RESUMEN
Ceramides are essential lipids for skin barrier function, and various classes and species exist in the human stratum corneum (SC). To date, the relationship between skin conditions and ceramide composition in healthy individuals has remained largely unclear. In the present study, we measured six skin condition parameters (capacitance, transepidermal water loss, scaliness, roughness, multilayer exfoliation, and corneocyte cell size) for the SC of the cheeks and upper arms of 26 healthy individuals and performed correlation analyses with their SC ceramide profiles, which we measured via liquid chromatography-tandem mass spectrometry. In the cheeks, high levels and/or ratios of two free ceramide classes containing an extra hydroxyl group in the long-chain moiety and a protein-bound ceramide class containing 6-hydroxysphingosine correlated with healthy skin conditions. In contrast, the ratios of two other free ceramide classes, both containing sphingosine, and a protein-bound ceramide class containing 4,14-sphingadiene correlated with unhealthy skin conditions, as did shortening of the carbon chain of the fatty acid portion of two ceramide classes containing non-hydroxy fatty acids. Thus, our findings help to elucidate the relationship between skin conditions and ceramide composition.
Asunto(s)
Ceramidas , Epidermis , Humanos , Ceramidas/metabolismo , Ceramidas/análisis , Femenino , Masculino , Epidermis/metabolismo , Adulto , Persona de Mediana Edad , Piel/metabolismo , Voluntarios Sanos , Espectrometría de Masas en Tándem , Adulto JovenRESUMEN
Bcl-rambo, also known as BCL2L13, has been reported to regulate apoptosis, mitochondrial fragmentation, and mitophagy. However, the molecular mechanisms by which Bcl-rambo regulates these processes currently remain unclear. In the present study, we identified phosphoglycerate mutase member 5 (PGAM5) as an emerging partner interacting with Bcl-rambo through phenotypic Drosophila screening. The rough eye phenotype induced by human Bcl-rambo was partly rescued by the knockdown of pgam5-2, a mammalian ortholog of PGAM5. Bcl-rambo bound to PGAM5, and their interaction required the Bcl-rambo transmembrane domain. The co-expression of Bcl-rambo and PGAM5 promoted effector caspase activity in human embryonic kidney 293T cells. The transient overexpression of Bcl-rambo increased LC3B-II levels, which had been decreased by the co-expression of PGAM5. These results suggest that PGAM5 promotes Bcl-rambo-dependent apoptosis, but conversely interferes with Bcl-rambo-dependent mitophagy.
Asunto(s)
Mitofagia , Fosfoglicerato Mutasa , Animales , Apoptosis/genética , Caspasas Efectoras/metabolismo , Drosophila/metabolismo , Humanos , Mamíferos/metabolismo , Mitocondrias/metabolismo , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Mitofagia/genética , Fosfoglicerato Mutasa/metabolismo , Fosfoproteínas Fosfatasas/genética , Fosfoproteínas Fosfatasas/metabolismoRESUMEN
We investigated the content of phenolic compounds and antioxidant capacity of two batches of non-heated and heated leaves of the yacon cultivar "Andes no yuki", grown in Japan. Lyophilized yacon leaves heated at 160°C for 20 min and 100°C for 60 min had a 1.96 to 9.69-times higher total phenolic content than that of the non-heated leaves. Heated leaves exhibited a 1.98 to 4.07-times higher antioxidant capacity than that of the non-heated leaves in three different free radical scavenging assays. Heated leaves were more efficient at attenuating the superoxide anion radical production in human granulocytic cells than the non-heated leaves. High-performance liquid chromatography analysis revealed that, in the heated leaves, the caffeic acid content was 2.13 to 3.64-times higher and the chlorogenic acid content was slightly lower than those in the non-heated leaves. Hence, heat processing may affect the active constituent contents in yacon leaves, potentiating its antioxidant capacity.Abbreviations: ABTS+: 2,2'-azinobis(2-ethylbenzothiazoline-6-sulfonic acid) cation; DPPH: 1,1-diphenyl-2-picrylhydrazyl, HPLC: high-performance liquid chromatography; NBT: nitroblue tetrazolium; O2-: superoxide anion; PMA: phorbol 12-myristate 13-acetate; PMS: phenazine methosulfate; TEAC: Trolox equivalent antioxidant capacity.
