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1.
Invest Ophthalmol Vis Sci ; 61(1): 1, 2020 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-31995153

RESUMO

Purpose: Vacuolar protein sorting 35 (Vps35) mutations and protein dysfunction have been linked to the hyperphosphorylation and accumulation of tau protein in a number of central neurodegenerative disorders. The aims of the present study were to investigate the mechanism underlying the tau hyperphosphorylation caused by Vps35 deficiency. Methods: The cells used in this study were primary retinal ganglion cells (RGCs). The rat retinal glutamate excitotoxicity model was used in vivo. Fresh retinal tissues or eyeballs were collected at different time points. The expression and interactions of Vps35, Cdk5/p35, tau hyperphosphorylation, LAMP1, EEA1 and UBE1 in RGCs were studied by immunofluorescence staining, Western blotting, and immunoprecipitation. Results: The downregulation and overexpression of Vps35 increased and decreased the expression of p35 and tau hyperphosphorylation, respectively. More important, roscovitine, a Cdk5 inhibitor, could effectively decrease the hyperphosphorylated tau level induced by Vps35 deficiency. Furthermore, this study confirmed that the inhibition of Vps35 could increase the activity of Cdk5/p35 by affecting the lysosomal degradation of p35 and lead to the degeneration of RGCs. Conclusions: These findings demonstrate the possibility that Cdk5/p35 acts as a "cargo" of Vps35 and provide new insights into the pathogenesis of RGC degeneration caused by hyperphosphorylated tau protein. Vps35 is a potential target for basic research and clinical treatment of RGC degeneration in many ocular diseases such as glaucoma.


Assuntos
Quinase 5 Dependente de Ciclina/metabolismo , Fosfotransferases/metabolismo , Células Ganglionares da Retina/metabolismo , Proteínas de Transporte Vesicular/deficiência , Proteínas tau/metabolismo , Animais , Western Blotting , Células Cultivadas , Quinase 5 Dependente de Ciclina/antagonistas & inibidores , Regulação para Baixo , Técnica Indireta de Fluorescência para Anticorpo , Ácido Glutâmico/toxicidade , Glicoproteínas de Membrana Associadas ao Lisossomo/metabolismo , Masculino , Camundongos , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Degeneração Retiniana/induzido quimicamente , Degeneração Retiniana/metabolismo , Roscovitina/farmacologia , Transfecção , Enzimas Ativadoras de Ubiquitina/metabolismo , Proteínas de Transporte Vesicular/metabolismo
2.
Biomed Pharmacother ; 118: 109295, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31545255

RESUMO

Mitochondrial dysfunction and inflammation contribute to the initiation and development of several brain pathological conditions, including Alzheimer's disease and cerebral ischemia. Linalool is an aromatic plant-derived monoterpene alcohol with reported anti-inflammatory, and anti-oxidant properties. We investigated the role of linalool on glutamate-induced mitochondrial oxidative stress in immortalized neuronal HT-22 cells. Glutamate induced oxidative stress in neuronal cells, as detected by real-time cell impedance measurements, MTT assay, and analysis of Annexin V/PI. Administration of linalool 100 µM reduced cell death mediated by glutamate. Staining of glutamate-stimulated mitochondria by MitoTracker revealed improved morphology in the presence of linalool. Furthermore, we demonstrated a potential neuroprotective effect of linalool in conditions of oxidative stress by a reduction of mitochondrial ROS and mitochondrial calcium levels, and by preserving mitochondrial membrane potential. Experiments using both high-resolution respirometry and Seahorse Extracellular flux analyzer showed that linalool was able to promote an increase in uncoupled respiration that could contribute to its neuroprotective capacity. Linalool protection was validated using organotypic hippocampal slices as ex vivo model with NMDA as a stimulus to induce excitotoxity cell damage. These results demonstrate that linalool is protective in an in vitro model of glutamate-induced oxidative stress and in an ex-vivo model for excitotoxity, proposing linalool as a potential therapeutic agent against neurodegenerative brain diseases where oxidative stress contributes to the pathology of the disease.


