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Biomedical subjects

H Grunz

Publications and source records attributed to H Grunz.

At least 37 records · Page 2Linked to original sources

Molecular mechanisms of tissue determination and pattern formation in amphibian embryos.

Factors of the TGF-beta superfamily (activin, vegetalizing factor) and the FGF family determine endoderm and mesoderm. The dorsoventral polarity of the mesoderm depends on additional factors (BMP-4, Wnt-8, noggin). Activin can directly activate gene transcription by signal transduction. Mesoderm is determined by factors prelocalized in the marginal zone. Its differentiation depends also on the animal ectoderm. Neural inducing factors have been partially purified. A masked neuralizing factor in the ectoderm is activated by induction of the ectoderm to the nervous system. Phorbolester can evoke neuralization signaling.

Amphibians↗

The role of vertical and planar signals during the early steps of neural induction.

The classical Einsteck-test (Spemann and Mangold, Roux Arch. Dev. Biol. 100: 599-638, 1924) and data from total exogastrulae (Holtfreter, 1933) suggest that vertical signals are transmitted between the chordamesoderm (organizer) and reacting ectoderm in the early phase of neural induction. In contrast to these results with Axoloti (urodeles), several authors observed the expression of neural specific genes in Xenopus exogastrulae, isolated dorsal blastopore lip with adjacent ectoderm (open-face explants) and Keller-sandwiches. Our data with Xenopus (anurans) also show that the expression of neural specific genes takes place in exogastrulae. However, when we prepared open face explants and exogastrula-like structures by microdissection at very early gastrula stage, the signal of a class II beta-tubulin, characteristic of terminal neural differentiation, is not found in the ectoderm. These results suggest that planar signals transmitted from the chordamesoderm into the ectodermal part can fairly be excluded under these experimental conditions. In similar experiments with Triturus alpestris we could not observe either the differentiation of neural structures in the ectodermal part of exogastrulae. These results confirm earlier experiments of Holtfreter performed with Ambystoma mexicanum (Axoltl) embryos. On the basis of the published data of different authors and our results, we cannot exclude the existence of planar signals for early and/or transient expressed genes before the onset of gastrulation in Xenopus, which make the neuroectoderm susceptible for the response to vertical signals during gastrulation. On the other hand our experiments with Triturus alpestris suggest that planar neural signals are unlikely in this species. These differences between Triturus and Xenopus embryos are discussed in the context of the peculiarities in morphological structure, competence and speed of development of the two species.

Ambystoma mexicanum↗

Partially purified factor from embryonic chick brain can provoke neuralization of Rana temporaria and Triturus alpestris but not Xenopus laevis early gastrula ectoderm.

A high neuralizing activity has been determined in forebrain of 7.5-day old chick embryos using Rana temporaria early gastrula ectoderm as reacting tissue (Mikhailov and Gorgolyuk, Soviet Scientific Reviews, Section of Physiology and General Biology, Vol. 1: 267-306, 1987). The corresponding protease-sensitive agent was extracted, partially purified by chromatography on DEAE-Toyopearl and Heparin-Ultragel columns, and its neuralizing activity was tested in vitro on ectoderm isolated from early gastrulae of R. temporaria, Triturus alpestris, and Xenopus laevis at different concentrations and for different periods of time (animal cap assay). Induction of neural structures was found in R. temporaria and T. alpestris explants (up to 100 and 60%, respectively), but not in cultures of X. laevis ectoderm. Under our experimental conditions, so-called "autoneuralization" of the ectoderm explants can safely be excluded. The results are discussed in relation to the neural competence of amphibian ectoderm and the mechanisms of neuralizing actions of different factors which might be involved in neural induction and patterning.

Animals↗

The four animal blastomeres of the eight-cell stage of Xenopus laevis are intrinsically capable of differentiating into dorsal mesodermal derivatives.

Mesoderm formation in the amphibian embryo is thought to be induced in the ectoderm of the animal region by signals emanating from the endoderm of the vegetal region after cleavage up to the mid-blastula. During this process the dorsal vegetal zone is thought to stimulate the dorsal animal zone to establish the Spemann organizer, which will in turn trigger the overlaying neuroectoderm during gastrulation resulting in the development of the central nervous system. In this concept it is assumed that the animal hemisphere is an uncommitted area, which receives its instructions from the vegetal region of the embryo. However, the experiments of this paper show that the 4 animal blastomeres of the eight-cell stage will form dorsal mesodermal structures in over 50% of the cases. The results support the view that developmental determinants are distributed in distinct gradients already in the early cleavage stages and that in the embryo the mesoderm is determined by factors prelocalized in the marginal zone. The spatial and temporal activation of certain genes in a distinct pattern is not simply emanating from certain areas or centers, but is a result of complex interactions between the vegetal and animal hemisphere and vice versa.

