Resumo:O processo de desenvolvimento embrionário e placentário está sob controle epigenético intenso, com os diversos processos epigenéticos regulando a diferenciação de células pluripotentes em células especializadas. Um desenvolvimento placentário anormal é observado com frequência em gestações de clones, e está associado a uma alteração na reprogramação epigenética da célula doadora e do embrião formado. O objetivo deste trabalho foi avaliar a expressão dos genes "imprinted" TSSC4 e PHLDA2 em tecido cotiledonário e intercotiledonário, aos 60 dias de gestação, de placentas produzidas por Transferência Embrionária (TE), Fecundação In Vitro (FIV) e Transferência Nuclear (TN) em bovinos, e também investigar o controle epigenético desses genes na placenta bovina, avaliando o perfil epigenético dos mesmos em tecidos cotiledonários dos grupos experimentais (TE, FIV e TN). A expressão do gene TSSC4 foi reduzida em 30% na região cotiledonária do grupo produzido por TN, no entanto a expressão do gene PHLDA2 aumentou em aproximadamente onze vezes nos grupos TN e FIV, quando comparados ao grupo TE. Análises de ChIP para a histona H3 na região promotora proximal dos genes TSSC4 e PHLDA2 mostraram um aumento para a marcação permissiva H3K4me2 e uma redução para a inibitória H3K9me2 em cotilédones de placentas clonadas em relação às placentas produzidas por TE. Interessantemente, ambas as marcações de histonas para o gene TSSC4 também estavam significativamente alteradas também em placentas produzidas por FIV, quando comparadas ao grupo TE. Esses resultados mostram que a expressão dos genes "imprinted" TSSC4 e PHLDA2 está alterada em placentas de gestações produzidas por transferência nuclear. Ainda, alterações na expressão do gene PHLDA2 e das modificações de histona no gene TSSC4, nos cotilédones produzidos por FIV, indicam que o cultivoo...(Resumo completo, clicar acesso eletrônico abaixo) / Abstract:The process of embryonic and placental development is under high epigenetic control, with different epigenetic processes regulating the differentiation of pluripotent cells into specialized cells. An abnormal placental development is observed frequently in pregnancies produced by nuclear transfer, and is associated with incomplete epigenetic reprogramming of the donor cell and formed embryo. The objective of this study was to evaluate the expression of the imprinted genes TSSC4 e PHLDA2 in cotiledonary and intercotiledonary tissues, after 60 days of pregnancy, in placentas produced by Embryo Transfer (ET) In Vitro Fertilization (IVF) and Nuclear Transfer (NT) in cattle, and also to investigate the epigenetic control of this gene in the bovine placenta evaluating the epigenetic profile of cotiledonary tissues of the experimental groups (ET, IVF and NT). The TSSC4 gene expression was reduced by 30% in the cotyledons in the NT group, however PHLDA2 expression of the gene was elevated by 11-fold in NT compared to ET. ChIP analysis for histone H3 at the proximal promoter region of TSSC4 and PHLDA2 genes showed an increase to the permissive mark H3K4me2 and a reduction for the inhibitory mark H3K9me2 in the cotyledons of cloned placentae, in relation to placentae produced by ET, for both genes. Interestingly, the inhibitory mark H3K9me2 for TSSC4 gene was also significantly reduced in placentae produced by IVF, compared to ET control group. These results show that expression of the "imprinted" genes TSSC4 and PHLDA2 is affected in placenta of pregnancies produced by nuclear transfer. Further, changes in expression of gene PHLDA2 and histone modifications in TSSC4 gene cotyledons produced by IVF indicate that the in vitro culture can contribute to the observed changes in NT group. / Orientador:Flavia Lombardi Lopes / Banca:Calie Castilho / Banca:Gisele Zoccal Mingoti / Mestre

Made available in DSpace on 2016-09-27T13:39:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2015-12-09. Added 1 bitstream(s) on 2016-09-27T13:45:08Z : No. of bitstreams: 1 000870620.pdf: 854848 bytes, checksum: 8c79744870f5da8cb69ae19583f3ce62 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / The process of embryonic and placental development is under high epigenetic control, with different epigenetic processes regulating the differentiation of pluripotent cells into specialized cells. An abnormal placental development is observed frequently in pregnancies produced by nuclear transfer, and is associated with incomplete epigenetic reprogramming of the donor cell and formed embryo. The objective of this study was to evaluate the expression of the imprinted genes TSSC4 e PHLDA2 in cotiledonary and intercotiledonary tissues, after 60 days of pregnancy, in placentas produced by Embryo Transfer (ET) In Vitro Fertilization (IVF) and Nuclear Transfer (NT) in cattle, and also to investigate the epigenetic control of this gene in the bovine placenta evaluating the epigenetic profile of cotiledonary tissues of the experimental groups (ET, IVF and NT). The TSSC4 gene expression was reduced by 30% in the cotyledons in the NT group, however PHLDA2 expression of the gene was elevated by 11-fold in NT compared to ET. ChIP analysis for histone H3 at the proximal promoter region of TSSC4 and PHLDA2 genes showed an increase to the permissive mark H3K4me2 and a reduction for the inhibitory mark H3K9me2 in the cotyledons of cloned placentae, in relation to placentae produced by ET, for both genes. Interestingly, the inhibitory mark H3K9me2 for TSSC4 gene was also significantly reduced in placentae produced by IVF, compared to ET control group. These results show that expression of the imprinted genes TSSC4 and PHLDA2 is affected in placenta of pregnancies produced by nuclear transfer. Further, changes in expression of gene PHLDA2 and histone modifications in TSSC4 gene cotyledons produced by IVF indicate that the in vitro culture can contribute to the observed changes in NT group.