Asunto(s)
Antioxidantes/farmacología , Asteraceae/química , Calor , Fenoles/análisis , Hojas de la Planta/química , Ácidos Cafeicos/análisis , Ácido Clorogénico/análisis , Cromatografía Líquida de Alta Presión , Depuradores de Radicales Libres/farmacología , Células HL-60 , Humanos , Extractos Vegetales/farmacología , Superóxidos/metabolismoRESUMEN
Carbamazepine (CBZ) binds adenosine receptors, but detailed effects of CBZ on astroglial transmission associated with adenosine receptor still need to be clarified. To clarify adenosinergic action of CBZ on astroglial transmission, primary cultured astrocytes were acutely or chronically treated with CBZ, proinflammatory cytokines (interferon γ (IFNγ) and tumor necrosis factor α (TNFα)), and adenosine A2A receptor (A2AR) agonist (CGS21680). IFNγ and TNFα increased basal, adenophostin-A (AdA)-evoked, and 2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid (AMPA)-evoked astroglial L-glutamate releases. In physiological condition, CGS21680 increased basal astroglial L-glutamate release but glutamate transporter inhibition prevented this CGS21680 action. CBZ did not affect basal release, whereas glutamate transporter inhibition generated CBZ-induced glutamate release. Furthermore, AdA-evoked and AMPA-evoked releases were inhibited by CBZ but were unaffected by CGS21680. Contrary to physiological condition, chronic administrations of IFNγ and TNFα enhanced basal, AdA-, and AMPA-evoked releases, whereas IFNγ and TNFα decreased and increased CGS21680-evoked releases via modulation A2AR expression. Both chronic administration of CGS21680 and CBZ suppressed astroglial L-glutamate release responses induced by chronic cytokine exposer. Especifically, chronic administration of CBZ and CGS21680 prevented the reduction and elevation of A2AR expression by respective IFNγ and TNFα. These findings suggest that A2AR agonistic effects of CBZ contribute to chronic prevention of pathomechanisms developments of several neuropsychiatric disorders associated with proinflammatory cytokines.
Asunto(s)
Astrocitos/efectos de los fármacos , Carbamazepina/farmacología , Ácido Glutámico/metabolismo , Receptor de Adenosina A2A/metabolismo , Adenosina/análogos & derivados , Adenosina/farmacología , Agonistas del Receptor de Adenosina A2/farmacología , Animales , Astrocitos/metabolismo , Células Cultivadas , Interferón gamma/farmacología , Fenetilaminas/farmacología , Ratas , Ratas Sprague-Dawley , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
Zebrafish have superior abilities to generate new neurons in the adult brain and to regenerate brain tissue after brain injury compared with mammals. There exist two types of neural stem cells (NSCs): neuroepithelial-like stem cells (NE) and radial glia (RG) in the optic tectum. We established an optic tectum stab injury model to analyze the function of NSCs in the regenerative condition and confirmed that the injury induced the proliferation of RG, but not NE and that the proliferated RG differentiated into new neurons after the injury. We then analyzed the involvement of Wnt signaling after the injury, using a Wnt reporter line in which canonical Wnt signaling activation induced GFP expression and confirmed that GFP expression was induced specifically in RG after the injury. We also analyzed the expression level of genes related to Wnt signaling, and confirmed that endogenous Wnt antagonist dkk1b expression was significantly decreased after the injury. We observed that Wnt signal inhibitor IWR1 treatment suppressed the proliferation and differentiation of RG after the injury, suggesting that up-regulation of Wnt signaling in RG after the stab injury was required for optic tectum regeneration. We also confirmed that Wnt activation by treatment with GSK3ß inhibitor BIO in uninjured zebrafish induced proliferation of RG in the optic tectum. This optic tectum stab injury model is useful for the study of the molecular mechanisms of brain regeneration and analysis of the RG functions in physiological and regenerative conditions.
