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Biomedical subjects

G Heinrich

Publications and source records attributed to G Heinrich.

At least 55 records · Page 3Linked to original sources
PubMed1993

Cloning and characterization of the gene encoding murine insulin-like growth factor-binding protein-2, mIGFBP-2.

We present a characterization of the single-copy gene, mIGFBP-2, encoding the murine insulin-like growth factor-binding protein-2 (mIGFBP-2). It consists of four exons with sizes of 470 +/- 2, 227, 141 and > 475 nucleotides (nt). The first intron spans 23 kb of genomic sequence, and the complete gene extends to more than 28 kb. Two kb of the 5'-flanking region were sequenced. This region has no TATA or CAAT boxes but is G+G-rich and contains several potential regulatory sequence motifs. A total of five GC boxes, which may serve as potential binding sites for a transcription factor, Sp1, are present immediately upstream of the transcription start point (tsp). By primer extension, we identified a single tsp at nt position -85 +/- 2. The murine IGFBP-2 locus was mapped to the proximal region of mouse chromosome 1, to a region of conserved synteny with human chromosome 2q. A comparison of the deduced amino acid sequences of mouse, rat and human IGFBP-2 reveals a high degree of homology between all three species.

Animals↗
PubMed1993

[Prenatal diagnosis of type II Arnold Chiari malformation].

During a 7-year-period 22 foetuses with an Arnold-Chiari deformity were diagnosed. The reason for referring to our centre were severe additional malformations. In 2 cases protrusion of the fetal brainstem is documented by sonography prenatally. Prenatal diagnosis of an ACM has not been documented in the literature. The cytogenetic and microbiological findings including the additional malformations are presented. Pathogenesis, outcome and treatment options are discussed.

Abnormalities, Multiple↗
PubMed1992

Differential regulation of the nerve growth factor and brain-derived neurotrophic factor genes in L929 mouse fibroblasts.

Nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are structurally related survival and differentiation factors for distinct sets of peripheral and central neurons. The regulation of NGF gene expression has been extensively studied in L929 mouse fibroblasts. L929 cells also express the BDNF gene. Northern blot hybridization analysis revealed 4 discrete BDNF mRNA species in L929 cells and rat hippocampus after induction of seizures with kainic acid. Serum as well as 12-O-tetradecanoyl phorbol-13-acetate (TPA) stimulated NGF and all 4 BDNF mRNAs in L929 cells. Treatment with both agents induced NGF mRNA to a much larger extent than the BDNF mRNAs. The induction of the BDNF mRNAs was rapid, with nearly maximal levels by 1 hr. In contrast, NGF mRNA induction occurred later and peaked at 4-6 hr. Both NGF and BDNF mRNA induction were inhibited by actinomycin D. Cycloheximide, on the other hand, inhibited only NGF but not BDNF mRNA induction. Corticosterone rapidly decreased NGF mRNA but not the BDNF mRNAs, and had no effect on seizure-induced NGF or BDNF mRNAs. Forskolin did not stimulate NGF or BDNF mRNAs. In contrast to NGF mRNA, forskolin did not interfere with the serum induction of BDNF mRNAs. These results demonstrate that 2 genes which encode closely related neurotrophic factors are differentially regulated in L929 cells. The molecular mechanisms which bring about this differential regulation remain to be elucidated.

Animals↗
PubMed1992

Uptake and transfer factors of 137Cs by mushrooms.

