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Estudio de las alfa-quimioquinas en la regulación del proceso de diferenciación de neuroesferas obtenidas de médula espinal de ratón
(Universidad de Chile, 2012)
RESUMEN
El recambio de elementos celulares en el tejido nervioso es sustentado por la existencia
de una célula troncal neural, responsable del proceso de neurogénesis a través del cual se
generan neuronas, astrocitos y oligodendrocitos. Recientemente...
aplicarse esta terapia celular. El trauma en la ME resulta en una serie de cambios fisiopatológicos en el tejido: muchas neuronas y oligodendrocitos mueren inmediata o progresivamente, se forman cavidades císticas que junto con los astrocitos reactivos...
aplicarse esta terapia celular. El trauma en la ME resulta en una serie de cambios fisiopatológicos en el tejido: muchas neuronas y oligodendrocitos mueren inmediata o progresivamente, se forman cavidades císticas que junto con los astrocitos reactivos...
Efecto sobre los sistemas sensoriales auditivo y visual a los 10 y 15 años en sujetos con antecedentes de anemia ferropénica en la infancia
(Universidad de Chile, 2016)
se produce en las células gliales denominadas oligodendrocitos y se
ha observado en estudios histológicos, tanto de modelos animales como
humanos, que son las que captan la mayor cantidad de hierro que se encuentra
en el cerebro, lo que sugiere que...
este elemento contribuye a su función de manera importante (Beard y Connor, 2003, Connor y Menzies, 1995, Todorich y cols., 2009). Aún no se tiene evidencia clara acerca de todas las funciones que puedan cumplir los oligodendrocitos, así como...
este elemento contribuye a su función de manera importante (Beard y Connor, 2003, Connor y Menzies, 1995, Todorich y cols., 2009). Aún no se tiene evidencia clara acerca de todas las funciones que puedan cumplir los oligodendrocitos, así como...
La señalización de angiopoyetinas a través de receptores TIE estimula la diferenciación de células troncales neurales obtenidas desde médula espinal de ratón
(Universidad de Chile, 2012-12)
células de linaje glial como también neuronal y así generar neuronas, astrocitos y
oligodendrocitos (Figura 1). In vivo, las CTN se encuentran en microambientes específicos,
zonas neurogénicas, caracterizadas por contener CTN, como también progenitores...
de los linajes obtenidos de CTN indican la presencia de progenitores intermediarios, ya sea un progenitor neural que dará origen a neuronas y, por otro lado, un progenitor glial, que tienen como destino diferenciarse a astrocito u oligodendrocito...
de los linajes obtenidos de CTN indican la presencia de progenitores intermediarios, ya sea un progenitor neural que dará origen a neuronas y, por otro lado, un progenitor glial, que tienen como destino diferenciarse a astrocito u oligodendrocito...
Modulación de la respuesta inmune durante la infección por virus distemper canino: implicancias terapéuticas y en el desarrollo de vacunas
(Universidad Austral de Chile, 2010)
SNC a través de la barrera
hematoencefálica ocurre en un escenario de severa
inmunosupresión, y a pesar de la infección restrictiva
de oligodendrocitos (menor al 10%) se desarrolla un
proceso temprano de desmielinización no inflamatoria,
asociada...
a fenómenos derivados de la replicación viral en astrocitos y microglia, que corresponden a las pobla ciones celulares responsables de mantener y facilitar la propagación viral en el SNC (Vandevelde 2004). En los escasos oligodendrocitos...
a fenómenos derivados de la replicación viral en astrocitos y microglia, que corresponden a las pobla ciones celulares responsables de mantener y facilitar la propagación viral en el SNC (Vandevelde 2004). En los escasos oligodendrocitos...
Leucoencefalopatía tóxica fatal: tres casos asociados al consumo de pasta base de cocaína
(Sociedad Médica de Santiago, 2015)
oligodendrocitos para mantener
la mielinización. Sin embargo, la alteración del
transporte axoplásmico no limita inicialmente la
trasmisión eléctrica de esos axones, postergando
la expresión clínica a la aparición de cambios del
entorno tisular y de la mielina...