Asunto(s)
Células Ependimogliales/fisiología , Regeneración Nerviosa/fisiología , Colículos Superiores/lesiones , Colículos Superiores/fisiopatología , Vía de Señalización Wnt/fisiología , Heridas Punzantes/fisiopatología , Animales , Animales Modificados Genéticamente , Diferenciación Celular/fisiología , Proliferación Celular/fisiología , Células Ependimogliales/patología , Glucógeno Sintasa Quinasa 3 beta/antagonistas & inhibidores , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Neuronas/patología , Neuronas/fisiología , Colículos Superiores/patología , Heridas Punzantes/patología , Pez Cebra , Proteínas de Pez Cebra/metabolismoRESUMEN
This study aimed to investigate the unique antioxidative effects of Japanese moringa products, herbal leaf tea and stem tea, using established free radical assays, focusing on superoxide anion (O2-) radical generation systems. Hot-water extracts from moringa teas resulted in different but lower scavenging activities than Trolox in four synthetic free radical models. Interestingly, these extracts further showed higher O2- radical scavenging effects than Trolox in the phenazine methosulfate-NADH-nitroblue tetrazolium and xanthine oxidase assay systems. Incubating human neutrophils in the presence of these tea extracts rather than Trolox effectively suppressed cellular O2- radical generation. Among the eight known phenolic constituents of moringa leaves, caffeic acid and chlorogenic acid may be responsible for the O2-specific radical scavenging capacity stronger than that of Trolox. These results suggest that moringa herbal teas are a good source of natural antioxidants for preventing O2- radical-mediated disorders. Abbreviations: O2-: superoxide anion; ROS: reactive oxygen species; H2O2: hydrogen peroxide; XOD: xanthine oxidase; DPPH: 1,1-diphenyl-2-picrylhydrazyl; ABTS+: 2,2'-azinobis(2-ethylbenzothiazoline-6-sulfonic acid) cation; CPZ+: chlorpromazine cation; PMS: phenazine methosulfate; NBT: nitroblue tetrazolium; PMA: phorbol 12-myristate 13-acetate.
Asunto(s)
Depuradores de Radicales Libres/farmacología , Moringa oleifera/química , Extractos Vegetales/farmacología , Hojas de la Planta/química , Tallos de la Planta/química , Superóxidos/metabolismo , Tés de Hierbas , Ácidos Cafeicos/análisis , Ácidos Cafeicos/farmacología , Ácido Clorogénico/análisis , Ácido Clorogénico/farmacología , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Polifenoles/análisisRESUMEN
AIM: The intracerebral antioxidant ability of mature rats after neonatal hypoxic-ischemic (HI) brain injury was estimated using the microdialysis-electron spin resonance method. MATERIAL AND METHODS: Seven-day-old Wistar rats were subjected to a modified Levine's procedure for producing HI brain injury. After HI insult, pups were returned and reared with their dams. Seven weeks after HI insult, their intracerebral antioxidant abilities were measured using the microdialysis-electron spin resonance method after the intraperitoneal injection of 3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl. Ascorbic acid, L-cysteine, and glutathione (GSH) were also determined. The rats without HI insult were used as a control. RESULTS: The decay rate of 3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl in the non-ligated side of the cerebral hemisphere of the HI group was significantly larger than that of the control group. The amounts of ascorbic acid in the perfusate from the non-ligated side of the HI group were about four times larger than those of the control group. The amounts of L-cysteine and GSH of the HI group were about 10 times larger than those of the control group. CONCLUSIONS: The antioxidant ability in the non-ligated sides of the cerebral hemispheres of the mature rats 7 weeks after neonatal HI insult was higher than that of the control group. Higher amounts of ascorbic acid and GSH supported the higher antioxidant ability. The increase of the intracerebral antioxidant ability of the non-ligated side indicates the compensation of motor function for the lost side. The present results should offer important insights into the prognosis for hypoxic-ischemic encephalopathy.