The 137Cs content of 118 species (668 samples) of higher fungi collected in the period from August 1984 to October 1989 at three different locations in Styria, Austria, was determined by gamma-spectrometry. The Cs-content of most mushrooms has been increasing since September 1986. In order to find out which factors determine the 137Cs-contamination of mushrooms and the transfer-value soil to mushroom, the concentration of total and plant-available radiocesium in soils as well as the pH-value, the content of humus, clay, silt, sand, exchangeable cations, the composition of the clay minerals, and the particle size distribution of the soils of two different locations were examined. The higher the 137Cs contamination of the soil, the thicker the layer of humus and the higher the content of humus, the lower the pH-value, and the lower the amount of essential cations, especially of K+, the higher the amount of 137Cs plant-available will be. Therefore, the contamination of the mushrooms in the coniferous forest of Koralpenblick (1000 m) is higher than in the mixed forest at the Rosenberg around Graz at approx. 500 m height. Of 26 different species of mushrooms measured at both sites, only 61% show the highest TF-values soil to mushrooms also at the Koralpenblick. In the spruce forest at Koralpenblick there are many species of mushrooms with high 137Cs-contamination which were not found at the Rosenberg. However, the properties of the species to which a mushroom belongs are more important than environmental conditions and soil properties. The transfer values of 40K stay within narrow bounds, whereas those of 137Cs differ widely.

Accidents↗
PubMed1992

The human nerve growth factor gene: structure of the promoter region and expression in L929 fibroblasts.

We previously studied the transcriptional mechanisms involved in expression of the murine nerve growth factor (NGF) gene. To investigate the regulation of transcription of the human NGF gene, the promoter region was cloned. The nucleotide sequences of the human and mouse genes are greater than 90% similar near their promoters. The cloned human promoter was transcriptionally active in mouse L929 fibroblasts. 5' Deletion analyses indicated that the -85 to -45 region stimulates basal transcription 6-fold. This segment is greater than 80% identical in human and mouse genes except for an AP-1 consensus sequence found only in the human gene. A second AP-1 consensus sequence at +34, previously shown to function as a regulatory element in the mouse gene, is identical in both genes. Gel shift analyses of L929 cell extracts revealed binding of protein to oligonucleotide probes spanning each of the two AP-1 consensus sequences of the human gene. The gel shift patterns differed, suggesting interaction of different proteins with the two probes. Our results demonstrate that the human NGF gene promoter is transcriptionally active in mouse fibroblasts, and implicate an upstream region in basal transcription.

Animals↗
PubMed1992

Structure of the human insulin-like growth factor binding protein-2 gene.

Insulin-like growth factors (IGFs) are polypeptide hormones with structural homology to proinsulin. IGFs circulate in blood bound to specific IGF binding proteins (IGFBPs). cDNA sequences of six members of a family of human and rat IGFBPs have been published. Here we present a partial characterization of the human IGFBP-2 gene. This single copy gene is located on chromosome 2 and spans a total of more than 32 kilobases (kb) of genomic sequence. It is organized in four exons with sizes of more than 568, 220, 141, and 496 nucleotides. The intron between exon one and exon two contributes 27 kb to the size of the IGFBP-2 gene. The second and the third introns comprise 1.1 kb and 1.95 kb, respectively. When the structure of the IGFBP-2 gene is compared to that of the IGFBP-1 and IGFBP-3 genes, the exon boundaries are found to be conserved in these three genes. A single transcriptional start site was localized to 113 +/- 2 nucleotides 5' of the ATG start codon of IGFBP-2 translation. Furthermore, the region between nucleotides -635 and -2 upstream of the ATG was demonstrated to exhibit promoter activity in human Jurkat K16 cells. This region is devoid of TATA or CAAT consensus sequence motifs and has a high content of dC and dG nucleotides. In this respect the putative IGFBP-2 promoter region resembles the promoters which are often associated with housekeeping genes.

Amino Acid Sequence↗
PubMed1991

Insulin-like growth factor binding protein expression in human small cell lung cancer cell lines.

Insulin-like growth factor binding proteins (IGF-BP) are secreted by several human small cell lung cancer cell lines (SCLC). In order to identify the IGF-BPs from SCLC cell lines the RNA from 10 different SCLC cell lines was analyzed by Northern blot analysis with the probes for three different IGF-BPs, IGFBP-1, IGFBP-2, and IGFBP-3. No hybridization signal could be detected with the probes encoding for IGFBP-1 and IGFBP-3. The hybridization with different IGFBP-2-specific oligodeoxynucleotide probes and with the corresponding full-length cDNA showed that all SCLC cell lines which secreted IGF-BPs express IGFBP-2.

