPH Oxidases/metabolismo , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Neutrófilos/patologia , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Traumatismo por Reperfusão/cirurgia , Resultado do Tratamento
3.
Neurochem Res ; 46(8): 2089-2096, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34008119

RESUMO

Repeated morphine administration results in analgesic tolerance. However, the underlying mechanism of morphine analgesic tolerance remains unclear. NADPH-oxidase 2 (NOX2) is the first discovered NADPH oxidase, which mainly functions to produce reactive oxygen species. Its specific role in morphine tolerance has not been fully investigated. In this work, we found that chronic morphine administration significantly increased the expression of NOX2 in spinal cord. Pretreatment of NOX2 inhibitor blocked the upregulation of NOX2 and autophagy markers, including LC3B and P62, and consequently the development of morphine tolerance. NOX2 and LC3B were both colocalized with NeuN in spinal dorsal horn in morphine-tolerant rats. Our results suggest that the increased autophagy activity in spinal neurons promoted by NOX2 activation contributes to the development of morphine tolerance. NOX2 may be considered as a new therapeutic target for morphine tolerance.


Assuntos
Analgésicos Opioides/farmacologia , Autofagia/efeitos dos fármacos , Tolerância a Medicamentos/fisiologia , Morfina/farmacologia , NADPH Oxidase 2/metabolismo , Neurônios/efeitos dos fármacos , Animais , Masculino , Proteínas Associadas aos Microtúbulos/metabolismo , NADPH Oxidase 2/antagonistas & inibidores , Oniocompostos/farmacologia , Ratos Sprague-Dawley , Corno Dorsal da Medula Espinal/citologia
4.
J Pharmacol Sci ; 146(1): 29-32, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33858652

RESUMO

Hydroxyl radical (•OH) production in the rat striatum during carbon monoxide (CO) poisoning, which inhibits complex IV, was enhanced synergistically by malonate, a mitochondrial complex II inhibitor, but not N-methyl-4-phenylpyridinium or NaCN, complex I and IV inhibitors, respectively. No such enhancement appeared in the case of NaCN combined with malonate. Intrastriatal dopamine, which is involved in •OH production by malonate, did not synergistically enhance CO-induced •OH production. Diphenyleneiodonium, a nonselective NADPH oxidase inhibitor, partly suppressed the potentiation of CO-induced •OH production by malonate. Impairment of mitochondrial functions might potentiate oxidative stress and intensify CO toxicity in the brain.


Assuntos
Intoxicação por Monóxido de Carbono/metabolismo , Corpo Estriado/metabolismo , Radical Hidroxila/metabolismo , Animais , Complexo IV da Cadeia de Transporte de Elétrons/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Masculino , Malonatos/farmacologia , Mitocôndrias/metabolismo , NADPH Oxidases/antagonistas & inibidores , Oniocompostos/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Ratos Sprague-Dawley
5.
Sci Rep ; 11(1): 6556, 2021 03 22.
Artigo em Inglês | MEDLINE | ID: mdl-33753859

RESUMO

Hyperactivation of ABC transporter ABCB1 and induction of epithelial-mesenchymal transition (EMT) are the most common mechanism of acquired cancer chemoresistance. This study describes possible mechanisms, that might contribute to upregulation of ABCB1 and synergistically boost the acquisition of doxorubicin (DOX) resistance in breast cancer MX-1 cell line. DOX resistance in MX-1 cell line was induced by a stepwise increase of drug concentration or by pretreatment of cells with an ABCB1 transporter activator tetraphenylphosphonium (TPP+) followed by DOX exposure. Transcriptome analysis of derived cells was performed by human gene expression microarrays and by quantitative PCR. Genetic and epigenetic mechanisms of ABCB1 regulation were evaluated by pyrosequencing and gene copy number variation analysis. Gradual activation of canonical EMT transcription factors with later activation of ABCB1 at the transcript level was observed in DOX-only treated cells, while TPP+ exposure induced considerable activation of ABCB1 at both, mRNA and protein level. The changes in ABCB1 mRNA and protein level were related to the promoter DNA hypomethylation and the increase in gene copy number. ABCB1-active cells were highly resistant to DOX and showed morphological and molecular features of EMT. The study suggests that nongenotoxic ABCB1 inducer can possibly accelerate development of DOX resistance.


