RESUMEN
Endothelial-mesenchymal transition (EndMT) is an important source of cancer-associated fibroblasts (CAFs), which facilitates tumour progression. PDAC is characterised by abundant CAFs and tumour necrosis factor-α (TNF-α). Here, we show that TNF-α strongly induces human endothelial cells to undergo EndMT. Interestingly, TNF-α strongly downregulates the expression of the endothelial receptor TIE1, and reciprocally TIE1 overexpression partially prevents TNF-α-induced EndMT, suggesting that TNF-α acts, at least partially, through TIE1 regulation in this process. We also show that TNF-α-induced EndMT is reversible. Furthermore, TNF-α treatment of orthotopic mice resulted in an important increase in the stroma, including CAFs. Finally, secretome analysis identified TNFSF12, as a regulator that is also present in PDAC patients. With the aim of restoring normal angiogenesis and better access to drugs, our results support the development of therapies targeting CAFs or inducing the EndMT reversion process in PDAC.
Asunto(s)
Fibroblastos Asociados al Cáncer/efectos de los fármacos , Carcinoma Ductal Pancreático/patología , Células Endoteliales/efectos de los fármacos , Transición Epitelial-Mesenquimal/efectos de los fármacos , Neoplasias Pancreáticas/patología , Factor de Necrosis Tumoral alfa/farmacología , Animales , Fibroblastos Asociados al Cáncer/metabolismo , Fibroblastos Asociados al Cáncer/patología , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Células Cultivadas , Citocina TWEAK/genética , Citocina TWEAK/metabolismo , Células Endoteliales/metabolismo , Células Endoteliales/patología , Humanos , Masculino , Ratones Transgénicos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Receptor TIE-1/genética , Receptor TIE-1/metabolismo , Factores de Transcripción de la Familia Snail/genética , Factores de Transcripción de la Familia Snail/metabolismo , Caja Homeótica 2 de Unión a E-Box con Dedos de Zinc/genética , Caja Homeótica 2 de Unión a E-Box con Dedos de Zinc/metabolismoRESUMEN
Endothelial-mesenchymal transition (EndMT) has a significant role in embryonic heart formation and in various pathologies. However, the molecular mechanisms that regulate EndMT induction remain to be elucidated. We show that suppression of receptor tyrosine kinase Tie1 but not Tie2 induces human endothelial cells to undergo EndMT and that Slug deficiency reverts this process. We find that Erk1/2, Erk5 and Akt cascades control Slug promoter activity induced by Tie1 deficiency. Interestingly, EndMT is present in human pancreatic tumour. We propose that EndMT associated with Tie1 downregulation participates in the pathological development of stroma observed in tumours.
Asunto(s)
Transdiferenciación Celular/fisiología , Endotelio/patología , Mesodermo/patología , Neovascularización Patológica/metabolismo , Receptor TIE-1/metabolismo , Adhesión Celular/genética , Adhesión Celular/fisiología , Movimiento Celular/genética , Movimiento Celular/fisiología , Transdiferenciación Celular/genética , Células Cultivadas , Endotelio/metabolismo , Humanos , Immunoblotting , Masculino , Mesodermo/metabolismo , Microscopía Confocal , Neovascularización Patológica/genética , Regiones Promotoras Genéticas/genética , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Interferencia de ARN , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptor TIE-1/genética , Transducción de Señal , Factores de Transcripción de la Familia Snail , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Embryonic stem (ES) cells differentiate in vitro into all cell lineages. We previously found that the p38 mitogen activated kinase (p38MAPK) pathway controls the commitment of ES cells toward either cardiomyogenesis (p38 on) or neurogenesis (p38 off ). In this study, we show that p38α knock-out ES cells do not differentiate into cardiac, endothelial, smooth muscle, and skeletal muscle lineages. Reexpression of p38MAPK in these cells partially rescues their mesodermal differentiation defects and corrects the high level of spontaneous neurogenesis of knock-out cells. Wild-type ES cells were treated with a p38MAPK-specific inhibitor during the differentiation process. These experiments allowed us to identify 2 early independent successive p38MAPK functions in the formation of mesodermal lineages. Further, the first one correlates with the regulation of the expression of Brachyury, an essential mesodermal-specific transcription factor, by p38MAPK. In conclusion, by genetic and biochemical approaches, we demonstrate that p38MAPK activity is essential for the commitment of ES cell into cardiac, endothelial, smooth muscle, and skeletal muscle mesodermal lineages.