Animals↗

Suramin prevents transcription of dorsal marker genes in Xenopus laevis embryos, isolated dorsal blastopore lips and activin A induced animal caps.

Suramin, a polyanionic compound which is known to interact with the receptors of growth factors inhibits the expression of dorsal marker genes in whole embryos and isolated dorsal blastopore lips. Suramin also prevents activin A induced dorsalization of animal cap explants from blastula stage embryos, but it simultaneously evokes a shift of the differentiation pattern from dorsal mesodermal structures (notochord, somites) to ventral mesodermal derivatives (mesothelium and erythroid precursor cells). The results are consistent with the assumption that the dorsal vegetal zone (Nieuwkoop center) primarily releases more general/ventral mesodermalization signals. They further suggest a dual role of activin A in early embryogenesis. While the maternal component may contribute to a more general/ventral type of induction, increasing concentrations of the zygotic component along with the activation of primary response genes may contribute to the dorsalization of the organizer.

Activins↗

Suramin changes the fate of Spemann's organizer and prevents neural induction in Xenopus laevis.

Suramin, a polyanionic compound, which has previously shown to dissociate platelet derived growth factor (PDGF) from its receptor, prevents the differentiation of neural (brain) structures of recombinants of dorsal blastopore lip (Spemann's organizer) and competent neuroectoderm. Furthermore, the suramin treatment changes the prospective differentiation pattern of isolated blastopore lip. While untreated dorsal blastopore lip will differentiate into dorsal mesodermal structures (notochord and somites), suramin treated dorsal blastopore lip will form ventral mesoderm structures, especially heart structures. The results are discussed in the context of the current opinion about the mode of action of different growth factor superfamilies.

Animals↗

Localization of a nervous system-specific class II beta-tubulin gene in Xenopus laevis embryos by whole-mount in situ hybridization.

A neural-specific beta-tubulin mRNA is expressed in the developing central nervous system shown by whole-mount in situ hybridization experiments. Of special interest is the fact that from the late blastula (stage 9; Nieuwkoop and Faber, 1967; Hausen and Ribesell, 1991) until the early neurula (stage 13) the signal can be found not only in the presumptive neural plate but also in the presumptive epidermis. Later in development (from stage 13) the specific mRNA becomes restricted to the presumptive brain and spinal cord area. The results are discussed in the context of predisposition and (pre)determination.

Animals↗

Extracellular matrix components prevent neural differentiation of disaggregated Xenopus ectoderm cells.

Neuralization (archencephalic brain formation) takes place after dissociation and delayed reaggregation of animal caps of early gastrula without inducer (Grunz, H. and L. Tacke: Cell Differ. Dev. 28, 211-218 (1989)). This autoneuralization can be prevented by the cell supernatant from dissociated ectoderm of Xenopus laevis, which contains extracellular matrix components. After phenol extraction of the supernatant, the aqueous phase does no longer show inhibitory activity. It can be concluded from these results that glycoconjugates responsible for the prevention of neuralization represent glycoproteins or proteoglycans which are loosely attached to integral plasma membrane components. Single early gastrula ectoderm cells mixed with non-competent late gastrula ectoderm or endoderm, which primarily form common aggregates, do not differentiate into neural derivatives. In these reaggregates the ectoderm cells remain separated from each other by heterologous cells (non-competent ectoderm or endodermal cells) during the period of competence. These data indicate that the quick recovery of extracellular matrix components together with the restoration of the former organization of the plasma membrane is responsible for the prevention of neuralization.

Animals↗

Mesoderm induction and blood island formation by angiogenic growth factors and embryonic inducing factors.

Factors which induce mesoderm, including endothelium lined cavities and primitive blood cells in omnipotent amphibian ectoderm, have been isolated from different sources. Recently it was shown that angiogenic factors, which belong to the protein families of the heparin binding growth factors (acidic and basic fibroblast growth factor) and the transforming growth factors (TGF-beta 1 and -beta 2), also induce mesodermal tissues in amphibian ectoderm. In triturus ectoderm, capillary like endothelial networks are induced preferentially by the transforming growth factors. The relationship between growth factors and inducing factors is discussed.

Amphibians↗

Neural differentiation of Xenopus laevis ectoderm takes place after disaggregation and delayed reaggregation without inducer.

