Navegando por Palavras-chave "Rho GTPases"
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- ItemSomente MetadadadosCaracterização dos efeitos celulares e moleculares de EHT- 1864, um inibidor de RAC, em células de leucemia mieloide aguda(Universidade Federal de São Paulo (UNIFESP), 2021) Ramos, Debora Felicia Vieira [UNIFESP]; Lazarini, Mariana [UNIFESP]; Universidade Federal de São PauloAcute myeloid leukemia (AML) is a hematological neoplasm caused by molecular changes in the hematopoietic stem- and progenitor cells, leading to the accumulation of undifferentiated myeloid cells with higher proliferation capacity (blasts). The presence of blasts in the bone marrow compromises hematopoiesis and blasts can affect peripheral blood and infiltrate other tissues and organs, such as lung and nervous system. AML is highly heterogeneous and standard treatment is nonspecific, resulting in low survival rates, especially in the elderly. Thus, it is essential to investigate new therapeutic targets that may result in the development of more effective and specific therapies. The Rho family of GTPase proteins is composed by twenty cytoskeleton regulators that have been shown to participate in the physiopathology of hematological neoplasms. Our preliminary results showed that the expression of Rac3 is increased in AML. In addition, the expression of Rho GTPase Rac2 is associated with lower survival of AML patients. However, little is known about the participation of these Rho GTPases in the physiopathology of this disease. The aim of this study was to evaluate the effects of EHT-1864, an inhibitor of the Rac subfamily of Rho GTPases (Rac1, Rac2 and Rac3) on AML cells. For this, we evaluated the processes of apoptosis, autophagy and cell cycle in a AML cell line treated in vitro with EHT-1864. The PI3K/AKT/mTOR signaling pathway was also evaluated, as well as the effects of the combination of EHT- 1864 and daunorubicin on the apoptosis. Pharmacological inhibition of Rac reduced the viability of OCI-AML3, KG1 and Kasumi1 cells in a time and dose-dependent manner. In OCI-AML3 cells, treatment with EHT-1864 for 48 and 72 hours induced apoptosis and autophagy processes and led to an accumulation of the percentage of cells in the G1 phase of the cell cycle, with consequent reduction in phase S. The changes in the cell cycle were concomitant with the change in the expression of p53 and cyclins (reduction of cyclins A and E and increase of cyclin B). In addition, we observed lower activation of the PI3K/AKT/mTOR pathway in OCI-AML3 cells treated with EHT-1864. Interestingly, the combined treatment of EHT-1864 and low doses of daunorrubicin in vitro enhanced OCI-AML3 cell apoptosis. This study demonstrates the antitumor capacity of EHT-1864 against AML cell lineage, including potentiating effect when9 used in combination with daunorubicin. Our results suggest that the observed effects are at least partly dependent on PI3K/AKT/mTOR inhibition.
- ItemSomente MetadadadosInvasion of MDCK epithelial cells with altered expression of Rho GTPases by Trypanosoma cruzi amastigotes and metacyclic trypomastigotes of strains from the two major phylogenetic lineages(Elsevier B.V., 2004-04-01) Fernandes, A. B.; Mortara, R. A.; Universidade Federal de São Paulo (UNIFESP)In order to invade mammalian cells, Trypanosoma cruzi infective forms cause distinct rearrangements of membrane and host cell cytoskeletal components. Rho GTPases have been shown to regulate three separate signal transduction pathways, linking plasma membrane receptors to the assembly of distinct actin filament structures. Here, we examined the role of Rho GTPases on the interaction between different T cruzi infective forms of strains from the two major phylogenetic lineages with nonpolarized MDCK cells transfected with different Rho GTPase constructs. We compared the infectivity of amastigotes isolated from infected cells (intracellular amastigotes) with forms generated from the axenic differentiation of trypomastigotes (extracellular amastigotes), and also with metacyclic trypomastigotes.No detectable effect of GTPase expression was observed on metacyclic trypomastigote invasion and parasites of Y and CL (T cruzi 11) strains invaded to similar degrees all MDCK transfectants, and were more infective than either G or Tulahuen (T cruzi I) strains. Intracellular amastigotes were complement sensitive and showed very low infectivity towards the different transfectants regardless of the parasite strain. Complement-resistant T cruzi I extracellular amastigotes, especially of the G strain, were more infective than T cruzi 11 parasites, particularly for the Rac1V12 constitutively active GTPase transfectant. the fact that in Rac1N17 dominant-negative cells, the invasion of G strain extracellular amastigotes was specifically inhibited suggested an important role for Rac1 in this process. (C) 2004 Elsevier SAS. All rights reserved.