Asunto(s)
Diferenciación Celular , Células Madre Embrionarias/citología , Mesodermo/citología , Proteína Quinasa 14 Activada por Mitógenos/metabolismo , Animales , Western Blotting , Células Cultivadas , Células Madre Embrionarias/efectos de los fármacos , Células Madre Embrionarias/metabolismo , Proteínas Fetales/genética , Proteínas Fetales/metabolismo , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Regulación de la Expresión Génica , Técnicas de Inactivación de Genes , Imidazoles/farmacología , Mesodermo/metabolismo , Ratones , Proteína Quinasa 14 Activada por Mitógenos/antagonistas & inhibidores , Desarrollo de Músculos , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Proteínas de Dominio T Box/genética , Proteínas de Dominio T Box/metabolismoRESUMEN
Pancreatic ductal adenocarcinoma remains one of the most lethal of all solid tumors with an overall five-year survival rate of only 3-5%. Its aggressive biology and resistance to conventional and targeted therapeutic agents lead to a typical clinical presentation of incurable disease once diagnosed. The disease is characterized by the presence of a dense stroma of fibroblasts and inflammatory cells, termed desmoplasia, which limits the oxygen diffusion in the organ, creating a strong hypoxic environment within the tumor. In this review, we argue that hypoxia is responsible for the highly aggressive and metastatic characteristics of this tumor and drives pancreatic cancer cells to oncogenic and metabolic changes facilitating their proliferation. However, the molecular changes leading to metabolic adaptations of pancreatic cancer cells remain unclear. Cachexia is a hallmark of this disease and illustrates that this cancer is a real metabolic disease. Hence, this tumor must harbor metabolic pathways which are probably tied in a complex inter-organ dialog during the development of this cancer. Such a hypothesis would better explain how under fuel source limitation, pancreatic cancer cells are maintained, show a growth advantage, and develop metastasis.
RESUMEN
Angiopoietins are ligands of the endothelial cell tyrosine kinase receptor Tie2. Angiopoietin-1 (Ang-1) is widely expressed in human normal adult tissues and promotes blood vessel maturation and stabilization by inducing Tie2 receptor phosphorylation. In contrast, the antagonistic ligand Angiopoietin-2 (Ang-2) is up-regulated by hypoxia, expressed only at sites of vascular remodeling and plays a crucial role in destabilizing vessels for normal or pathological angiogenesis. Ang-2 expression is tightly regulated at transcriptional and post-transcriptional levels. To characterize the regulatory sequences of the human Ang-2 gene we cloned a fragment of around 8.5 kb upstream of the Ang-2 coding sequence and analyzed the luciferase reporter activity of constructs of various lengths in endothelial and non-endothelial cells. We isolated a minimal promoter sequence sufficient to promote significant Ang-2 non-cell type specific transcription. Moreover, we identified sequences conferring endothelial specificity. Indeed, sequence analysis of the fragment revealed the presence of several potential binding sites for specific endothelial regulatory factors like GATA or Ets. Using GATA-2 and Ets-1 co-transfection and overexpression assay, we showed that these two factors are able to induce Ang-2 promoter activation. We also show that hypoxic regulation of Ang-2 is HIF-dependent and demonstrate that HIF-1alpha binds in human microvascular endothelial cells (HMVEC) to an evolutionary conserved Hypoxia-Responsive Element (HRE) located in the first intron of the Ang-2 gene. In conclusion, our study provides new elements in favor of HIF involvement in Ang-2 hypoxic regulation and identifies Ets-1 and particularly GATA-2 as central factors in endothelial specific Ang-2 expression.
Asunto(s)
Angiopoyetina 2/genética , Células Endoteliales/metabolismo , Factor de Transcripción GATA2/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Intrones/genética , Proteína Proto-Oncogénica c-ets-1/metabolismo , Regulación hacia Arriba/genética , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Sitios de Unión , Bovinos , Hipoxia de la Célula , Clonación Molecular , Células HeLa , Humanos , Ratones , Células 3T3 NIH , Unión Proteica , Elementos de Respuesta/genética , Transcripción GenéticaRESUMEN
Tie2, an endothelial cell-specific receptor kinase, has an important role in tumour angiogenesis. In an attempt to identify peptides that specifically interact with and block the Tie2 pathway, a phage-displayed peptide library was screened on a recombinant Tie2 receptor. One peptide, NLLMAAS, completely abolished the binding to Tie2 of both angiopoietin 2 and angiopoietin 1 (Ang1). We further show that NLLMAAS specifically suppresses both Ang1-induced ERK activity and migration in human umbilical endothelial cells. Moreover, in vivo, this peptide inhibits angiogenesis in the chick chorioallantoic membrane assay. NLLMAAS is the first peptide described to interact with Tie2. Our results demonstrate that it is an efficient and specific antagonist of the binding of Tie2 ligands, and suggest that this peptide or its derivates may have potential applications in the treatment of angiogenesis diseases. It also represents a potent tool to dissect the molecular mechanisms involved in the Tie2 pathway.