Base Sequence↗
PubMed1991

Structural and functional identification of regulatory regions and cis elements surrounding the nerve growth factor gene promoter.

The transcriptional mechanisms which contribute to the regulation of nerve growth factor (NGF) production are still largely unknown. We previously expressed the NGF promoter region in transgenic mice to localize cis regulatory elements to within 5 kb of the promoter. To further map these elements, and to begin to study the corresponding transacting factors, we here assayed the effects of 5' deletions and point mutations and examined the binding of nuclear factors to the NGF promoter region using L929 cell fibroblasts. Sequential deletions delineated regions upstream from the promoter which stimulated and inhibited transcription. DNAse-1 footprinting experiments identified four upstream segments, designated F2, F4, F6 and F8, which bound L929 cell nuclear proteins. F2 and F4 mapped to stimulatory and F6 and F8 to inhibitory regions. Competition experiments using a heptanucleotide present in both F2 and F4 segments suggested that they may be bound by related factors. Gel shift assays showed that the F8 binding proteins are less abundant in L929 cells than in NIH 3T3 fibroblasts and B16 melanoma cells. In addition to the upstream segments, a downstream AP-1 consensus sequence bound L929 nuclear proteins. Mutation of the AP-1 consensus sequence eliminated binding of nuclear proteins and reduced transcriptional activity. Our results indicate that transcriptional activator as well as suppressor regions surround the NGF gene promoter. The regulation of NGF production is likely to involve cis elements within these regions and transacting factors that bind to them.

Animals↗
PubMed1991

Multiple signalling pathways interact in the regulation of nerve growth factor production in L929 fibroblasts.

Fibroblasts are one of several cell types producing nerve growth factor (NGF) in neuronal targets. In previous studies we found that NGF production is up-regulated by 12-O-tetradecanoylphorbol 13-acetate (TPA) and serum, down-regulated by corticosterone, and unaffected by dibutyryl-cyclic AMP (db-cyclic AMP) in fibroblasts. As fibroblasts in vivo are likely to be exposed to regulatory effects by more than one of these agents at any given time, we examined the effects of combinations of them on NGF production using L929 fibroblasts as a model system. TPA and serum together stimulated NGF production 10-fold more than either agent alone. Corticosterone reduced NGF mRNA and NGF production to less than 10% of basal levels whether or not TPA or serum, or both, were present but not in the presence of the glucocorticoid antagonist RU486. Corticosterone did not increase the rate of NGF mRNA degradation. Forskolin and db-cyclic AMP prevented NGF mRNA induction by TPA and serum without changing basal levels. TPA induced c-fos and junB mRNAs transiently and preceding NGF mRNA induction but c-jun mRNA remained undetectable. Forskolin enhanced the induction of both junB and c-fos mRNA whereas corticosterone prolonged junB mRNA induction. Thus, TPA induction of NGF mRNA is modulated differentially by corticosterone and cyclic AMP. c-fos and junB may play a role in the underlying mechanisms.

Animals↗
PubMed1990

Induction of nerve growth factor gene expression by 12-O-tetradecanoyl phorbol 13-acetate.

Nerve injury leads to activation of fibroblasts, including stimulation of nerve growth factor (NGF) gene expression. Although interleukin-1 has been implicated as a mediator of NGF gene induction, the underlying mechanisms are not known. We investigated whether 12-O-tetradecanoyl phorbol 13-acetate (TPA), also a known stimulator of protein kinase C, regulates NGF gene expression. We show here that TPA stimulates NGF mRNA in mouse kidney and L929 fibroblasts but not in dispersed salivary cells. NGF mRNA stimulation in L929 cells is delayed by 2 h, is transient, and is followed by a parallel increase in NGF secretion. The induction of NGF mRNA is inhibited by cycloheximide, NGF mRNA levels decrease to similar values after 4 h of incubation with actinomycin D alone or in combination with TPA. These results indicate that the TPA response is cell specific and suggest that it is mediated at the transcriptional level via newly synthesized protein.