Asunto(s)
Antioxidantes/metabolismo , Traumatismos del Nacimiento/metabolismo , Corteza Cerebral/metabolismo , Modelos Animales de Enfermedad , Hipoxia-Isquemia Encefálica/metabolismo , Neuronas/metabolismo , Estrés Oxidativo , Animales , Antioxidantes/análisis , Ácido Ascórbico/sangre , Ácido Ascórbico/metabolismo , Traumatismos del Nacimiento/sangre , Traumatismos del Nacimiento/fisiopatología , Corteza Cerebral/irrigación sanguínea , Corteza Cerebral/lesiones , Óxidos N-Cíclicos/análisis , Cisteína/sangre , Cisteína/metabolismo , Progresión de la Enfermedad , Espectroscopía de Resonancia por Spin del Electrón , Glutatión/sangre , Glutatión/metabolismo , Humanos , Hipoxia-Isquemia Encefálica/sangre , Hipoxia-Isquemia Encefálica/fisiopatología , Recién Nacido , Microdiálisis , Pirrolidinas/análisis , Ratas Wistar , Marcadores de SpinRESUMEN
Traumatic brain injury occasionally causes posttraumatic epilepsy. To elucidate the molecular events responsible for posttraumatic epilepsy, we established a rodent model that involved the injection of microliter quantities of FeCl3 solution into the amygdalar nuclear complex. We previously compared hippocampal gene expression profiles in the traumatic epilepsy model and normal rats at 5 days after brain injury (acute phase) to determine the role of inflammation. In this study, we focused on later stages of epileptogenesis. We compared gene expression profiles at 5, 15 (sub-chronic phase), and 30 days (chronic phase) after brain injury to identify temporal changes in molecular networks involved in epileptogenesis. A total of 81 genes were significantly (at least twofold) up- or downregulated over the course of disease progression. We found that genes related to lipid metabolism, namely, Apoa1, Gh, Mc4r, Oprk1, and Pdk4, were temporarily upregulated in the sub-chronic phase. Changes in lipid metabolism regulation might be related to seizure propagation during epileptogenesis. This temporal description of hippocampal gene expression profiles throughout epileptogenesis provides clues to potential markers of disease phases and new therapeutic targets.
Asunto(s)
Modelos Animales de Enfermedad , Epilepsia Postraumática/genética , Perfilación de la Expresión Génica , Hipocampo/metabolismo , Metabolismo de los Lípidos/genética , Regulación hacia Arriba , Animales , RatasRESUMEN
The effect of the oral administration of mimosa tannin (MMT) on the rat intra-hippocampal antioxidant ability was examined. Wistar rats at the age of 6 weeks were reared for 8 weeks with the rodent diet (RD) consisting of 0.1 g/kg of MMT (RD-MMT). The antioxidant ability of rat brain was evaluated from the decay of a brain-blood-barrier permeable stable nitroxide, 3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl (PCAM) measured by the microdialysis-electron spin resonance system under a freely moving state. The decay rate of PCAM in the brain of rats fed RD-MMT was significantly larger than that of rats fed control rodent diet, which indicates the increase of the antioxidant ability in the brain of rats fed RD-MMT. In vitro study showed that MMT did not reduce PCAM directly but enhanced the reduction of PCAM by ascorbic acid. These results indicate that MMT is a potent antioxidant in vitro and in vivo.
Asunto(s)
Antioxidantes/metabolismo , Encéfalo/metabolismo , Taninos/administración & dosificación , Animales , Ácido Ascórbico/química , Barrera Hematoencefálica , Óxidos N-Cíclicos/química , Óxidos N-Cíclicos/farmacocinética , Espectroscopía de Resonancia por Spin del Electrón , Microdiálisis , Pirrolidinas/química , Pirrolidinas/farmacocinética , Ratas , Ratas WistarRESUMEN
Orexin (hypocretin) is a neuropeptide secreted from hypothalamic neurons that is known to be activated during motivated behaviors and active waking. Presently, our knowledge of orexin is mainly limited to the central nervous system, and the involvement of the orexin system in peripheral tissues has received little attention. In the present study, we analyzed the existence of the orexin system in the adrenal medulla, which is part of the sympathetic nervous system. Orexin and its receptors are expressed in the bovine adrenal medulla. Orexins stimulated intracellular calcium changes and epinephrine release from cultured bovine adrenal medullary cells. Applied orexin decreased expression of prepro-orexin, orexin receptor-1 and orexin receptor-2, suggesting negative feedback regulation in the adrenal gland. Our results indicate involvement of the orexin system in the sympathetic regulation of the adrenal medulla.
