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Biomedical subjects

S Werner

Publications and source records attributed to S Werner.

At least 199 records · Page 11Linked to original sources
PubMed1992

Two FGF receptor genes are differentially expressed in epithelial and mesenchymal tissues during limb formation and organogenesis in the mouse.

Fibroblast growth factors (FGFs) can influence the growth and differentiation of cultured cells derived from neuroectoderm, ectoderm or mesenchyme. The FGFs interact with a family of at least four closely related receptor tyrosine kinases that are products of individual genes. To investigate the role of FGFs in the growth and differentiation of embryonic tissues and to determine whether the individual FGF receptor genes might have specific functions, we compared the localization of mRNA for two FGF receptor genes, FGFR1 (the flg gene product) and FGFR2 (the bek gene product), during limb formation and organogenesis in mouse embryos (E9.5-E16.5). Although the two genes were coexpressed in some tissues, the differential expression of FGFR1 and FGFR2 in most embryonic tissues was striking. FGFR1 was expressed diffusely in mesenchyme of limb buds, somites and organ rudiments. In contrast, FGFR2 was expressed predominantly in the epithelial cells of embryonic skin and of developing organs. The differential expression of FGFR1 and FGFR2 in mesenchyme and epithelium respectively, suggests the receptor genes are independently regulated and that they mediate different functions of FGFs during development.

Animals↗
PubMed1992

Primary structure and in vitro expression of the N. crassa phosphoglycerate kinase.

The primary structure of 3-phosphoglycerate kinase (PGK) from Neurospora crassa was determined by sequencing a full-length cDNA. The deduced 418 amino acids protein shows a considerable identity to PGKs of other organisms with all the residues thought to be important for the function of the yeast enzyme conserved. The cloned PGK cDNA could be efficiently expressed in vitro resulting in a product with the expected molecular weight.

Amino Acid Sequence↗
PubMed1991

Primary structures of two subunits of NADH: ubiquinone reductase from Neurospora crassa concerned with NADH-oxidation. Relationship to a soluble NAD-reducing hydrogenase of Alcaligenes eutrophus.

The primary structures of the nuclear-encoded 51 kDa and 78 kDa subunits of the respiratory chain NADH: ubiquinone reductase (complex I) from Neurospora crassa mitochondria were determined by sequencing cDNA and the N-terminus of the mature proteins. Both subunits are related to the soluble NAD-reducing hydrogenase of the bacterium Alcaligenes eutrophus. Sequence comparison between these subunits, the corresponding subunits of the bovine complex I and the bacterial NAD-reducing hydrogenase further confirms the binding sites of NAD(H), FMN and three iron-sulfur clusters.

Alcaligenes↗
PubMed1991

External irradiation of growth hormone producing pituitary adenomas: prolactin as a marker of hypothalamic and pituitary effects.

Fifty-six patients with acromegaly were treated with external irradiation, 50 Gy, after unsuccessful pituitary surgery. A 50% reduction of pre-irradiation growth hormone levels was obtained in 51/56 patients. This level was reached after 26 +/- 14 months in 33 patients with prolactin levels less than 25 micrograms/l at diagnosis, after 21 +/- 17 months in 18 patients with prolactin greater than or equal to 25 micrograms/l, and after 20 +/- 21 months in 12 patients with prolactin greater than 40 micrograms/l at diagnosis. A further 50% decrease of growth hormone levels was obtained in 40/51 patients 42 +/- 22 months after radiotherapy, indicating that in clearly responsive patients, the growth hormone depression after radiotherapy follows a first order reaction. Four patients did not reach a 50% reduction of growth hormone levels 48-80 months after radiotherapy. During 10 years of follow-up, the growth hormone depression tended to be more pronounced in patients with mixed secretion of growth hormone and prolactin. The reduction of growth hormone levels was not correlated with the irradiated volume or the cumulative radiation effect. Within the first year, prolactin increased within the normal range in normoprolactinemic patients and remained so during follow-up. In hyperprolactinemic patients, prolactin decreased successively but to a lesser extent than growth hormone. Pituitary insufficiencies increased over time and three patients developed GH-insufficiency. Hypothalamic damage as indicated by prolactin changes was a regular phenomenon after radiotherapy.

