Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos
Autor(a) principal: | |
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Data de Publicação: | 2018 |
Tipo de documento: | Dissertação |
Idioma: | por |
Título da fonte: | Repositório Institucional da UFSCAR |
Texto Completo: | https://repositorio.ufscar.br/handle/ufscar/10578 |
Resumo: | Fertility is directly related to oocyte developmental competence. In this context, oocyte mitochondria play a crucial role and have become an important target in assisted reproduction therapies. However, the molecular mechanisms underlying these mitochondrial effects are unknown. Therefore, the implications of the deletion of Mitofusin 1 (Mfn1 cKO) in murine oocytes was investigated in this project. Infertility of Mfn1 cKO females was characterized by interruption in oocyte growth and total failure in ovulation. Compared with wild type oocytes (WT), Mfn1 cKO oocytes showed mitochondria with abnormal ultrastructure, fewer cristae and a vesicular internal structure. Assays with BrdU incorporation by the granulosa cells reinforced the phenotype of blocked folliculogenesis, since the Mfn1 cKO group presented a decrease in the number of cells in replication. In addition, it was verified that this group shows a decrease, both in the oocyte and in the ovary, in the amount of transcripts of most genes analyzed, involved with oocyte-granulosa communication and folliculogenesis control. Important factors for the progression of folliculogenesis had their mRNA levels restored when Mfn1 cKO granulosa-oocyte complexes (GOCs) were co-cultured with denuded WT oocytes. To test whether were due to an imbalance in the expression of Mitofusin 2 (MFN2) relative to MFN1, MFN2 or MFN1 and MFN2 were overexpressed during GOC culture. It was observed that MFN2 overexpression inhibited oocyte growth, which was rescued by overexpression of both Mitofusins. Therefore, the infertility of females Mfn1 cKO seems to be related to an imbalance in the expression of MFN1 and MFN2. Consequently, Mfn1 deletion led to the inhibition of PI3K/AKT signaling pathways in the oocyte as well as the expression of GDF9, Bmp15 and Fgf8b, resulting in folliculogenesis blockage, as it affected oocyte-granulosa communication. |
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Carvalho, Karen FreireChiaratti, Marcos Robertohttp://lattes.cnpq.br/0392078007500289http://lattes.cnpq.br/02932350261259722018-10-15T13:11:08Z2018-10-15T13:11:08Z2018-08-31CARVALHO, Karen Freire. Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos. 2018. Dissertação (Mestrado em Genética Evolutiva e Biologia Molecular) – Universidade Federal de São Carlos, São Carlos, 2018. Disponível em: https://repositorio.ufscar.br/handle/ufscar/10578.https://repositorio.ufscar.br/handle/ufscar/10578Fertility is directly related to oocyte developmental competence. In this context, oocyte mitochondria play a crucial role and have become an important target in assisted reproduction therapies. However, the molecular mechanisms underlying these mitochondrial effects are unknown. Therefore, the implications of the deletion of Mitofusin 1 (Mfn1 cKO) in murine oocytes was investigated in this project. Infertility of Mfn1 cKO females was characterized by interruption in oocyte growth and total failure in ovulation. Compared with wild type oocytes (WT), Mfn1 cKO oocytes showed mitochondria with abnormal ultrastructure, fewer cristae and a vesicular internal structure. Assays with BrdU incorporation by the granulosa cells reinforced the phenotype of blocked folliculogenesis, since the Mfn1 cKO group presented a decrease in the number of cells in replication. In addition, it was verified that this group shows a decrease, both in the oocyte and in the ovary, in the amount of transcripts of most genes analyzed, involved with oocyte-granulosa communication and folliculogenesis control. Important factors for the progression of folliculogenesis had their mRNA levels restored when Mfn1 cKO granulosa-oocyte complexes (GOCs) were co-cultured with denuded WT oocytes. To test whether were due to an imbalance in the expression of Mitofusin 2 (MFN2) relative to MFN1, MFN2 or MFN1 and MFN2 were overexpressed during GOC culture. It was observed that MFN2 overexpression inhibited oocyte growth, which was rescued by overexpression of both Mitofusins. Therefore, the infertility of females Mfn1 cKO seems to be related to an imbalance in