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Master's Dissertation
DOI
https://doi.org/10.11606/D.25.2010.tde-17082010-152714
Document
Author
Full name
Aline de Lima Leite
Institute/School/College
Knowledge Area
Date of Defense
Published
Bauru, 2010
Supervisor
Committee
Buzalaf, Marilia Afonso Rabelo (President)
Labate, Carlos Alberto
Pessan, Juliano Pelim
Title in Portuguese
Análise proteômica em rim de ratos submetidos à exposição aguda de flúor
Keywords in Portuguese
Falha renal
Fluoreto
Proteômica
Toxicidade aguda
Abstract in Portuguese
O flúor é comprovadamente um agente terapêutico contra a cárie dentária, porém, quando consumido de maneira inadequada pode causar reações indesejáveis, tanto crônicas como agudas. A proteômica é uma ferramenta que permite analisar o perfil proteico completo dos sistemas biológicos, permitindo um melhor entendimento da fisiologia normal do organismo, bem como dos mecanismos de doenças, descoberta de biomarcadores para detecção precoce de doenças, identificação de novas terapias e descobertas de fármacos. Assim, o presente estudo teve como trabalho realizar um estudo proteômico diferencial em amostras de tecido renal de ratos submetidos à intoxicação aguda por fluoreto. Para isso, 27 animais, receberam desde o desmame água deionizada e ração AIN-93 por 50 dias. Ao completarem 75 dias de vida, e, após jejum por 12 horas, os animais receberam via gástrica as seguintes doses agudas de F: 0 (controle), 50 e 100 mgF/Kg peso corporal. Após 2 horas foram, os animais foram anestesiados e tiveram o sangue e os rins coletados. Inicialmente foi realizada a dosagem de flúor no plasma e tecidos renais, após difusão facilitada por hexamethyldisiloxano. Em um segundo momento, as proteínas do tecido renal foram extraídas e submetidas à eletroforese bidimensional. Os géis obtidos foram analisados através do software ImageMaster 2D-platinum v. 7.0. Os spots que se apresentaram diferencialmente expressos foram submetidos à identificação por espectrometria de massas. As proteínas identificadas foram classificadas em 5 categorias funcionais. A categoria metabolismo e energia reuniu a maior parte das proteínas (40%), seguida pelas categorias de transporte e processos celulares com 20% e 13% respectivamente. Na categoria estrutura e organização estrutural, que reúne proteínas com funções relacionadas ao citoesqueleto e membrana celular, foram identificadas 17% das proteínas. Por fim, os 10% restantes das proteínas pertenciam à categoria vias de informação, que abriga proteínas envolvidas nos processos de síntese/degradação de DNA e RNA. Com base nos resultados obtidos pode se concluir que doses agudas de fluoreto alteram a expressão a nível renal de diversas proteínas relacionadas a distintos processos celulares.
Keywords in English
Fluoride
Kidney fail
Proteomic analysis
Abstract in English
Fluoride has been widely used in dentistry as a caries prophylactic agent. However, if consumed in high concentrations, it can cause side effects as dental fluorosis, skeletal fluorosis and, in extreme situations, death. The recent development of proteomic techniques has allowed the analysis of the entire protein profile of biological systems, contributing to the understanding of the normal physiology of the organism, as well as the mechanisms of diseases and the investigation of biomarkers for their early detection, identification of new therapies and drugs. This study used proteomic techniques to analyze the differential protein expression in kidney of rats submitted to acute fluoride treatments. Three groups of five 75-day-old Wistar rats received, by gastric gavage, the following single doses of fluoride: 0 (control), 50 and 100 mgF/Kg body weight. After the treatments, the animals were killed and the left kidney and plasma were collected for fluoride analysis. For proteomic analysis, the right kidney was collected. Fluoride in plasma and renal tissues was analyzed with the electrode, after hexamethyldisiloxanefacilitated diffusion. Proteins from kidney were profiled by two-dimensional gel electrophoresis. Gels from control and treated groups were digitalized using the ImageScanner III and analyzed with ImageMaster 2D Platinum version 7.0 for statistical differences (ANOVA, p <0.05). The proteins identified were classified into five functional categories. The category of metabolism and energy had the majority of the proteins (40%), followed by the categories of "transport" and cell processes, with 20% and 13% of the proteins, respectively. In the category "structure and cell organization, which brings together proteins with functions related to the cytoskeleton and plasma membrane were identified 17% of the protein. Finally, 10% of the proteins belonged to the category "information pathways", which comprises the proteins involved in the processes of synthesis and degradation of DNA and RNA. Based on the results obtained, it can be concluded that acute doses of fluoride alters the expression of several proteins related to different cellular processes in the kidneys.
 
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AlineLimaLeite.pdf (12.73 Mbytes)
Publishing Date
2010-08-19
 
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