Asunto(s)
Médula Suprarrenal/metabolismo , Células Cromafines/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neuropéptidos/metabolismo , Receptores de Orexina/metabolismo , Animales , Calcio/metabolismo , Bovinos , Células Cultivadas , Epinefrina/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Factores de Transcripción NFATC/metabolismo , Neuropéptidos/genética , Receptores de Orexina/genética , Orexinas , ARN Mensajero/metabolismoRESUMEN
Allantopyrone A is an α-pyrone metabolite that was originally isolated from the endophytic fungus Allantophomopsis lycopodina KS-97. We previously demonstrated that allantopyrone A exhibits anti-cancer, anti-inflammatory, and neuroprotective activities. In the present study, we showed that allantopyrone A up-regulated the protein expression of hypoxia-inducible factor (HIF)-1α in human fibrosarcoma HT-1080 cells. It also up-regulated the mRNA expression of BNIP3 and ENO1, but not other HIF target genes or HIF1A. Allantopyrone A did not inhibit the prolyl hydroxylation of HIF-1α, but enhanced the ubiquitination of cellular proteins. Consistent with this result, chymotrypsin-like and trypsin-like proteasome activities were reduced, but not completely inactivated by allantopyrone A. Allantopyrone A decreased the amount of proteasome catalytic subunits. Therefore, the present results showed that allantopyrone A interfered with the degradation of HIF-1α protein by reducing proteasome activity in human fibrosarcoma HT-1080 cells.
Asunto(s)
Fibrosarcoma , Complejo de la Endopetidasa Proteasomal , Humanos , Pironas/farmacología , Fibrosarcoma/tratamiento farmacológico , Hipoxia , Subunidad alfa del Factor 1 Inducible por HipoxiaRESUMEN
The anti-epileptic drug zonisamide is reported to exert beneficial effects in patients with Parkinson's disease. To elucidate the pathophysiological mechanisms underlying the anti-parkinsonism effects of zonisamide, we examined the effect of zonisamide co-administered with levodopa in the striata of rats with 6-hydoroxydopamine hemiparkinsonism by using a DNA microarray for genome-wide gene expression profiling. We found that the expression of some genes related to metabolism and nervous system development and function were upregulated by zonisamide; expression of these genes was downregulated by levodopa. Furthermore, many genes related to the immune system and inflammation were downregulated by zonisamide, and their expression was upregulated by levodopa. These results indicate that zonisamide has a protective effect when co-administered with levodopa.
Asunto(s)
Anticonvulsivantes/administración & dosificación , Antiparkinsonianos/administración & dosificación , Cuerpo Estriado/efectos de los fármacos , Expresión Génica/efectos de los fármacos , Isoxazoles/administración & dosificación , Levodopa/administración & dosificación , Enfermedad de Parkinson Secundaria/tratamiento farmacológico , Enfermedad de Parkinson/tratamiento farmacológico , Animales , Cuerpo Estriado/metabolismo , Cuerpo Estriado/patología , Modelos Animales de Enfermedad , Quimioterapia Combinada , Perfilación de la Expresión Génica , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , Oxidopamina/farmacología , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/patología , Enfermedad de Parkinson Secundaria/genética , Enfermedad de Parkinson Secundaria/patología , Ratas , Ratas Wistar , ZonisamidaRESUMEN
Acetaminophen (APAP) and p-aminophenol (p-AP) are the analogous simple phenolic compounds that undergo sulfate conjugation (sulfation) by cytosolic sulfotransferases. Sulfation is generally thought to lead to the inactivation and disposal of endogenous as well as xenobiotic compounds. This study aimed to investigate the antioxidative effects of O-sulfated form of APAP and p-AP, i.e., APAPS and p-APS, in comparison with their unsulfated counterparts. Using a 1,1-diphenyl-2-picrylhydrazyl radical scavenging assay, the antioxidant capacity of APAPS was shown to be approximately 126-times lower than that of APAP. In contrast, p-APS displayed comparable activity as unsulfated p-AP. Similar trends concerning the suppressive effects of these chemicals on cellular O2- radical generation were found using an activated granulocytic neutrophil cell model. Collectively, these results indicated that, depending on the presence of an additional "active site", sulfation may not always decrease the antioxidant activities of phenolic compounds.