Assuntos
/farmacologia , Ácido Glutâmico/toxicidade , Mitocôndrias/patologia , N-Metilaspartato/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Encéfalo/metabolismo , Respiração Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Mitocôndrias/efeitos dos fármacos , Substâncias Protetoras/farmacologia
3.
Biomed Res Int ; 2019: 6593125, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31467905

RESUMO

Methanolic extract of Clinacanthus nutans Lindau leaves (MECN) has been reported to exert antinociceptive activity. The present study aimed to elucidate the possible antinociceptive mechanisms of a lipid-soluble fraction of MECN, which was obtained after sequential extraction in petroleum ether. The petroleum ether fraction of C. nutans (PECN), administered orally to mice, was (i) subjected to capsaicin-, glutamate-, phorbol 12-myristate 13-acetate-, bradykinin-induced nociception model; (ii) prechallenged (intraperitoneal (i.p.)) with 0.15 mg/kg yohimbine, 1 mg/kg pindolol, 3 mg/kg caffeine, 0.2 mg/kg haloperidol, or 10 mg/kg atropine, which were the respective antagonist of α 2-adrenergic, ß-adrenergic, adenosinergic, dopaminergic, or muscarinic receptors; and (iii) prechallenged (i.p.) with 10 mg/kg glibenclamide, 0.04 mg/kg apamin, 0.02 mg/kg charybdotoxin, or 4 mg/kg tetraethylammonium chloride, which were the respective inhibitor of ATP sensitive-, small conductance Ca2+-activated-, large conductance Ca2+-activated-, or nonselective voltage-activated-K+ channel. Results obtained demonstrated that PECN (100, 250, and 500 mg/kg) significantly (P<0.05) inhibited all models of nociception described earlier. The antinociceptive activity of 500 mg/kg PECN was significantly (P<0.05) attenuated when prechallenged with all antagonists or K+ channel blockers. However, only pretreatment with apamin and charybdotoxin caused full inhibition of PECN-induced antinociception. The rest of the K+ channel blockers and all antagonists caused only partial inhibition of PECN antinociception, respectively. Analyses on PECN's phytoconstituents revealed the presence of antinociceptive-bearing bioactive compounds of volatile (i.e., derivatives of γ-tocopherol, α-tocopherol, and lupeol) and nonvolatile (i.e., cinnamic acid) nature. In conclusion, PECN exerts a non-opioid-mediated antinociceptive activity involving mainly activation of adenosinergic and cholinergic receptors or small- and large-conductance Ca2+-activated-K+ channels.


Assuntos
Acanthaceae/química , Analgésicos/farmacologia , Dor Nociceptiva/tratamento farmacológico , Extratos Vegetais/farmacologia , Alcanos/química , Analgésicos/química , Analgésicos não Entorpecentes/química , Analgésicos não Entorpecentes/farmacologia , Animais , Bradicinina/toxicidade , Capsaicina/toxicidade , Ácido Glutâmico/toxicidade , Humanos , Metanol/química , Camundongos , Dor Nociceptiva/induzido quimicamente , Dor Nociceptiva/patologia , Extratos Vegetais/química , Folhas de Planta/química , Canais de Potássio/genética , Acetato de Tetradecanoilforbol/análogos & derivados , Acetato de Tetradecanoilforbol/toxicidade
4.
Biosci Biotechnol Biochem ; 83(11): 2016-2026, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31272310

RESUMO

Exposure of PC12 cells to 10 mM glutamate caused significant viability loss, cell apoptosis, decreased activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) as well as increased levels of malondialdehyde (MDA). In parallel, glutamate significantly increased the intracellular levels of ROS and intracellular calcium. However, pretreatment of the cells with acteoside and isoacteoside significantly suppressed glutamate-induced cellular events. Moreover, acteoside and isoacteoside reduced the glutamate-induced increase of caspase-3 activity and also ameliorated the glutamate-induced Bcl-2/Bax ratio reduction in PC12 cells. Furthermore, acteoside and isoacteoside significantly inhibited glutamate-induced DNA damage. In the mouse model, acteoside significantly attenuated cognitive deficits in the Y maze test and attenuated neuronal damage of the hippocampal CA1 regions induced by glutamate. These data indicated that acteoside and isoacteoside play neuroprotective effects through anti-oxidative stress, anti-apoptosis, and maintenance of steady intracellular calcium.


Assuntos
Antioxidantes/química , Antioxidantes/farmacologia , Ácido Glutâmico/toxicidade , Glicosídeos/química , Glicosídeos/farmacologia , Neurotoxinas/toxicidade , Álcool Feniletílico/química , Animais , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Hipocampo/citologia , Hipocampo/efeitos dos fármacos , Hipocampo/fisiologia , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Malondialdeído/metabolismo , Memória/efeitos dos fármacos , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/farmacologia , Células PC12 , Ratos , Espécies Reativas de Oxigênio/metabolismo
5.
Elife ; 82019 07 18.
Artigo em Inglês | MEDLINE | ID: mdl-31318331

