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Dye-coupling and the formation and fate of the hypoblast in the teleost fish embryo, Barbus conchonius.

The present report describes Lucifer Yellow (LY) transfer between the syncytial layer of the yolk cell (YSL) and blastodermal cells during epiboly in the teleost fish Barbus conchonius. The fate of a group of labeled cells is described until germ layer formation. At the onset of epiboly, LY seems to be transferred from the YSL to all blastodermal cells. Between 10% and 40% epiboly, dye-coupling appears to be restricted to the marginal region. Within 60 min individually labeled cells are distributed among unlabeled cells within the blastoderm. Between 40% and 60% epiboly, we observed a ring-shaped group of labeled cells, which probably have involuted during early gastrulation. Consequently, this cell group may correlate with the leading edge of the hypoblast layer within the germ ring. At 60% epiboly and later, the blastodermal cells are dye-uncoupled from the YSL. A gradual translocation of the ring-shaped hypoblast towards a dorsally located bar-like structure is observed between 50% and 100% epiboly. At 100% epiboly, fluorescent cells were located in contact with the YSL within the embryo proper, with the brightest fluorescence in the future head region. The translocation is due to dorsalwards convergent cell movements during the gastrulation process. The appearance of the hypoblast as a dye-coupled cell layer may correlate with some restriction in cell fate since the hypoblast differs in fate from the epiblast.

Animals↗

Vertebrate anteroposterior patterning: the Xenopus neurectoderm as a paradigm.

This review discusses formation of the vertebrate anteroposterior (AP) axis, focusing on the dorsal ectoderm, which gives rise to the nervous system, using the frog Xenopus as a model. After summarizing classical models of AP neural patterning, we describe recent molecular studies that are encouraging re-examination of these models. Such studies have shown that AP ectodermal patterning occurs by the onset of gastrulation, much earlier than previously thought. The identity of tissues that determine AP pattern is discussed, and the definition of the Organizer is reconsidered. The activity of factors secreted by inducing tissues in early patterning decisions is assessed and formulated into a revised model for Xenopus AP neural patterning. Finally, AP ectodermal patterning in Xenopus dorsal ectoderm is compared to that of other germ layers, and to other vertebrates.

Animals↗

Generation of embryoid bodies from mouse embryonic stem cells cultured on STO feeder cells.

Embryoid bodies, which are similar to post-implantation egg-cylinder stage embryos, provide a model for the study of embryo development and stem cell differentiation. We describe here a novel method for generating embryoid bodies from murine embryonic stem (ES) cells cultured on the STO feeder layer. The ES cells grew into compact aggregates in the first 3 days of coculture, then became simple embryoid bodies (EBs) possessing primitive endoderm on the outer layer. They finally turned into cystic embryoid bodies after being transferred to Petri dishes for 1-3 days. Evaluation of the EBs in terms of morphology and differentiating potential indicates that they were typical in structure and could generate cells derived from the three germ layers. The results show that embryoid bodies can form not only in suspension culture but also directly from ES cells cultured on the STO feeder layer.

Animals↗

Characterization of the unusually rapid cell cycles during rat gastrulation.

The onset of gastrulation in rodents is associated with the start of differentiation within the embryo proper and a dramatic increase in the rate of growth and proliferation. We have determined the duration of the cell cycle for mesodermal and ectodermal cells of rat embryos during gastrulation (days 8.5 to 9.5 of gestation) using a stathmokinetic analysis. These embryonic cells are the most rapidly dividing mammalian cells yet described. Most cells of the ectoderm and mesoderm had a cell cycle time of 7 to 7.5 hours, but the cells of the primitive streak divided every 3 to 3.5 hours. Total cell cycle time was reduced by shortening S and G2, as well as G1, in contrast to cells later in development, when cell cycle duration is modulated largely by varying the length of G1. In the ectoderm and mesoderm, G1 was 1.5 to 2 hours, S was 3.5 to 4 hours, and G2 was 30 to 40 minutes. G1, S and G2 were shortened even further in the cells of the primitive streak: G1 was less than 30 minutes, S was 2 to 2.75 hours, and G2 was less than 20 minutes. Thus, progress of cells through all phases of the cell cycle is extensively modified during rodent embryogenesis. Specifically, the increased growth rate during gastrulation is associated with radical changes in cell cycle structure and duration. Further, the commitment of cells to become mesoderm and endoderm by entering the primitive streak is associated with expression of a very short cell cycle during transit of the primitive streak, such that developmental decisions determining germ layer fate are reflected in differences in cell cycle regulation.