Papel de la glicoproteína neuronal THY-1 en la migración y proliferación de astrocitos
(Universidad de Chile, 2007)
.
15
1. INTRODUCCIÓN
Las células gliales del sistema nervioso de vertebrados se pueden dividir en
dos grandes grupos: microglia y macroglia. La microglia corresponde a los
fagocitos del cerebro. La macroglia incluye: oligodendrocitos...
, células de Schwann y astrocitos (Figura 1). Los oligodendrocitos están a cargo de la formación de la vaina de mielina, la cual forma una barrera alrededor de las fibras nerviosas...
, células de Schwann y astrocitos (Figura 1). Los oligodendrocitos están a cargo de la formación de la vaina de mielina, la cual forma una barrera alrededor de las fibras nerviosas...
Rol del receptor P2X7 en la respuesta de los astrocitos a Thy-1 en un contexto de inflamación por daño celular
(Universidad de Chile, 2014)
los astrocitos que forman la cicatriz glial, la proteína Nogo presente en la membrana de los oligodendrocitos y, muy importantemente para este estudio, la glicoproteína Thy-1, presente en la superficie neuronal. Por muchos años, nuestro laboratorio ha...
Astrocytes respond to inflammatory stimuli by modifying their morphology and gene expression pattern, and participate in the formation of the glial scar in the case of a traumatic lesion. This glial scar represents a non-permissive environment for axonal regeneration that harbors many inhibitory molecules thought to be responsible for the poor capacity of neurons in the Central Nervous System to repair themselves. In this context, molecules like Chondroitin sulfate proteoglycans secreted by astrocytes, the Nogo protein present on the surface of oligodendrocytes and, importantly for our studies, the neuronal surface glycosyl phosphatidylinositol protein Thy-1, all are highly relevant. For many years, our laboratory has focused on the study of Thy-1 function and signaling. We have shown that Thy-1 inhibits axonal growth and engages in cell-cell interactions. We have also demonstrated that these effects are elicited through the interaction with two receptors present in DITNC1 astrocytes: αvβ3 Integrin and Syndecan-4. We have observed that short-term interaction of Thy-1 with these two receptors enhances astrocyte adhesion; however, following prolonged stimulation, Thy-1 induces astrocyte migration. Thy-1 interaction with its receptors induces the activation of signal transduction pathways in astrocytes that increase RhoA and Rac1 GTPase activity with different kinetics. Thy-1 also induces ATP release from the astrocytes, resulting in the activation of the P2X7 receptor and entry of extracellular calcium. We know that these signals are necessary for astrocyte adhesion. However, how ATP is released in response to Thy-1 remains to be determined. Also whether extracellular ATP and P2X7 receptor activation are required for DITNC1 astrocyte migration induced by Thy-1 is unknown. These and related questions we sought to answer with the studies described in this thesis. On the other hand, results from our laboratory have shown that, in contrast to DITNC1 cells, primary astrocytes in culture do not respond to Thy-1 and express very low levels of β3 integrin, which however increase under pro-inflammatory conditions, such as those produced in the brain by a stroke. Because astrocytes undergo morphological changes and migrate under these conditions in an attempt to limit and repair the damage, we hypothesized that the different responses observed in DITNC1 cells and primary astrocytes, after Thy-1 stimulation, are due to the different levels of integrin expression observed in the two types of cells. Therefore, in this study we propose that DITNC1 cell migration induced by Thy-1 requires ATP release into the extracellular space – by still unknown mechanisms – and P2X7 receptor activation. Furthermore, we hypothesize that primary astrocytes do not respond to Thy-1 due to their low levels of β3 Integrin expression. In order to study ATP release from the astrocytes, and the signaling pathways involved, we measured extracellular ATP by luminescence techniques, and evaluated calcium kinetics by loading the cells with fluorescent probes, and follow fluorescent dye uptake. To assess the requirements