Acromegaly↗
PubMed1991

Regulation of glucocorticoid receptor expression in cultured fibroblasts from a patient with familial glucocorticoid resistance.

The thermolabile glucocorticoid receptor (GR) in fibroblasts from a patient with familial glucocorticoid resistance (FGR) was characterized by solution hybridization, Northern blot analysis and Western immunoblotting using an hGR and cRNA probe and a GR specific monoclonal antibody. Specific DNA binding was measured by binding of cytosolic GR to mouse mammary tumour virus (MMTV) DNA. Northern blot analysis of total cellular RNA isolated from the fibroblasts showed hybridization of the hGR probe to 7.0 and 6.1 kb RNA species. Basal expression of hGR mRNA was 1.8 times higher in fibroblasts derived from the patient compared to control fibroblasts as assayed by solution hybridization. Even though nonsignificant, dexamethasone treatment maximally caused at 60% down-regulation of GR mRNA in normal fibroblasts after 12 h but only a 40% down-regulation in fibroblasts from the patient. In both cases, the initial mRNA values were restored after 72 h. No difference in GR mRNA stability was observed between fibroblasts from the patient and from controls. The induction of the glucocorticoid-regulated gene metallothionein IIA (MTIIA) by dexamethasone and cadmium sulphate was studied at different temperatures using a cRNA probe for human MTIIA. At elevated temperatures, cadmium sulphate but not dexamethasone increased MTIIA mRNA levels approximately three-fold in fibroblasts from the patient, whereas in normal fibroblasts regardless of temperature both cadmium sulphate and dexamethasone increased MTIIA mRNA levels approximately three- and two-fold, respectively. Cytosolic GR from FGR-fibroblasts showed an increased specific binding to MMTV DNA at 4 degrees C. These data support our previous findings of a thermolabile GR, probably due to a defect intrinsic to the GR protein, in this patient with primary cortisol resistance and indicate a compensatory mechanism at the transcriptional level of GR expression. The data also indicate a receptor defect affecting specific DNA binding in vitro.

Binding Sites↗
PubMed1991

Long-term effects of radiotherapy and bromocriptine treatment in patients with previous surgery for macroprolactinomas.

The long-term effect of radiotherapy and bromocriptine treatment was retrospectively evaluated in 25 patients who had previously undergone transsphenoidal surgery for treatment of macroprolactinomas. Surgery had reduced the median serum prolactin (PRL) value from 613 micrograms/l, a reduction of 53%. Postoperative bromocriptine was administered to 21 of the 25 patients. In 14 of these patients, serum PRL values became normal or almost normal with medication. There were no radiological or ophthalmological signs of progressive tumor growth during bromocriptine treatment. Fourteen patients received postoperative radiotherapy. After withdrawal of bromocriptine in 13 of these patients an average of 7 years after radiotherapy, the median serum PRL value had further decreased by 95%. The PRL reduction was similar for all doses applied, 38 to 52 Gy. After withdrawal of bromocriptine in 8 patients not receiving radiotherapy an average of 7 years after operation, the median serum PRL level had further decreased by 75%. At follow-up, 18 additional instances of pituitary insufficiency had developed in the group receiving radiotherapy, compared with 8 cases of insufficiency in the group not receiving radiotherapy. Thus, because bromocriptine has a long-standing effect on prolactin secretion, and radiotherapy is associated with a notably high incidence of pituitary insufficiency, we propose that photon irradiation should be considered mainly for patients who are not candidates for surgical or medical treatment.

Adult↗
PubMed1991

Stereotactic radiosurgery with the cobalt-60 gamma unit in the treatment of growth hormone-producing pituitary tumors.