Assuntos
Antineoplásicos/farmacologia , Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Oniocompostos/farmacologia , Compostos Organofosforados/farmacologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/agonistas , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Biologia Computacional/métodos , Metilação de DNA , Resistencia a Medicamentos Antineoplásicos/genética , Epigênese Genética/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Feminino , Dosagem de Genes , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Modelos Biológicos , Transcriptoma
6.
Plant Signal Behav ; 16(5): 1889251, 2021 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-33632064

RESUMO

Heterotrimeric G-protein α and ß-subunits regulate H2O2-mediated aerenchyma formation. The rice G-protein γ-subunit, dense and erect panicle 1 (DEP1), is known to interact with the α-subunit and regulate nitrogen utilization and yield. However, it is unclear whether DEP1 regulates cell death for aerenchyma formation in rice roots. Using wild-type WYJ8 and its transgenic line WYJ8(DEP1), we confirmed that DEP1 is involved in H2O2-mediated aerenchyma formation. The rates of aerenchyma formation varied in different parts of the roots in both varieties, with the highest rate in the 4-7 cm segments, reaching a plateau in the 7-8 cm segments. Compared with WYJ8, the aerenchyma area and H2O2 content in WYJ8(DEP1) were increased by 55.98% and 53.37%, respectively; however, the responses of aerenchyma formation to exogenous H2O2 were basically the same in the two varieties. Diphenylene iodonium (DPI) treatment had no effect on H2O2 production and elimination processes in WYJ8, but significantly reduced the activity of the key enzyme that catalyzes H2O2 biosynthesis in WYJ8(DEP1). Importantly, exogenous H2O2 treatment did not offset the effect of the decrease in endogenous H2O2 level caused by DPI on aerenchyma formation. These results indicated that DEP1 enhanced H2O2 biosynthesis and promoted the cell death of the root cortex, thus contributing to aerenchyma development in WYJ8(DEP1).


Assuntos
Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Peróxido de Hidrogênio/metabolismo , Oryza/metabolismo , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , Transdução de Sinais , Amitrol (Herbicida)/farmacologia , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Oniocompostos/farmacologia , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética
7.
Food Chem Toxicol ; 150: 112055, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33577942

RESUMO

Patulin (PAT) is a kind of mycotoxins that commonly found in decayed fruits and their products. Our previous studies have shown that PAT induced cell apoptosis and the overproduction of reactive oxygen species (ROS) in human embryonic kidney (HEK293) cells. The present study aimed to further investigate the functional role of NADPH oxidase, one of the main cellular sources of ROS, in PAT-induced apoptosis and oxidative damage in HEK293 cells. We demonstrated that the protein and mRNA expression levels of NADPH oxidase catalytic subunit NOX2 and regulatory subunit p47phox were up-regulated under PAT stress. Inhibiting of NADPH oxidase with the specific antagonist diphenyleneiodonium (DPI) suppressed cytotoxicity and apoptosis induced by PAT as evidenced by the increase of cell viability, the decrease of LDH release and the inhibition of caspase activities. Furthermore, DPI re-established mitochondrial membrane potential (MMP) and enhanced cellular ATP content. Importantly, DPI supplementation elevated endogenous GSH contents as well as the ratio of GSH/GSSG. Meanwhile, the antioxidant-enzyme activities of GPx, GR, CAT and SOD were significantly promoted. Collectively, our results suggested that NADPH oxidase played a critical role in PAT-induced nephrotoxicity, and inhibition of NADPH oxidase by DPI attenuated cell injury and apoptosis via regulation of oxidative damage.


Assuntos
Compostos de Bifenilo/farmacologia , Sobrevivência Celular/efeitos dos fármacos , NADPH Oxidases/metabolismo , Oniocompostos/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Patulina/toxicidade , Trifosfato de Adenosina/metabolismo , Caspases/genética , Caspases/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Glutationa/metabolismo , Células HEK293 , Humanos , Lactato Desidrogenases/genética , Lactato Desidrogenases/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mutagênicos/toxicidade , NADPH Oxidases/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
8.
Cell Death Dis ; 12(2): 189, 2021 02 16.
Artigo em Inglês | MEDLINE | ID: mdl-33594044