of astrocyte migration induced by Thy-1, we performed wound healing and cell polarity assays. The participation of the different molecules in the mechanisms under study was analyzed by the use of pharmacological inhibitors, channel blockers or receptor antagonists. Also, expression of some of these molecules was modified using shRNA and siRNA or other approaches. Primary astrocyte responses to Thy-1 were monitored in cortical astrocytes treated or not treated with pro-inflammatory cytokines, such as TNFα. Then, changes in β3 Integrin expression were analyzed by Western Blotting, and astrocyte migration was evaluated using the wound healing assay under different conditions, including silencing or overexpression of β3 Integrin. The results shown in this study indicate that ATP release induced by Thy-1 required the interaction with both αvβ3 Integrin and Syndecan-4 receptors. Mutation of the integrin-binding site completely abolished the response, while mutation in the heparin-binding domain decreased and delayed ATP release. We further demonstrate that ATP release occurred through Connexin 43 and Pannexin 1 hemichannels, which are opened by a mechanism that required an increase in intracellular calcium stored in the endoplasmic reticulum. This calcium was released to the cytoplasm through IP3R receptor activation. ATP release was necessary for the increase of both average focal adhesion area and the number of focal adhesions per cell induced by stimulation for short periods with Thy-1. Prolonged stimulation induced DITNC1 cell polarization and migration, and both of these effects required the opening of hemichannels, the presence of extracellular ATP and P2X7 receptor activation. Treatment of astrocytes with BzATP, a P2X7 receptor agonist, stimulated cell migration at high concentrations. However, our study shows that DITNC1 astrocytes treated with low concentrations of BzATP, which did not induce cell migration, were able to migrate upon co-stimulation with a mutant form of Thy-1 lacking the integrin-binding site, but with an intact heparin-binding domain, suggesting that increases in extracellular ATP requires the integrin-binding site of Thy-1. These results also indicate that activation of the P2X7 receptor by ATP (or BzATP) and downstream signaling induced by Thy-1 through Syndecan-4, are necessary for astrocyte migration. Primary astrocytes changed their phenotype after treatment with TNFα, increased the expression of β3 Integrin, GFAP and P2X7 receptor, and migrated in response to Thy-1. Furthermore, experiments in which β3 Integrin was overexpressed or silenced indicated that after treatment with pro-inflammatory agents, the observed responses to Thy-1 depended on the expression of this integrin. Migration of TNFα-treated astrocytes induced by Thy-1 also required extracellular ATP and the activation of the P2X7receptor. These results improve our understanding of the role that astrocytes play during an inflammatory response after damage to the Central Nervous System. They also uncover mechanisms relevant to how neuron-astrocyte interactions, established via neuronal Thy-1 and its astrocytic receptors, regulates astrocyte migration, modifies intracellular signaling, cell permeability and generates changes in cell behavior. These results set the stage for future studies that will yield a deeper understanding of how astrocytes change their behavior after interacting with Thy-1. These insights may eventually help in generating therapeutic opportunities and pharmacological targets to counteract the inhibition of axonal growth ascribed to interaction with astrocytes. With our results, new questions arise. For example, which pathways are related to the opening of hemichannels? How do these pathways regulate changes in adhesion and migration of astrocytes? What role does Syndecan-4 play in ATP release induced by Thy-1? What is the role of Syndecan-4 in primary astrocytes stimulated with Thy-1 under inflammatory conditions? Finally, it would be very interesting to compare these results with the responses that are observed in animal models. In conjunction, these approaches may eventually help to define new approaches for the treatment of patients with post-trauma brain damage...