RESUMO

Overproduction of reactive oxygen species (ROS) is known to mediate glutamate excitotoxicity in neurological diseases. However, how ROS burdens can influence neural circuit integrity remains unclear. Here, we investigate the impact of excitotoxicity induced by depletion of Drosophila Eaat1, an astrocytic glutamate transporter, on locomotor central pattern generator (CPG) activity, neuromuscular junction architecture, and motor function. We show that glutamate excitotoxicity triggers a circuit-dependent ROS feedback loop to sculpt the motor system. Excitotoxicity initially elevates ROS, thereby inactivating cholinergic interneurons and consequently changing CPG output activity to overexcite motor neurons and muscles. Remarkably, tonic motor neuron stimulation boosts muscular ROS, gradually dampening muscle contractility to feedback-enhance ROS accumulation in the CPG circuit and subsequently exacerbate circuit dysfunction. Ultimately, excess premotor excitation of motor neurons promotes ROS-activated stress signaling that alters neuromuscular junction architecture. Collectively, our results reveal that excitotoxicity-induced ROS can perturb motor system integrity through a circuit-dependent mechanism.


Assuntos
Drosophila melanogaster/fisiologia , Retroalimentação Fisiológica , Ácido Glutâmico/toxicidade , Neurônios Motores/fisiologia , Neurotoxinas/toxicidade , Espécies Reativas de Oxigênio/metabolismo , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Neurônios Colinérgicos/efeitos dos fármacos , Neurônios Colinérgicos/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/efeitos dos fármacos , Interneurônios/efeitos dos fármacos , Interneurônios/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Neurônios Motores/efeitos dos fármacos , Mutação/genética , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Junção Neuromuscular/efeitos dos fármacos , Junção Neuromuscular/metabolismo , Estresse Oxidativo/efeitos dos fármacos
6.
Cell Mol Neurobiol ; 39(8): 1177-1186, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31270710

RESUMO

Oxidative glutamate toxicity plays a vital role in the neurodegeneration diseases, including Alzheimer's diseases (AD). This study set out with the aim to investigate the beneficial effects of fangchinoline (FAN), a natural alkaloid, against glutamate-induced oxidative damage, and to clarify the underlying cellular and biochemical mechanisms. FAN prevented HT22 cells death from oxidative glutamate cytotoxicity in a dose-dependent manner, and significantly attenuated the overproduction of intracellular reactive oxygen species (ROS) and reversed the reduction of superoxide dismutase (SOD) activity induced by glutamate. Further investigations on the underlying mechanisms demonstrated that FAN potently up-regulated the protein level of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase (HO-1), in glutamate-exposed HT22 cells. The protective effects of FAN were almost completely antagonized by inhibitor of Nrf2. Subsequent studies revealed that FAN could down-regulate Kelch-like ECH-associated protein 1 (Keap1) in both mRNA level and protein level. To sum up, our result demonstrated the protective effects of FAN against glutamate-induced oxidative neuronal damage, and for the first time clarified the anti-oxidative mechanisms of FAN involve activating endogenous antioxidant defense system including enhancing SOD activity and regulating Keap1/Nrf-2 antioxidation signaling through modulation of Keap1 expression. Above results shed more light on the molecular mechanisms of FAN's neuroprotective effects, and may provide important clues for the drug development in preventing oxidative stress-associated neurodegenerative diseases.


Assuntos
Alcaloides/farmacologia , Benzilisoquinolinas/farmacologia , Ácido Glutâmico/toxicidade , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Fármacos Neuroprotetores/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais , Animais , Morte Celular/efeitos dos fármacos , Linhagem Celular , Heme Oxigenase-1/metabolismo , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Superóxido Dismutase/metabolismo
7.
Int J Mol Sci ; 20(14)2019 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-31323752

RESUMO

The dried flowers of Chionanthus retusus were extracted with 80% MeOH, and the concentrate was divided into EtOAc, n-BuOH, and H2O fractions. Repeated SiO2, octadecyl SiO2 (ODS), and Sephadex LH-20 column chromatography of the EtOAc fraction led to the isolation of four flavonols (1-4), three flavones (5-7), four flavanonols (8-11), and one flavanone (12), which were identified based on extensive analysis of various spectroscopic data. Flavonoids 4-6 and 8-11 were isolated from the flowers of C. retusus for the first time in this study. Flavonoids 1, 2, 5, 6, 8, and 10-12 significantly inhibited NO production in RAW 264.7 cells stimulated by lipopolysaccharide (LPS) and glutamate-induced cell toxicity and effectively increased HO-1 protein expression in mouse hippocampal HT22 cells. Flavonoids with significant neuroprotective activity were also found to recover oxidative-stress-induced cell damage by increasing HO-1 protein expression. This article demonstrates that flavonoids from C. retusus flowers have significant potential as therapeutic materials in inflammation and neurodisease.


