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PubMed2004

Signals derived from the underlying mesoderm are dispensable for zebrafish neural crest induction.

Signals from the non-neural ectoderm, the neural ectoderm, and the underlying mesoderm have all been implicated in the induction of neural crest. Bone morphogenetic protein (BMP) signaling in particular has an important role in this process; however, it is unclear whether this activity of BMP is due to its effects on patterning the underlying mesoderm, to its ability to establish a competent neural plate boundary zone, or to the direct specification of neural crest at intermediate levels of activity within a BMP gradient. We show neural crest induction occurs in zebrafish in the absence of involuted mesoderm, indicating that this tissue and signals derived from it are dispensable for the formation of neural crest. Dorsal-involuted mesoderm is a major source of secreted BMP antagonists, and the activity of BMP signaling is thought to depend on the presence of the opposing activity of these antagonists. We find that the three BMP antagonists known to be expressed during gastrulation in zebrafish, noggin1, follistatin, and chordin, are dispensable for neural crest induction. These results suggest that mechanisms for restricting the spatio-temporal pattern of BMP expression may compensate for the loss of secreted BMP antagonist activity in establishing dorso-ventral patterning, neural induction, and the neural crest.

Animals↗
PubMed1992

Ventral ectoderm of Xenopus forms neural tissue, including hindbrain, in response to activin.

The peptide growth factor Activin A has been shown to induce complete axial structures in explanted blastula animal caps. However, it is not understood how much this response to activin depends upon early signals that prepattern the ectoderm. We have therefore asked what tissues can be induced in blastula animal caps by activin in the absence of early dorsal signals. Using whole-mount in situ hybridization, we compare the expression of three neural markers, N-CAM, En-2 and Krox-20 in activin-treated ectoderm from control and ventralized embryos. In response to activin, both normal and ventralized animal caps frequently form neural tissue (and express N-CAM) and express the hindbrain marker Krox-20. However, the more anterior marker, En-2, is expressed in only a small fraction of normal animal caps and rarely in ventralized animal caps; the frequency of expression does not increase with higher doses of activin. In all cases En-2 and Krox-20 are expressed in coherent patches or stripes in the induced caps. Although mesoderm is induced in both control and ventralized animal caps, notochord is found in response to activin at moderate frequency in control caps, but rarely in ventralized animal caps. These results support the idea that in the absence of other signals, activin treatment elicits hindbrain but not notochord or anterior neural tissue; and thus, the anterior and dorsal extent of tissues formed in response to activin depends on a prior prepatterning or previous inductions.

Activins↗
PubMed1993

Anterior mesendoderm induces mouse Engrailed genes in explant cultures.

We have developed germ layer explant culture assays to study the role of mesoderm in anterior-posterior (A-P) patterning of the mouse neural plate. Using isolated explants of ectodermal tissue alone, we have demonstrated that the expression of Engrailed-1 (En-1) and En-2 genes in ectoderm is independent of mesoderm by the mid- to late streak stage, at least 12 hours before their onset of expression in the neural tube in vivo at the early somite stage. In recombination explants, anterior mesendoderm from headfold stage embryos induces the expression of En-1 and En-2 in pre- to early streak ectoderm and in posterior ectoderm from headfold stage embryos. In contrast, posterior mesendoderm from embryos of the same stage does not induce En genes in pre- to early streak ectoderm but is able to induce expression of a general neural marker, neurofilament 160 x 10(3) M(r). These results provide the first direct evidence for a role of mesendoderm in induction and regionalization of neural tissue in mouse.

Animals↗
PubMed1989

Mechanisms underlying the organizer formation in Bufo arenarum embryos.

In the early gastrula of Bufo arenarum the prospective mesoderm was previously identified as a marginal belt of grey cells. To analyze their differentiation capacity explants of these cells were cultured within ectodermal vesicles, in isolation and in combination with vegetal components. When cultured in isolation, dorsal and ventral fragments from the deep marginal zone behaved differently. Whilst ventral explants produced blood cells, dorsal explants failed to differentiate, remaining as masses of yolk-laden cells. On the other hand, both cultures were drastically modified when associated with superficial cells from the blastoporal zone, which caused the following effects: a) Promotion of differentiation in dorsal marginal explants, able now to produce notochordal and somitic structures, in addition to mesenchymatic cells. b) Promotion of dorsalization in ventral marginal explants, which changed their expected destiny developing axial components, similar to those furnished by "activated" dorso marginal explants. On the contrary, combined cultures of animal and vegetal pieces were unable to generate mesodermal structures. These studies suggest that the axial mesoderm, identified as the "organizer", develops from a marginal substrate of genuine mesodermal cells through a dorsalizing inductive stimulus originated in superficial periblastoporal cells.