RESUMO

Oncogenic RAS is a critical driver for the initiation and progression of several types of cancers. However, effective therapeutic strategies by targeting RAS, in particular RASG12D and RASG12V, and associated downstream pathways have been so far unsuccessful. Treatment of oncogenic RAS-ravaged cancer patients remains a currently unmet clinical need. Consistent with a major role in cancer metabolism, oncogenic RAS activation elevates both reactive oxygen species (ROS)-generating NADPH oxidase (NOX) activity and ROS-scavenging glutathione biosynthesis. At a certain threshold, the heightened oxidative stress and antioxidant capability achieve a higher level of redox balance, on which cancer cells depend to gain a selective advantage on survival and proliferation. However, this prominent metabolic feature may irrevocably render cancer cells vulnerable to concurrent inhibition of both NOX activity and glutathione biosynthesis, which may be exploited as a novel therapeutic strategy. In this report, we test this hypothesis by treating the HRASG12V-transformed ovarian epithelial cells, mutant KRAS-harboring pancreatic and colon cancer cells of mouse and human origins, as well as cancer xenografts, with diphenyleneiodonium (DPI) and buthionine sulfoximine (BSO) combination, which inhibit NOX activity and glutathione biosynthesis, respectively. Our results demonstrate that concomitant targeting of NOX and glutathione biosynthesis induces a highly potent lethality to cancer cells harboring oncogenic RAS. Therefore, our studies provide a novel strategy against RAS-bearing cancers that warrants further mechanistic and translational investigation.


Assuntos
Carcinoma Ductal Pancreático/tratamento farmacológico , Neoplasias do Colo/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Genes ras , Glutationa/biossíntese , Metionina/análogos & derivados , Mutação , NADPH Oxidases/antagonistas & inibidores , Oniocompostos/farmacologia , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Pancreáticas/tratamento farmacológico , Sulfóxidos/farmacologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Ductal Pancreático/enzimologia , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Morte Celular/efeitos dos fármacos , Neoplasias do Colo/enzimologia , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Feminino , Genes p53 , Glutamato-Cisteína Ligase/antagonistas & inibidores , Glutamato-Cisteína Ligase/metabolismo , Células HCT116 , Humanos , Metionina/farmacologia , Camundongos Nus , Camundongos Transgênicos , NADPH Oxidases/metabolismo , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Estresse Oxidativo , Neoplasias Pancreáticas/enzimologia , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Transdução de Sinais , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
9.
J Integr Plant Biol ; 63(3): 583-596, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33017089

RESUMO

Salicylic acid (SA) plays a crucial role in plant immunity. However, its function in plant development is poorly understood. The quiescent center (QC), which maintains columella stem cells (CSCs) in the root apical meristem and typically exhibits low levels of cell division, is critical for root growth and development. Here, we show that the Arabidopsis thaliana SA overaccumulation mutant constitutively activated cell death 1 (cad1), which exhibits increased cell division in the QC, is rescued by additional mutations in genes encoding the SA biosynthetic enzyme SALICYLIC ACID INDUCTION DEFFICIENT2 (SID2) or the SA receptor NONEXPRESSER OF PR GENES1 (NPR1), indicating that QC cell division in the cad1 mutant is promoted by the NPR1-dependent SA signaling pathway. The application of exogenous SA also promoted QC cell division in wild-type plants in a dose-dependent manner and largely suppressed the expression of genes involved in QC maintenance, including those encoding the APETALA2 (AP2) transcription factors PLETHORA1 (PLT1) and PLT2, as well as the homeodomain transcription factor WUSCHEL-RELATED HOMEOBOX5 (WOX5). Moreover, we showed that SA promotes reactive oxygen species (ROS) production, which is necessary for the QC cell division phenotype in the cad1 mutant. These results provide insight into the function of SA in QC maintenance.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Divisão Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Proteínas de Homeodomínio/genética , Espécies Reativas de Oxigênio/metabolismo , Ácido Salicílico/farmacologia , Fatores de Transcrição/genética , Arabidopsis/citologia , Arabidopsis/efeitos dos fármacos , Proteínas de Arabidopsis/metabolismo , Diferenciação Celular/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Homeodomínio/metabolismo , Ácidos Indolacéticos/metabolismo , Mutação/genética , Oniocompostos/farmacologia , Fenótipo , Plantas Geneticamente Modificadas , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição/metabolismo
10.
Plant Signal Behav ; 16(2): 1848086, 2021 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-33210579

