RESUMEN
miR-34a is involved in the regulation of the fate of different cell types. However, the mechanism by which it controls the differentiation programme of neural cells remains largely unknown. Here, we investigated the role of miR-34a in neurogenesis and maturation of developing neurons and identified Doublecortin as a new miR-34a target. We found that the overexpression of miR-34a in vitro significantly increases precursor proliferation and influences morphology and function of developing neurons. Indeed, miR-34a overexpressing neurons showed a decreased expression of several synaptic proteins and receptor subunits, a decrement of NMDA-evoked current density and, interestingly, a more efficient response to synaptic stimulus. In vivo, miR-34a overexpression showed stage-specific effects. In neural progenitors, miR-34a overexpression promoted cell proliferation, in migratory neuroblasts reduced the migration and in differentiating newborn neurons modulated process outgrowth and complexity. Importantly, we found that rats overexpressing miR-34a in the brain have better learning abilities and reduced emotionality.
Asunto(s)
Conducta Animal , Forma de la Célula , MicroARNs/metabolismo , Neurogénesis , Neuronas/citología , Neuronas/metabolismo , Animales , Secuencia de Bases , Bromodesoxiuridina/metabolismo , Diferenciación Celular , Movimiento Celular , Proliferación Celular , Células Cultivadas , Corteza Cerebral/citología , Cognición , Dependovirus/metabolismo , Proteínas de Dominio Doblecortina , Proteína Doblecortina , Emociones , Femenino , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Mitosis , Datos de Secuencia Molecular , Neuritis/metabolismo , Neuropéptidos/genética , Neuropéptidos/metabolismo , Fenotipo , Ratas Wistar , Células Madre/citologíaRESUMEN
Neocortical networks play a major role in the genesis of generalized spike-and-wave (SW) discharges associated with absence seizures in humans and in animal models, including genetically predisposed WAG/Rij rats. Here, we tested the hypothesis that alterations in GABA(B) receptors contribute to neocortical hyperexcitability in these animals. By using Real-Time PCR we found that mRNA levels for most GABA(B(1)) subunits are diminished in epileptic WAG/Rij neocortex as compared with age-matched non-epileptic controls (NEC), whereas GABA(B(2)) mRNA is unchanged. Next, we investigated the cellular distribution of GABA(B(1)) and GABA(B(2)) subunits by confocal microscopy and discovered that GABA(B(1)) subunits fail to localize in the distal dendrites of WAG/Rij neocortical pyramidal cells. Intracellular recordings from neocortical cells in an in vitro slice preparation demonstrated reduced paired-pulse depression of pharmacologically isolated excitatory and inhibitory responses in epileptic WAG/Rij rats as compared with NECs; moreover, paired-pulse depression in NEC slices was diminished by a GABA(B) receptor antagonist to a greater extent than in WAG/Rij rats further suggesting GABA(B) receptor dysfunction. In conclusion, our data identify changes in GABA(B) receptor subunit expression and distribution along with decreased paired-pulse depression in epileptic WAG/Rij rat neocortex. We propose that these alterations may contribute to neocortical hyperexcitability and thus to SW generation in absence epilepsy.
Asunto(s)
Epilepsia Tipo Ausencia/fisiopatología , Neocórtex/fisiología , Receptores de GABA-B/genética , Animales , Modelos Animales de Enfermedad , Electrofisiología , Neocórtex/citología , Inhibición Neural/fisiología , Técnicas de Cultivo de Órganos , Células Piramidales/fisiología , ARN Mensajero/metabolismo , Ratas , Ratas Mutantes , Reacción en Cadena de la Polimerasa de Transcriptasa InversaAsunto(s)
Daño del ADN/fisiología , Reparación del ADN/fisiología , Mitosis , Neuronas/fisiología , Animales , Apoptosis , Células Cultivadas , Cerebelo/citología , Cerebelo/metabolismo , Daño del ADN/genética , Reparación del ADN/genética , Genes Reporteros , Vectores Genéticos , Proteínas Fluorescentes Verdes/genética , Operón Lac , Ratones , Ratones SCID , Neuronas/ultraestructura , Técnicas de Cultivo de TejidosRESUMEN
Survivin, a dimeric baculovirus inhibitor of apoptosis repeat (BIR) motif protein that is principally expressed in G2 and mitosis, has been associated with protection against apoptosis of cells that exit mitosis aberrantly. Mammalian survivin has been reported to associate with centrosomes and with the mitotic spindle. We have expressed a human hemagglutinin-tagged survivin plasmid to determine its localization, and find instead that it clearly acts as a passenger protein. In HeLa cells, survivin first associates with the kinetochores, and then translocates to the spindle midzone during anaphase and, finally, to the midbody during cell cleavage. Its localization is similar to that of TD-60, a known passenger protein. Both a point mutation in the baculovirus IAP repeat motif (C84A) and a COOH-terminal deletion mutant (Delta106) of survivin fail to localize to either kinetochores or midbodies, but neither interferes with cell cleavage. The interphase localization of survivin is cell cycle regulated since in permanently transfected NIH3T3 cells it is excluded from the nuclei until G2, where it localizes with centromeres. Survivin remains associated with mitotic kinetochores when microtubule assembly is disrupted and its localization is thus independent of microtubules. We conclude that human survivin is positioned to have an important function in the mechanism of cell cleavage.