Astrocytes respond to inflammatory stimuli by modifying their morphology and gene expression pattern, and participate in the formation of the glial scar in the case of a traumatic lesion. This glial scar represents a non-permissive environment for axonal regeneration that harbors many inhibitory molecules thought to be responsible for the poor capacity of neurons in the Central Nervous System to repair themselves. In this context, molecules like Chondroitin sulfate proteoglycans secreted by astrocytes, the Nogo protein present on the surface of oligodendrocytes and, importantly for our studies, the neuronal surface glycosyl phosphatidylinositol protein Thy-1, all are highly relevant. For many years, our laboratory has focused on the study of Thy-1 function and signaling. We have shown that Thy-1 inhibits axonal growth and engages in cell-cell interactions. We have also demonstrated that these effects are elicited through the interaction with two receptors present in DITNC1 astrocytes: αvβ3 Integrin and Syndecan-4. We have observed that short-term interaction of Thy-1 with these two receptors enhances astrocyte adhesion; however, following prolonged stimulation, Thy-1 induces astrocyte migration. Thy-1 interaction with its receptors induces the activation of signal transduction pathways in astrocytes that increase RhoA and Rac1 GTPase activity with different kinetics. Thy-1 also induces ATP release from the astrocytes, resulting in the activation of the P2X7 receptor and entry of extracellular calcium. We know that these signals are necessary for astrocyte adhesion. However, how ATP is released in response to Thy-1 remains to be determined. Also whether extracellular ATP and P2X7 receptor activation are required for DITNC1 astrocyte migration induced by Thy-1 is unknown. These and related questions we sought to answer with the studies described in this thesis. On the other hand, results from our laboratory have shown that, in contrast to DITNC1 cells, primary astrocytes in culture do not respond to Thy-1 and express very low levels of β3 integrin, which however increase under pro-inflammatory conditions, such as those produced in the brain by a stroke. Because astrocytes undergo morphological changes and migrate under these conditions in an attempt to limit and repair the damage, we hypothesized that the different responses observed in DITNC1 cells and primary astrocytes, after Thy-1 stimulation, are due to the different levels of integrin expression observed in the two types of cells. Therefore, in this study we propose that DITNC1 cell migration induced by Thy-1 requires ATP release into the extracellular space – by still unknown mechanisms – and P2X7 receptor activation. Furthermore, we hypothesize that primary astrocytes do not respond to Thy-1 due to their low levels of β3 Integrin expression. In order to study ATP release from the astrocytes, and the signaling pathways involved, we measured extracellular ATP by luminescence techniques, and evaluated calcium kinetics by loading the cells with fluorescent probes, and follow fluorescent dye uptake. To assess the requirements of astrocyte migration induced by Thy-1, we performed wound healing and cell polarity assays. The participation of the different molecules in the mechanisms under study was analyzed by the use of pharmacological inhibitors, channel blockers or receptor antagonists. Also, expression of some of these molecules was modified using shRNA and siRNA or other approaches. Primary astrocyte responses to Thy-1 were monitored in cortical astrocytes treated or not treated with pro-inflammatory cytokines, such as TNFα. Then, changes in β3 Integrin expression were analyzed by Western Blotting, and astrocyte migration was evaluated using the wound healing assay under different conditions, including silencing or overexpression of β3 Integrin. The results shown in this study indicate that ATP release induced by Thy-1 required the interaction with both αvβ3 Integrin and Syndecan-4 receptors. Mutation of the integrin-binding site completely abolished the response, while mutation in the heparin-binding domain decreased and delayed ATP release. We further demonstrate that ATP release occurred through Connexin 43 and Pannexin 1 hemichannels, which are opened by a mechanism that required an increase in intracellular calcium stored in the endoplasmic reticulum. This calcium was released to the cytoplasm through IP3R receptor activation. ATP release was necessary for the increase of both average focal adhesion area and the number of focal adhesions per cell induced by stimulation for short periods with Thy-1. Prolonged stimulation induced DITNC1 cell polarization and migration, and both of these effects required the opening of hemichannels, the