RESUMO
Abnormal trophoblast function is associated with human fetal growth restriction (FGR). Targeted disruption of homeobox gene transforming growth ß-induced factor (TGIF-1) results in placental dysfunction in the mouse. The role of human TGIF-1 in placental cell function is unknown. The aims of this study were to determine the expression of TGIF-1 in human idiopathic FGR-affected placentae compared with gestation-matched controls (GMC), to elucidate the functional role of TGIF-1 in trophoblasts and to identify its downstream targets. Real-time PCR and immunoblotting revealed that TGIF-1 mRNA and protein expression was significantly increased in FGR-affected placentae compared with GMC (n = 25 in each group P < 0.05). Immunoreactive TGIF-1 was localized to the villous cytotrophoblasts, syncytiotrophoblast, microvascular endothelial cells and in scattered stromal cells in both FGR and GMC. TGIF-1 inactivation in BeWo cells using two independent siRNA resulted in significantly decreased mRNA and protein of trophoblast differentiation markers, human chorionic gonadotrophin (CGB/hCG), syncytin and 3ß-hydroxysteroid dehydrogenase/3ß-honest significant difference expression. Our data demonstrate that homeobox gene TGIF-1 is a potential up-stream regulator of trophoblast differentiation and the altered TGIF-1 expression may contribute to aberrant villous trophoblast differentiation in FGR.
Assuntos
Retardo do Crescimento Fetal/metabolismo , Proteínas de Homeodomínio/metabolismo , Proteínas Repressoras/metabolismo , Trofoblastos/citologia , Trofoblastos/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Adulto , Animais , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Gonadotropina Coriônica/genética , Gonadotropina Coriônica/metabolismo , Feminino , Retardo do Crescimento Fetal/genética , Produtos do Gene env/genética , Produtos do Gene env/metabolismo , Proteínas de Homeodomínio/genética , Humanos , Camundongos , Gravidez , Proteínas da Gravidez/genética , Proteínas da Gravidez/metabolismo , Proteínas Repressoras/genéticaRESUMO
Human idiopathic fetal growth restriction (FGR) is associated with placental insufficiency. Previously, we reported that the expression of homeobox gene Distal-less 3 (DLX3) is increased in idiopathic FGR placentae and is a regulator of villous trophoblast differentiation. Here, we identify the downstream targets of DLX3 in trophoblast-derived cell lines. We modelled the high levels of DLX3 in FGR using an over-expression plasmid construct and complemented this using short-interference RNA (siRNA) for inactivation in cultured cells. Using a real-time PCR-based gene profiling, candidate target genes of DLX3 over-expression and inactivation were identified as regulators of trophoblast differentiation; GATA2 and PPARγ. The expression of GATA2 and PPARγ were further assessed in placental tissues and showed increased mRNA and protein levels in FGR-affected tissues compared with gestation-matched controls. We conclude that DLX3 orchestrates the expression of multiple regulators of trophoblast differentiation and that expression of these regulatory genes is abnormal in FGR.
Assuntos
Retardo do Crescimento Fetal/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Proteínas de Homeodomínio/metabolismo , Placenta/metabolismo , Transdução de Sinais/genética , Fatores de Transcrição/metabolismo , Adulto , Linhagem Celular , Feminino , Fator de Transcrição GATA2/metabolismo , Estudos de Associação Genética , Proteínas de Homeodomínio/genética , Humanos , PPAR gama/metabolismo , Placenta/patologia , Plasmídeos/metabolismo , Gravidez , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Reprodutibilidade dos Testes , Fatores de Transcrição/genética , Trofoblastos/metabolismo , Trofoblastos/patologiaRESUMO
Human idiopathic foetal growth restriction (FGR) is frequently associated with placental insufficiency. In our previous studies, we have reported the isolation and characterisation of the homeobox gene Distal-less 3 (DLX3) in the human placenta. In this study, we have investigated the level of DLX3 expression in idiopathic FGR-affected placentae and determined its functional role in villous trophoblast differentiation. FGR-affected placentae (n = 25) were collected based on well-defined clinical criteria and matched for gestation with control uncomplicated pregnancies (n = 25). Real-time polymerase chain reaction and immunoblotting showed increased DLX3 mRNA and protein expression in FGR-affected placentae compared with gestation-matched controls. Qualitative immunohistochemistry revealed DLX3 localisation in the syncytiotrophoblast, cytotrophoblasts and endothelial cells surrounding the foetal capillaries in both FGR-affected and control placentae. Down-regulation of DLX3 in primary villous trophoblast cells and a trophoblast-derived cell line showed decreased expression of differentiation markers, 3ßHSD, ßhCG and syncytin. Therefore, we conclude that increased DLX3 expression in FGR may contribute to trophoblast dysfunction observed in FGR.