Stereotactic radiosurgery on the pituitary given with the cobalt-60 gamma unit was used in the treatment of 21 patients with growth hormone (GH)-producing pituitary adenomas and acromegaly. All but one patient had locally invasive macroadenomas, and in the majority of cases, there was parasellar growth. Radiosurgery was the initial treatment for seven patients. Fourteen patients were previously treated by pituitary surgery, eight of whom had undergone conventional external pituitary irradiation as well. All patients had clinical signs of active acromegaly before radiosurgery. The radiation doses given to the previously untreated patients or those who only underwent surgery were 40 to 70 Gy in each of one to three irradiations. The patients with previous external irradiation received a lower dose of 30 to 50 Gy in each of one or two irradiations. The patients were observed during a period of 1 to 21 years from the first radiosurgical session. Two young patients had a clinical remission with a substantial decline of GH levels to near normal serum profiles. Another eight patients obtained reduction of GH levels and clinical activity. More than half of the patients (11 of 21) had minor or no effects from the treatment. There were no complications from the radiosurgery except the development of pituitary insufficiency in 2 of 13 patients who did not undergo previous conventional external irradiation. The remission rates were lower than the results previously reported by us for radiosurgery for Cushing's disease. This may be a result of the predominance in the present study of invasive macroadenomas and single treatments and to the lower radiation doses used in the patients who underwent conventional irradiation previously.(ABSTRACT TRUNCATED AT 250 WORDS)

Acromegaly↗
PubMed1991

Somatostatin levels in plasma in nonsmoking and smoking breast-feeding women.

The objective of the study was to record how somatostatin levels in plasma are altered in response to breast-feeding during the lactation period and to relate somatostatin levels to the success of the lactational performance and to smoking habits. Fifty-two women were investigated 4 days post partum and 3-4 months later. Blood samples were collected and the levels of somatostatin-like immunoreactivity (below referred to as SLI) were measured with radioimmunoassay. The periods of exclusive breast-feeding and of mixed feeding were assessed as well as the milk yield. Smoking habits were noted. SLI levels were found to be significantly lower on day 4 after delivery, compared to 3-4 months later. Also the type of response to breast-feeding was different. Thus, a significant fall of SLI was seen during breast-feeding at the maternity unit, but not 3-4 months later. Smoking women breast-fed fully for a significantly shorter time than nonsmokers and had significantly higher SLI levels at onset of breast-feeding day 4 post partum. Whether the high somatostatin levels recorded in connection with breast-feeding in smokers are related to the shorter period of breast-feeding seen in this group remains to be established. In addition, the highest levels of somatostatin were seen the day after the very last breast-feeding and a possible role for somatostatin in the weaning process should be explored.

Adolescent↗
PubMed1991

The human fibroblast growth factor receptor genes: a common structural arrangement underlies the mechanisms for generating receptor forms that differ in their third immunoglobulin domain.

To determine the mechanisms by which multiple forms of fibroblast growth factor (FGF) receptors are generated, we have mapped the arrangement of exons and introns in the human FGF receptor 1 (FGFR 1) gene (flg). We found three alternative exons encoding a portion of the third immunoglobulin (Ig)-like domain of the receptor. One of these alternatives encodes a sequence that is part of a secreted form of FGFR 1. The other two encode sequences that are likely part of transmembrane forms of FGFR 1. One of these forms has not been previously reported in published cDNAs. Also, we have determined the structural organization of a portion of the human FGFR 2 gene (bek) and found a similar arrangement of alternative exons for the third Ig-like domain. The arrangement of these genes suggests that there are conserved mechanisms governing the expression of secreted FGF receptors as well as the expression of at least two distinct membrane-spanning forms of the FGF receptors. The diverse forms appear to be generated by alternative splicing of mRNA and selective use of polyadenylation signals.

Amino Acid Sequence↗
PubMed1991

Serum levels and molecular sizes of growth hormone during pregnancy in relation to levels of lactogens, insulin-like growth factor I and insulin-like growth factor binding protein-1.