Assuntos
Flavonoides/farmacologia , Flores/química , Ácido Glutâmico/toxicidade , Oleaceae/química , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Heme Oxigenase-1/metabolismo , Lipopolissacarídeos/farmacologia , Camundongos , Células RAW 264.7 , Transdução de Sinais/efeitos dos fármacos
8.
J Vet Med Sci ; 81(7): 1047-1054, 2019 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-31092742

RESUMO

Quercetin is a plant flavonoid that has anti-oxidant, anti-inflammatory, anti-cancer, and anti-ischemic properties. Moreover, quercetin exerts neuroprotective effects against focal cerebral ischemia. Protein phosphatase 2A (PP2A) is a form of serine/threonine phosphatase that modulates various biological functions. Among PP2A subunit types, subunit B exists abundantly in brain tissue and plays an essential function in nervous system. We previously reported the decrease of PP2A subunit B in focal cerebral animal model. This study explored the change of PP2A subunit B expression by quercetin treatment in cerebral ischemic animal model and glutamate-treated hippocampal-derived (HT22) cell culture. Quercetin (10 mg/kg) or vehicle was injected intraperitoneally into male rats before 30 min of middle cerebral artery occlusion (MCAO), and cerebral cortices were isolated 24 hr after MCAO. MCAO induced the neurological behavioral deficit and increased infarct volume. However, quercetin treatment attenuated the increase of neurological deficit and infarction. We detected the alleviation of MCAO-induced the decrease in PP2A subunit B by quercetin treatment using a proteomic approach. Reverse-transcription PCR and Western blot analyses confirmed lower PP2A subunit B expression levels in MCAO group with vehicle. However, quercetin treatment attenuated MCAO-induced this reduction. We also observed the neuroprotective effect of quercetin and the change of PP2A subunit B expression in glutamate-exposed HT22 cells. Glutamate exposure dramatically reduced cell viability and PP2A subunit B expression, and quercetin treatment significantly improved these decreases. We clearly showed that quercetin performs a neuroprotective function and modulates down-regulation of PP2A subunit B against MCAO injury and glutamate toxicity. Thus, our finding suggests that the regulation of PP2A subunit B by quercetin contributes to neuroprotective function in ischemic brain injury.


Assuntos
Isquemia Encefálica/prevenção & controle , Ácido Glutâmico/toxicidade , Fármacos Neuroprotetores/farmacologia , Proteína Fosfatase 2/metabolismo , Quercetina/farmacologia , Animais , Isquemia Encefálica/enzimologia , Isquemia Encefálica/etiologia , Linhagem Celular Transformada , Modelos Animais de Doenças , Infarto da Artéria Cerebral Média/complicações , Infarto da Artéria Cerebral Média/metabolismo , Masculino , Camundongos , Proteína Fosfatase 2/genética , Ratos Sprague-Dawley
9.
J Neuroinflammation ; 16(1): 91, 2019 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-30995916

RESUMO

BACKGROUND: During excitotoxic damage, neuronal death results from the increase in intracellular calcium, the induction of oxidative stress, and a subsequent inflammatory response. NADPH oxidases (NOX) are relevant sources of reactive oxygen species (ROS) during excitotoxic damage. NADPH oxidase-2 (NOX-2) has been particularly related to neuronal damage and death, as well as to the resolution of the subsequent inflammatory response. As ROS are crucial components of the regulation of inflammatory response, in this work, we evaluated the role of NOX-2 in the progression of inflammation resulting from glutamate-induced excitotoxic damage of the striatum in an in vivo model. METHODS: The striata of wild-type C57BL/6 J and NOX-2 KO mice (gp91Cybbtm1Din/J) were stereotactically injected with monosodium glutamate either alone or in combination with IL-4 or IL-10. The damage was evaluated in histological sections stained with cresyl violet and Fluoro-Jade B. The enzymatic activity of caspase-3 and NOX were also measured. Additionally, the cytokine profile was identified by ELISA and motor activity was verified by the tests of the cylinder, the adhesive tape removal, and the inverted grid. RESULTS: Our results show a neuroprotective effect in mice with a genetic inhibition of NOX-2, which is partially due to a differential response to excitotoxic damage, characterized by the production of anti-inflammatory cytokines. In NOX-2 KO animals, the excitotoxic condition increased the production of interleukin-4, which could contribute to the production of interleukin-10 that decreased neuronal apoptotic death and the magnitude of striatal injury. Treatment with interleukin-4 and interleukin-10 protected from excitotoxic damage in wild-type animals. CONCLUSIONS: The release of proinflammatory cytokines during the excitotoxic event promotes an additional apoptotic death of neurons that survived the initial damage. During the subsequent inflammatory response to excitotoxic damage, ROS generated by NOX-2 play a decisive role in the extension of the lesion and consequently in the severity of the functional compromise, probably by regulating the anti-inflammatory cytokines production.