the expression of MFN1 and MFN2. Consequently, Mfn1 deletion led to the inhibition of PI3K/AKT signaling pathways in the oocyte as well as the expression of GDF9, Bmp15 and Fgf8b, resulting in folliculogenesis blockage, as it affected oocyte-granulosa communication.A fertilidade está diretamente relacionada à competência de desenvolvimento do oócito. Nesse contexto, as mitocôndrias oocitárias desempenham um papel crucial e se tornaram um importante alvo em terapias de reprodução assistida. No entanto, os mecanismos moleculares subjacentes a estes efeitos mitocondriais são desconhecidos. Assim sendo, as implicações da deleção da Mitofusina 1 (Mfn1 cKO) em oócitos murinos foi investigada nesse projeto. A infertilidade de fêmeas Mfn1 cKO foi caracterizada por interrupção no crescimento do oócito e total falha na ovulação. Em comparação com oócitos selvagens (WT), oócitos Mfn1 cKO apresentaram mitocôndrias com ultraestrutura anormal, menor quantidade de cristas e estrutura interna vesiculada. Ensaios com incorporação de BrdU pelas células da granulosa reforçaram o fenótipo de foliculogênese bloqueada, pois o grupo Mfn1 cKO apresentou diminuição no número de células em replicação. Além disso, foi verificado que este grupo apresenta diminuição, tanto no oócito como no ovário, na quantidade de transcritos da maior parte dos genes analisados, envolvidos com comunicação oócito-granulosa e com o controle da foliculogênese. Fatores importantes para a progressão da foliculogênese tiveram seus níveis de mRNA restaurados quando complexos granulosa-oócito (GOCs) Mfn1 cKO foram co-cultivados com oócitos WT desnudos. Para testar a hipótese de que o efeito do cKO de Mfn1 foi devido a um desbalanço na expressão da Mitofusina 2 (MFN2) em relação a MFN1, MFN2 ou MFN1 e MFN2 foram superexpressados durante o cultivo de GOCs. Observou-se que a superexpressão de MFN2 inibiu o crescimento do oócito, o qual foi resgatado pela superexpressão de ambas as Mitofusinas. Portanto, a infertilidade de fêmeas Mfn1 cKO parece estar relacionada a um desbalanço na expressão de MFN1 e MFN2. Consequentemente, a deleção somente de Mfn1 levou à inibição de vias de sinalização PI3K/AKT no oócito e também da expressão de GDF9, Bmp15 e Fgf8b, resultando em bloqueio na foliculogênese, por afetar a comunição oócito-granulosa.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)FAPESP 2016/11942-5porUniversidade Federal de São CarlosCâmpus São CarlosPrograma de Pós-Graduação em Genética Evolutiva e Biologia Molecular - PPGGEvUFSCarFertilidadeMitocôndriaMitofusinaFertilityMitochondriaMitofusinCIENCIAS BIOLOGICAS::BIOQUIMICA::BIOLOGIA MOLECULARCIENCIAS BIOLOGICASCIENCIAS BIOLOGICAS::GENETICA::GENETICA ANIMALEfeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinosFertility effect of the knockout of Mitofusin 1 on murine oocytesinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisOnlineinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UFSCARinstname:Universidade Federal de São Carlos (UFSCAR)instacron:UFSCARORIGINALDissertação_Karen_versão final.pdfDissertação_Karen_versão final.pdfArquivo contendo a versão final da dissertação.application/pdf2077786https://{{ getenv "DSPACE_HOST" "repositorio.ufscar.br" }}/bitstream/ufscar/10578/1/Disserta%c3%a7%c3%a3o_Karen_vers%c3%a3o%20final.pdf630e790e5769c24c95a17727a301fc98MD51LICENSElicense.txtlicense.txttext/plain; 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dc.title.por.fl_str_mv |
Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos |
dc.title.alternative.eng.fl_str_mv |
Fertility effect of the knockout of Mitofusin 1 on murine oocytes |
title |
Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos |
spellingShingle |
Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos Carvalho, Karen Freire Fertilidade Mitocôndria Mitofusina Fertility Mitochondria Mitofusin CIENCIAS BIOLOGICAS::BIOQUIMICA::BIOLOGIA MOLECULAR CIENCIAS BIOLOGICAS CIENCIAS BIOLOGICAS::GENETICA::GENETICA ANIMAL |
title_short |
Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos |
title_full |
Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos |
title_fullStr |
Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos |
title_full_unstemmed |
Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos |
title_sort |
Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos |
author |
Carvalho, Karen Freire |
author_facet |
Carvalho, Karen Freire |
author_role |
author |