Animals↗
PubMed1990

Development of a high performance liquid chromatography method for the simultaneous measurement of prednisone and prednisolone.

An improved high performance liquid chromatographic assay (HPLC) is presented for the quantitative determination of prednisone and its main metabolite prednisolone. This HPLC assay, employing a UV-detector and dexamethasone as an internal standard, provides a selective and sensitive determination of glucocorticoids in plasma. The advantage of this method is the possibility of simultaneously measuring exogenic and endogenic glucocorticoids. The lower limit of quantification is 5 ng/ml.

Chromatography, High Pressure Liquid↗
PubMed1989

Characterization of a human T cell-specific chimeric antibody (CD7) with human constant and mouse variable regions.

A chimeric human-mouse anti-T lymphocyte mAb (CHT2; SDZ 214-380) has been constructed by cloning the variable region exons of both the L and H chains from the murine hybridoma RFT2 which have CD7 specificity and reactivity with a 40-kDa Ag. The variable L chain exon was joined to the human C kappa, and the variable H chain exon was joined to the human IgG1 region exon encoding the human allotype nGlm(z), nGlm(a). The gene constructs were introduced by electroporation into SP2/0, a non-Ig-producing murine myeloma. The identical tissue reactivity of the newly made CHT2 and the original murine RFT2 mAb (CD7) was confirmed by blocking experiments as well as by immunohistology and flow cytometry. Because this new mAb may have clinical use, the CD7 Ag expression of T lineage cells has also been quantitated in double and triple immunofluorescence assays in combinations with mAb to restricted forms of leukocyte common Ag that designate unprimed (CD45R+) and primed T lymphocyte populations (UCHL1+). CHT2 shows very strong reactivity with large thymic blast cells and cortical thymocytes from which T-ALL originates. Strong staining is seen on CD45R+ unprimed "virgin" T lymphocytes, whereas the expression on UCHL1+ primed "memory" cell types is weaker unless these cells are reactivated by mitogens or Ag. Thus CHT2 may spare a substantial population of resting memory T cells which is relevant to its potential therapeutic use. In addition the chimeric antibody had a greater in vitro antibody dependent cytotoxicity and a prolonged half-life (4.2 to 5.0 days) in Rhesus monkeys.

Amino Acid Sequence↗
PubMed1989

Cloning, sequence analysis and expression of a cDNA encoding a novel insulin-like growth factor binding protein (IGFBP-2).

Insulin-like growth factors bind with high affinity to specific binding proteins in extracellular fluids. To identify structural characteristics of IGF-binding proteins that might define their physiological roles, we determined the complete primary structure of a novel human IGF-binding protein (IGFBP-2) from a cloned cDNA. The cDNA encodes a 328 amino acid IGF-binding protein precursor which contains a 39-residue signal peptide. The mature 289 amino acid IGFBP-2 has a predicted Mr of 31,325. Chinese hamster ovary (CHO) cells stably transformed with the IGFBP-2 cDNA secreted a 36 kd protein which bound, with different affinities, IGFII and IGFI, but did not bind insulin. The predicted protein sequence of this IGF-binding protein shares extensive amino acid homology (greater than 85%) with the IGF-binding protein secreted by rat BRL-3A cells, but less than 40% homology with human IGFBP-1. Therefore IGFBP-2, and not IGFBP-1 as previously suggested, represents the human homologue of the rat BRL-BP (alpha IGFBP-2). Moreover, from alignment of the predicted protein sequences of IGFBP-1 and IGFBP-2, extensive conservation of the distribution of cysteine residues is observed. Although the overall amino acid homology shared by these proteins is not high, we suggest that they represent a family of structurally related human IGFBPs. Southern blot analysis of human DNA demonstrates that IGFBP-2 is encoded by a single-copy gene, different from that of IGFBP-1.

Amino Acid Sequence↗
PubMed1989

Cell-specific and developmental regulation of a nerve growth factor-human growth hormone fusion gene in transgenic mice.