RESUMO

Cutting is a frequently used model to study the process of adventitious root formation, and excision of cuttings leads to rapid wound response signaling. We recently showed that as a wound signal, reactive oxygen species (ROS, mainly hydrogen peroxide) participate in adventitious root induction of hypocotyl cuttings through regulation of auxin biosynthesis and transport. Here, superoxide anion (O2 -•), an early type of ROS, exhibited rapid burst at the cutting site immediately in response to wounding in Arabidopsis hypocotyl cuttings. Diphenylene iodonium chloride (DPI, inhibitor of NADPH oxidase) overwhelmingly suppressed O2 -• propagation through the hypocotyl. Compared to wild type, O2 -• burst only occur in cut base, and upward transduction were inhibited completely in NADPH oxidase mutant AtRbohD. These results indicate O2 -• generation and propagation in response to wound and via NADPH oxidase in adventitious root induction of hypocotyl cuttings.


Assuntos
Arabidopsis/metabolismo , Superóxidos/metabolismo , Arabidopsis/efeitos dos fármacos , Oniocompostos/farmacologia , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/metabolismo , Espécies Reativas de Oxigênio/metabolismo
11.
Plant Signal Behav ; 16(3): 1856546, 2021 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-33315520

RESUMO

The plasma membrane NADPH Oxidase-derived ROS as signaling molecules play crucial roles in salt stress response. As the motor organelle of cells, mitochondria are also important for salt tolerance. However, the possible interaction between NADPH Oxidase-derived ROS and mitochondria is not well studied. Here, a transgenic Arabidopsis expressing mitochondrial matrix-targeted pH-sensitive indicator cpYFP was used to monitor the pH dynamics in root cells under salt stress. A significant alkalization in mitochondria was observed when the root was exposed to NaCl or KCl, but not osmotic stress such as isotonic mannitol. Interestingly, when pretreated with the NADPH Oxidase inhibitor DPI, the mitochondrial alkalization in root cells was largely abolished. Genetic evidence further showed that salt-induced mitochondrial alkalization was significantly reduced in the loss of function mutant atrbohF . Pretreatment with endocytosis-related inhibitor PAO or TyrA23, which inhibited the ROS accumulation under salt treatment, almost abolished this effect. Furthermore, [Ca2+]cyt increase might also play important roles by affecting ROS generation to mediate salt-induced mitochondrial alkalization as indicated by treatment with plasma membrane Ca2+ channel inhibitor LaCl3 and mitochondrial Ca2+ uniporter inhibitor Ruthenium Red. Together, these results suggest that the plasma membrane NADPH Oxidase-derived ROS promote the mitochondrial alkalization under salt treatment, providing a possible link between different cellular compartments under salt stress.


Assuntos
Álcalis/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Mitocôndrias/metabolismo , NADPH Oxidases/metabolismo , Raízes de Plantas/citologia , Espécies Reativas de Oxigênio/metabolismo , Estresse Salino/fisiologia , Arabidopsis/citologia , Arabidopsis/efeitos dos fármacos , Proteínas de Arabidopsis/antagonistas & inibidores , Cálcio/metabolismo , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Endocitose/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Mitocôndrias/efeitos dos fármacos , NADPH Oxidases/antagonistas & inibidores , Oniocompostos/farmacologia , Estresse Salino/efeitos dos fármacos , Cloreto de Sódio/farmacologia
12.
Int J Mol Sci ; 21(20)2020 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-33081375

