RESUMEN
We previously showed that the cell adhesion molecule Nectin-4 is overexpressed in ovarian cancer tumors, and its cleaved extracellular domain can be detected in the serum of ovarian cancer patients. The ADAM (adisintegrin and metalloproteinase) proteases are involved in ectodomain cleavage of transmembrane proteins, and ADAM17 is known to cleave Nectin-4 in breast cancer. However, the mechanism of Nectin-4 cleavage in ovarian cancer has not yet been determined. Analysis of ovarian cancer gene microarray data showed that higher expression of Nectin-4, ADAM10, and ADAM17 is associated with significantly decreased progression-free survival. We quantified Nectin-4 shedding from the surface of ovarian cancer cells after stimulation with lysophosphatidic acid. We report that ADAM17 and ADAM10 cleave Nectin-4 and release soluble Nectin-4 (sN4). Small molecule inhibitors and siRNA knockdown of both ADAM proteases confirmed these results. In matched samples from 11 high-grade serous ovarian cancer patients, we detected 2-20-fold more sN4 in ascites fluid than serum. Co-incubation of ovarian cancer cells with ascites fluid significantly increased sN4 shedding, which could be blocked using a dual inhibitor of ADAM10 and ADAM17. Furthermore, we detected RNA for Nectin-4, ADAM10, and ADAM17 in primary ovarian carcinoma tumors, secondary omental metastases, and ascites cells isolated from serous ovarian cancer patients. In a signaling pathway screen, lysophosphatidic acid increased phosphorylation of AKT, EGF receptor, ERK1/2, JNK1/2/3, and c-Jun. Understanding the function of Nectin-4 shedding in ovarian cancer progression is critical to facilitate its development as both a serum biomarker and a therapeutic target for ovarian cancer.
Asunto(s)
Proteína ADAM10/metabolismo , Proteína ADAM17/metabolismo , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Moléculas de Adhesión Celular/metabolismo , Proteínas de la Membrana/metabolismo , Neoplasias Ováricas/metabolismo , Proteína ADAM10/genética , Proteína ADAM17/genética , Secretasas de la Proteína Precursora del Amiloide/genética , Moléculas de Adhesión Celular/genética , Línea Celular Tumoral , Quinasas MAP Reguladas por Señal Extracelular/genética , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Humanos , Sistema de Señalización de MAP Quinasas/genética , Proteínas de la Membrana/genética , Metástasis de la Neoplasia , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Dominios Proteicos , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-jun/genética , Proteínas Proto-Oncogénicas c-jun/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Neoplásico/genética , ARN Neoplásico/metabolismoRESUMEN
Bone morphogenetic proteins (BMPs) are involved in embryonic mammary gland (MG) development and can be dysregulated in breast cancer. However, the role BMPs play in the postnatal MG remains virtually unknown. BMPs are potent morphogens that are involved in cell fate determination, proliferation, apoptosis and adult tissue homeostasis. Twisted gastrulation (TWSG1) is a secreted BMP binding protein that modulates BMP ligand availability in the extracellular space. Here we investigate the consequences of TWSG1 deletion on development of the postnatal MG. At puberty, Twsg1 is expressed in the myoepithelium and in a subset of body cells of the terminal end buds. In the mature duct, Twsg1 expression is primarily restricted to the myoepithelial layer. Global deletion of Twsg1 leads to a delay in ductal elongation, reduced secondary branching, enlarged terminal end buds, and occluded lumens. This is associated with an increase in luminal epithelial cell number and a decrease in apoptosis. In the MG, pSMAD1/5/8 level and the expression of BMP target genes are reduced, consistent with a decrease in BMP signaling. GATA-3, which is required for luminal identity, is reduced in Twsg1(-/-) MGs, which may explain why K14 positive cells, which are normally restricted to the myoepithelial layer, are found within the luminal compartment and shed into the lumen. In summary, regulation of BMP signaling by TWSG1 is required for normal ductal elongation, branching of the ductal tree, lumen formation, and myoepithelial compartmentalization in the postnatal MG.
Asunto(s)
Proteínas Morfogenéticas Óseas/metabolismo , Regulación del Desarrollo de la Expresión Génica/fisiología , Glándulas Mamarias Animales/crecimiento & desarrollo , Proteínas/metabolismo , Transducción de Señal/fisiología , Animales , Apoptosis/fisiología , Western Blotting , Bromodesoxiuridina , Línea Celular , Proliferación Celular , Epitelio/metabolismo , Femenino , Galactósidos , Regulación del Desarrollo de la Expresión Génica/genética , Técnicas de Sustitución del Gen , Hematoxilina , Inmunohistoquímica , Indoles , Glándulas Mamarias Animales/metabolismo , Glándulas Mamarias Animales/trasplante , Ratones , Ratones Noqueados , Proteínas/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Pediatricians and child health providers face with situation in which families disagree about recommended treatments. Managing these disagreements is more challenging during periods of divorce or separation particularly when parents are in dispute over custody or medical decision-making. Parental disagreement exists along a continuum. General principles apply 1) the worse the conflict, the worse the outcome for children, 2) when conflict is combined with other factors such as separation, the outcome is often worse, and 3) the pediatrician/primary health care provider can mitigate this. This manuscript provides a review of the subject and suggestions for the practicing provider.