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Biomedical subjects

H Tiedemann

Publications and source records attributed to H Tiedemann.

At least 19 recordsLinked to original sources

The vegetalizing factor. A member of the evolutionarily highly conserved activin family.

The mesoderm and endoderm inducing vegetalizing factor was partially sequenced after BrCN cleavage. A sequence which is highly conserved in activin A near the C-terminal end was identified. This shows that the factor belongs to the activin family. The activins are not confined to embryos and gonads, but widely distributed in other tissues like calf kidney and calf liver. Functional aspects are discussed.

Activins

Bone morphogenetic protein 4 (BMP-4), a member of the TGF-beta family, in early embryos of Xenopus laevis: analysis of mesoderm inducing activity.

We have screened a Xenopus ovary cDNA library using a synthetic oligonucleotide derived from that part of the inhibin beta A sequence, which is highly conserved within the TGF-beta family. Out of several clones yielding autoradiographic signals four turned out to represent Xenopus counterparts to the human bone morphogenetic protein 4 (BMP-4). Each two of the four sequences are nearly identical and probably account for different alleles whereas the two pairs showing 5% divergence may have arisen by genome duplication in this tetraploid species. The amino acid sequence of the Xenopus protein is 80% homologous to the human sequence showing no single exchange within the last 100 amino acids at the C-terminus. This region, which constitutes the main part of the mature, biologically active protein, also exhibits substantial homologies to other representatives of the TGF-beta family, especially to the Drosophila DPPC protein. Transfection of COS-1 cells with the Xenopus BMP-4 sequence under control of the CMV-promoter leads to the secretion of a protein which exhibits mesoderm inducing activity when tested with animal cap explants from Xenopus blastula stage embryos.

Amino Acid Sequence

Isolation of a vegetalizing inducing factor after extraction with acid ethanol. Concentration-dependent inducing capacity of the factor.

A vegetalizing factor has been isolated from chicken embryos by an improved method. The factor is extracted with acid/ethanol and finally purified by four consecutive steps of reversed phase HPLC. The molecular mass is about 25 kDa. The factor dissociates after reduction with dithiothreitol into two subunits of about 13 kDa. The factor was tested on Triturus alpestris by the implantation method, and on isolated ectoderm of Xenopus laevis in solution. The factor induces as the crude fractions all types of mesodermal tissues dependent on the concentration of the factor.

Acids

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

Activation of masked neural determinants in amphibian eggs and embryos and their release from the inducing tissue.

The neural-archencephalic (forehead) inducing activity of the microsomes and the high speed supernatant from Xenopus laevis ovaries and eggs was tested on gastrula ectoderm of Triturus alpestris. Both fractions have a very small inducing activity which increases considerably after autolysis or treatment with dissociating agents. A comparison with the inducing activity of the high speed supernatant from gastrulae suggests that neural inducing factors are synthesized in the ovary, stored in a masked form and activated, in part, during gastrulation. When the supernatant proteins from eggs and gastrulae were subjected to size-exclusion HPLC the neural inducing activity was eluted in different size classes, suggesting a limited proteolysis of a precursor during early embryogenesis. Further experiments have shown that treatment of the blastoporal lip from early Triturus gastrulae with actinomycin D or cycloheximide which inhibit the synthesis of RNA and protein respectively, diminishes the neural-archencephalic (forehead) inducing activity of the blastoporal lip. Treatment with actinomycin D or cycloheximide followed by ethanol, which, besides other structures, impairs the plasma membrane, does, however, not reduce the inducing activity. This may suggest that components which are needed for the transport of the neuralizing factor are diminished, when the synthesis of mRNA and protein is inhibited. Such components are probably needed for the release of the neuralizing factor from the blastoporal lip, the inducer tissue.

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

Affinity chromatography of embryonic inducing factors on heparin-Sepharose.

Mesoderm-inducing factors were extracted from chicken embryos and partially purified by chromatography on DEAE-cellulose. The DEAE-cellulose eluate was applied to heparin-Sepharose. Most of the proteins are not bound to heparin. The adsorbed proteins were eluted with a linear NaCl gradient. In totipotent gastrula ectoderm of amphibians the eluted proteins induce the differentiation of muscle and notochord as well as of large masses of renal tubules and blood cells. A possible relationship to fibroblast growth factors and angiogenesis factor is discussed.

Amphibians

Covalent coupling of neuralizing factors from Xenopus to Sepharose beads: no decrease of inducing activity.

Two neural inducing factors extracted from Xenopus gastrulae, a basic protein from ribonucleoprotein particles and an acidic protein from the high speed supernatant were covalently bound to CNBr-Sepharose or cross-linked CNBr-Sepharose particles. The protein-Sepharose complexes cannot be taken up by the competent ectoderm cells, but both factors remain fully active. The inducing activity is not due to a release of the bound factors. The experiments suggest that both neural inducing factors act on the cell surface of the competent ectoderm cells.

Animals

Embryonic induction and cation concentrations in amphibian embryos.

Explanted ectoderm from early gastrulae of Triturus alpestris was treated with the Na-K ionophore gramicidin (10(-9) to 10(-5) M) and the Ca-ionophore A 23187 (10(-7) to 10(-5) M). The ectoderm developed almost exclusively to atypical epidermis as in the control explants. When the ectoderm was treated with ouabain (10(-4) M), intracellular Na+ increased about 4.4-fold and K+ was reduced by half. Mesenchyme cells in small number differentiated in about 40% of the ouabain-treated explants. The time course of total Na+ and K+ ion concentrations was measured over a period of 72 h in ectoderm of T. alpestris after induction with vegetalizing factor and in control explants. In the first 15 h after explantation, no significant differences between control and induced explants were found. Thereafter, the steady state concentration of K+ decreased in the induced explants, whereas the steady-state concentration of Na+ slightly increased. The membrane resting potential recorded intracellularly of ectoderm sandwiches from early gastrula stages was found to be -41.3 mV in control and -59.3 mV in induced explants. From the specific conductances and permeabilities of non-induced and induced cells it is concluded that the induction process leads to a differentiation of the cell membrane, which acquires the characteristics of ionic selectivity. Ectoderm from Ambystoma mexicanum forms neural or neuroid tissue, mesenchyme and melanophores after explantation in salt solution in up to 50% of the explants without any additions. Isolated Ambystoma ectoderm is therefore not suitable for test experiments.

Ambystoma