RESUMO

Y-27632 is known as a selective Rho-associated coiled coil-forming kinase (ROCK) inhibitor. Y-27632 has been shown to induce neurite outgrowth in several neuronal cells. However, the precise molecular mechanisms linking neurite outgrowth to Y-27632 are not completely understood. In this study, we examined the ability of Y-27632 to induce neurite outgrowth in PC12 cells and evaluated the signaling cascade. The effect of Y-27632 on the neurite outgrowth was inhibited by reactive oxygen species (ROS) scavengers such as N-acetyl cysteine (NAC) and trolox. Furthermore, Y-27632-induced neurite outgrowth was not triggered by NADPH oxidase 1 (NOX1) knockdown or diphenyleneiodonium (DPI), a NOX inhibitor. Suppression of the Rho-family GTPase Rac1, which is under the negative control of ROCK, with expression of the dominant negative Rac1 mutant (Rac1N17) prevented Y-27632-induced neurite outgrowth. Moreover, the Rac1 inhibitor NSC23766 prevented Y-27632-induced AKT and p21-activated kinase 1 (PAK1) activation. AKT inhibition with MK2206 suppressed Y-27632-induced PAK1 phosphorylation and neurite outgrowth. In conclusion, our results suggest that Rac1/NOX1-dependent ROS generation and subsequent activation of the AKT/PAK1 cascade contribute to Y-27632-induced neurite outgrowth in PC12 cells.


Assuntos
Amidas/farmacologia , Crescimento Neuronal/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Transdução de Sinais , Acetilcisteína/farmacologia , Animais , Cromanos/farmacologia , Sequestradores de Radicais Livres/farmacologia , NADPH Oxidase 1/metabolismo , Oniocompostos/farmacologia , Células PC12 , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Quinases Ativadas por p21/metabolismo , Proteínas rac1 de Ligação ao GTP/metabolismo
13.
PLoS One ; 15(10): e0240610, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33049001

RESUMO

Presynaptic mitochondrial Ca2+ plays a critical role in the regulation of synaptic transmission and plasticity. The presynaptic bouton of the hippocampal mossy fiber (MF) is much larger in size than that of the Schaffer collateral (SC) synapse. Here we compare the structural and physiological characteristics of MF and SC presynaptic boutons to reveal functional and mechanistic differences between these two synapses. Our quantitative ultrastructural analysis using electron microscopy show many more mitochondria in MF presynaptic bouton cross-section profiles compared to SC boutons. Consistent with these results, post-tetanic potentiation (PTP), a form of presynaptic short-term plasticity dependent on mitochondrial Ca2+, is reduced by inhibition of mitochondrial Ca2+ release at MF synapses but not at SC synapses. However, blockade of mitochondrial Ca2+ release results in reduction of PTP at SC synapses by disynaptic MF stimulation. Furthermore, inhibition of mitochondrial Ca2+ release selectively decreases frequency facilitation evoked by short trains of presynaptic stimulation at MF synapses, while having no effect at SC synapses. Moreover, depletion of ER Ca2+ stores leads to reduction of PTP at MF synapses, but PTP is unaffected by ER Ca2+ depletion at SC synapses. These findings show that MF and SC synapses differ in presynaptic mitochondrial content as well as mitochondrial Ca2+ dependent synaptic plasticity, highlighting differential regulatory mechanisms of presynaptic plasticity at MF and SC synapses.


Assuntos
Cálcio/metabolismo , Hipocampo/metabolismo , Fibras Musgosas Hipocampais/metabolismo , Plasticidade Neuronal/fisiologia , Animais , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/fisiologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Hipocampo/citologia , Hipocampo/efeitos dos fármacos , Hipocampo/ultraestrutura , Masculino , Camundongos , Microscopia Eletrônica , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Fibras Musgosas Hipocampais/efeitos dos fármacos , Fibras Musgosas Hipocampais/ultraestrutura , Plasticidade Neuronal/efeitos dos fármacos , Oniocompostos/farmacologia , Compostos Organofosforados/farmacologia , Técnicas de Patch-Clamp , Tiazepinas/farmacologia
14.
Environ Toxicol Pharmacol ; 80: 103507, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33007436

RESUMO

Tetrachlorobisphenol A (TCBPA) can promote intracellular reactive oxygen species (ROS) accumulation. However, limited attention has been given to mechanisms underlying TCBPA exposure-associated ROS accumulation. Here, such mechanisms were explored in the simple eukaryotic model organism Saccharomyces cerevisiae exposed to multiple concentrations of TCBPA. Addition of diphenyleneiodonium, a specific inhibitor of NADPH oxidase, blocked TCBPA treatment-associated intracellular ROS accumulation. NADPH oxidase can be activated by calcineurin, mitogen-activated protein kinase (MAPK), and tyrosine kinase. Therefore, corresponding specific inhibition respectively on these three kinases was performed and results suggested that the Ca2+ signaling pathway, MAPK pathway, and tyrosine kinase pathway all contributed to the TCBPA exposure-associated intracellular ROS accumulation. In addition, TCBPA exposure-associated up-regulation of genes involved in ROS production and down-regulation of catalase promoted ROS accumulation in S. cerevisiae. To sum up, our current results provide insights into the understanding of TCBPA exposure-associated ROS accumulation.


