The organizer and neural induction in birds and mammals.

In avian and mammalian embryos the "organizer" property associated with neural induction of competent ectoderm into a neural plate and its subsequent patterning into rostro-caudal domains resides at the tip of the primitive streak before neurulation begins, and before a morphological Hensen's node is discernible. The same region and its later derivatives (like the notochord) also have the ability to "dorsalize" the adjacent mesoderm, for example by converting lateral plate mesoderm into paraxial (pre-somitic) mesoderm. Both neural induction and dorsalization of the mesoderm involve inhibition of BMP, and the former also requires other signals. This review surveys the key experiments done to elucidate the functions of the organizer and the mechanisms of neural induction in amniotes. We conclude that the mechanisms of neural induction in amniotes and anamniotes are likely to be largely the same; apparent differences are likely to be due to differences in experimental approaches dictated by embryo topology and other practical constraints. We also discuss the relationships between "neural induction" assessed by grafts of the organizer and normal neural plate development, as well as how neural induction relates to the generation of neuronal cells from embryonic and other stem cells in vitro.

Competence for neural crest induction is controlled by hydrostatic pressure through Yap.

Embryonic induction is a key mechanism in development that corresponds to an interaction between a signalling and a responding tissue, causing a change in the direction of differentiation by the responding tissue. Considerable progress has been achieved in identifying inductive signals, yet how tissues control their responsiveness to these signals, known as competence, remains poorly understood. While the role of molecular signals in competence has been studied, how tissue mechanics influence competence remains unexplored. Here we investigate the role of hydrostatic pressure in controlling competence in neural crest cells, an embryonic cell population. We show that neural crest competence decreases concomitantly with an increase in the hydrostatic pressure of the blastocoel, an embryonic cavity in contact with the prospective neural crest. By manipulating hydrostatic pressure in vivo, we show that this increase leads to the inhibition of Yap signalling and impairs Wnt activation in the responding tissue, which would be required for neural crest induction. We further show that hydrostatic pressure controls neural crest induction in amphibian and mouse embryos and in human cells, suggesting a conserved mechanism across vertebrates. Our work sets out how tissue mechanics can interplay with signalling pathways to regulate embryonic competence.

The organizer: What it meant, and still means, to developmental biology.

This article is about how the famous organizer experiment has been perceived since it was first published in 1924. The experiment involves the production of a secondary embryo under the influence of a graft of a dorsal lip from an amphibian gastrula to a host embryo. The early experiments of Spemann and his school gave rise to a view that the whole early amphibian embryo was "indifferent" in terms of determination, except for a special region called "the organizer". This was viewed mainly as an agent of neural induction, also having the ability to generate an anteroposterior body pattern. Early biochemical efforts to isolate a factor emitted by the organizer were not successful but culminated in the definition of "neuralizing (N)" and "mesodermalizing (M)" factors present in a wide variety of animal tissues. By the 1950s this view became crystallized as a "two gradient" model involving the N and M factors, which explained the anteroposterior patterning effect. In the 1970s, the phenomenon of mesoderm induction was characterized as a process occurring before the commencement of gastrulation. Reinvestigation of the organizer effect using lineage labels gave rise to a more precise definition of the sequence of events. Since the 1980s, modern research using the tools of molecular biology, combined with microsurgery, has explained most of the processes involved. The organizer graft should now be seen as an experiment which involves multiple interactions: dorsoventral polarization following fertilization, mesoderm induction, the dorsalizing signal responsible for neuralization and dorsoventral patterning of the mesoderm, and additional factors responsible for anteroposterior patterning.

Spemann-Mangold organizer and mesoderm induction.

The discovery of the Spemann-Mangold organizer strongly influenced subsequent research on embryonic induction, with research aiming to elucidate the molecular characteristics of organizer activity being currently underway. Herein, we review the history of research on embryonic induction, and describe how the mechanisms of induction phenomena and developmental processes have been investigated. Classical experiments investigating the differentiation capacity and inductive activity of various embryonic regions were conducted by many researchers, and important theories of region-specific induction and the concept for chain of induction were proposed. The transition from experimental embryology to developmental biology has enabled us to understand the mechanisms of embryonic induction at the molecular level. Consequently, many inducing substances and molecules such as transcriptional factors and peptide growth factors involved in the organizer formation were identified. One of peptide growth factors, activin, acts as a mesoderm- and endoderm-inducing substance. Activin induces several tissues and organs from the undifferentiated cell mass of amphibian embryos in a concentration-dependent manner. We review the extent to which we can control in vitro organogenesis from undifferentiated cells, and discuss the application to stem cell-based regenerative medicine based on insights gained from animal experiments, such as in amphibians.