When Xenopus blastula or early gastrula ectoderm is disaggregated and cells are kept dispersed for up to 5 h prior to reaggregation, the resulting spheres will differentiate into large neural structures. In contrast, dissociated and immediately reaggregated ectoderm will only differentiate into ciliated epidermis (so-called 'atypical epidermis'). Ectoderm treated with mesoderm-inducing XTC-conditioned medium during the period of reaggregation immediately after disaggregation will only form one- or two-cell types (notochord and somites) only. Ectoderm treated with XTC-factor prior to disaggregation will differentiate into a large variety of cell types.

Animals↗

Accumulation and decay of DG42 gene products follow a gradient pattern during Xenopus embryogenesis.

The DG42 gene is expressed during a short window during embryogenesis of Xenopus laevis. The mRNA for this gene can be first detected just after midblastula, peaks at late gastrula, and decays by the end of neurulation. The sequence of the DG42 cDNA and genomic DNA predicts a 70,000-Da protein that is not related to any other known protein. Antibodies prepared against portions of the DG42 open reading frame that had been expressed in bacteria detected a 70,000-Da protein in the embryo with a temporal course of appearance and decay that follows that of the RNA by several hours. Localization of the mRNA in dissected embryos and immunohistochemical detection of the protein showed that DG42 expression moves as a wave or gradient through the embryo. The RNA is first detected in the animal region of the blastula, and by early gastrula is found everywhere except in the outer layer of the dorsal blastopore lip. By midgastrula DG42 protein is present in the inner ectodermal layer and the endoderm; it disappears from dorsal ectoderm as the neural plate is induced and later decays in a dorsoventral direction. The last remnants of DG42 protein are seen in ventral regions of the gut at the tailbud stage.

Animals↗

Induction of mesodermal tissues by acidic and basic heparin binding growth factors.

The inducing activity of two heparin binding growth factors HBGF-1 (prostate epithelial cell growth factor; acidic pI) and HBGF-2 (fibroblast growth factor; basic pI) from bovine brain has been tested on totipotent ectoderm from early amphibian (Xenopus laevis, Ambystoma mexicanum) embryos. Both factors induced, at high concentrations, mostly compact spheres surrounded by a non-epidermal epithelium. When the concentration or time of incubation was reduced, large muscle inductions frequently organized as somites were formed besides endothelial vesicles, mesenchyme and smaller areas of intestine-like epithelium. Further reduction of the concentrations or the time of incubation led to an increase in size and number of endothelium-lined vesicles and of mesenchyme, whereas the induction of muscle decreased. At still lower concentrations the overall rate of inductions decreased. The relationship of the growth factors to the vegetalizing factor from chicken embryos, dilution of which shows a similar shift in induced organs, is discussed. The present and previous experiments suggest that different mesodermal and endodermal tissues are induced by secondary interactions in which additional factors are involved. The induced organs derive from dorsal as well as from ventral mesoderm.

Animals↗

Close juxtaposition between inducing chordamesoderm and reacting neuroectoderm is a prerequisite for neural induction in Xenopus laevis.

The results of this study indicate that the induction of the central nervous system in Xenopus laevis depends on the close juxtaposition of inducing chordamesoderm and reacting ectoderm, which is necessary for the short distance migration of neural inducing factors. The examination of the neuroectoderm-chordamesoderm interface at intervals of 1 h up to 5 h showed that the onset of neural induction is correlated to the degree of contact formation between ectodermal and mesodermal cells. In the ectoderm cells the number of coated pits, a feature of receptor-mediated endocytosis, is increased. Furthermore there exist telophase bridges between some ectoderm cells, which are possibly correlated to secondary cell interactions.

Animals↗

Gene expression in the embryonic nervous system of Xenopus laevis.

Development of the nervous system in the amphibian embryo is initiated during gastrulation by an inductive interaction between chordamesoderm and dorsal ectoderm. The induced ectoderm forms the neural plate while uninduced ectoderm generates epidermis. We screened for genes activated during gastrulation and expressed specifically in the nervous system of Xenopus laevis in the expectation that clones representing such genes will constitute useful markers for the study of early neurogenesis. Probes were prepared from adult brain RNA by subtraction with RNA from ovary and from different combinations of adult kidney, muscle, and skin; cDNA libraries prepared from early to late neurula embryo RNA were screened with these probes. Six clones were chosen for further study. Three of these clones are not represented in the maternal RNA population but are activated at the late gastrula stage; the other three increase from a maternal base. Expression of five of the genes is restricted to the neural plate during embryogenesis, and all six are restricted to the central nervous system in premetamorphic tadpoles and adults. One of the clones encodes an apparently neurospecific isoform of beta-tubulin; the identity of the other clones is unknown. Expression of all six genes is suppressed in axis-deficient embryos that lack dorsal structures including the brain.

Animals↗