dc.contributor.authorlattes.por.fl_str_mv |
http://lattes.cnpq.br/0293235026125972 |
dc.contributor.author.fl_str_mv |
Carvalho, Karen Freire |
dc.contributor.advisor1.fl_str_mv |
Chiaratti, Marcos Roberto |
dc.contributor.advisor1Lattes.fl_str_mv |
http://lattes.cnpq.br/0392078007500289 |
contributor_str_mv |
Chiaratti, Marcos Roberto |
dc.subject.por.fl_str_mv |
Fertilidade Mitocôndria Mitofusina |
topic |
Fertilidade Mitocôndria Mitofusina Fertility Mitochondria Mitofusin CIENCIAS BIOLOGICAS::BIOQUIMICA::BIOLOGIA MOLECULAR CIENCIAS BIOLOGICAS CIENCIAS BIOLOGICAS::GENETICA::GENETICA ANIMAL |
dc.subject.eng.fl_str_mv |
Fertility Mitochondria Mitofusin |
dc.subject.cnpq.fl_str_mv |
CIENCIAS BIOLOGICAS::BIOQUIMICA::BIOLOGIA MOLECULAR CIENCIAS BIOLOGICAS CIENCIAS BIOLOGICAS::GENETICA::GENETICA ANIMAL |
description |
Fertility is directly related to oocyte developmental competence. In this context, oocyte mitochondria play a crucial role and have become an important target in assisted reproduction therapies. However, the molecular mechanisms underlying these mitochondrial effects are unknown. Therefore, the implications of the deletion of Mitofusin 1 (Mfn1 cKO) in murine oocytes was investigated in this project. Infertility of Mfn1 cKO females was characterized by interruption in oocyte growth and total failure in ovulation. Compared with wild type oocytes (WT), Mfn1 cKO oocytes showed mitochondria with abnormal ultrastructure, fewer cristae and a vesicular internal structure. Assays with BrdU incorporation by the granulosa cells reinforced the phenotype of blocked folliculogenesis, since the Mfn1 cKO group presented a decrease in the number of cells in replication. In addition, it was verified that this group shows a decrease, both in the oocyte and in the ovary, in the amount of transcripts of most genes analyzed, involved with oocyte-granulosa communication and folliculogenesis control. Important factors for the progression of folliculogenesis had their mRNA levels restored when Mfn1 cKO granulosa-oocyte complexes (GOCs) were co-cultured with denuded WT oocytes. To test whether were due to an imbalance in the expression of Mitofusin 2 (MFN2) relative to MFN1, MFN2 or MFN1 and MFN2 were overexpressed during GOC culture. It was observed that MFN2 overexpression inhibited oocyte growth, which was rescued by overexpression of both Mitofusins. Therefore, the infertility of females Mfn1 cKO seems to be related to an imbalance in the expression of MFN1 and MFN2. Consequently, Mfn1 deletion led to the inhibition of PI3K/AKT signaling pathways in the oocyte as well as the expression of GDF9, Bmp15 and Fgf8b, resulting in folliculogenesis blockage, as it affected oocyte-granulosa communication. |
publishDate |
2018 |
dc.date.accessioned.fl_str_mv |
2018-10-15T13:11:08Z |
dc.date.available.fl_str_mv |
2018-10-15T13:11:08Z |
dc.date.issued.fl_str_mv |
2018-08-31 |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/masterThesis |
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masterThesis |
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publishedVersion |
dc.identifier.citation.fl_str_mv |
CARVALHO, Karen Freire. Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos. 2018. Dissertação (Mestrado em Genética Evolutiva e Biologia Molecular) – Universidade Federal de São Carlos, São Carlos, 2018. Disponível em: https://repositorio.ufscar.br/handle/ufscar/10578. |
dc.identifier.uri.fl_str_mv |
https://repositorio.ufscar.br/handle/ufscar/10578 |
identifier_str_mv |
CARVALHO, Karen Freire. Efeito do knockout da Mitofusina 1 sobre a fertilidade de oócitos murinos. 2018. Dissertação (Mestrado em Genética Evolutiva e Biologia Molecular) – Universidade Federal de São Carlos, São Carlos, 2018. Disponível em: https://repositorio.ufscar.br/handle/ufscar/10578. |
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https://repositorio.ufscar.br/handle/ufscar/10578 |
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openAccess |
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Universidade Federal de São Carlos Câmpus São Carlos |
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Programa de Pós-Graduação em Genética Evolutiva e Biologia Molecular - PPGGEv |
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UFSCar |
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Universidade Federal de São Carlos Câmpus São Carlos |
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