RESUMO
Ovarian cancer is characterized by being highly metastatic, a feature that represents the main cause of failure of the treatment. This study investigated the effects of γ-secretase inhibition on the TGF-ß-induced epithelial-mesenchymal transition (EMT) process in ovarian cancer cell lines. SKOV3 cells incubated in the presence of TGF-ß showed morphological and biochemical changes related to EMT, which were blocked by co-stimulation with TGF-ß and the γ-secretase inhibitor DAPT. In SKOV3 and IGROV1 cells, the co-stimulation blocked the cadherin switch and the increase in the transcription factors Snail, Slug, Twist and Zeb1 induced by TGF-ß. DAPT impaired the translocation of phospho-ß-catenin to the inner cell compartment observed in TGF-ß-treated cells, but was not able to block the induction at protein level induced by TGF-ß. Moreover, the inhibitor blocked the increased cell migration and invasiveness ability of both cell lines induced by TGF-ß. Notch target genes (Hes1 and Hey1) were induced by TGF-ß, decreased by DAPT treatment and remained low in the presence of both stimuli. However, DAPT alone caused no effects on most of the parameters analyzed. These results demonstrate that the γ-secretase inhibitor used in this study exerted a blockade on TGF-ß-induced EMT in ovarian cancer cells.
Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Dipeptídeos/farmacologia , Inibidores Enzimáticos/farmacologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Neoplasias Ovarianas/patologia , Fator de Crescimento Transformador beta/efeitos adversos , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Forma Celular/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Invasividade Neoplásica , Neoplasias Ovarianas/genética , Fosforilação/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Notch/metabolismo , beta Catenina/metabolismoRESUMO
In this study, we investigated the interaction between the Notch pathway and progesterone to maintain the functionality of the corpus luteum (CL). When Notch signaling is activated, the γ-secretase complex releases the active intracellular domains (NICD) of their receptors, which exert survival effects. We designed studies to analyze whether the in vitro inhibition of Notch affects progesterone production, steroidogenic regulators, apoptotic parameters, and signaling transduction pathways in the cultures of CL isolated from pregnant and superovulated rats. We detected a decrease in progesterone production when corpora lutea (CL) were incubated with N-(N-(3,5-difluorophenacetyl-l-alanyl))-S-phenylglycine t-butyl ester (DAPT), a γ-secretase inhibitor. This effect could be in part due to the decrease detected in the CL protein levels of P450scc because STAR and 3ß-hydroxysteroid dehydrogenase were not affected by Notch inhibition. Besides, the addition of aminoglutethimide to the CL culture medium decreased NICD of NOTCH1. We observed an increase in the expression of active CASPASE3 (CASP3) after inhibition by Notch, which was reversed by the presence of progesterone. The BAX:BCLXL ratio was increased in CL treated with DAPT and the presence of progesterone reversed this effect. In addition, phosphorylation of AKT was inhibited in CL treated with DAPT, but had no effect on ERK activation. To demonstrate that the action of DAPT is specifically related with the inhibition of Notch, CLs were incubated with DLL4 antibody and a decrease in progesterone production was detected. These results suggest the existence of a novel link between progesterone and the Notch signaling pathway to maintain the functionality of the CL.