Animals↗

Somatic mosaicism for a DMD gene deletion.

Mosaicism is a mixed state, with two cell populations of different genetic origins caused by a cell mutation occurring after fertilization. In the present case, DNA analysis of lymphocytes led to a DMD diagnosis before death. Postmortem immunocytochemical and DNA analysis showed somatic mosaicism. At age 18 years, blood lymphocyte DNA analysis showed a DMD gene deletion, upstream from exon 7 to the 5' end containing both muscle and brain promoters. As the patient's mother and elder sister had no deletions, he was considered to have a new mutation. Immunocytochemical studies of postmortem tissues showed that dystrophin was absent from the tongue, deltoid, intercostal, psoas and rectus femoris muscles, but there was a mix of dystrophin-positive and negative fibers in the rectus abdominis, cardiac, temporalis and sternocleidomastoid muscles. All diaphragm cells were dystrophin positive. Polymerase chain reaction (PCR) amplification from all tissues except the temporalis and sternocleidomastoid muscles, diaphragm and kidney, in which no deletion was found, showed the deletion from at least exon 6 to the 5' end containing both muscle and brain promoters. In this case, a genomic deletion of the DMD gene contributed to the formation of tissues derived from both ectoderm and endoderm, and cells of mesodermal origin showed genotypic and phenotypic heterogeneity. Our results indicate a mutation of the present case may have occurred just before the period of germ layer formation.

Brain Chemistry↗

A digital image-based method for computational tissue fate mapping during early avian morphogenesis.

The early stages of vertebrate development, encompassing gastrulation, segmentation, and caudal axis formation, presumably involve large (finite) morphogenetic deformations; however, there are few quantitative biomechanical data available for describing such large-scale or tissue-level deformations in the embryo. In this study, we present a new method for automated computational "tissue fate mapping," by combining a recently developed high-resolution time-lapse digital microscopy system for whole-avian embryo imaging with particle image velocimetry (PIV), a well-established digital image correlation technique for measuring continuum deformations. Tissue fate mapping, as opposed to classical cell fate mapping or other cell tracking methods, is used to track the spatiotemporal trajectories of arbitrary (virtual) tissue material points in various layers of the embryo, which can then be used to calculate finite morphogenetic deformation or strain maps. To illustrate the method, we present representative tissue fate and strain mapping data for normal early-stage quail embryos. These data demonstrate, to our knowledge, for the first time, large tissue-level deformations that are shared between different germ layers in the embryo, suggesting a more global morphogenetic patterning mechanism than had been previously appreciated.

Animals↗

A morphological study of primordial germ cells at pregastrular stages in the chick embryo.

The spatial distribution, germ layer location, number and appearance of primordial germ cells (PGCs) are studied. Blastoderms from stage X (Eyal-Giladi and Kochav, 1976) to stage 2 (Hamburger and Hamilton, 1951) were serially sectioned and distribution patterns of PGCs were reconstructed. The presence of PGCs in clusters was noted at all stages examined. PGCs were found in relation to the epiblast, the primary hypoblast and in the blastocoel, during the whole period under examination. These observations show that PGCs can be directly studied at blastula stages. The results are consistent with observations on an early segregation of chick germ line that begins as early as stage X.

Animals↗

[General classification and limited nosology of the phacomatoses].