Influence of the neural tube/notochord complex on MyoD expression and cellular proliferation in chicken embryos

Important advances have been made in understanding the genetic processes that control skeletal muscle formation. Studies conducted on quails detected a delay in the myogenic program of animals selected for high growth rates. These studies have led to the hypothesis that a delay in myogenesis would allow somitic cells to proliferate longer and consequently increase the number of embryonic myoblasts. To test this hypothesis, recently segmented somites and part of the unsegmented paraxial mesoderm were separated from the neural tube/notochord complex in HH12 chicken embryos. In situ hybridization and competitive RT-PCR revealed that MyoD transcripts, which are responsible for myoblast determination, were absent in somites separated from neural tube/notochord (1.06 and 0.06 10-3 attomol MyoD/1 attomol á-actin for control and separated somites, respectively; P<0.01). However, reapproximation of these structures allowed MyoD to be expressed in somites. Cellular proliferation was analyzed by immunohistochemical detection of incorporated BrdU, a thymidine analogue. A smaller but not significant (P = 0.27) number of proliferating cells was observed in somites that had been separated from neural tube/notochord (27 and 18 for control and separated somites, respectively). These results confirm the influence of the axial structures on MyoD activation but do not support the hypothesis that in the absence of MyoD transcripts the cellular proliferation would be maintained for a longer period of time

La transferencia de embriones. Una técnica de mejoramiento animal en ganado de lidia / Embryo transfer. A technique of animal improvement in fighting livestock

Dos explotaciones del estado de Tlaxcala (uno productor de ganado de lidia y otro productor de leche), se utilizaron para evaluar dos tipos de hormonas, la gonadotropina de suero de yegua gestante (PMSG) y la hormona folículo estimulante (FSH), en relación con la cantidad y calidad de embriones que se obtuvieron por superovulación en el programa de transferencia de embriones. Para la superovulación, se formaron dos grupos de cinco hembras de lidia; administrando al primero, FSH en dosis descendentes de 100 U.I. a 37.5 U.I. durante cuatro días consecutivos, y al segundo grupo con la administración de PMSG, en una sola dosis de 2,000 U.I. Se obtuvieron con FSH, 25 embriones transferibles, 20 degenerados o inmaduros y 15 ovocitos no fertilizados, sin embargo con PMSG, se obtuvieron cinco embriones transferibles, cinco degenerados o inmaduros y cinco ovocitos no fertilizados, lo que indica que el tratamiento con FSH, es más efectivo que el de PMSG para superovular a hembras de lidia con una probabilidad estadísticamente significativa (P < 0.05).Los 30 embriones que se obtuvieron para ser transferidos, fueron transportados a la explotación de leche, donde se llevó a cabo la transferencia de los mismos a 30 vaquillas respectivamente, mismas que fueron consideradas como receptoras. Del total de las transferencias, sólo cinco vaquillas se diagnosticaron como gestantes (16.6 por ciento), coincidiendo con embriones obtenidos de hembras superovuladas con FSH; cinco (16.6 por ciento) repitieron el estro a los 21 días de ciclo; otras cinco vaquillas (16.6 por ciento) repitieron el estro hasta los 37 días de ciclo; diez (33.3 por ciento) sólo presentaron un cuerpo lúteo en el ovario derecho y las cinco restantes (16.6 por ciento) además de presentar también un cuerpo lúteo en el ovario derecho, presentaban infección uterina. Esto se corroboró estadísticamente con una probabilidad estadística (P < 0.05).

Analysis of the mechanisms involved in neural induction

La determinación del área neural presuntiva en el embrión de vertebrado tiene lugar durante la gastrullación bajo la influencia inductora del cordomesoblasto en el curso de su invaginación. A pesar de los extensivos estudios destinados a esclarecer este complejo proceso, el problema no ha llegado a ser resuelto. En el presente artículo se consiera principalmente la competencia de la célula efectora y la naturaleza de la señal mediadora en este proceso de interación celular. Ambos puntos son discutidos en base a datos experimentales obtenidos principalmente de embriones de anfibios

Mechanisms underlying the organizer formation in Bufo arenarum embryos

El mesodermo prospectivo en la gástula temprana de Bufo arenarum fue previamente identificado ocmo una banda marginal de células grises. Para analizar su capacidad de diferenciación, explantos de estas células, fueron cultivados en el interior de vesículas ectodermales, en aislamiento y en combinación con componentes vegetativos. Cuando se cultivaron en aislamiento, fragmentos dorsales y ventrales de la zona marginal profunda se comportaron diferentemente. Mientras que los explantos ventrales produjeron células sanguíneas, los explantos dorsales fallaron en diferenciar permenecidendo como masas de células ricas en vitelo. Por otra parte, ambos cultivos fueron drásticamente modificados cuando se asociaron con células superficiales de la zona blastoporal, las que causaron los siguiente s efectos. a) Promoción de diferenciación en exploantos marginales dorsales, capaces ahora de generar estructuras notocordales y somáticas en adición a células mesenquimáticas. b) Promoción de dorsalización en explantos marginales ventrales, los cuales cambiaron su destino esperando desarrollando componentes axiales, similares a aquellos producidos por explantos dorso marginales "activados". Por el contrario, cultivos combinados de piezas animales y vegetativas fueron incapaces de generar estructuras mesodermales. Estos estudios sugieren que el mesodermo axial, identificado como el "organizador", se desarrolla a partir de un sustrato marginal de células mesodérmicas genuinas, mediante un estímulo inductivo dorsalizante originado en células periblastoporales superficiais