Assuntos
Clorofenóis/toxicidade , Retardadores de Chama/toxicidade , Saccharomyces cerevisiae/efeitos dos fármacos , Cálcio/metabolismo , Catalase/genética , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NADPH Oxidases/antagonistas & inibidores , Oniocompostos/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Transdução de Sinais/efeitos dos fármacos , Superóxido Dismutase-1/genética
15.
Br J Cancer ; 123(12): 1737-1748, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32958825

RESUMO

BACKGROUND: Peroxisome proliferator-activated receptor γ (PPARγ) agonists frequently induce cell death in human non-small-cell lung cancer (NSCLC) cells. However, majority of NSCLC patients acquire resistance after cancer therapy, and it is still unclear. METHODS: In this study we investigated the apoptotic mechanism and the anti-cancer effects of a novel purine-based PPARγ agonist, CB11 (8-(2-aminophenyl)-3-butyl-1,6,7-trimethyl-1H-imidazo[2,1-f]purine-2,4(3H,8H)-dione), on human NSCLC cells. CB11 mediates PPARγ-dependent cell death, reactive oxygen species (ROS) production, mitochondrial membrane potential (MMP) collapse, cell cycle arrest, lactate dehydrogenase (LDH) cytotoxicity, and caspase-3 activity in human NSCLC cells. RESULTS: CB11 causes cell death via ROS-mediated ATM-p53-GADD45α signalling in human NSCLC cells, and diphenyleneiodonium (DPI), an NADPH oxidase inhibitor, decreases cell death by inhibiting CB11-mediated ATM signalling. In a xenograft experiment, CB11 dramatically reduced tumour volume when compared to a control group. Furthermore, CB11 induced cell death by inhibiting epithelial-to-mesenchymal transition (EMT) under radiation exposure in radiation-resistant human NSCLC cells. However, PPARγ deficiency inhibited cell death by blocking the ATM-p53 axis in radiation/CB11-induced radiation-resistant human NSCLC cells. CONCLUSIONS: Taken together, our results suggest that CB11, a novel PPARγ agonist, may be a novel anti-cancer agent, and it could be useful in a therapeutic strategy to overcome radio-resistance in radiation-exposed NSCLC.


Assuntos
Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/radioterapia , Imidazóis/uso terapêutico , Neoplasias Pulmonares/radioterapia , PPAR gama/agonistas , Purinas/uso terapêutico , Tolerância a Radiação/efeitos dos fármacos , Células 3T3 , Adipócitos/citologia , Anilidas/farmacologia , Animais , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Compostos Azo , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Caspase 3/metabolismo , Caspase 9/metabolismo , Pontos de Checagem do Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Morte Celular , Diferenciação Celular , Linhagem Celular Tumoral , Dano ao DNA , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos da radiação , Feminino , Humanos , L-Lactato Desidrogenase , Ligantes , Luciferases/metabolismo , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Potencial da Membrana Mitocondrial , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Oniocompostos/farmacologia , PPAR gama/deficiência , PPAR gama/metabolismo , RNA Interferente Pequeno , Tolerância a Radiação/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Tiazolidinedionas/farmacologia , Carga Tumoral/efeitos dos fármacos , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/metabolismo
16.
Int J Mol Sci ; 21(17)2020 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-32867341