The authors suggest a new classification of phacomatoses based on the dysembryoplastic concept of these disorders, which are blastomas of the germ layers. Three main groups are described : ectoblastic, mesoblastic and entoblastic phacomatoses. The diffuse forms are often familial and very evolutive and are contrasted with the regional forms which are commonly sporadic and not very evolutive. The limits of the phacomatoses are finally specified.

Angiomatosis↗

Tetraploid embryonic stem cells contribute to the inner cell mass of mouse blastocysts.

The demonstration that mouse somatic cells can be reprogrammed following fusion with embryonic stem (ES) cells may provide an alternative to somatic cell nuclear transfer (therapeutic cloning) to generate autologous stem cells. In an attempt to produce cells with an increased pool of reprogramming factors, tetraploid ES cells were produced by polyethylene glycol mediated fusion of two ES cell lines transfected with plasmids carrying puromycin or neomycin resistance cassettes, respectively, followed by double antibiotic selection. Tetraploid ES cells retain properties characteristic of diploid ES cells, including the expression of pluripotent gene markers Oct4 and Rex1. On injection into the testis capsule of severe combined immunodeficient (SCID) mice, tetraploid ES cells are able to form teratomas containing cells representative of all three germ layers. Further, these cells demonstrated the ability to integrate into the inner cell mass of blastocysts. This study indicates that tetraploid ES cells are promising candidates as cytoplasm donors for reprogramming studies.

Animals↗

ERYTHROPOIESIS IN THE CHICK EMBRYO: THE ROLE OF ENDODERM.

The endoderm can be separated from the ectomesoderm of the area vasculosa of the early chick embryo. The development of the separated germ layers in organ cultures shows that the endoderm is necessary for the normal formation of blood islands by mesodermal cells. In the absence of endoderm the mesodermal cells undergo erythropoiesis in dispersed groups of cells but do not form endothelium.

Animals↗

The terminology of early development: history, concepts, and current usage.

The meaning of some terms used in describing the development of embryos is discussed in the light of their historical origin and current usage. Attention is focused on the terminology associated with the early development of animals, particularly that concerned with gastrulation and germ layer theory. It is suggested that terms linked with outdated concepts are not always appropriate for, and unless defined with care may limit, our appreciation of the significance of subsequent observation and experiment. Recommendations are made concerning the use of the terms mesoblast, mesoderm, and mesenchyme.

Animals↗

Gastrulation movements: the logic and the nuts and bolts.

Gastrulation, the period during the early development of animals when major cell and tissue movements remodel an initially unstructured group of cells, requires coordinated control of different types of cellular activities in different cell populations. A hierarchy of genetic control mechanisms, involving cell signaling and transcriptional regulation, sets up the embryonic axes and specify the territories of the future germ layers. Cells in these territories modulate their cytoskeleton and their adhesive behavior, resulting in shape changes and movement. Similarities among different species in patterning and cell biological mechanisms are beginning to allow us to recognize general, conserved principles and speculate on possible ancestral mechanisms of gastrulation.

Animals↗

Generation and characterization of rabbit embryonic stem cells.

We described the derivation of four stable pluripotent rabbit embryonic stem cell (ESC) lines, one (RF) from blastocysts fertilized in vivo and cultured in vitro and three (RP01, RP02, and RP03) from parthenogenetic blastocysts. These ESC lines have been cultivated for extended periods (RF >1 year, RP01 >8 months, RP02 >8 months, and RP03 >6 months) in vitro while maintaining expression of pluripotent ESC markers and a normal XY or XX karyotype. The ESCs from all lines expressed alkaline phosphatase, transcription factor Oct-4, stage-specific embryonic antigens (SSEA-1, SSEA-3, and SSEA-4), and the tumor-related antigens (TRA-1-60 and TRA-1-81). Similar to human and mouse ESCs, rabbit ESCs expressed pluripotency (Oct-4, Nanog, SOX2, and UTF-1) and signaling pathway genes (fibroblast growth factor, WNT, and transforming growth factor pathway). Morphologically, rabbit ESCs resembled primate ESCs, whereas their proliferation characteristics were more like those seen in mouse ESCs. Rabbit ESCs were induced to differentiate into many cell types in vitro and formed teratomas with derivatives of the three major germ layers in vivo when injected into severe combined immunodeficient mice. Our results showed that pluripotent, stable ESC lines could be derived from fertilized and parthenote-derived rabbit embryos.