Animals↗
PubMed1978

Biochemical investigation of lens induction in vitro. II. Demonstration of the induction substance.

An organ culture method to study the process of lens induction in the chick is described. To see whether or not a direct contact between the participating tissues is required for lens formation, agar slices (0.5 mm thickness) were cultured between the eye cup and the ectoderm. New lenses were formed in 43% of the transplants. Both the eye cup and the ectoderm originated from 72 hours old embryos (stage 18). By culturing agar slices on the eye cups during 2--24 hours and afterwards culturing these slices separately in combination with ectoderm, it was proved that lens inducing substance (s) penetrate into the agar slices. 3-4 hours of culturing on the eye cup is sufficient to obtain "inducing" agar slices. The same eye cups were shown to be able to induce a lens for more than one time. By using millipore filters with a 0.65 mu pore size, induction occurs in 36% of the cultures. The possibility of a restraining influence of the formed lens on the induction capacity of the eye cup was noticed.

Agar↗
PubMedDate not supplied

CD 2394, a novel synthetic retinoid, initiates an embryonic type of differentiation in hyperproliferative skin.

In human skin, there are 2 types of epidermal differentiation: normal differentiation, characterized by keratin 10 expression, and alternative differentiation. Alternative differentiation may be regeneration-associated differentiation (keratin 6 and 16) or re-induction of embryonic differentiation (expression of keratin 13, 15 and 19). The purpose of this study was to investigate the effect of the novel synthetic retinoid CD 2394 on hyperproliferative human skin, with respect to embryonic differentiation in particular. The effects of CD 2394 were compared with untreated and vehicle-treated skin 48 h after tape-stripping. In a multiparameter flow cytometric assay, parameters of proliferation, normal differentiation, embryonic differentiation and inflammation were assessed. With respect to proliferation, treatment with CD 2394 resulted in a decreased number of cells in the G2M-phase. Normal differentiation was decreased in CD 2394 treated skin. Furthermore, most of the CD 2394 treated samples showed expression of keratin 13, which was not seen in the otherwise treated skin. A correlation between keratin 10 and keratin 13 expression could not be demonstrated. This study showed that CD 2394 is capable of inducing an embryonic pathway of differentiation, which is distinct from normal differentiation or regeneration-associated differentiation.

Acne Vulgaris↗
PubMed2001

The avian organizer.

The development of avian embryos is characterized by the large amount of yolk present from the one-cell stage until late phases of organogenesis. In the chick, an axis of bilateral symmetry is established already before egg laying, when the egg rotates in the uterus. There is evidence for an active Wnt-catenin pathway in the vegetal cells in the periphery of the multi-cellular embryo. It overlaps with the posteriorly restricted expression of genes characterizing the vegetal hemisphere in amphibia. The zone of overlap bears several functional characteristics of a Nieuwkoop center, which is first apparent in the posterior marginal zone, but continues into the early primitive streak. Only the anterior part of the late streak is capable of direct neural induction, and only its tip, Hensen's node, can induce an anterior neural identity. This latter activity leaves the node together with the cells representing the anterior mesendoderm. Thus, although the constraints and dynamics of avian development make comparisons with the amphibian situation a complex undertaking, Hensen's node comes as close as possible to an organizer in Spemann and H. Mangold's definition.

Animals↗
PubMed2000

Evidence for dual mechanisms of mesoderm establishment in Xenopus embryos.

In Xenopus embryos, the maternally encoded transcription factor VegT (also known as Xombi, Antipodean, Brat, and Xtbx6) is essential for normal endoderm and mesoderm formation. This finding and the localization of VegT mRNA in the vegetal hemisphere of the oocyte are consistent with several models of germ layer patterning. Specific models have been proposed in which (1) combinations of cytoplasmic determinants, (2) inductive signals, or (3) intracellular concentration (morphogen effects) predominate. We test predictions of these models. We show that contrary to previous proposals, FGF does not suppress endoderm formation and so cannot be an anti-endoderm mesodermal determinant. We further show that, at the right dose, VegT can induce mesodermal marker expression cell autonomously and that it induces mesoderm at concentrations below those that induce endoderm. These results are consistent with a dual mechanism of mesoderm establishment in which both VegT-initiated inductive signals and an intracellular VegT morphogen gradient play a part.

Animals↗
PubMed1993

Inductive events in the patterning of the Xenopus laevis hatching and cement glands, two cell types which delimit head boundaries.

