Estudo do papel do óxido nítrico na ativação da GTPase Rac1 em células endoteliais e de melanoma murino
Data
2015-11-25
Autores
Borges, Roberta Eller 
Orientadores
Monteiro, Hugo Pequeno
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
As proteínas Rac pertencem à família Rho-GTPases que controlam uma ampla variedade de processos celulares como, por exemplo, a formação de fibras de estresse e de adesão focal, a diferenciação celular, migração celular e apoptose. Elas atuam como chaves moleculares, e quando ativadas se encontram associadas ao nucleotídeo GTP e nesta conformação interagem com outras proteínas sinalizadoras participando ativamente dos processos de transdução de sinais originários de vários estímulos extracelulares. Muitos estudos mostraram que a família Rho-GTPases desempenha um papel importante em processos como a migração celular e produção de espécies reativas de oxigênio (ERO). Além disso, também é conhecido que a GTPase Rac-l, está envolvida na migração de células endoteliais associada a angiogênese com as ERO promovendo estes eventos (L. Moldovan et ai, 2006). A GTPase Rac-l também participa na migração de células tumorais relacionada a processos de invasão e de metástase. Entretanto, pouco se sabe sobre o papel do óxido nítrico (NO) nestes processos. Assim, o objetivo deste trabalho foi avaliar a participação do NO na regulação da atividade de Rac-l emcélulas endoteliais de aorta de coelho (RAEC), bem como em modelo de melanoma murino. Inicialmente foi observado que tanto NO derivado de estimulação por bradicinina (BK) como o S-nitrosotiol nitrosoglutationa (GSNO) foram capazes de ativar Rac-l em células RAEC. Este processo levou a igração das células e mostrou-se dependente da S-nitrosilação de Ras, com participação de PI3K. Nas células tumorais B16FIO, foram isoladas duas sublinhagens com capacidades metastáticas polares, sendo a sublinhagem Nex8H (células metastáticas), e a sublinhagem NexlOC (mimetiza tumor primário). Observamos que a ativação de Rac- 1 está envolvida na inibição da migração e invasão celular do melanoma murino metastático Nex8H, quando a produção endógena de NO foi inibida. Curiosamente, o 1 resultado inverso foi observado em células NexlOC (não metastáticas). Esses resultados mostram que a GTPase Rac-I desempenha papéis antagônicos frente a um mesmo estímulo dependendo do estágio de desenvolvimento em que se encontra a célula tumoral, e os mesmos pcidem contribuir para melhor compreender os mecanismos que envolvem a relação entre NO, Rac-I e processos de invasão e migração celular.
The proteins Rac1 of the Rho family proteins that control a variety of cellular processes such as the formation of stress fibers and focal adhesion, cell differentiation, cell migration and apoptosis. They act as molecular switches, when activated and are associated with the nucleotide GTP, and this conformation interact with other signaling proteins participating actively in signal transduction processes originate from various extracellular stimuli. Many studies have shown that Rho-family GTPases plays an important role in processes such as cell migration and production of reactive oxygen species (ROS). Furthermore, it is also known that the Rac GTPase-1 is involved in migration of endothelial cells to angiogenesis associated with ROS promote these events (L. Moldovan et al, 2006). However, not known about the role of nitric oxide (NO) in this processo The objective of this work was to evaluate the role of NO in the regulation of Rac-l activity in Rabbit Aorta Endothelial Cells (RAEC), as well in murine melanoma model. Initially was observed that both the derived stimulation by bradykinin (BK) nitrosothiol such as S-nitrosoglutathione (GSNO) were able to activate Rac-I in RAEC cells. This process led to the migration of cells and was shown to be dependent on the S-nitrosylation ofRas, with the participation ofPI3K. In the B16FIO tumor cells, they were isolated sublineages two polar metastatic capacity, with Nex8H subline (metastatic cells), and NexlOC subline (mimicking primary tumor). This second part has been observed that the activation of Rac-l is involved in inhibiting cell migration and invasion of metastatic murine melanoma Nex8H, when the endogenous NO production was inhibited. Interestingly, the opposite effect was observed in NexlOC cells (non-metastatic). These results show that this has GTPase front antagonistic roles to the same stimulus depending on the metastatic capacity of the tumor cell, and they can contribute to better understand the mechanisms that involve the relationship between NO, Rac-l and invasion and cellular migration.
The proteins Rac1 of the Rho family proteins that control a variety of cellular processes such as the formation of stress fibers and focal adhesion, cell differentiation, cell migration and apoptosis. They act as molecular switches, when activated and are associated with the nucleotide GTP, and this conformation interact with other signaling proteins participating actively in signal transduction processes originate from various extracellular stimuli. Many studies have shown that Rho-family GTPases plays an important role in processes such as cell migration and production of reactive oxygen species (ROS). Furthermore, it is also known that the Rac GTPase-1 is involved in migration of endothelial cells to angiogenesis associated with ROS promote these events (L. Moldovan et al, 2006). However, not known about the role of nitric oxide (NO) in this processo The objective of this work was to evaluate the role of NO in the regulation of Rac-l activity in Rabbit Aorta Endothelial Cells (RAEC), as well in murine melanoma model. Initially was observed that both the derived stimulation by bradykinin (BK) nitrosothiol such as S-nitrosoglutathione (GSNO) were able to activate Rac-I in RAEC cells. This process led to the migration of cells and was shown to be dependent on the S-nitrosylation ofRas, with the participation ofPI3K. In the B16FIO tumor cells, they were isolated sublineages two polar metastatic capacity, with Nex8H subline (metastatic cells), and NexlOC subline (mimicking primary tumor). This second part has been observed that the activation of Rac-l is involved in inhibiting cell migration and invasion of metastatic murine melanoma Nex8H, when the endogenous NO production was inhibited. Interestingly, the opposite effect was observed in NexlOC cells (non-metastatic). These results show that this has GTPase front antagonistic roles to the same stimulus depending on the metastatic capacity of the tumor cell, and they can contribute to better understand the mechanisms that involve the relationship between NO, Rac-l and invasion and cellular migration.
Descrição
Citação
BORGES, Roberta Eller. Estudo do papel do óxido nítrico na ativação da GTPase Rac1 em células endoteliais e de melanoma murino. 2015. 133 f. Tese (Doutorado em Biologia Molecular) - Escola Paulista de Medicina, Universidade Federal de São Paulo (UNIFESP), São Paulo, 2015.