Asunto(s)
Acetaminofén , Sulfatos , Aminofenoles , Antioxidantes/farmacología , Fenoles , Sulfotransferasas , XenobióticosRESUMEN
Endoplasmic reticulum (ER) is an organelle responsible for correct folding and sorting of proteins contributing to neurogenesis and neuronal cell death. We used rapid kindling to analyze specific ER stress marker expression underlying focal epileptogenesis. Seven-week-old rats were divided into three groups: sham (n = 6), partially kindled (n = 8), and over-kindled rats (n = 9). Over kindled rats received over100 stimuli. Partially kindled animals had stimuli halted at stage 2. Protein from ipsilateral hippocampus was electrophoresed on SDS-PAGE, followed by hybridization with primary antibodies, anti-KDEL (-Lys-Asp-Glu-coo-), Bcl-2, BDNF (brain-derived neurotrophic factor), CHOP (C/EBP-homolog protein), C/EBP (CCAAT/enhancer-binding protein), NMDA (N-methyl-D-aspartate; -R1 &2A), GluR1 (glutamate receptor), and ß-tubulin. Western blotting revealed that the ER stress marker BiP (immunoglobulin heavy chain-binding protein) was markedly increased in both partially- and over-kindled groups. BiP expression was ninefold greater than control in partially kindling while twofold greater than control in over-kindled animals. Although ER stress response was accelerated, CHOP expression, which upregulates when apoptosis signaling is accelerated by ER stress, was suppressed. Bcl-2, which acts as an anti-apoptotic molecule, was upregulated in the over-kindled group. Remarkable elevation of BiP was found in partially kindled animals, but not in over-kindled. Over-kindled rats had spontaneous generalized seizure, while partially kindled ones had only partial seizures. Elevation of markers of ER stress in partial seizures might reflect transfer of discharge to contralateral limbic structures. We observed indications of functional changes and neurogenesis in limbic structure during kindling. Widespread indications of functional changes in several membrane and secreted proteins, including NMDA-R1 & R2A and BDNF, for mossy fiber re-construction on the CA3 area, which are related to protein synthesis in the ER, may be important in epileptogenesis.
Asunto(s)
Amígdala del Cerebelo/fisiología , Estrés del Retículo Endoplásmico/fisiología , Excitación Neurológica/fisiología , Animales , Biomarcadores/metabolismo , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Ciclina B1/metabolismo , Electroencefalografía , Chaperón BiP del Retículo Endoplásmico , Proteínas de Choque Térmico/metabolismo , Masculino , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/metabolismo , Convulsiones/fisiopatología , Factor de Transcripción CHOP/metabolismoRESUMEN
In the present study, we performed comprehensive gene expression and gene network analyses using a DNA microarray to elucidate the molecular events responsible for the pathology of posttraumatic epilepsy at the partial seizure stage. We used an experimental posttraumatic epilepsy model of amygdalar focal FeCl(3)-injected rats and compared gene expression profiles in the hippocampus at the partial seizure stage (less than stage 3 on Racine's convulsion scale) and that of sham-operated animals. At the partial seizure stage, upregulation of phospholipase A2 (PLA2) and lipid metabolism were observed, which have been reported to be caused by brain injury and seizures in previous studies. Furthermore, significant upregulation of genes related to inflammation and the immune system was observed. These molecular changes in PLA2 and lipid metabolism may be related to seizure propagation.
Asunto(s)
Epilepsia Postraumática/fisiopatología , Perfilación de la Expresión Génica , Redes Reguladoras de Genes , Hipocampo/metabolismo , Convulsiones/inducido químicamente , Animales , Cloruros/toxicidad , Compuestos Férricos/toxicidad , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , Fosfolipasas A2/biosíntesis , Ratas , Ratas Wistar , Regulación hacia ArribaRESUMEN
The antiepileptic/anticonvulsive action of brivaracetam is considered to occur via modulation of synaptic vesicle protein 2A (SV2A); however, the pharmacological mechanisms of action have not been fully characterised. To explore the antiepileptic/anticonvulsive mechanism of brivaracetam associated with SV2A modulation, this study determined concentration-dependent effects of brivaracetam on astroglial L-glutamate release associated with connexin43 (Cx43), tumour-necrosis factor-α (TNFα) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/glutamate receptor of rat primary cultured astrocytes using ultra-high-performance liquid chromatography. Furthermore, interaction among TNFα, elevated extracellular K+ and brivaracetam on expression of SV2A and Cx43 was determined using capillary immunoblotting. TNFα and elevated extracellular K+ predominantly enhanced astroglial L-glutamate release associated with respective AMPA/glutamate receptor and hemichannel. These effects were enhanced by a synergistic effect of TNFα and elevated extracellular K+ in combination. The activation of astroglial L-glutamate release, and expression of SV2A and Cx43 in the plasma membrane was suppressed by subchronic brivaracetam administration but were unaffected by acute administration. These results suggest that migration of SV2A to the astroglial plasma membrane by hyperexcitability activates astroglial glutamatergic transmission, perhaps via hemichannel activation. Subchronic brivaracetam administration suppressed TNFα-induced activation of AMPA/glutamate receptor and hemichannel via inhibition of ectopic SV2A. These findings suggest that combined inhibition of vesicular and ectopic SV2A functions contribute to the antiepileptic/anticonvulsive mechanism of brivaracetam action.