Animals↗

Protein synthetic patterns of tissues in the early chick embryo.

Tissues dissected from early chick embryos were labelled in vitro with [35S]methionine, and their patterns of polypeptide synthesis investigated using the technique of two-dimensional (2-D) polyacrylamide gel electrophoresis. Apart from providing a preliminary description of the molecular changes associated with the processes of gastrulation and segmentation in the chick embryo, this study has revealed a number of polypeptides that may be useful as markers of cell type or function. The protein synthetic patterns of hypoblast from early and late gastrulae (stages 2 and 4, respectively: Hamburger & Hamilton, 1951) and of definitive endoblast and junctional endoblast from late gastrulae all resemble one another closely, but differ markedly from that of the epiblast at either stage. The lower layer tissues are characterized by the presence of eleven polypeptides that are largely absent from the epiblast. These findings are discussed with reference to current theories on the origins of the lower layer tissues. Comparisons between the 2-D patterns for tissues dissected from gastrulae and from embryos undergoing segmentation (stage 12) have revealed ten polypeptides showing stage-specific rather than tissue-specific expression. Apart from these ten polypeptides, the 2-D patterns for epiblast and ectoderm were practically identical, and distinguishable from those of other tissues by a lack of any unique polypeptides. On the other hand, stage-4 endoblast and stage-12 endoderm differed in the expression of many polypeptides. One polypeptide was found that may be considered as a marker of mesodermal cell type, as it was present in lateral plate, segmental plate and somitic mesoderm, but not in tissues of the other germ layers. Lateral plate could be distinguished from the other mesodermal tissues in the expression of a number of polypeptides, but the similarity in the 2-D patterns for segmental plate and somites suggest that the separation of somites from the anterior end of the segmental plate is not accompanied by the synthesis of new polypeptides.

Animals↗

The effect of ACTH and cortisone on the carbohydrate-protein complexes of different types of epithelium in relation to their embryological origin.

Using histochemical methods the authors studied the effect of ACTH and cortisone on carbohydrate-protein complexes of epithelial cells derived from 3 different germ layers. It has been shown that cortisone increased the intensity of PAS reaction in all types of the epithelium investigated. Changes in acid mucosubstances after treatment with cortisone were different. After the administration of ACTH the observed changes in epithelium were not parallel to the changes seen after treatment with cortisone.

Adrenocorticotropic Hormone↗

The effect of ACTH and cortisone on basement membrane in relation to the embryological origin of the epithelium.

The changes of intensity of PAS reaction in basement membranes of epithelial cells derived from 3 different germ layers in rats after treatment with ACTH and cortisone were observed. After treatment with cortisone an increase of PAS reaction in all investigated basement membranes was observed. Influence of cortisone on all basement membranes was independent of the embryological origin. The results after treatment with ACTH were not uniform.

Adrenocorticotropic Hormone↗

Intraocular growth and differentiation of tissue fragments isolated from primitive streak-stage mouse embryos.

Small tissue fragments were isolated from 5 different regions of mouse embryos at the primitive streak-stage and were grafted ectopically into the anterior eye chambers of syngeneic adult mice. Of the grafts, 22-66% became attached to the iris and were vascularised within 2-3 days. The attached grafts grew in size and formed loose aggregates of embryonic cells. During the fourth week in oculo, fully differentiated tissues representing derivatives of all three germ layers were found within the grafts. No differences in the composition of tissues derived were observed in grafts obtained from different regions of the embryo except that the endodermal grafts formed endodermal cysts only. The size of the original fragments did not affect their in oculo differentiation but smaller fragments had a lower chance of intraocular survival.

Animals↗