presence of extracellular ATP and P2X7 receptor activation. Treatment of astrocytes with BzATP, a P2X7 receptor agonist, stimulated cell migration at high concentrations. However, our study shows that DITNC1 astrocytes treated with low concentrations of BzATP, which did not induce cell migration, were able to migrate upon co-stimulation with a mutant form of Thy-1 lacking the integrin-binding site, but with an intact heparin-binding domain, suggesting that increases in extracellular ATP requires the integrin-binding site of Thy-1. These results also indicate that activation of the P2X7 receptor by ATP (or BzATP) and downstream signaling induced by Thy-1 through Syndecan-4, are necessary for astrocyte migration. Primary astrocytes changed their phenotype after treatment with TNFα, increased the expression of β3 Integrin, GFAP and P2X7 receptor, and migrated in response to Thy-1. Furthermore, experiments in which β3 Integrin was overexpressed or silenced indicated that after treatment with pro-inflammatory agents, the observed responses to Thy-1 depended on the expression of this integrin. Migration of TNFα-treated astrocytes induced by Thy-1 also required extracellular ATP and the activation of the P2X7receptor. These results improve our understanding of the role that astrocytes play during an inflammatory response after damage to the Central Nervous System. They also uncover mechanisms relevant to how neuron-astrocyte interactions, established via neuronal Thy-1 and its astrocytic receptors, regulates astrocyte migration, modifies intracellular signaling, cell permeability and generates changes in cell behavior. These results set the stage for future studies that will yield a deeper understanding of how astrocytes change their behavior after interacting with Thy-1. These insights may eventually help in generating therapeutic opportunities and pharmacological targets to counteract the inhibition of axonal growth ascribed to interaction with astrocytes. With our results, new questions arise. For example, which pathways are related to the opening of hemichannels? How do these pathways regulate changes in adhesion and migration of astrocytes? What role does Syndecan-4 play in ATP release induced by Thy-1? What is the role of Syndecan-4 in primary astrocytes stimulated with Thy-1 under inflammatory conditions? Finally, it would be very interesting to compare these results with the responses that are observed in animal models. In conjunction, these approaches may eventually help to define new approaches for the treatment of patients with post-trauma brain damage...
Papel de CoREST durante el desarrollo de la corteza cerebral in vivo
(Universidad de Chile, 2012)
(intermedios), astrocitos
y oligodendrocitos, de acuerdo a un programa intrínseco recapitulado aún en cultivos
primarios corticales in vitro [Qian y cols., 2000]. Los tres tipos celulares se producen en
un patrón temporalmente distinto, aunque superpuesto...
[Sauvageot y cols., 2002]. Poco antes del nacimiento (E18.5-E19.5), las glias radiales se transforman en astrocitos. Los oligodendrocitos son generados en dos oleadas. Durante el desarrollo (alrededor de 27 E13.5), progenitores en la eminencia ganglionar...
[Sauvageot y cols., 2002]. Poco antes del nacimiento (E18.5-E19.5), las glias radiales se transforman en astrocitos. Los oligodendrocitos son generados en dos oleadas. Durante el desarrollo (alrededor de 27 E13.5), progenitores en la eminencia ganglionar...
Posible tráfico de ATP mediado por lanzaderas de ATP en el cilio olfatorio y movimiento de glucosa a través de la mucosa olfatoria
(2019-03)
disponible para que los axones lo tomen y metabolicen oxidativamente (Brown et al. 2003a).
En el SNC, los astrocitos y los oligodendrocitos liberan lactato (Brown et al. 2003a; Lee et al.
2012). De forma similar, el glicógeno contenido en células de Schwann...
través de transportadores de monocarboxilato (Tekkok et al. 2005). Recientemente se ha demostrado que los oligodendrocitos expresan la proteína MCT1, lo que indica que los oligodendrocitos también son capaces de suministrar un sustrato extra a los...
través de transportadores de monocarboxilato (Tekkok et al. 2005). Recientemente se ha demostrado que los oligodendrocitos expresan la proteína MCT1, lo que indica que los oligodendrocitos también son capaces de suministrar un sustrato extra a los...
La respuesta a proteínas mal plegadas como blanco terapéutico en la enfermedad de Alzheimer
(Sociedad Médica de Santiago, 2020)
específico para determinados tipos
celulares (es decir, astrocitos, oligodendrocitos,
neuronas, etc.71). Esto, combinado con promo-
tores celular o tejido específico, puede asegurar
la expresión controlada y restringida del transgen.
Conclusiones
La EA es...