Asunto(s)
Astrocitos/metabolismo , Ácido Glutámico/metabolismo , Glicoproteínas de Membrana/antagonistas & inhibidores , Glicoproteínas de Membrana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Pirrolidinonas/farmacología , Animales , Animales Recién Nacidos , Anticonvulsivantes/farmacología , Astrocitos/efectos de los fármacos , Células Cultivadas , Relación Dosis-Respuesta a Droga , Antagonistas de Aminoácidos Excitadores/farmacología , Femenino , Masculino , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Embarazo , Ratas , Ratas Sprague-Dawley , Receptores AMPA/antagonistas & inhibidores , Receptores AMPA/metabolismo , Receptores de Glutamato/metabolismo , Vesículas Sinápticas/efectos de los fármacos , Vesículas Sinápticas/metabolismo , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
This study explored the effect of coral calcium hydride (CCH) on rat intrahippocampal antioxidant ability by measuring the PCAM nitroxide radical decay ratio when CCH was (a) co-perfused into the hippocampus and (b) fed orally to the rats for 4 weeks under a freely moving state. Estimation of the in vivo antioxidant effect was obtained by administration of the blood-brain barrier-permeable PCAM nitroxide radical and the measured PCAM radical decay ratio then correlated to the amount of antioxidant in the brain using electron spin resonance (ESR) spectroscopy combined with microdialysis. The half-life periods of PCAM in rats treated with CCH in both the co-perfusion and orally fed groups were significantly shorter compared to the control group. These results clarify the mechanism that CCH may exert antioxidant activity by significantly enhancing the basal endogenous antioxidant ability in the hippocampus through a synergistic effect with α-tocopherol and ascorbic acid.
Asunto(s)
Antozoos , Antioxidantes/farmacología , Compuestos de Calcio/administración & dosificación , Hipocampo/efectos de los fármacos , Hidrógeno/metabolismo , Animales , Antioxidantes/metabolismo , Ácido Ascórbico/farmacología , Compuestos de Calcio/metabolismo , Compuestos de Calcio/farmacología , Dieta , Sinergismo Farmacológico , Hipocampo/metabolismo , Masculino , Microdiálisis , Ratas , Ratas Wistar , alfa-Tocoferol/farmacologíaRESUMEN
Recent studies using the genetic partial epilepsy model have demonstrated that hyperfunction of astroglial hemichannels contributes to pathomechanism of epileptic seizure. Therefore, to explore the novel anticonvulsive mechanisms, the present study determined the effects of voltage-dependent Na+ channel (VDSC)-inhibiting anticonvulsants, carbamazepine (CBZ), lacosamide (LCM), and zonisamide (ZNS) on the astroglial release of l-glutamate and adenosine triphosphate (ATP). The effects of subchronic administration of therapeutic-relevant dose of three anticonvulsants on the release of l-glutamate and ATP in the orbitofrontal cortex (OFC) were determined using microdialysis. The concentration-dependent effects of acute and subchronic administrations of anticonvulsants on astroglial gliotransmitter release were determined using primary cultured astrocytes. The concentration-dependent effects of subchronic administrations of anticonvulsants on connexin43 (Cx43) expression in the plasma membrane of primary cultured astrocytes were determined using the Simple Western system. An increase in the levels of extracellular K+ resulted in a concentration-dependent increase in the astroglial release of l-glutamate and ATP. The depleted levels of extracellular Ca2+ alone did not affect astroglial gliotransmitter release but did accelerate K+-evoked gliotransmitter release via activation of astroglial hemichannels. Both non-selective hemichannel inhibitor carbenoxolone (CBX) and selective Cx43 inhibitor GAP19 prevented both gliotransmitter release through activated astroglial hemichannels and the hemichannel-activating process induced by elevation of the levels of extracellular K+ with depletion of the levels of extracellular Ca2+. ZNS subchronically decreased Cx43 expression and acutely/subchronically inhibited Cx43 hemichannel activity. LCM acutely inhibited hemichannel activity but did not subchronically affect Cx43 expression. Therapeutic-relevant concentration of CBZ did not affect hemichannel activity or Cx43 expression, but supratherapeutic concentration of CBZ decreased Cx43 expression and hemichannel activity. Therefore, the present study demonstrated the distinct effects of CBZ, LCM, and ZNS on gliotransmitter release via modulation of astroglial hemichannel function. The different features of the effects of three VDSC-inhibiting anticonvulsants on astroglial transmission associated with hemichannels, at least partially, possibly contributing to the formation of the properties of these three anticonvulsants, including the antiepileptic spectrum and adverse effects regarding mood and cognitive disturbance.
