RESUMEN
The vitamin A metabolite, all-trans retinoic acid (atRA), is a regulator of nervous system development. Using a subtracted cDNA library constructed from neuroblastoma cells, the atRA-responsive gene calmin (Clmn) was identified (Merrill et al. [2004] Biol Chem 385:605-614). The Clmn transcript is detected very early in rat embryonic development and is sensitive to retinoid status. In vitamin A-deficient embryos, Clmn mRNA is dramatically down-regulated in the neuroepithelium adjacent to the somites, and this expression can be rescued with the addition of atRA. In embryonic day 18.5 embryos, CLMN is detected in regions where newly differentiated neurons are found, including the neural retina and the cortical plate; and in the adult brain, CLMN is most highly expressed in the neuron cell bodies of the hippocampus, cerebellum, and olfactory bulb. Thus, Clmn is sensitive to retinoid status during early gestational stages, and its expression is relegated to postmitotic neuronal cells in the adult rat brain.
Asunto(s)
Encéfalo/metabolismo , Proteínas de la Membrana/metabolismo , Tubo Neural/metabolismo , Neuronas/metabolismo , Tretinoina/metabolismo , Animales , Línea Celular Tumoral , Regulación hacia Abajo , Biblioteca de Genes , Immunoblotting , Inmunohistoquímica , Hibridación in Situ , Proteínas de la Membrana/genética , Ratones , Proteínas de Microfilamentos , ARN Mensajero/metabolismo , RatasRESUMEN
Mammalian eye development requires vitamin A (retinol, ROL). The role of vitamin A at specific times during eye development was studied in rat fetuses made vitamin A deficient (VAD) after embryonic day (E) 10.5 (late VAD). The optic fissure does not close in late VAD embryos, and severe folding and collapse of the retina is observed at E18.5. Pitx2, a gene required for normal optic fissure closure, is dramatically downregulated in the periocular mesenchyme in late VAD embryos, and dissolution of the basal lamina does not occur at the optic fissure margin. The addition of ROL to late VAD embryos by E12.5 restores Pitx2 expression, supports dissolution of the basal lamina, and prevents coloboma, whereas supplementation at E13.5 does not. Surprisingly, ROL given as late as E13.5 completely prevents folding of the retina despite the presence of an open fetal fissure, showing that coloboma and retinal folding represent distinct VAD-dependent defects. Retinal folding due to VAD is preceded by an overall reduction in the percentage of cyclin D1 positive cells in the developing retina, (initially resulting in retinal thinning), as well as a dramatic reduction in the cell adhesion-related molecules, N-cadherin and beta-catenin. Reduction of retinal cell number combined with a loss of the normal cell-cell adhesion proteins may contribute to the collapse and folding of the retina that occurs in late VAD fetuses.
Asunto(s)
Retina/citología , Retina/embriología , Vitamina A/metabolismo , Animales , Membrana Basal/efectos de los fármacos , Membrana Basal/patología , Cadherinas/metabolismo , Adhesión Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Coloboma/complicaciones , Coloboma/embriología , Coloboma/genética , Ciclina D1/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Embrión de Mamíferos/anomalías , Embrión de Mamíferos/efectos de los fármacos , Feto/anomalías , Feto/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Retina/anomalías , Retina/efectos de los fármacos , Células Ganglionares de la Retina/citología , Células Ganglionares de la Retina/efectos de los fármacos , Factores de Tiempo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Vitamina A/farmacología , Deficiencia de Vitamina A/embriología , Deficiencia de Vitamina A/genética , beta Catenina/metabolismo , Proteína del Homeodomínio PITX2RESUMEN
Vitamin A plays an essential role in vertebrate embryogenesis. In the present study, pregnant vitamin A-deficient (VAD) rats were maintained during early pregnancy on the short half-life vitamin A metabolite, all-trans retinoic acid (atRA), in an amount sufficient to support normal development to E10.5, with a higher level of atRA (250 microg atRA/g diet) provided from embryonic day (E) 8.5-10.5 to prevent mid-gestational resorption. When limiting amounts of atRA (1.5 or 12 microg/g diet) were provided after E10.5, a highly reproducible and penetrant state of late fetal vitamin A deficiency (late VAD) was induced in the organs of developing fetuses. In addition, late VAD fetuses displayed both anteriorization of cervical regions and novel posteriorization events at the thoracic and sacral levels of the skeleton, and showed sternal and pelvic malformations not previously observed in early VAD or genetic models. The expression of several Hox genes (Hoxd3 and Hoxb4) was altered in late VAD embryos, with a reduction in Hoxd3 noted as early as 1 day after instituting deficiency. All late VAD-induced malformations were prevented by the addition of retinol starting at E10.5, whereas provision of a high level of atRA throughout pregnancy improved but could not completely rescue the development of all organ systems. This work defines a nutritional model in which vitamin A deficiency can be induced during fetal development, and reveals new functions for the vitamin in the development of the axial and appendicular skeleton.
Asunto(s)
Fenómenos Fisiológicos Nutricionales de los Animales , Desarrollo Óseo/fisiología , Desarrollo Fetal/fisiología , Organogénesis/fisiología , Deficiencia de Vitamina A/embriología , Animales , Modelos Animales de Enfermedad , Gástrula/fisiología , Hibridación in Situ , Ratas , Ratas Sprague-Dawley , Tretinoina/uso terapéutico , Deficiencia de Vitamina A/tratamiento farmacológicoRESUMEN
The vitamin A metabolite, all-trans retinoic acid (atRA), plays an essential role in vertebrate embryogenesis, including development of the nervous system. In the human neuroblastoma cell line, SH-SY5Y, atRA rapidly induces (within 4 hr) the expression of the Crk-associated substrate (Cas) family member, neural precursor cell-expressed, developmentally down-regulated gene 9 (NEDD9) also called the human enhancer of filamentation (HEF1). NEDD9 is expressed in the developing hindbrain (5-somite stage) in the presumptive rhombomeres 2, 3, and 5 before the onset of overt segmentation. Exposure of rat embryos to excess atRA at times ranging from E9.25 to E12 leads to altered NEDD9 expression in the developing hindbrain within 6 hr. NEDD9 expression is also perturbed in vitamin A-deficient embryos. A putative retinoic acid response element in the 5' region of the NEDD9 promoter binds specifically to a RXR/RAR heterodimer and forms a higher molecular weight complex upon addition of a retinoic acid receptor-specific antibody. Regulation of NEDD9 may be an important means whereby atRA promotes cell spreading and neurite outgrowth in SH-SY5Y human neuroblastoma cells, and NEDD9 represents a new downstream target of atRA and its receptors in the developing hindbrain.