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Previous issue date: 2015-02-23 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / Iron is essential in the neonatal brain for normal neurological development and for establishment of the concentration of iron in the adult brain, as iron absorption is greatest during the neonatal period. It is believed that iron overload contributes to the development of neurodegeneration, the exacerbation of the normal apoptosis rates, largely due to its participation in the Fenton reaction and production of reactive oxygen species. Previous studies in our laboratory have shown that treatment with iron in the neonatal period worsens the memory, as well as an increase in oxidative stress parameters and levels of apoptotic proteins. Recently, we have also demonstrated that this treatment reduces synaptophysin levels (a synaptic marker) in the rats hippocampus. Mitochondria are involved in this process because they are considered the main intracellular source of superoxide anion and the main target of attack by free radicals. They are highly dynamic organelles that bind (fusion) and divide (fission) in response to environmental stimuli, developmental status and energy needs of the cells. Mitochondria are widely distributed in all neurons, but mitochondria subpopulations are enriched pre-synaptically in nerve terminals and post-synaptically in dendrites. In addition, they have been implicated in the pathogenesis of a wide variety of neurodegenerative diseases where synapses are known to be the main target. Sulforaphane is a natural compound that has been studied since the 1980s and considered to possess antioxidant and anti-inflammatory properties. This study aims to investigate the effects of treatment with Sulforaphane in adulthood on memory deficits and changes in markers of mitochondrial function, DNML1 and OPA1, and synaptic marker synaptophysin induced by neonatal iron treatment. Male Wistar rats received, orally, vehicle or carbonyl iron (30mg / kg) from the 12th to the 14th postnatal day. In order to evaluate the effects of acute Sulforaphane treatment on memory, rats were trained in the object recognition task and received one single intraperitoneal injection of Vehicle or Sulforaphane (0.5 or 5 mk/kg) immediately after training. To evaluate the effects of Sulforaphane chronic treatment on memory, rats received vehicle or Sulforaphane (0.5 or 5 mg / kg) for 14 days every other day, and were trained in the object recognition task 24 h after the last injection. The memory analysis was performed using the recognition index, expressed as the ratio between the amount of time spent in exploring the new object on the total time spent exploring both objects. DNML1, OPA1, and synaptophysin levels in the hippocampus were quantified by Western blotting. The quantification of protein was performed by measuring the density of the individual bands, normalized by ?-actin density. Data were analyzed by comparing the averages for analysis of variance (ANOVA), with p <0.05 considered statistically significant. Our results showed that acute treatment with Sulforaphane (0.5 mg / kg) did not improve memory but at a dose of 5 mg / kg it was able to restore the memory deficits induced by iron. Chronic treatment with Sulforaphane, at both doses, also recovers the memory in rats treated with iron in the neonatal period. Sulforaphane at the dose of 5 mg/kg also recovers DNM1L mitochondrial alteration and synaptophysin in the hippocampus, confirming a possible neuroprotective role for this compound. / O ferro ? essencial no c?rebro neonatal para o desenvolvimento neurol?gico normal e para o estabelecimento da concentra??o de ferro no c?rebro adulto, j? que a absor??o de ferro ? m?xima durante o per?odo neonatal. Acredita-se que a sobrecarga de ferro contribua para o desenvolvimento da neurodegenera??o, na exacerba??o das taxas normais de apoptose, em grande parte devido ? sua participa??o na rea??o de Fenton e produ??o de esp?cies reativas de oxig?nio. Estudos pr?vios em nosso laborat?rio demonstraram que o tratamento com