RESUMEN
In Caenorhabditis elegans, Wnt signaling regulates many asymmetric cell divisions. During embryogenesis, the C. elegans Dishevelled (Dsh) homolog, DSH-2, regulates asymmetric neuroblast division of the ABpl/rpppa blast cell. Dsh is a key intracellular component of both beta-catenin dependent and beta-catenin independent Wnt pathways. In C. elegans, most of the well-characterized asymmetric cell divisions regulated by Wnts are dependent on beta-catenin. In the ABpl/rpppa neuroblast division, however, we determined that DSH-2 regulates cell polarity through a beta-catenin independent Wnt pathway. We also established that the C. elegans Wnt homolog, cwn-1, functions to regulate asymmetric division of the ABpl/rpppa blast cell. Our results indicated that cwn-1 does not act alone in this process, and it functions with another redundant ligand that appears not to be a Wnt. Finally, we show widespread requirements for DSH-2 during embryogenesis in the generation of many other neurons. In particular, DSH-2 function is necessary for the correct production of the embryonic ventral cord motor neurons. This study demonstrates a role for DSH-2 and Wnt signaling in neuronal specification during C. elegans embryogenesis.
Asunto(s)
Proteínas de Caenorhabditis elegans/fisiología , Caenorhabditis elegans/fisiología , Proteínas de Ciclo Celular/fisiología , Neuronas Motoras/fisiología , Proteínas Wnt/fisiología , beta Catenina/fisiología , Animales , Animales Modificados Genéticamente , Tipificación del Cuerpo/fisiología , Caenorhabditis elegans/citología , Caenorhabditis elegans/embriología , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/genética , División Celular , Linaje de la Célula , Polaridad Celular/fisiología , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/fisiología , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/fisiología , Proteínas Dishevelled , Proteínas del Grupo de Alta Movilidad/genética , Proteínas del Grupo de Alta Movilidad/fisiología , Neuronas Motoras/citología , Neurogénesis/fisiología , Estructura Terciaria de Proteína , Transducción de Señal , Factores de Transcripción/genética , Factores de Transcripción/fisiología , Proteínas Wnt/genéticaRESUMEN
To examine the tumor microregional effects after gemcitabine administration to mice, we mapped the location of proliferating and hypoxic cells relative to vasculature in human colon cancer xenografts. The S-phase marker bromodeoxyuridine was used as a surrogate of drug effect and administered 2 hours before tumor excision, whereas vessel position and perfusion were assessed via staining for CD31 and intravenous injection of carbocyanine, respectively. Hypoxia was detected using pimonidazole. Images of the four markers were overlaid to reveal the spatial relationship between proliferation, vasculature, and hypoxia and to examine the microregional effects. Within 1 day after administration of 240 mg/kg of gemcitabine, proliferation throughout the tumor was completely inhibited. Over time, a reemergence of dividing cells occurred in relation to the distance from vasculature. Microregional analysis revealed that cells located distal to vasculature commenced cycling sooner than cells located proximal to vasculature. A similar trend was seen after multiple doses of gemcitabine (40 mg/kg on days 1, 4, 7, and 10). The possibility that the effect of gemcitabine could be attributed to changes in oxygenation was discounted after examining the vessel perfusion and patterns of hypoxia. The effect of gemcitabine was examined in multilayered cell culture, and at doses <30 micromol/L, a gradient in proliferation between the exposed and unexposed sides was observed. We show a differential effect on cell proliferation in relation to vasculature and conclude that cells distal to blood vessels are less affected by gemcitabine probably because of limited penetration.