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Biomedical subjects

H Tiedemann

Publications and source records attributed to H Tiedemann.

At least 37 records · Page 2Linked to original sources
PubMed1987

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↗
PubMed1986

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↗
PubMed1985

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↗
PubMed1985

An embryonic inducing factor: isolation by high performance liquid chromatography and chemical properties.

A protein (vegetalizing factor) which induces amphibian gastrula ectoderm to tissues which in normal development are derived from endoderm and mesoderm has been isolated from chicken embryo trunks by a combination of size exclusion and reversed phase HPLC. An amount of 0.5 ng factor per gastrula evokes inductions in 80-100% of the cases. The protein (apparent Mr approximately 13 000) is split by NaBH4 to polypeptide chains of about half the size under conditions in which disulfide bridges are reduced. The biological activity is lost.

Amino Acids↗
PubMed1984

Ribonucleoprotein particles from Xenopus eggs and embryos. Neural-archencephalic-inducing activity of the protein moiety.

Ribonucleoprotein particles were isolated from unfertilized eggs and gastrula stages of Xenopus laevis. The particles from both stages induce in gastrula ectoderm the formation of large foreheads (neural-archencephalic-inducing activity), whereas ribosomal subunits have no inducing activity. The inducing activity of particles from both stages is largely abolished after treatment with proteolytic enzymes and to some extent with ribonuclease. The protein moiety of gastrula ribonucleoprotein particles was extracted with phenol and the protein reduced with 2-mercaptoethanol. The protein induces foreheads, but at a lower rate than the intact particles. The protein was fractionated by high-performance liquid size-exclusion chromatography on a derivatized silica gel with 75% formic acid as eluent. The fraction which includes proteins from 10 000 to 16 000 Da has the highest neural-archencephalic-inducing activity.

Animals↗
PubMed1982

Radioiodination with 125I and reductive methylation with tritium of a vegetalizing inducer protein. Specific radio-activities and effect on biological activity.

An inducer protein, the vegetalizing factor, which causes amphibian ectoderm to differentiate into endoderm was labelled with carrier-free 125I (2000 Ci/m-Atom) to specific activities of 17-32 micronCi/microgram protein. The partial loss of the biological inducing activity is not caused by the iodination of the factor but is due to the oxidation and reduction steps in the iodination procedure. By reductive methylation with formaldehyde and [3H]NaBH4 under mild conditions the factor was labelled to specific activities of 0.8-2.3 micronCi/microgram protein depending on the specific activity of the [3H]NaBH4 (5-15 Ci/mmol). Under these conditions up to 25% of the amino groups of the protein were methylated without a reduction of the biological activity of the factor. The inducing activity of the protein was however considerably reduced, when under more stringent conditions 60-70% of the amino groups were methylated. It was shown by NMR spectroscopy that about one third of protein amino groups is dimethylated under these conditions. The factor could be labelled with N-(propionyloxy)succinimide without a loss of the biological activity, when very few amino groups were substituted. The experiments show that the factor can be labelled to high specific activity without loss of its inducing activity by reductive methylation, which is therefore the most suitable method.

Animals↗
PubMed1981

High performance gel permeation chromatography of proteins. Application to embryonic inducing factors.

Proteins in the molecular weight range of 10 000-170 000 were separated by high performance gel permeation chromatography. Silica particles with 30 nm or 50 nm pores were derivatized with glycidoxypropyltrimethoxysilane and used as support. The proteins were eluted with 50% formic acid. A protein fraction which induces endodermal and mesodermal tissues in amphibian gastrula ectoderm was purified by this method.

Animals↗
PubMed1981

A vegetalizing inducing factor. Isolation and chemical properties.

A vegetalizing factor which induces the formation of endodermal and mesodermal organs in amphibian gastrula ectoderm was purified from chicken embryos. Preparative sodium dodecyl sulfate polyacrylamide electrophoresis and gel permeation chromatography on sephadex with different eluants were employed. In buffer containing 6 M urea the molecular weight of the factor was estimated to about 28 000-30 000. In buffer containing sodium dodecyl sulfate (SDS) the factor partially dissociates to smaller polypeptide chains. Because an equilibrium between molecules of different size is established, SDS-containing buffers are not suitable for preparative purposes. In 50%-70% formic acid the factor completely dissociates into smaller peptide chains (Mr about 13 000-15 000). Furthermore, very little absorption of the factor to the gel matrices or glass surfaces is observed in formic acid. The final purification can be achieved by high-performance gel permeation chromatography with glycerolpropyl-treated silica gel as column packing and 50% formic acid as eluant.

Animals↗
PubMed1980

R-loop hybridization of collagen DNA: separation in Cs2SO4 gradients.

High molecular weight poly(A)RNA (26-35 S) from chicken embryo trunks which is enriched for collagen mRNA was iodinated and hybridized to DNA under conditions of R-loop formation. The R-loops were separated in Cs2SO4 gradients from the bulk of DNA yielding double stranded DNA enriched for collagen genes.

Animals↗
PubMed1979

Chicken ribosomal DNA: gene frequency and purification by R-loop hybridization.

The genes for ribosomal RNA of chicken are present in about 200--240 copies per haploid genome. The ribosomal genes were separated from the bulk of DNA by different methods. In CsCl-actinomycin D gradients the GC-rich ribosomal DNA is shifted to the lower density side of the bulk of DNA. After hybridization with iodinated ribosomal RNA under conditions where R-loops are formed ribosomal DNA has an increased density. The R-loops can be separated from the bulk of DNA in Cs2-SO4 gradients.

Animals↗
PubMed1978

A proteoglycan with affinity for the vegetalizing factor: characterization by density gradient centrifugation.

A crude proteoglycan fraction isolated from chicken embryos has an affinity for the vegetalizing factor, which induces mesodermal and endodermal tissues in gastrula ectoderm of Triturus alpestris. Binding of the factor to the crude proteoglycan results in inactivation of the vegetalizing factor. The crude proteoglycan was centrifuged in CsCl and CsCl-urea density gradients. Most of the inactivating material was recovered from the gradients in the high density proteoglycan fraction. Part of the inactivating material was found in glycoproteins of lower density. It is concluded that not all of the polysaccharide moiety of the proteoglycan is involved in binding of the vegetalizing factor. The proteoglycan is inactivated by incubation with hyaluronidase.

Animals↗
PubMed1977

Template active chromatin structures: degradation by deoxyribonuclease I.

Chicken embryos were pulse-labelled in vivo with [3H]uridine (10 min), the chromatin isolated and treated with DNAse I. The residual chromatin was separated from the degradation products by centrifugation. The nascent pulse-labelled RNA is completely recovered in the residual chromatin even after prolonged incubation with DNAase I, whereas the DNA is completely degraded to 80 base polynucleotide fragments and smaller fragments.

Animals↗