RESUMEN
Elevated oxidative stress has been suggested to be associated with the features of Down's syndrome (DS). We previously reported increased oxidative stress in cultured cells from the embryonic brain of Ts1Cje, a mouse genetic DS model. However, since in vivo evidence for increased oxidative stress is lacking, we here examined lipid peroxidation, a typical marker of oxidative stress, in the brains of Ts1Cje and another DS mouse model Ts2Cje with an overlapping but larger trisomic segment. Accumulations of proteins modified with the lipid peroxidation-derived products, 13-hydroperoxy-9Z,11E-octadecadienoic acid and 4-hydroxy-2-nonenal were markedly increased in Ts1Cje and Ts2Cje brains. Analysis with oxidation-sensitive fluorescent probe also showed that reactive oxygen species themselves were increased in Ts1Cje brain. However, electron spin resonance analysis of microdialysate from the hippocampus of Ts1Cje showed that antioxidant activity remained unaffected, suggesting that the reactive oxygen species production was accelerated in Ts1Cje. Proteomics approaches with mass spectrometry identified the proteins modified with 13-hydroperoxy-9Z,11E-octadecadienoic acid and/or 4-hydroxy-2-nonenal to be involved in either ATP generation, the neuronal cytoskeleton or antioxidant activity. Structural or functional impairments of these proteins by such modifications may contribute to the DS features such as cognitive impairment that are present in the Ts1Cje mouse.
Asunto(s)
Encéfalo/metabolismo , Síndrome de Down/metabolismo , Síndrome de Down/fisiopatología , Peroxidación de Lípido/fisiología , Factores de Edad , Aldehídos/metabolismo , Animales , Encéfalo/patología , Modelos Animales de Enfermedad , Síndrome de Down/genética , Síndrome de Down/patología , Electroforesis en Gel Bidimensional/métodos , Femenino , Regulación de la Expresión Génica/genética , Humanos , Ácidos Linoleicos/metabolismo , Peróxidos Lipídicos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Microdiálisis , Especies Reactivas de Oxígeno/metabolismo , Trisomía/genéticaRESUMEN
Traumatic brain injury causes development of posttraumatic epilepsy. Bleeding within neuropil is followed by hemolysis and deposition of hemoglobin in neocortex. Iron from hemoglobin and transferring is deposited in brains of patients with posttraumatic epilepsy. Iron compounds form reactive free radical oxidants. Microinjection of ferric ions into rodent brain results in chronic recurrent seizures and liberation of glutamate into the neuropil, as is observed in humans with epilepsy. Termination of synaptic effects of glutamate is by removal via transporter proteins. EAAC-1 is within neurons while GLT-1 and GLAST are confined to glia. Persistent down regulation of GLAST production is present in hippocampal regions in chronic seizure models. Down regulation of GLAST may be fundamental to a sequence of free radical reactions initiated by brain injury with hemorrhage. Administration of antioxidants to animals causes interruption of the sequence of brain injury responses induced by hemorrhage, suggesting that such a strategy needs to be evaluated in patients with traumatic brain injury.