Assuntos
Corpo Estriado/enzimologia , Corpo Estriado/patologia , Inflamação/enzimologia , Inflamação/patologia , NADPH Oxidase 2/metabolismo , Animais , Corpo Estriado/imunologia , Progressão da Doença , Ácido Glutâmico/toxicidade , Inflamação/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
10.
Neurotox Res ; 36(3): 551-562, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31016690

RESUMO

Mitochondria are the major site of adenosine triphosphate (ATP) production in mammalian cells. Moreover, mitochondria produce most of the reactive oxygen species (ROS) in nucleated cells. Redox and bioenergetic abnormalities have been seen in mitochondria during the onset and progression of neurodegenerative diseases. In that context, excitotoxicity induced by glutamate (GLU) plays an important role in mediating neurotoxicity. Several drugs have been used in the treatment of diseases involving excitotoxicity. Nonetheless, some patients (20-30%) present drug resistance. Thus, it is necessary to find chemicals able to attenuate mitochondrial dysfunction in the case of excitotoxicity. In this work, we treated the human neuroblastoma SH-SY5Y cell line with the diterpene carnosic acid (CA) at 1 µM for 12 h prior to the exposure to GLU for further 24 h. We found that CA prevented the GLU-induced mitochondrion-related redox impairment and bioenergetic decline in SH-SY5Y cells. CA also downregulated the pro-apoptotic stimulus elicited by GLU in this experimental model. CA exerted mitochondrial protection by a mechanism associated with the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2), since silencing of this protein with small interfering RNA (siRNA) suppressed the CA-induced protective effects. Future directions include investigating whether CA would be able to modulate mitochondrial function and/or dynamics in in vivo experimental models of excitotoxicity.


Assuntos
/farmacologia , Ácido Glutâmico/toxicidade , Mitocôndrias/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Neuroblastoma , Óxido Nítrico/metabolismo , Oxirredução/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismo
11.
J Asian Nat Prod Res ; 21(5): 409-418, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30924351

RESUMO

Seven natural compounds, including new compounds hyperascyrins L-N (1-3) and four known compounds (4-7), were acquired from the aerial parts of Hypericum ascyron, that were all identified as methylated polycyclic polyprenylated acylphloroglucinol derivatives (mPPAPs). The structures of these compounds were established by NMR spectroscopy, experimental and calculated electronic circular dichroism (ECD) data. The neuroprotective activities and hepatoprotective activity of these compounds (10 µM) were evaluated. Compounds 1, 2 and 3 exhibited neuroprotection activity. Compounds 1 and 3 show hepatoprotective activity.


Assuntos
Hypericum/química , Floroglucinol/análogos & derivados , Componentes Aéreos da Planta/química , Acetaminofen/toxicidade , Analgésicos/toxicidade , Linhagem Celular , Ácido Glutâmico/toxicidade , Hepatócitos/efeitos dos fármacos , Humanos , Modelos Moleculares , Estrutura Molecular , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/farmacologia , Floroglucinol/química , Floroglucinol/farmacologia
12.
J Ethnopharmacol ; 236: 302-315, 2019 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-30872169

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Zhi-zi-chi Decoction(ZZCD), a traditional Chinese medicine formula, has been reported its potential protective effect on psychological sub-health diseases. However, there still remains a lack of molecular mechanism interpretation. AIM OF THE STUDY: This study was aimed at investigating the mechanism of glutamate-induced toxicity in PC12 cells and the neuroprotective effect of ZZCD based on a novel strategy of the combination of cell metabolomics and pharmacology. MATERIALS AND METHODS: The PC12 cells were treated with glutamate to simulate neurotoxic cell model. Gas chromatography coupled with mass spectrometry based on cell metabolomics approach was performed to comprehensively investigate the molecular mechanism of glutamate-induced toxicity The cell viability and cytotoxicity analysis, the determination of glutathione reductase(GR), superoxide dismutase(SOD) and reactive oxygen species(ROS), apoptosis analysis and western blot analysis were performed to evaluate the neuroprotection of ZZCD. RESULTS: Forty metabolites were identified as potential biomarkers in model cells by principal components analysis(PCA) and orthogonal partial least squares-discriminant analysis(OPLS-DA). Glutamate decreased the GR and SOD activities, increased the level of intracellular ROS, activated the apoptotic pathway, and induced the changes of energy metabolism, amino acid metabolism and lipid metabolism. In addition, the extract of ZZCD could reverse the disturbed metabolic pathways by regulating those potential biomarkers and exerted anti-oxidation and anti-apoptosis. CONCLUSION: ZZCD has neuroprotective effect and the novel strategy can be applicable for other traditional Chinese medicine formulas.


Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Gardenia/química , Fármacos Neuroprotetores/farmacologia , Soja/química , Animais , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Frutas/química , Ácido Glutâmico/toxicidade , Glutationa Redutase/metabolismo , Medicina Tradicional Chinesa , Redes e Vias Metabólicas/efeitos dos fármacos , Metabolômica/métodos , Células PC12 , Ratos , Espécies Reativas de Oxigênio/metabolismo , Sementes/química , Superóxido Dismutase/metabolismo
14.
Int J Mol Med ; 43(5): 2144-2152, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30896788

RESUMO

The aim of this study was to investigate the antioxidant and anti­apoptotic activities, as well as the underlying mechanisms of action, of Scrophularia buergeriana (S. buergeriana) extract (SBE) in glutamate­induced SH­SY5Y cell death. The roots of S. buergeriana were extracted with 70% ethanol, and standardized SBE was used in this study. To induce cytotoxicity, the SH­SY5Y cells were exposed to glutamate for 3 h, or pre­treated with SBE for 1 h, and subsequently incubated with glutamate for 3 h. The neuroprotective effects were assessed by measuring cell viability and the total glutathione contents using commercial kits. The antioxidant and anti­apoptotic mechanisms of action of SBE were evaluated by western blot analysis. The results confirmed that glutamate­induced toxicity was caused by reactive oxygen species (ROS) production, leading to oxidative stress and DNA damage, thus leading to cell death. However, treatment of the SH­SY5Y cells with SBE significantly increased the viability of the cells exposed to glutamate by upregulating the levels of antioxidant proteins, such as superoxide dismutase (SOD)1, SOD2 and glutathione peroxidase­1 (GPx­1), and directly enhancing the total glutathione contents. Furthermore, SBE attenuated DNA impairment and decreased B­cell lymphoma-2 (Bcl­2)­associated X protein (Bax), cleaved caspase­3 and cleaved poly(adenosine diphosphate (ADP)­ribose) polymerase (PARP) activation. In addition, SBE upregulated Bcl­2 expression via p38 mitogen­activated protein kinases (MAPKs). On the whole, the findings of this study demonstrated that SBE exerts neuroprotective effects against glutamate­induced cell toxicity through its antioxidant and anti­apoptotic activities.


Assuntos
Ácido Glutâmico/toxicidade , Fármacos Neuroprotetores/farmacologia , Extratos Vegetais/farmacologia , Scrophularia/química , Acetilcolinesterase/metabolismo , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Fragmentação do DNA/efeitos dos fármacos , Glutationa/metabolismo , Humanos , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
15.
Front Neural Circuits ; 13: 11, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30858799

RESUMO

Parkinson's disease (PD) is a neurodegenerative disease associated with progressive and inexorable loss of dopaminergic cells in Substantia Nigra pars compacta (SNc). Although many mechanisms have been suggested, a decisive root cause of this cell loss is unknown. A couple of the proposed mechanisms, however, show potential for the development of a novel line of PD therapeutics. One of these mechanisms is the peculiar metabolic vulnerability of SNc cells compared to other dopaminergic clusters; the other is the SubThalamic Nucleus (STN)-induced excitotoxicity in SNc. To investigate the latter hypothesis computationally, we developed a spiking neuron network-model of SNc-STN-GPe system. In the model, prolonged stimulation of SNc cells by an overactive STN leads to an increase in 'stress' variable; when the stress in a SNc neuron exceeds a stress threshold, the neuron dies. The model shows that the interaction between SNc and STN involves a positive-feedback due to which, an initial loss of SNc cells that crosses a threshold causes a runaway-effect, leading to an inexorable loss of SNc cells, strongly resembling the process of neurodegeneration. The model further suggests a link between the two aforementioned mechanisms of SNc cell loss. Our simulation results show that the excitotoxic cause of SNc cell loss might initiate by weak-excitotoxicity mediated by energy deficit, followed by strong-excitotoxicity, mediated by a disinhibited STN. A variety of conventional therapies were simulated to test their efficacy in slowing down SNc cell loss. Among them, glutamate inhibition, dopamine restoration, subthalamotomy and deep brain stimulation showed superior neuroprotective-effects in the proposed model.


