RESUMO
Interleukin-11 (IL-11) up-regulates the proliferative and invasive capacity of many cancers. Coexpression of glycoprotein 130 (GP130) and IL-11 receptor alpha (IL-11Ralpha) is necessary for high-affinity binding of IL-11 to IL-11Ralpha. This study investigated the expression of IL-11 and role of prostaglandin F(2alpha)-F-prostanoid receptor (FP receptor) signaling in the modulation of IL-11 expression in endometrial adenocarcinoma cells. Localization of IL-11, IL-11Ralpha, and GP130 expression was performed by immunohistochemistry. IL-11 and regulator of calcineurin 1 isoform 4 (RCAN1-4) mRNA and protein expression were determined by real-time RT-PCR and/or enzyme-linked immunosorbent assay/Western blot analysis using Ishikawa endometrial adenocarcinoma cells stably expressing the FP receptor (FPS cells) and endometrial adenocarcinoma explants. IL-11 mRNA expression was significantly elevated in endometrial adenocarcinoma samples compared with normal endometrium and increased with tumor grade. IL-11 protein expression localized with FP receptor, IL-11Ralpha, and GP130 in the neoplastic glandular epithelium of endometrial adenocarcinomas. Prostaglandin F(2alpha)-FP receptor signaling significantly elevated the expression of IL-11 mRNA and protein in a Gq-protein kinase C-calcium-calcineurin-nuclear factor of activated T cells-dependent manner in FPS cells. The calcineurin signaling pathway is known to be controlled by the RCAN (RCAN1-4). Indeed, RCAN1-4 expression was significantly elevated in well-differentiated endometrial adenocarcinoma compared with normal endometrium and was found to decrease with tumor grade and negatively regulate IL-11 expression in vitro. This study has highlighted a new mechanism regulating IL-11 expression in endometrial adenocarcinoma cells by the FP receptor via the calcium-calcineurin-nuclear factor of activated T cells pathway.
Assuntos
Calcineurina/metabolismo , Cálcio/metabolismo , Neoplasias do Endométrio/genética , Interleucina-11/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Musculares/metabolismo , Fatores de Transcrição NFATC/metabolismo , Receptores de Prostaglandina/metabolismo , Idoso , Diferenciação Celular , Proliferação de Células , Receptor gp130 de Citocina/genética , Receptor gp130 de Citocina/metabolismo , Proteínas de Ligação a DNA , Neoplasias do Endométrio/patologia , Endométrio/metabolismo , Endométrio/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Interleucina-11/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Pessoa de Meia-Idade , Modelos Biológicos , Proteínas Musculares/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Interleucina-11/genética , Receptores de Interleucina-11/metabolismoRESUMO
Prokineticin-1 (PROK1) is a multifunctional secreted protein which signals via the G-protein coupled receptor, PROKR1. Previous data from our laboratory using a human genome survey microarray showed that PROK1-prokineticin receptor 1 (PROKR1) signalling regulates numerous genes important for establishment of early pregnancy, including the cytokine interleukin (IL)-11. Here, we have shown that PROK1-PROKR1 induces the expression of IL-11 in PROKR1 Ishikawa cells and first trimester decidua via the calcium-calcineurin signalling pathway in a guanine nucleotide-binding protein (G(q/11)), extracellular signal-regulated kinases, Ca(2+) and calcineurin-nuclear factor of activated T cells dependent manner. Conversely, treatment of human decidua with a lentiviral miRNA to abolish endogenous PROK1 expression results in a significant reduction in IL-11 expression and secretion. Importantly, we have also shown a regulatory role for the regulator of calcineurin 1 isoform 4 (RCAN1-4). Overexpression of RCAN1-4 in PROKR1 Ishikawa cells using an adenovirus leads to a reduction in PROK1 induced IL-11 indicating that RCAN1-4 is a negative regulator in the calcineurin-mediated signalling to IL-11. Finally, we have shown the potential for both autocrine and paracrine signalling in the human endometrium by co-localizing IL-11, IL-11Ralpha and PROKR1 within the stromal and glandular epithelial cells of non-pregnant endometrium and first trimester decidua. Overall we have identified and characterized the signalling components of a novel PROK1-PROKR1 signalling pathway regulating IL-11.
Assuntos
Calcineurina/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Interleucina-11/metabolismo , Fatores de Transcrição NFATC/fisiologia , Receptores Acoplados a Proteínas G/fisiologia , Fator de Crescimento do Endotélio Vascular Derivado de Glândula Endócrina/fisiologia , Calcineurina/metabolismo , Inibidores de Calcineurina , Linhagem Celular Tumoral , Ciclosporina/farmacologia , Decídua/metabolismo , Ácido Egtázico/farmacologia , Endométrio/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Flavonoides/farmacologia , Humanos , Imuno-Histoquímica , Interleucina-11/genética , Fatores de Transcrição NFATC/antagonistas & inibidores , Peptídeos Cíclicos/farmacologia , Gravidez , Primeiro Trimestre da Gravidez , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Fator de Crescimento do Endotélio Vascular Derivado de Glândula Endócrina/genética , Fator de Crescimento do Endotélio Vascular Derivado de Glândula Endócrina/farmacologiaRESUMO
BACKGROUND: Combretastatin A-4-phosphate (CA-4-P) is a microtubule depolymerising agent currently in clinical trial as a tumour vascular-targeting agent. In vivo, CA-4-P causes rapid shutdown of tumour blood flow (within minutes) and a significant neutrophil infiltration at later times. MATERIALS AND METHODS: Using an in vitro flow-cell assay, we investigated neutrophil-endothelial cell interactions and associated mechanisms, following endothelial cell exposure to CA-4-P. Cellular adhesion molecule (CAM) expression was examined using immunoblotting and immunofluorescence, and the role of CAM in neutrophil recruitment was investigated using specific blocking antibodies. RESULTS: Exposure of HUVEC to CA-4-P, resulted in significant neutrophil recruitment, and increased expression of endothelial CAM. Results of antibody studies demonstrated that endothelial CAM expression induced by CA-4-P is responsible for the observed neutrophil recruitment. DISCUSSION: This study demonstrated that the tumour vascular targeting agent, CA-4-P, directly induces endothelial CAM expression and subsequent neutrophil recruitment. In vivo, neutrophil infiltration probably contributes to CA-4-P-induced tumour cell kill. Therefore, increasing neutrophil recruitment into tumours may have potential for optimising vascular-targeted strategies for cancer therapy.