Asunto(s)
Cinetocoros/metabolismo , Proteínas Asociadas a Microtúbulos , Proteínas/metabolismo , Células 3T3 , Secuencias de Aminoácidos , Sustitución de Aminoácidos/genética , Animales , División Celular , Técnica del Anticuerpo Fluorescente , Células HeLa , Humanos , Proteínas Inhibidoras de la Apoptosis , Ratones , Mutación/genética , Proteínas de Neoplasias , Unión Proteica , Transporte de Proteínas , Proteínas/química , Proteínas/genética , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/metabolismo , Huso Acromático/metabolismo , Survivin , Dedos de ZincRESUMEN
Are the microtubule-organising centers of the different cell types of a metazoan interchangeable? If not, what are the differences between them? Do they play any role in the differentiation processes to which these cells are subjected? Nearly one hundred years of centrosome research has established the essential role of this organelle as the main microtubule-organising center of animal cells. But only now are we starting to unveil the answers to the challenging questions which are raised when the centrosome is studied within the context of a pluricellular organism. In this review we present some of the many examples which illustrate how centrosomes and microtubule organisation changes through development in Drosophila and discuss some of its implications.
Asunto(s)
Centrosoma/fisiología , Drosophila/crecimiento & desarrollo , Microtúbulos/fisiología , Animales , Centrosoma/ultraestructura , Drosophila/fisiología , Drosophila/ultraestructura , Femenino , Masculino , Mitosis , Mutación , Oogénesis , Espermatogénesis , Huso Acromático/fisiologíaRESUMEN
Schistosoma mansoni infection in adult mice is known to cause granulomas in the liver and intestine. Using a specific enzyme-linked immunoassay, it was found that Schistosoma mansoni infection enhances the level of nerve growth factor in the liver and surprisingly also in the hypothalamus. Exogenous administration of purified NGF antibodies inhibits NGF biological activity both in the hypothalamus and liver and drastically reduces the number of NGF-responsive cells, the mast cells, present in liver granuloma. These findings and those reported by others showing the effect of NGF on cells of the immune system support the hypothesis that this molecule plays a role in neuroendocrine-immune interactions.
Asunto(s)
Hipotálamo/metabolismo , Hígado/metabolismo , Factores de Crecimiento Nervioso/metabolismo , Esquistosomiasis mansoni/metabolismo , Animales , Anticuerpos/inmunología , Células Cultivadas , Ensayo de Inmunoadsorción Enzimática , Femenino , Granuloma/patología , Hipotálamo/patología , Inmunohistoquímica , Hígado/patología , Mastocitos/inmunología , Mastocitos/metabolismo , Ratones , Factores de Crecimiento Nervioso/inmunología , Ratas , Esquistosomiasis mansoni/patologíaRESUMEN
We have recently reported that nerve growth factor (NGF) increases in the synovium of patients affected by arthritis, as well as in animal models. We report here that the synovium of transgenic arthritic mice expressing human tumour necrosis factor (TNF) contains numerous mast cells (MC) and that their appearance is a phenomenon which was correlated to the local increase in NGF level. These findings provide further evidence that NGF plays a role in inflammation and suggest a functional link between NGF and MC.