RESUMO

The reactive oxygen species (ROS) burst is the most common plant immunity mechanism to prevent pathogen infection, although the exact role of ROS in plant immunity has not been fully elucidated. We investigated the expression and translocation of Oryza sativa respiratory burst oxidase homologue B (OsRBOHB) during compatible and incompatible interactions between rice epidermal cells and the pathogenic fungus Pyricularia oryzae (syn. Magnaporthe oryzae). We characterized the functional role of ROS focal accumulation around invading hyphae during P. oryzae infection process using the OsRBOHB inhibitor diphenyleneiodonium (DPI) and the actin filament polymerization inhibitor cytochalasin (Cyt) A. OsRBOHB was strongly induced during incompatible rice-P. oryzae interactions, and newly synthesized OsRBOHB was focally distributed at infection sites. High concentrations of ROS focally accumulated at the infection sites and suppressed effector biotrophy-associated secreted (BAS) proteins BAS4 expression and invasive hyphal growth. DPI and Cyt A abolished ROS focal accumulation and restored P. oryzae effector BAS4 expression. These results suggest that ROS focal accumulation is able to function as an effective immune mechanism that blocks some effectors including BAS4-expression during P. oryzae infection. Disruption of ROS focal accumulation around invading hyphae enables successful P. oryzae colonization of rice cells and disease development.


Assuntos
Ascomicetos/fisiologia , Proteínas Fúngicas/genética , Oryza/metabolismo , Doenças das Plantas/microbiologia , Proteínas de Plantas/genética , Espécies Reativas de Oxigênio/metabolismo , Citocalasinas/farmacologia , Resistência à Doença , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Oniocompostos/farmacologia , Oryza/microbiologia , Doenças das Plantas/prevenção & controle , Imunidade Vegetal
17.
Cell Tissue Bank ; 21(4): 675-684, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32607683

RESUMO

Sperm cryopreservation leads to various structural and functional damages, some of which induce by oxidative stress. The reactive oxygen species (ROS) generates by mitochondria and membrane NADPH oxidases (NOXs). Among the NOXs, only NOX5 has been identified in the cell membrane of human sperm. This study was designed to clarify the possible role of NOX5 on sperm cryoinjury. Forty human semen samples were washed and randomly divided into fresh and cryopreserved groups. Each group was divided into 4 subgroups containing Ham's F10 (control), 0.1% DMSO (vehicle), 100 nM of PMA (phorbol 12-myristate 13-acetate) and 1 µM of DPI (diphenyleneiodonium), as NOX5 activator and inhibitor. The samples of cryopreserved groups were preserved in liquid nitrogen for 1 month. The sperm kinematics, membrane integrity, ROS production, apoptosis rate, mitochondrial membrane potential (MMP), intracellular ATP and calcium concentration [Ca2+]i were evaluated. The percent of sperm with intact membrane and motile sperm reduced significantly after thawing (p ≤ 0.01). The ROS production (p ≤ 0.01) and the apoptotic rate increased, MMP dissipated, and the percentage of live cells with high [Ca2+]i decreased significantly in the cryopreserved control group relative to the fresh control group. DPI, in contrast to PMA, improved sperm progressive motility (p ≤ 0.01), membrane integrity in fresh and cryopreserved groups and reduced the ROS amount in cryopreserved group (p ≤ 0.01). Apoptotic rate, [Ca2+]i, ATP, and MMP did not change with DPI and PMA in cryopreserved groups. We conclude that NOX5 activity in fresh sperm is low, and it increases during cryopreservation. NOX5 inhibition improves the cryopreserved sperm quality.


Assuntos
Criopreservação , NADPH Oxidase 5/metabolismo , Espermatozoides/enzimologia , Espermatozoides/patologia , Trifosfato de Adenosina/metabolismo , Adulto , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Humanos , Espaço Intracelular/metabolismo , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Oniocompostos/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Motilidade Espermática/efeitos dos fármacos , Acetato de Tetradecanoilforbol/farmacologia , Adulto Jovem
18.
Exp Dermatol ; 29(8): 749-758, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32640089

RESUMO

Emerging evidence suggests oxidative stress plays a role in the pathophysiology of both atopic dermatitis (AD) and psoriasis (PSO). We established in vitro models of AD and PSO skin, and characterized these models in regard to their oxidative stress state. Both AD and PSO model keratinocytes exhibited elevated reactive oxygen species (ROS) levels and accumulated more DNA damage than control cells after oxidative stress induced by 250 µmol/L H2 O2 . Elevated ROS levels and DNA damage accumulation could be inhibited by the NADPH oxidase (NOX) inhibitor diphenyleneiodonium (DPI). Further, immunofluorescence analysis revealed the presence of both NOX1 and NOX4 in keratinocytes. By inhibiting NOX1, stress-related signalling cascades and elevated ROS levels could be abrogated, and survival of AD and PSO cells improved. Taken together, this study reveals that inhibition of NOX inhibition could abrogate elevated oxidative stress in a 2D model of AD and PSO.