Assuntos
Simulação por Computador , Neurônios Dopaminérgicos/efeitos dos fármacos , Agonistas de Aminoácidos Excitatórios/toxicidade , Ácido Glutâmico/toxicidade , Modelos Neurológicos , Doença de Parkinson/patologia , Neurônios Dopaminérgicos/fisiologia , Humanos
16.
J Agric Food Chem ; 67(7): 1831-1838, 2019 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-30742443

RESUMO

Roots of Glehnia littoralis have been used to heal stroke as a traditional medicine. Even though many studies on this plant have been conducted, the secondary metabolites produced by its endophytes and their bioactivities have not been investigated thus far. Therefore, a new meroditerpenoid named sartorypyrone E (1) and eight known compounds (2-9) were isolated from extracts of cultured Neosartorya fischeri JS0553, an endophyte of G. littoralis. The isolated metabolites were identified using spectroscopic methods and chemical reaction, based on a comparison to literature data. Relative and absolute stereochemistries of compound 1 were also elucidated. To identify the protective effects of isolated compounds (1-9) in HT22 cells against glutamate-induced cytotoxicity, we assessed inhibition of cell death, intracellular reactive oxygen species (ROS) accumulation, and calcium ion (Ca2+) influx. Among the isolates, compound 8, identified as fischerin, showed significant neuroprotective activity on glutamate-mediated HT22 cell death through inhibition of ROS, Ca2+ influx, and phosphorylation of mitogen-activated protein kinase, including c-Jun N-terminal kinase, extracellular signal-regulated kinase, and p38. The results suggested that the metabolites produced by the endophyte N. fischeri JS0553 might be related to the neuroprotective activity of its host plant, G. littoralis.


Assuntos
Apiaceae/microbiologia , Neosartorya/metabolismo , Fármacos Neuroprotetores/metabolismo , Animais , Cálcio/metabolismo , Morte Celular/efeitos dos fármacos , Linhagem Celular Transformada , Ácido Glutâmico/toxicidade , Hipocampo , Camundongos , Proteínas Quinases Ativadas por Mitógeno/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Estrutura Molecular , Naftalenos/isolamento & purificação , Naftalenos/farmacologia , Neosartorya/isolamento & purificação , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/farmacologia , Fosforilação/efeitos dos fármacos , Piridonas/isolamento & purificação , Piridonas/farmacologia , Pironas/metabolismo , Pironas/farmacologia , Espécies Reativas de Oxigênio/antagonistas & inibidores
17.
Mol Cell Neurosci ; 95: 71-78, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30738184

RESUMO

Interleukin 16 (IL-16) is a cytokine that is primarily associated with CD4+ T cell function, but also exists as a multi-domain PDZ protein expressed within cerebellar and hippocampal neurons. We have previously shown that lymphocyte-derived IL-16 is neuroprotective against excitotoxicity, but evidence of how it affects neuronal function is limited. Here, we have investigated whether IL-16 modulates neuronal excitability and synaptic activity in mouse primary hippocampal cultures. Application of recombinant IL-16 impairs both glutamate-induced increases in intracellular Ca2+ and sEPSC frequency and amplitude in a CD4- and CD9-independent manner. We examined the mechanisms underlying these effects, with rIL-16 reducing GluA1 S831 phosphorylation and inhibiting Na+ channel function. Taken together, these data suggest that IL-16 reduces neuronal excitability and synaptic activity via multiple mechanisms and adds further evidence that alternative receptors may exist for IL-16.


Assuntos
Potenciais Pós-Sinápticos Excitadores , Interleucina-16/farmacologia , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Receptores de AMPA/metabolismo , Canais de Sódio/metabolismo , Animais , Antígenos CD4/metabolismo , Cálcio/metabolismo , Células Cultivadas , Ácido Glutâmico/toxicidade , Hipocampo/citologia , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Neurônios/fisiologia , Fosforilação , Tetraspanina 29/metabolismo
18.
Neurosci Lett ; 701: 58-64, 2019 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-30790645