Growth hormone (GH), placental lactogen (PL), prolactin (PRL), insulin-like growth factor I (IGF-I) and IGF binding protein-1 (IGFBP-1) were determined in serum by radioimmunoassays (RIAs) in 12 women during pregnancy. GH and PL were analyzed by two monoclonal antibodies (Mab 3 and Mab 1) raised against pituitary GH. Serum IGFBP-1 had reached maximum levels at midpregnancy while PRL, PL and IGF-I increased continuously during pregnancy. Mab 1, which cross-reacts with PL, measured consistently higher levels of PL in serum than a commercial PL RIA (p less than 0.01) due to interference of cross-reacting serum proteins in the Mab 1 RIA. The GH-specific Mab 3 showed decreasing GH levels in unfractionated serum throughout gestation, but detected GH-immunoreactive proteins of approximately 40-200 kD after molecular sieve chromatography of pooled serum from late pregnancy. It is suggested that the formation of GH complexes of large molecular mass account for the successive disappearance of monomeric GH during pregnancy.

Adult↗
PubMed1991

Growth hormone specific stimulation of mitosis by size-fractionated serum from patients with growth hormone insufficiency: a study on the rat lymphoma cell line, Nb2.

The aim was to investigate the bioactivity of a high-molecular weight human growth hormone, identified following molecular sieve chromatography of serum. Nine patients with pituitary disease and GH insufficiency were studied. All patients had non-detectable levels of immunoreactive GH, less than 0.2 micrograms/l, in diurnal serum profiles. GH bioactivity was determined before and after size-fractionation of serum. The bioassay is based on the finding that a rat lymphoma cell line, Nb2, proliferates in the presence of lactogens. GH and PRL immunoreactivities were measured by radioimmunoassays. Pronounced GH immunoreactivity was found in fractions of sera from 7 out of the 9 patients and of 2 of 4 control sera, particularly in fractions corresponding to the elution volume of high-molecular weight proteins (greater than 160 kD). PRL immunoreactivity was only detected in fractions corresponding to the elution volume of monomeric PRL. Unfractionated serum had a dose-dependent mitogenic effect on the Nb2 cells. GH-antibodies could not inhibit this effect. Fractions of serum obtained from the patients stimulated Nb2 cell division as well. The mitogenic effect of serum fractions could be inhibited by GH-antibodies. Thus, high-molecular weight GH circulating in patients with GH insufficiency were shown to exert a GH-specific bioactivity in vitro after size-fractionation.

Animals↗
PubMed1991

Fibroblast growth factor 5 proto-oncogene is expressed in normal human fibroblasts and induced by serum growth factors.

Fibroblast growth factor 5 (FGF-5) is a member of the fibroblast growth factor family with transforming potential. It has been found to be expressed in several human tumor cell lines, but nothing is known about expression of this growth factor in normal cells and its biological functions. Here we show that the FGF-5 gene is expressed in exponentially growing normal human fibroblasts. In quiescent fibroblasts, expression of FGF-5 is strongly induced by serum and several growth factors such as platelet-derived growth factor (PDGF), epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alpha). This induction can be mediated by at least two different pathways involving protein kinase C or cAMP-dependent kinases. Since the effect is independent of de novo protein synthesis, FGF-5 represents the product of a primary response gene. In addition our data suggest that FGF-5 is mitogenic for human fibroblasts, indicating the existence of an FGF-5-mediated positive feedback in these cells which could amplify and prolong the cellular response to the initial stimulus.

Animals↗
PubMed1991

Low mitogenic response to EGF and TGF-alpha: a characteristic feature of cultured Kaposi's sarcoma derived cells.

We have investigated the mitogenicity of Epidermal Growth Factor (EGF) and Transforming Growth Factor alpha (TGF-alpha) for cultured Kaposi's sarcoma derived cells (KS cells). In contrast to control fibroblasts from the same patients, KS cells revealed only a weak mitogenic response to these growth factors. Neither EGF nor TGF-alpha were able to substitute for Platelet-derived growth factor (PDGF) when KS cells were grown in PDGF-depleted Platelet-Poor-Plasma serum (PPPS)-supplemented medium. The low mitogenicity of EGF and TGF-alpha for KS cells is not based upon a reduced expression of EGF receptor mRNA and protein in KS cells. However, the binding of EGF to KS cells is about 50% lower than that to fibroblasts. This reduced binding is not due to an occupation of the receptors by TGF-alpha since the expression level of this mitogen in different KS cell lines does not correlate with their capacity to bind EGF. In contrast, the low EGF binding seems to be an intrinsic feature of the EGF receptors of KS cells. In spite of the low mitogenicity of EGF for the tumor cells, the expression of c-myc, PDGF-A and Fibroblast growth factor 5 (FGF-5) mRNA is equally induced by purified EGF in KS cells and fibroblasts. This shows that at least the signal transduction pathways which lead to the expression of these genes are functional in KS cells.