In Xenopus, the hatching and cement glands form at important boundaries in the head. We have examined induction of these glands in order to see the basis for this ectodermal patterning. Ectoderm can be induced to form hatching and cement glands by dorsal mesoderm and by the neural plate. Because the neural plate is sufficient to induce hatching and cement glands and lies adjacent to presumptive hatching and cement glands at the time of induction, it is the best candidate for the in vivo inducer of these tissues. The cement gland is restricted to the front of the head in part because the anterior but not posterior neural plate is capable of inducing it. The hatching gland is also restricted to the head, but can be induced by both anterior and posterior neural plates. Therefore, some factor suppresses hatching gland differentiation in the trunk. Transplanted neural plate pieces induced hatching gland cells in the ectoderm of embryos, suggesting that the inductive signal is planar. Treatment with retinoic acid or lithium at the start of gastrulation caused a loss of head structures. The presence of hatching gland in lithium-treated embryos suggests that these two agents have distinct effects and supports the idea that the induction of hatching and cement glands involves different pathways.

Animals↗
PubMed1999

Animal-vegetal asymmetries influence the earliest steps in retina fate commitment in Xenopus.

An individual retina descends from a restricted and invariant group of nine animal blastomeres at the 32-cell stage. We tested which molecular signaling pathways are responsible for the competence of animal blastomeres to contribute to the retina. Inactivation of activin/Vg1 or fibroblast growth factor (FGF) signaling by expression of dominant-negative receptors does not prevent an animal blastomere from contributing to the retina. However, increasing bone morphogenetic protein (BMP) signaling in the retina-producing blastomeres significantly reduces their contribution. Conversely, reducing BMP signaling by expression of a dominant-negative BMP receptor or Noggin allows other animal blastomeres to contribute to the retina. Thus, the initial step in the retinal lineage is regulated by position within the BMP/Noggin field of epidermal versus neural induction. Vegetal tier blastomeres, in contrast, cannot contribute to the retina even when given access to the appropriate position and signaling fields by transplantation to the dorsal animal pole. We tested whether expression of molecules within the mesoderm inducing (activin, FGF), mesoderm-modifying (Wnt), or neural-inducing (BMP, Noggin) pathways impart a retinal fate on vegetal cell descendants. None of these, several of which induce secondary head structures, caused vegetal cells to contribute to retina. This was true even if the injected blastomeres were transplanted to the dorsal animal pole. Two pathways that specifically induce head tissues also were investigated. The simultaneous blockade of Wnt and BMP signaling, which results in the formation of a complete secondary axis with head and eyes, did not cause the vegetal clone to give rise to retina. However, Cerberus, a secreted protein that also induces an ectopic head with eyes, redirected vegetal progeny into the retina. These experiments indicate that vegetal blastomere incompetence to express a retinal fate is not due to a lack of components of known signaling pathways, but relies on a specific pathway of head induction.

Activins↗
PubMed2000

Pharmacokinetic data support pharmacologically induced embryonic dysrhythmia as explanation to Fetal Hydantoin Syndrome in rats.

New studies suggest that the teratogenicity of phenytoin (PHT) is linked to its membrane-stabilizing pharmacological action via the rapid component of the delayed rectified potassium channel (lkr), resulting in embryonic cardiac dysrhythmia during a restricted sensitive period. In order to further elucidate this theory, PHT was administered to Sprague-Dawley rats on gestation day (GD) 11 with either a single dose of 150 or 100 mg/kg ip or 150 mg/kg po and developmental toxicity at term (GD 21) was studied. In satellite animals blood samples were withdrawn (0.5-24 h after dose) and total and free maternal plasma concentrations of PHT were measured. Pharmacokinetic data correlated well with pregnancy outcome data. At 150 mg/kg ip high concentrations of long duration (C(max) 240 microM and AUC 5300 microMhl(-1) - total) and marked developmental toxicity (embryonic death, decreased fetal weights, and orofacial clefts) were observed. After 100 mg/kg ip (C(max) 150 microM, AUC 2600 microMhl(-1) - total) only slight developmental toxicity (decreased fetal weights) was recorded and after 150 mg/kg po the plasma concentrations were even lower (C(max) 63 microM and AUC 1100 microMhl(-1) - total) and no adverse effects at all were observed. In separate experiments the effect of different concentrations of PHT on the embryonic heart was studied by adding PHT to GD 11 rat embryos cultured in vitro or by culturing GD 11 embryos from exposed dams. The decrease in heart rates was 3, 16, and 32% after culture with 50, 100, and 200 microM of PHT, respectively. After maternal administration of 150 mg/kg ip or po, the embryonic heart rate in vitro decreased by 25 and 7%, respectively, compared to controls. Altogether the results suggest that the development toxicity of PHT is caused by concentration-dependent induction of embryonic dysrhythmia and hypoxia related damage.

Abnormalities, Drug-Induced↗
PubMed1992

Intracellular signalling pathways involved in mesoderm induction by FGF.