RESUMO

Prolactin (PRL) is a pleiotropic hormone secreted by several cells and tissues in the body, such as mammary glands, T-lymphocytes, hypothalamus, among others. This hormone possess neuroprotective properties against glutamate-excitotoxicity through the activation of NF-kB, suggesting it could exert an antioxidant action. However, the role of PRL on the antioxidant defense during glutamate-induced excitotoxicity is not clear to date. Therefore, in the present study, we have evaluated the effect of PRL on SOD activity and protein content of both of its isoforms (Mn2+-SOD and Cu2+/Zn2+-SOD), as well as, its action on mitochondrial activity in primary culture of hippocampal neurons of rats. Additionally, we have evaluated the possible antioxidant effect of PRL through the determination of lipid peroxidation products (LPO), measured as malondialdehyde (MDA). Results show that PRL enhances the activity and the protein content of Mn2+-SOD and Cu2+/Zn2+-SOD in neurons exposed to glutamate-induced excitotoxicity. Moreover, our results demonstrate that PRL prevents mitochondrial dysfunction induced by glutamate and significantly decreases the levels of LPO products. To our knowledge, this is the first time that a potential antioxidant effect of PRL has been described in hippocampal neurons exposed to glutamate excitotoxicity, opening questions of its potentiality for therapeutics.


Assuntos
Ácido Glutâmico/toxicidade , Hipocampo/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Prolactina/farmacologia , Animais , Antioxidantes/farmacologia , Autofagia/efeitos dos fármacos , Hipocampo/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Malondialdeído/metabolismo , Neuroproteção/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Cultura Primária de Células , Ratos , Ratos Wistar , Superóxido Dismutase/metabolismo
19.
J Nat Med ; 73(3): 672-678, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30778894

RESUMO

Neurological disorders are a public health problem worldwide for which there is currently no direct treatment of the cause of the disorder. The goal of this study was to investigate the potential in vitro neuroprotective property of plants used in Mayan traditional medicine. Plant ethanolic extracts were prepared and tested on models in which neuronal damage was induced by glutamate, i.e., a human neuroblastoma cell line (SH-SY5Y) and rat cortical neurons. HPLC profiles from active extracts were also obtained. A total of 51 plant species were identified in the literature as plant species used in Mayan traditional medicine for the treatment of symptoms suggestive of neurological disorders, and we studied 34 of these in our analysis. Six extracts had a neuroprotective effect on SH-SY5Y cells, with the most active extract being that from Schwenckia americana roots (half maximal effective concentration [EC50] 11.3 ± 2.9 µg/mL), and three extracts exhibited a neuroprotective effect in the rat neuron cortical model, with the most active extract being that from Elytraria imbricata aerial parts (EC50 6.8 ± 3.1 µg/mL). These results suggest that the active extracts from such plants have the potential to be a great resource. Future studies should be performed that are more extensive and which isolate the active constituents.


Assuntos
Ácido Glutâmico/toxicidade , Fármacos Neuroprotetores/uso terapêutico , Extratos Vegetais/química , Plantas Medicinais/química , Animais , Humanos , Fármacos Neuroprotetores/farmacologia , Ratos , Ratos Wistar
20.
Neurosci Lett ; 701: 106-111, 2019 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-30807795

RESUMO

Oxidative damage in neurons including glutamate excitotoxicity has been linked to increasing numbers of neuropathological conditions. Under these conditions, cells trigger several different cellular responses such as autophagy, apoptosis, necrosis and senescence. However, the connection between these responses is not well understood. In this study, we found that the 60-kDa BECN1 was specifically degraded to a 40-kDa fragment in hippocampal HT22 cells treated with 5 mM glutamate. Increased BECN1 cleavage was specifically associated with a decrease in cell viability under oxidative stress. Interestingly, this BECN1 cleavage was specifically inhibited by a calpain inhibitor ALLN but was not affected by other protease inhibitors. Also, the BECN1 cleavage was not detected in calpain-4-deficient cell lines. Furthermore, calpain cleaved BECN1 at a specific site between the coiled-coil domain and Bcl2 homology 3 domain, which is associated with the anti-apoptotic protein Bcl-2. Moreover, some cellular senescence markers, including ß-galactosidase, p21, p27Kip1, p53 and p16INK4A, increased proportionally to those of BECN1 cleaved fragments. These results suggest that calpain-mediated BECN1 cleavage under oxidative conditions is specifically associated with cell death induced by cellular senescence.


Assuntos
Proteína Beclina-1/metabolismo , Calpaína/metabolismo , Hipocampo/metabolismo , Estresse Oxidativo/fisiologia , Animais , Apoptose/fisiologia , Calpaína/antagonistas & inibidores , Calpaína/deficiência , Caspases/metabolismo , Linhagem Celular , Sobrevivência Celular/fisiologia , Senescência Celular/fisiologia , Ácido Glutâmico/toxicidade , Células HeLa , Hipocampo/patologia , Humanos , Leupeptinas/farmacologia , Camundongos , Células NIH 3T3 , Neurônios/metabolismo , Neurônios/patologia , Espécies Reativas de Oxigênio/metabolismo
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