RESUMO
Glutathione (GSH) is an important antioxidant that plays a critical role in neuroprotection. GSH depletion in neurons induces oxidative stress and thereby promotes neuronal damage, which in turn is regarded as a hallmark of the early stage of neurodegenerative diseases. The neuronal GSH level is mainly regulated by cysteine transporter EAAC1 and its inhibitor, GTRAP3-18. In this study, we found that the GTRAP3-18 level was increased by up-regulation of the microRNA miR-96-5p, which was found to decrease EAAC1 levels in our previous study. Since the 3'-UTR region of GTRAP3-18 lacks the consensus sequence for miR-96-5p, an unidentified protein should be responsible for the intermediate regulation of GTRAP3-18 expression by miR-96-5p. Here, we discovered that RNA-binding protein NOVA1 functions as an intermediate protein for GTRAP3-18 expression via miR-96-5p. Moreover, we show that intra-arterial injection of a miR-96-5p-inhibiting nucleic acid to living mice by a drug delivery system using microbubbles and ultrasound decreased the level of GTRAP3-18 via NOVA1 and increased the levels of EAAC1 and GSH in the dentate gyrus of the hippocampus. These findings suggest that the delivery of a miR-96-5p inhibitor to the brain would efficiently increase the neuroprotective activity by increasing GSH levels via EAAC1, GTRAP3-18 and NOVA1.
Assuntos
Giro Denteado/efeitos dos fármacos , Glutationa/metabolismo , MicroRNAs/antagonistas & inibidores , Fármacos Neuroprotetores/farmacologia , Proteínas de Ligação a RNA/metabolismo , Animais , Linhagem Celular Tumoral , Giro Denteado/metabolismo , Transportador 3 de Aminoácido Excitatório/genética , Transportador 3 de Aminoácido Excitatório/metabolismo , Células HEK293 , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Injeções Intra-Arteriais , Masculino , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/metabolismo , Microbolhas , Antígeno Neuro-Oncológico Ventral , Fármacos Neuroprotetores/administração & dosagem , Proteínas de Ligação a RNA/genética , Ultrassom , Regulação para CimaRESUMO
Glutathione (GSH) is a key antioxidant that plays an important neuroprotective role in the brain. Decreased GSH levels are associated with neurodegenerative diseases such as Parkinson's disease and Alzheimer's disease. Here we show that a diurnal fluctuation of GSH levels is correlated with neuroprotective activity against oxidative stress in dopaminergic cells. In addition, we found that the cysteine transporter excitatory amino acid carrier 1 (EAAC1), which is involved in neuronal GSH synthesis, is negatively regulated by the microRNA miR-96-5p, which exhibits a diurnal rhythm. Blocking miR-96-5p by intracerebroventricular administration of an inhibitor increased the level of EAAC1 as well as that of GSH and had a neuroprotective effect against oxidative stress in the mouse substantia nigra. Our results suggest that the diurnal rhythm of miR-96-5p may play a role in neuroprotection by regulating neuronal GSH levels via EAAC1.
Assuntos
Antioxidantes/metabolismo , Transportador 3 de Aminoácido Excitatório/genética , Glutationa/metabolismo , MicroRNAs/genética , Fármacos Neuroprotetores/metabolismo , Animais , Encéfalo/patologia , Linhagem Celular , Ritmo Circadiano , Neurônios Dopaminérgicos/metabolismo , Transportador 3 de Aminoácido Excitatório/biossíntese , Glutationa/biossíntese , Células HEK293 , Humanos , Masculino , Camundongos , MicroRNAs/antagonistas & inibidores , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/metabolismo , Estresse Oxidativo , Substância Negra/metabolismoRESUMO
Glutathione (GSH) is an important neuroprotective molecule in the brain. The strategy to increase neuronal GSH level is a promising approach to the treatment of neurodegenerative diseases. However, the regulatory mechanism by which neuron-specific GSH synthesis is facilitated remains elusive. Glutamate transporter-associated protein 3-18 (GTRAP3-18) is an endoplasmic reticulum protein interacting with excitatory amino acid carrier 1 (EAAC1), which is a neuronal glutamate/cysteine transporter. To investigate the potential regulatory mechanism to increase neuronal GSH level in vivo, we generated GTRAP3-18-deficient (GTRAP3-18(-/-)) mice using a gene-targeting approach. Disruption of the GTRAP3-18 gene resulted in increased EAAC1 expression in the plasma membrane, increased neuronal GSH content and neuroprotection against oxidative stress. In addition, GTRAP3-18(-/-) mice performed better in motor/spatial learning and memory tests than wild-type mice. Therefore, the suppression of GTRAP3-18 increases neuronal resistance to oxidative stress by increasing GSH content and also facilitates cognitive function. The present results may provide a molecular basis for the development of treatments for neurodegenerative diseases.