- ItemAcesso aberto (Open Access)Investigação dos efeitos do silenciamento das proteínas RhoA e RhoC na proliferação clonal de linhagens mieloides(Universidade Federal de São Paulo, 2023-11-24) Mendonça, Guilherme Ramos Sales de [UNIFESP]; Lazarini, Mariana [UNIFESP]; http://lattes.cnpq.br/7419529682193778; http://lattes.cnpq.br/3572076966097043A leucemia mieloide aguda (LMA) é um câncer de medula óssea altamente heterogêneo, caracterizado por desregulação bioquímica resultante de alterações moleculares em células-tronco e progenitoras do sistema hematopoiético. Estas alterações causam um bloqueio na diferenciação celular com acúmulo de células imaturas na medula óssea e/ou sangue periférico, comprometendo a produção de células sanguíneas. O tratamento da LMA em geral é inespecífico e está associado a baixas taxas de sobrevida. RhoA e RhoC são proteínas homólogas da família de Rho GTPases envolvidas em processos relacionados à organização do citoesqueleto, como proliferação celular, e estão desreguladas em cânceres humanos. Porém, as funções de RhoA e RhoC foram pouco exploradas em LMA. O objetivo deste trabalho foi investigar a participação das proteínas RhoA e RhoC no processo de proliferação de células leucêmicas. Para isso, foram avaliados os efeitos do silenciamento de RhoA e RhoC na proliferação clonal das linhagens celulares das séries mieloides U937 e OCI-AML3. O ensaio de proliferação clonal foi primeiramente padronizado nas células parentais e então realizado em células silenciadas com lentivírus específicos para inibição de RhoA ou RhoC. O silenciamento de RhoA causou redução significativa na capacidade de proliferação clonal de ambas as linhagens leucêmicas, enquanto o silenciamento de RhoC não causou mudanças significativas na proliferação clonal destas linhagens.
- ItemAcesso aberto (Open Access)Rac1/WAVE2 and Cdc42/N-WASP Participation in Actin-Dependent Host Cell Invasion by Extracellular Amastigotes of Trypanosoma cruzi(Frontiers Media Sa, 2018) Bonfim-Melo, Alexis [UNIFESP]; Ferreira, Eden R. [UNIFESP]; Mortara, Renato A. [UNIFESP]This study evaluated the participation of host cell Rho-family GTPases and their effector proteins in the actin-dependent invasion by Trypanosoma cruzi extracellular amastigotes (EAs). We observed that all proteins were recruited and colocalized with actin at EA invasion sites in live or fixed cells. EA internalization was inhibited in cells depleted in Rac1, N-WASP, and WAVE2. Time-lapse experiments with Rac1, N-WASP and WAVE2 depleted cells revealed that EA internalization kinetics is delayed even though no differences were observed in the proportion of EA-induced actin recruitment in these groups. Overexpression of constitutively active constructs of Rac1 and RhoA altered the morphology of actin recruitments to EA invasion sites. Additionally, EA internalization was increased in cells overexpressing CA-Rac1 but inhibited in cells overexpressing CA-RhoA. WT-Cdc42 expression increased EA internalization, but curiously, CA-Cdc42 inhibited it. Altogether, these results corroborate the hypothesis of EA internalization in non-phagocytic cells by a phagocytosis-like mechanism and present Rac1 as the key Rho-family GTPase in this process.