Identificação de genes diferencialmente expressos no endométrio de mulheres durante o ciclo menstrual
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Arquivos
Data
2006
Autores
Otsuka, Audrey Yumi [UNIFESP]
Orientadores
Silva, Ismael Dale Cotrim Guerreiro da [UNIFESP]
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
As variações fisiológicas dos hormônios sexuais ao longo do ciclo menstrual
promovem alterações histomorfológicas e funcionais no endométrio. Para melhor
entender as variações de expressão genética responsáveis por tais eventos,
motivamo-nos a avaliar as eventuais variações na expressão gênica nas fases
proliferativa e secretora do ciclo menstrual. Para tanto, estudamos o endométrio
normal de 24 pacientes submetidas à histerectomia por causas benignas.
Retiraram-se fragmentos do endométrio para extração de RNA, os quais foram
imediatamente acondicionados em gelo seco para futura utilização. As amostras
foram submetidas a ensaios do tipo cDNA microarray em lâminas contendo 10.000
genes. A análise da expressão diferencial foi realizada pelo software SAM e os
resultados confirmados, individualmente, por PCR em tempo real para RNA
mensageiro, além de imunohistoquímica para a correta confirmação em nível
protéico. Dos 10.000 genes estudados, obtivemos um total de 1127 genes
diferencialmente expressos. Dentre estes, 364 genes estavam superexpressos na
fase proliferativa e 763 genes tinham seu RNA mensageiro em maior quantidade na
fase secretora. A fase secretora apresentou-se, do ponto de vista de variedade de
expressão gênica, como uma fase marcadamente mais rica, em especial quando
analisamos os genes envolvidos na resposta imune, e os genes que respondem aos
hormônios tireoideanos, bem como aqueles envolvidos no metabolismo de cálcio.
In order to further comprehend the genetic events underlying the endometrial steroids response we undertook a 10.000 genes microarray analysis between human normal endometrium in the proliferative and secretory phases of the menstrual cycle. We selected 24 patients out of 72 women who were diagnosed as having uterine leiomyomata. Neither patient had received any hormonal treatment during the six months previously the diagnosis of leiomyomata, endometrial fragments were divided in proliferative and secretory after histological analysis. Total RNA was extracted from samples by using prior to reverse transcription, two pools of RNA were obtained by mixing 3 µg of total RNA from each individual case in two separated tubes representing endometrial tissues in proliferative (group 1) and secretory phase (group 2). These two pools were then hybridized two times each to four CodeLink UniSet Human I Bioarrays (Amersham Biosciences, Piscataway, NJ). After analysing the results by the SAM software 1127 genes were found differentially expressed between secretory and proliferative phases when a 2,5 difference was used as a cut-off criteria. 364 genes were up-regulated in the proliferative phase and 763 genes were up-regulated in the secretory phase. Among them, genes involved in immune response, thyroid and calcium metabolism were found differentially expressed specially in the secretory phase. It is hypothesize that overexpression of calcium related, thyroid and immune responsive genes might be one of the hallmarks of the second phase of menstrual cycle.
In order to further comprehend the genetic events underlying the endometrial steroids response we undertook a 10.000 genes microarray analysis between human normal endometrium in the proliferative and secretory phases of the menstrual cycle. We selected 24 patients out of 72 women who were diagnosed as having uterine leiomyomata. Neither patient had received any hormonal treatment during the six months previously the diagnosis of leiomyomata, endometrial fragments were divided in proliferative and secretory after histological analysis. Total RNA was extracted from samples by using prior to reverse transcription, two pools of RNA were obtained by mixing 3 µg of total RNA from each individual case in two separated tubes representing endometrial tissues in proliferative (group 1) and secretory phase (group 2). These two pools were then hybridized two times each to four CodeLink UniSet Human I Bioarrays (Amersham Biosciences, Piscataway, NJ). After analysing the results by the SAM software 1127 genes were found differentially expressed between secretory and proliferative phases when a 2,5 difference was used as a cut-off criteria. 364 genes were up-regulated in the proliferative phase and 763 genes were up-regulated in the secretory phase. Among them, genes involved in immune response, thyroid and calcium metabolism were found differentially expressed specially in the secretory phase. It is hypothesize that overexpression of calcium related, thyroid and immune responsive genes might be one of the hallmarks of the second phase of menstrual cycle.
Descrição
Citação
OTSUKA, Audrey Yumi. Identificação de genes diferencialmente expressos no endométrio de mulheres durante o ciclo menstrual. 2006. 116 f. Tese (Doutorado em Ciências) - Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, 2006.