ferro no per?odo neonatal piora a mem?ria, bem como aumento em par?metros de estresse oxidativo e em n?veis de prote?nas apopt?ticas. Recentemente, tamb?m demonstramos que esse tratamento reduz os n?veis de sinaptofisina (um marcador sin?ptico) em hipocampo de ratos. As mitoc?ndrias est?o envolvidas neste processo por serem consideradas a fonte intracelular principal do ?nion super?xido bem como o alvo principal de ataque de radicais livres. S?o organelas altamente din?micas que se unem (fus?o) e se dividem (fiss?o) em resposta a est?mulos ambientais, status de desenvolvimento, e as necessidades energ?ticas das c?lulas. As mitoc?ndrias s?o amplamente distribu?das por todos os neur?nios, por?m subpopula??es de mitoc?ndrias s?o enriquecidas pr?-sinapticamente nos terminais nervosos e p?s-sinapticamente nos dendritos. Al?m disso, estas t?m sido implicadas na patog?nese de uma ampla variedade de doen?as neurodegenerativas, onde sinapses s?o conhecidas por serem o alvo principal. O Sulforafano ? um composto natural que vem sendo estudado desde a d?cada de 1980 e considerado por possuir propriedades antioxidantes e anti-inflamat?rias. O presente trabalho visa investigar os efeitos do tratamento com Sulforafano na idade adulta sobre os d?ficits de mem?ria e sobre altera??es em marcadores da fun??o mitocondrial, DNML1 e OPA1, e marcador sin?ptico sinaptofisina, induzidos pelo tratamento neonatal com ferro. Os ratos Wistar machos receberam, por via oral, ve?culo ou ferro carbonila (30mg/kg) do 12? ao 14? dia p?s-natal. Na idade adulta, para a avalia??o dos efeitos do tratamento agudo com Sulforafano sobre a mem?ria, os animais foram treinados na tarefa de reconhecimento de objeto e receberam uma ?nica inje??o, por via intraperitoneal, de ve?culo ou Sulfarafano (0,5 ou 5 mg/kg) imediatamente ap?s o treino. Para a avalia??o dos efeitos do tratamento cr?nico com Sulforafano sobre a mem?ria, os animais foram tratados com ve?culo ou Sulfarafano (0,5 ou 5 mg/kg) durante 14 dias, em dias intercalados e treinados na tarefa de reconhecimento de objeto 24 horas ap?s a ?ltima inje??o. A an?lise de mem?ria foi realizada atrav?s do ?ndice de reconhecimento, expresso pela raz?o entre a quantidade de tempo gasto na explora??o do objeto novo sobre o tempo total gasto explorando ambos os objetos. Os n?veis prot?icos de DNML1, OPA1 e sinaptofisina no hipocampo foram quantificados atrav?s de Western blotting. A quantifica??o das prote?nas foi realizada atrav?s da medida das densidades das bandas individuais, normalizadas pela densidade de ?-actina. Os dados foram analisados atrav?s das compara??es entre as m?dias por an?lise de vari?ncia de uma via (ANOVA), sendo o p<0,05 considerado estatisticamente significativo. Nossos resultados mostraram que o tratamento agudo com Sulforafano (0,5 mg / kg) n?o melhora a mem?ria mas na dose de 5 mg/kg foi capaz de restaurar a mem?ria e no tratamento cr?nico com Sulforafano, ambas as doses recuperam a mem?ria em ratos tratados com ferro no per?odo neonatal. O Sulforafano na dose de 5 mg/kg tamb?m recupera os danos mitocondriais causados na DNM1L e sinaptofisina no hipocampo, corroborando com um poss?vel papel neuroprotetor para esse composto.
| Identifer | oai:union.ndltd.org:IBICT/oai:tede2.pucrs.br:tede/5998 |
| Date | 23 February 2015 |
| Creators | Lavich, Isabela Cavalheiro |
| Contributors | Schr?der, Nadja |
| Publisher | Pontif?cia Universidade Cat?lica do Rio Grande do Sul, Programa de P?s-Gradua??o em Biologia Celular e Molecular, PUCRS, Brasil, Faculdade de Bioci?ncias |
| Source Sets | IBICT Brazilian ETDs |
| Language | Portuguese |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, info:eu-repo/semantics/masterThesis |
| Format | application/pdf |
| Source | reponame:Biblioteca Digital de Teses e Dissertações da PUC_RS, instname:Pontifícia Universidade Católica do Rio Grande do Sul, instacron:PUC_RS |
| Rights | info:eu-repo/semantics/openAccess |
| Relation | 8198246930096637360, 600, 600, 600, 600, 36528317262667714, -1634559385931244697, 2075167498588264571 |
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