Agrobacterium rhizogenes vs auxinic induction for in vitro rhizogenesis of Prosopis chilensis and Nothofagus alpina

The induction and improvement of in vitro rhizogenesis of microshoots of Prosopis chilensis (Mol.) Stuntz and Nothofagus alpina (Poep. et Endl. Oerst.) were compared using Agrobacterium rhizogenes (Ar) versus indole-3-butyric acid (IBA) in the culture media. Microshoots of P. chilensis (1-2 cm length), coming from in vitro grown seedlings, were cultivated in a modified Broadleaved Tree Medium (BTMm) containing half salt concentration of macronutrients and 0.05 mg x L(-1) benzilaminopurine (BAP). After 30 days, microshoots with 2-4 leaves were selected and cultured in BTMm-agar in presence or abscense of Ar and in combination with IBA. For N. alpina, the apical shoots with the first 2 true leaves, from 5 weeks old seedlings, were cultured in the abovementioned medium, but with 0.15 mg x L(-1) of BAP. After 2 months, microshoots with 2-3 leaves were selected and cultured in BTMm-agar, supplemented with 5 mg x L(-1) IBA or in liquid BTMm on perlite and, in the presence or absence of A. rhizogenes (Ar) and in combination with 3 mg x L(-1) IBA. Rooting in P. chilensis reached 100.0% when Ar infection was produced in the presence of IBA, increasing both, the number and dry weight of roots. In N. alpina, 90.0% of rooting efficiency was obtained when Ar infection was produced in liquid culture and in the absence of auxin.

Aspectos médico-legales de la fecundación in vitro y transferencia embrionaria FIVET: primera parte / Medicolegal aspects on In-vitro fertilization and embryonic transference FIVET: first part

La fecundación in vitro o fecundación extrauterina con transferencia embrionaria (FIVET) fue una realidad en julio de 1978 cuando Edwards y Steptoe lograron en Inglaterra el nacimiento de una niña concebida fuera del claustro materno. Desde entonces no se han aclarado problemas legales como los derechos del embrión, el deber del Estado de protegerlo y el interés público en la prevención de niños genéticamente defectuosos. Se considera que solamente deben fecundarse aquellos óvulos que serán implantados, a fin de evitar el problema de embriones sobrantes. El procedimiento debe estar bajo control de un comité consultor de médicos, biólogos, moralistas, abogados, genetistas y autoridades del Ministerio de Salud. El médico practicante de la FIVET tiene responsabilidad penal y civil por su deber de diligencia en el examen de los componentes germinales y de los progenitores, así como en la custodia del embrión. El lavado del útero para remover un embrión que se trate de implantar en otra mujer debe constituir delito de aborto. Las donaciones de gametos deben limtar se por ley para prevenir el riesgo de relaciones incestuosas. La dignidad del ser humano como valor fundamental de la sociedad es enfatizado. PALABRAS CLAVES: Fecundación in vitro, transferencia embrionaria, genética y ley, medicina legal, derechos del embrión.

Transferencia embrionaria en día 3 versus día 5 bajo sistema de cultivo secuencial / Day 3 versus Day 5 Embryonic transference under sequential culture system

El presente tiene como objeto validar la hipótesis que el cultivo embrionario en día 5 y la transferencia en blastocisto es superior a la convencional en día 3. Se analizan los resultados obtnidos sobre un total de 39 ciclios con recuperación ovacitaria en los que se realizaron 36 transferencias, 22 en día 3 y 14 en día 5 en población no seleccionada y bajo las mismas condicions de cultivo. Ambsa muestras no presentan diferencias en su perfil poblaconal y parámetros de respusta a la estimulación

Los inositoles de la membrana celular y el efecto del Li+ sobre el desarrollo embrionario / The inositols of the cellular membrane and the effect of Li+ in the development of the embryo

El Li+ tiene la capacidad, en embriones sanos, de inducir la formación supernumeraria de estructuras cefálicas y de revertir las deficiencias del eje neural que se evidencian en embriones expuestos a la luz ultravioleta. Por otra parte, la hidrólisis de los fosfoinositodos de la membrana se ha implicado en los aumentos de Ca2 que proceden a los eventos que se suceden durante la fertilización y el desarrollo embrionario y el Li+ impide que los niveles de fosfoinositol (4,5), bifosfato retornen a sus niveles normales. La conclusión a la cual se debe llegar es que el Li+ altera la reespecificación dorsoanterior de los embriones y que esto debe depender de su capacidad de interferir con el ciclo de los fosfoinositidos. Estas observaciones tienen relevancia particular puesto que los efectos específicos del Li+ sobre el ciclo de los fosfoinositidos permiten suponer que alguno de los intermediarios de este ciclo debe jugar un papel importante en la diferenciación y especificación dorsoanterior del embrión. Es preocupante el hecho de que estos efectos pueden obtenerse utilizando concentraciones de Li+ similares a las usadas terapéuticamente en el manejo de las enfermedades maniacodepresivas