We have examined the possible role of two signal transducing mechanisms, tyrosine phosphorylation and activation of protein kinase C (PKC), during fibroblast growth factor (FGF)-induced mesoderm induction in Xenopus. Tyrosine phosphorylation was examined through the use of a monoclonal anti-phosphotyrosine antibody. This antibody was shown to recognize the FGF receptor crosslinked to radioiodinated FGF. We also studied the response of Xenopus ectodermal explants to sodium orthovanadate, a compound that has been shown to elevate intracellular phosphotyrosine levels. Thirty percent of explants cultured in 100 microM vanadate were induced. In addition, vanadate synergized with FGF to give inductions that were more dorsal in nature than either vanadate or FGF alone. The role of PKC was evaluated by measuring PKC activity during mesoderm induction by FGF and by examining the effect of the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) on explants. TPA did not induce mesoderm, however, activation of PKC was detected in FGF-treated explants. Therefore, activation of the PKC pathway alone is not sufficient for mesoderm induction. Simultaneous treatment with TPA and FGF resulted in a significant inhibition of mesoderm induction by FGF, suggesting that activation of PKC could be part of a negative feedback mechanism. In contrast, TPA had no effect on induction by activin A.

Activins↗
PubMed2006

Sensitivity of expression of perivitelline membrane glycoprotein ZP1 mRNA in the liver of Japanese quail (Coturnix japonica) to estrogenic compounds.

Avian perivitelline membrane protein, ZP1, is synthesized and secreted by the liver with the stimulation of estrogens. In the present study, we measured the expression of ZP1 gene in the liver of immature male quail treated with various estrogenic compounds and in the liver of male quail embryos that were developed in the fertilized eggs laid by mother quail injected with various estrogenic compounds during vitellogenesis. Total RNA extracted from the liver was reverse-transcribed and cDNA was subjected to real-time PCR. Both diethylstilbestrol and ethinyl estradiol caused significant effect on the increase in mRNA in immature male quail. In contrast, diethylstilbestrol administered via the route of maternal injection was not effective for induction of embryonic mRNA, although the effect of ethinyl estradiol administered via the same route was prominent. These results showed that direct administration of estrogenic compounds, diethylstilbestrol and ethinyl estradiol, stimulates the induction of ZP1 gene, but the rate of accumulation of these compounds in the yolk is different during vitellogenesis. The present studies suggest that although ZP1 gene is a sensitive biomarker to evaluate the effects of endocrine disruptors, the route of administration is an important factor to compare the effectiveness.

Animals↗
PubMed2005

Gli function is essential for motor neuron induction in zebrafish.

The Gli family of zinc-finger transcription factors mediates Hedgehog (Hh) signaling in all vertebrates. However, their roles in ventral neural tube patterning, in particular motor neuron induction, appear to have diverged across species. For instance, cranial motor neurons are essentially lost in zebrafish detour (gli1(-)) mutants, whereas motor neuron development is unaffected in mouse single gli and some double gli knockouts. Interestingly, the expression of some Hh-regulated genes (ptc1, net1a, gli1) is mostly unaffected in the detour mutant hindbrain, suggesting that other Gli transcriptional activators may be involved. To better define the roles of the zebrafish gli genes in motor neuron induction and in Hh-regulated gene expression, we examined these processes in you-too (yot) mutants, which encode dominant repressor forms of Gli2 (Gli2(DR)), and following morpholino-mediated knockdown of gli1, gli2, and gli3 function. Motor neuron induction at all axial levels was reduced in yot (gli2(DR)) mutant embryos. In addition, Hh target gene expression at all axial levels except in rhombomere 4 was also reduced, suggesting an interference with the function of other Glis. Indeed, morpholino-mediated knockdown of Gli2(DR) protein in yot mutants led to a suppression of the defective motor neuron phenotype. However, gli2 knockdown in wild-type embryos generated no discernable motor neuron phenotype, while gli3 knockdown reduced motor neuron induction in the hindbrain and spinal cord. Significantly, gli2 or gli3 knockdown in detour (gli1(-)) mutants revealed roles for Gli2 and Gli3 activator functions in ptc1 expression and spinal motor neuron induction. Similarly, gli1 or gli3 knockdown in yot (gli2(DR)) mutants resulted in severe or complete loss of motor neurons, and of ptc1 and net1a expression, in the hindbrain and spinal cord. In addition, gli1 expression was greatly reduced in yot mutants following gli3, but not gli1, knockdown, suggesting that Gli3 activator function is specifically required for gli1 expression. These observations demonstrate that Gli activator function (encoded by gli1, gli2, and gli3) is essential for motor neuron induction and Hh-regulated gene expression in zebrafish.

Animals↗