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Estudio De Las Vías De Señalización E Inflamatorias Del Canal P2X7 Y Su Variante Polimórfica Gln460arg En Células Del Sistema Nervioso Central

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Estudio De Las Vías De Señalización E Inflamatorias Del Canal P2X7 Y Su Variante Polimórfica Gln460arg En Células Del Sistema Nervioso Central Tesis Doctoral Estudio de las vías de señalización e inflamatorias del canal P2X7 y su variante polimórfica Gln460Arg en células del sistema nervioso central Aprile García, Fernando 2014-04-11 Este documento forma parte de la colección de tesis doctorales y de maestría de la Biblioteca Central Dr. Luis Federico Leloir, disponible en digital.bl.fcen.uba.ar. Su utilización debe ser acompañada por la cita bibliográfica con reconocimiento de la fuente. This document is part of the doctoral theses collection of the Central Library Dr. Luis Federico Leloir, available in digital.bl.fcen.uba.ar. It should be used accompanied by the corresponding citation acknowledging the source. Cita tipo APA: Aprile García, Fernando. (2014-04-11). Estudio de las vías de señalización e inflamatorias del canal P2X7 y su variante polimórfica Gln460Arg en células del sistema nervioso central. Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Cita tipo Chicago: Aprile García, Fernando. "Estudio de las vías de señalización e inflamatorias del canal P2X7 y su variante polimórfica Gln460Arg en células del sistema nervioso central". Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. 2014-04-11. Dirección: Biblioteca Central Dr. Luis F. Leloir, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires. Contacto: [email protected] Intendente Güiraldes 2160 - C1428EGA - Tel. (++54 +11) 4789-9293 UNIVERSIDAD DE BUENOS AIRES Facultad de Ciencias Exactas y Naturales Estudio de las vías de señalización e inflamatorias del canal P2X7 y su variante polimórfica Gln460Arg en células del sistema nervioso central Tesis presentada para optar al título de Doctor de la Universidad de Buenos Aires en el área CIENCIAS BIOLOGICAS Lic. Fernando Aprile García Director de tesis: Dr Eduardo Arzt Director Asistente: Dra Ana C Liberman Consejero de Estudios: Dr Eduardo Arzt Lugar de trabajo: a. Instituto de Investigación en Biomedicina de Buenos Aires – CONICET – Instituto Partner de la Sociedad Max Planck. b. Laboratorio de Fisiología y Biología Molecular – Departamento de Fisiología, Biología Molecular y Celular – Facultad de Ciencias Exactas y Naturales. Buenos Aires, 2014 Fernando Aprile García Índice Índice ÍNDICE ............................................................................................................................................. 2 ÍNDICE DE FIGURAS Y TABLAS ................................................................................................ 8 RESUMEN ....................................................................................................................................... 11 ABSTRACT .................................................................................................................................... 13 AGRADECIMIENTOS ................................................................................................................. 15 ABREVIATURAS .......................................................................................................................... 19 INTRODUCCIÓN ........................................................................................................................ 20 1. DEPRESIÓN ................................................................................................................... 21 1.1. Introducción............................................................................................................... 21 1.2. Epidemiología ............................................................................................................ 22 1.3. Neurobiología de la depresión .................................................................................. 22 1.3.1. Glutamato ............................................................................................................... 23 1.3.2. BDNF ...................................................................................................................... 24 1.3.3. Hormonas del estrés y depresión ............................................................................ 24 1.3.4. Inflamación y depresión.......................................................................................... 26 1.4. Astrocitos y depresión ............................................................................................... 29 1.4.1. Tipos de células gliales ........................................................................................... 29 1.4.2. Astrocitos ................................................................................................................ 30 1.5. Componente genético en depresión ......................................................................... 31 2. RECEPTORES PURINÉRGICOS .............................................................................. 32 2.1. Receptores P1 ............................................................................................................ 33 2.2. Receptores P2 ............................................................................................................ 34 2.3. Receptores P2Y .......................................................................................................... 35 2.4. Receptores P2X .......................................................................................................... 36 3. RECEPTORES P2X7 .................................................................................................... 39 3.1. Expresión de los receptores P2X7 ............................................................................ 40 3.2. Estequiometría de los receptores P2X7 ................................................................... 41 3.3. Activación y propiedades farmacológicas de P2X7 ................................................ 41 3.4. Mecanismos de transducción de señales asociados a P2X7 ................................... 42 3.5. Funciones en SNC ..................................................................................................... 45 3.6. Liberación de neurotransmisores ............................................................................ 46 3.7. Rol funcional de P2X7 en glía .................................................................................. 47 3.8. Efectos transcripcionales de P2X7 ........................................................................... 48 2 Fernando Aprile García Índice 3.8.1. Egr........................................................................................................................... 49 3.8.2. NFAT ....................................................................................................................... 50 3.8.3. NFκB ...................................................................................................................... 51 3.8.4. CREB ...................................................................................................................... 52 3.8.5. AP-1 ........................................................................................................................ 52 4. PARP1 ............................................................................................................................. 53 4.1. PARPs ......................................................................................................................... 53 4.2. PARP1 ........................................................................................................................ 55 4.3. PARP1 e inflamación ................................................................................................ 57 5. VARIANTE POLIMÓRFICA DEL RECEPTOR P2X7: GLN460ARG .................. 59 5.1. Polimorfismos de P2X7 ............................................................................................. 59 5.2. Asociación de la variante polimórfica de P2X7 Gln460Arg con depresión…….. 60 5.3. Antecedentes experimentales: Función de la variante polimórfica P2X7- Gln460Arg ..…. .................................................................................................................................... 62 OBJETIVOS .................................................................................................................................. 65 MATERIALES Y MÉTODOS .................................................................................................... 67 1. APARATOS, ESTÍMULOS, ANTICUERPOS E INHIBIDORES............................ 68 1.1. Aparatos o dispositivos especiales ............................................................................ 68 1.2. Estímulos y agentes farmacológicos......................................................................... 68 1.3. Anticuerpos ................................................................................................................ 68 1.4. Otros reactivos o materiales ..................................................................................... 69 2. MANEJO DE PLÁSMIDOS ........................................................................................ 70 2.1. Plásmidos utilizados .................................................................................................. 70 2.2. Subclonado de plásmidos .......................................................................................... 72 2.3. Purificación de fragmentos de ADN ........................................................................ 74 2.4. Reacción de ligación .................................................................................................
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