Blotting, Northern↗
PubMed1991

[Enzyme immunoassay for the determination of albumin in urine].

Two modifications of a double antibody enzyme immunoassay for the determination of urinary albumin content are described. The method is simple, rapid and precise and can be carried out in test tubes and on microtiter plates as well. In 1:10 diluted urine samples albumin concentrations of 1.25 to 20 mg/l (corresponding to the normal range) can be determined. For a control sample with 0.3 mg/l albumin the intra- and interassay coefficients of variation were 4.9% (n = 11) and 10.4% (n = 21), respectively, on microtiter plates.

Albuminuria↗
PubMed1990

Primary structure and expression of a nuclear-coded subunit of complex I homologous to proteins specified by the chloroplast genome.

A 31-kDa subunit of complex I from Neurospora crassa, of nuclear origin, was cloned. The precursor polypeptide (33 kDa) could be efficiently expressed in an in vitro system for transcription and translation. The processing of the precursor to the mature protein was also obtained in vitro. An open reading frame coding for a precursor protein of 283 amino acids (32247 Da) was found by DNA sequencing. The predicted primary structure shows significant homology with proteins made in chloroplast. This supports the hypothesis that an enzyme similar to respiratory chain NADH dehydrogenase might exist in these organelles.

Amino Acid Sequence↗
PubMed1990

Molecular cloning of subunits of complex I from Neurospora crassa. Primary structure and in vitro expression of a 22-kDa polypeptide.

A lambda gt11 cDNA expression library was screened with antibodies directed against individual subunits of complex I from Neurospora crassa. Clones encoding cytoplasmically synthesized polypeptides with apparent molecular masses of 22, 29, 31, and 33 kDa were isolated. Northern blot analysis revealed that the corresponding genes are transcribed into mRNA species of about 0.85, 0.95, 1.3, and 1.4 kilobases, respectively. Further characterization of clones encoding the 22-kDa subunit was performed. A cDNA insert of 755 base pairs containing the complete coding sequence was used to express the polypeptide in vitro. A precursor of the protein is synthesized on cytoplasmic ribosomes without a cleavable signal sequence. Our data indicate that after import into the organelle and before assembly into complex I, the 22-kDa polypeptide forms intramolecular disulfide bridge(s). Nucleotide sequencing revealed an open reading frame coding for a protein of 183 amino acids. A molecular mass of 20,828 daltons was calculated. The polypeptide is hydrophilic and contains no obvious membrane-spanning domains. Eight cysteine residues arranged in a regular pattern are found in the primary structure of the protein. Therefore, this subunit is a good candidate to bind at least one of the iron-sulfur centers present in complex I of the respiratory chain.

Amino Acid Sequence↗
PubMed1990

The primary structure of the gamma-subunit of the ATPase from Synechocystis 6803.

The nucleotide sequence of the gene coding for the F0F1-ATPase gamma-subunit (atpC) from the transformable cyanobacterium Synchocystis 6083 has been determined. The deduced translation product consists of 314 amino acid residues and is highly homologous (72% identical residues) to the sequences of other cyanobacterial gamma-subunits. The Synechocystis 6803 sequence is also homologous to the chloroplast gamma-sequence. Like in the other cyanobacterial subunits, only the first of the 3 cysteine residues, which are involved in energy-linked functions of the gamma-subunit in spinach chloroplasts, is conserved in Synechocystis 6803.

Adenosine Triphosphatases↗