We recently showed that a nerve growth factor-human growth hormone (NGF-hGH) fusion gene containing the promoter and 750 bp of 5' flanking region is transcriptionally active in the NGF-secreting L929 mouse fibroblast cell line. For the present experiments, we extended the 5' flank by 5 kb and constructed transgenic mice. These mice began to secrete hGH into saliva at puberty. hGH was detected immunocytochemically in the granulated convoluted tubular cells of the submandibular gland (SMG). SMG levels of hGH mRNA were 10-fold higher in adult males than in females. hGH mRNA was very abundant in SMG, moderately abundant in heart, brain, and kidney, rare in skin and adrenal gland, and undetectable in lung, liver, and spleen. Thus, the NGF-hGH gene reflects NGF gene expression. We conclude that basal NGF gene transcription is regulated by sequences in the cloned NGF gene fragment.

Animals↗
PubMed1989

A low molecular weight insulin-like growth factor binding protein from rat: cDNA cloning and tissue distribution of its messenger RNA.

Rat serum contains two major forms of insulin-like growth factor (IGF) binding proteins (BPs) that have apparent mol wts of about 35,000 and 150,000. We have isolated a cDNA clone encoding an IGF-BP whose N-terminal sequence is completely homologous to the NH2-terminal of the Buffalo rat liver cells-3A BP. The 270 amino acid mature protein has a predicted mol wt of 29,500. It contains a cysteine rich domain at each end of the molecule and an Arg-Gly-Asp (RGD) tripeptide motif near its C-terminus which suggests that this BP might associate with integrin cell surface receptors. The mature protein shares only partial homology with two published human IGF-BPs. Northern blot analysis shows that its mRNA is abundant in several fetal tissues, in adult brain, testes, ovaries, and kidney. Expression in the liver is high in fetal life but decreases to a barely detectable level in adulthood. However, upon hypophysectomy, the mRNA level increases at least 20-fold which suggests a hormonal regulation for the hepatic production of this small IGF-BP.

Amino Acid Sequence↗
PubMed1988

Nerve growth factor (NGF) is present in human placenta and semen, but undetectable in normal and Paget's disease blood: measurements with an anti-mouse-NGF enzyme immunoassay using a recombinant human NGF reference.

An enzyme immunoassay (EIA) originally developed for mouse beta-nerve growth factor (NGF) and commercially available was validated for human NGF. Cell culture medium containing bioactive recombinant human NGF was used as a reference, and mouse 2.5S beta-NGF as a standard. One of three human placentas contained measurable NGF (70 pg/g of tissue of mouse beta-NGF equivalents), a second detectable, and a third undetectable NGF. In three human semen samples NGF content ranged from 0.13 - 1.4 ng/ml. NGF could not be detected in normal human serum and in plasma from patients with Paget's disease, although mouse 2.5S beta-NGF added to human blood could be completely recovered from the serum.

Animals↗
PubMed1988

Structural and functional analysis of the promoter region of the nerve growth factor gene.

Molecular clones containing the NGF gene promoter regions and exons I were isolated from mouse and rat genomic libraries with synthetic oligonucleotide hybridization probes that corresponded to the 5' end of mouse submandibular gland NGF mRNA. The nucleotide sequences of the 5' flanking regions and exons I were determined and compared. There was 95% similarity in exons I and the adjacent promoter regions between mouse and rat sequences. Further upstream, the similarity decreased to 76%. Both mouse and rat promoter regions were only 33% similar to the presumptive human NGF gene promoter region. Upstream from the capsite of submandibular gland NGF mRNA as determined by an S1-nuclease protection assay, two promoter-like TATA-boxes were found at positions -28 and -49, resp., and two CAAT-like boxes at -379 and -546, resp. Both promoter regions contained a cluster of conserved CpG dinucleotide sequences in a GC-rich island whereas less conserved upstream regions contained only one CpG sequence. The promoter regions were fused to the human growth hormone gene reporter function. Transient expression in L929 cells yielded appropriate fusion mRNAs and secretion of hGH, demonstrating that the cloned promoters are functional.

Animals↗