Assuntos
Dermatite Atópica/metabolismo , Queratinócitos/metabolismo , NADPH Oxidases/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Psoríase/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Células Cultivadas , Dano ao DNA/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Humanos , Peróxido de Hidrogênio/farmacologia , Técnicas In Vitro , NADPH Oxidase 1/metabolismo , NADPH Oxidase 4/metabolismo , NADPH Oxidases/antagonistas & inibidores , Oniocompostos/farmacologia , Oxidantes/farmacologia , Transdução de Sinais
19.
Theranostics ; 10(15): 6743-6757, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32550901

RESUMO

Long-term inflammatory stimulation is considered one of the most important causes of colorectal cancer. Diphenyleneiodonium (DPI), an NADPH oxidase inhibitor, can inhibit a variety of inflammatory responses. However, the systemic toxicity of DPI limits its clinical application. Whether DPI can inhibit colitis-associated colorectal cancer (CAC) at ultralow concentrations remains unknown. Methods: CAC was induced by azoxymethane (AOM) injection followed by treatment with dextran sulfate sodium (DSS), and DPI was intraperitoneally injected (i.p.) in the first cycle for 21 days. Colon tissue was collected and analyzed by western blotting. Immune cell infiltration and macrophage polarization were examined by immunohistochemistry, immunofluorescence, or real-time polymerase-chain reaction (PCR). Reactive oxygen species (ROS) production was measured by flow cytometry. Results: Ultralow dose DPI significantly ameliorated the DSS-induced colitis and attenuated the colon tumorigenesis in the mouse model of AOM/ DSS-induced CAC. Mechanistically, an ultralow dose of DPI inhibited the production of pro-inflammatory cytokines, (tumor necrosis factor (TNF)-α and interleukin (IL)-6), reduced the macrophage infiltration and classical polarization, and induced the ROS generation. These effects were found to be related to the inhibition of the phosphorylation of signal transducer and activator of transcription 3 (STAT3), mitogen-activated protein kinase (MAPK), and nuclear factor kappa B (NF -κB). Conclusion: The present study revealed that an ultralow dose of DPI, with no significant systemic toxicity involved, may be an effective way to prevent the occurrence and development of CAC.


Assuntos
Azoximetano/toxicidade , Neoplasias Associadas a Colite/tratamento farmacológico , Colite/complicações , Neoplasias Colorretais/tratamento farmacológico , NADPH Oxidases/antagonistas & inibidores , Oniocompostos/farmacologia , Animais , Carcinógenos/toxicidade , Linhagem Celular , Colite/induzido quimicamente , Neoplasias Associadas a Colite/etiologia , Neoplasias Associadas a Colite/patologia , Neoplasias Colorretais/etiologia , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NADPH Oxidases/metabolismo , Espécies Reativas de Oxigênio/metabolismo
20.
ACS Comb Sci ; 22(9): 457-467, 2020 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-32589005

RESUMO

New 8-chloro-2-phenyl-2,7-naphthyridin-1(2H)-one building blocks bearing diverse substitutes on the 2-phenyl group were synthesized via an efficient diaryliodonium salt-based N-arylation strategy with the advantage of mild conditions, short reaction times, and high yields. A small combinatorial library of 8-amino substituted 2-phenyl-2,7-naphthyridin-1(2H)-one was further conveniently constructed based on the above chlorinated naphthyridinones and substituted aniline. Preliminary biochemical screening resulted in the discovery of the new 2,7-naphthyridone-based MET/AXL kinase inhibitors. More importantly, 17c (IC50,MET of 13.8 nM) or 17e (IC50,AXl of 17.2 nM) and 17i (IC50,AXl of 31.8 nM) can efficient selectively inhibit MET or AXL kinase, respectively, while commercial cabozantinib showed no selectivity. The further exploration of the 8-substituted 2-phenyl-2,7-naphthyridin-1(2H)-one combinatorial library would significantly accelerate the discovery of more potent and selective inhibitors against diverse kinases.


Assuntos
Descoberta de Drogas , Oniocompostos/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Oniocompostos/química , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Sais/química , Sais/farmacologia , Relação Estrutura-Atividade
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