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Biomedical subjects

M Nielsen

Publications and source records attributed to M Nielsen.

At least 163 records · Page 9Linked to original sources
PubMed1992

[Determination of estrogen receptor status in breast cancer by immunocytochemical examination of fine-needle aspirates].

Estrogen receptor status was determined by immunocytochemical examination of fine-needle aspirates in 109 cases of primary invasive breast carcinoma. The results were compared with those from biochemical determination performed on tumor cytosols. Positive agreement was found between qualitative determination (positive/negative) by the two methods in 83% of the cases, and after semiquantitative stratification into three groups the results corresponded in 76%. The predictive value of a positive result on fine-needle aspirate was 96%, and 57% of a negative result. It is concluded that immunocytochemical determination of estrogen receptor status of breast carcinomas on fine-needle aspirate with the applied technique cannot replace the current standard biochemical method.

Aged↗
PubMed1992

Target size analysis of prostaglandin endoperoxide synthase. Radiation inactivation of both cyclooxygenase and peroxidase correlated with the monomer of 72 kDa.

To determine the size of the functional catalytic unit of prostaglandin endoperoxide (prostaglandin H) synthase, radiation inactivation experiments were performed. Both microsomes from ovine seminal vesicles and purified enzyme were irradiated with 10 MeV electrons. The enzymic activities of prostaglandin H synthase, cyclooxygenase and peroxidase, showed mono-exponential inactivation curves dependent on radiation dose, indicating molecular masses of approximately 72 kDa. The enzyme in microsomes, in its native environment, as well as in its purified state after solubilisation with nonionic detergent showed identical molecular masses. The results clearly demonstrate that the monomer of the enzyme with an apparent molecular mass of 72 kDa (SDS/PAGE) is the functional unit for catalysis of both activities. Hence the two active sites of cyclooxygenase and peroxidase reside on the same polypeptide chain.

Animals↗
PubMed1992

Identification, paracrine generation, and possible function of human breast carcinoma myofibroblasts in culture.

Myofibroblasts from human breast carcinomas were identified and experimentally generated in culture, and a possible function was examined. The frequency of alpha-smooth muscle actin immunoreactive cells was evaluated as a measure of myofibroblast differentiation in primary culture. Few or no alpha-smooth muscle actin-positive stromal cells (6.1 +/- 8.4%) were identified in primary cultures from normal breast tissue (n = 9). In contrast, high frequencies (68.8 +/- 15.1%) were observed in primary cultures from carcinomas (n = 19). The frequencies of myofibroblasts in primary cultures were almost identical to those obtained in the corresponding cryostat sections (69.1 vs. 68.8%). A possible precursor cell to the myofibroblast was looked for among typical fibroblasts and vascular smooth muscle cells. Purified blood vessels containing both fibroblasts and vascular smooth muscle cells were embedded in collagen gel and incubated with medium conditioned by breast epithelial cells. Fibroblasts rather than smooth muscle cells were recruited from the blood vessels. In medium conditioned by carcinoma cell lines or in co-cultures of carcinoma cell lines and purified fibroblasts, alpha-smooth muscle actin and the typical myofibroblast phenotype were induced in otherwise alpha-smooth muscle actin-negative fibroblasts. The effect of myofibroblasts on cellular movement--essential to neoplastic cells--was analyzed. Spontaneous motility of tumor cells (MCF-7) was entirely suppressed in a collagen gel assay. Under these conditions tumor cell motility was selectively mediated by direct cell-to-cell interaction between tumor cells and myofibroblasts. Under chemically defined conditions, interaction was dependent on the presence of plasminogen. Anti-plasminogen, soybean trypsin inhibitor, and anti-fibronectin partly neutralized the effect of plasminogen. It is concluded that elements of myofibroblast differentiation and function may be studied in culture.

Actins↗
PubMed1992

Characterisation of an allosteric modulatory protein associated with alpha-[3H]amino-3-hydroxy-5-methylisoxazolepropionate binding sites in chick telencephalon: effects of high-energy radiation and detergent solubilisation.

alpha-[3H]Amino-3-hydroxy-5-methylisoxazolepropionate ([3H]AMPA) binds to 1-day-old chick telencephalon membranes with KD and Bmax values of 138 nM and 2.56 pmol/mg of protein, respectively. High-energy radiation bombardment of intact frozen telencephalon resulted in a biphasic inactivation curve for [3H]AMPA binding. At a 5.8-Mrad radiation dose, the affinity of [3H]AMPA binding was increased (54 nM), but there was no apparent alteration in the Bmax value (2.76 pmol/mg of protein). We attribute this phenomenon to the inactivation of a high molecular weight modulatory protein that down-regulates the affinity of [3H]AMPA binding. The estimated molecular masses of the AMPA binding site and of the modulatory component were 59 and 108 kDa, respectively. Solubilisation with n-octyl-beta-glucopyranoside resulted in an increase in the Bmax (4.7 pmol/mg of protein) with no pronounced alteration in the affinity (109 nM) of [3H]AMPA binding. However, the solubilisation-induced increase in Bmax did not occur in telencephalon irradiated before solubilisation. In contrast, the increase in affinity induced by radiation treatment was still detected in solubilised extracts. These results suggest that the number and affinity of [3H]AMPA sites in chick telencephalon are closely regulated and that the modulatory systems involved are affected by both irradiation and solubilisation.

Allosteric Regulation↗
PubMed1991

Pharmacological and biochemical properties of the gamma-aminobutyric acid-benzodiazepine receptor protein from codfish brain.

The gamma-aminobutyric acidA (GABAA) receptor of codfish brain has been purified to homogeneity and contains a single polypeptide band of 56 kDa molecular mass. Polyacrylamide gel electrophoresis in sodium dodecyl sulfate (SDS-PAGE) of codfish GABA receptor photoaffinity-labeled by both [3H]flunitrazepam ([3H]Flu) and [3H]muscimol showed a single radioactive peak with molecular mass of 56 kDa, in contrast to the multiple subunits found in other vertebrate species. The codfish receptor, purified using benzodiazepine (BZ, Ro 7-1986/1) affinity chromatography, contains an apparent single band both by isoelectric focussing and on a silver-stained SDS gel. The receptor density and affinity constants for [3H]muscimol and [3H]Flu binding are comparable to those in mammalian brain, and the specific activity (greater than 1,000 pmol/mg of protein) is comparable to that of preparations purified from those sources. The pharmacological specificity of the codfish GABA-BZ receptor is generally similar to that of mammalian brain, including GABA-BZ coupling. The BZ binding exhibits homogeneous kinetic properties resembling those of the mammalian BZ2 receptor type, and shows strong GABA enhancement of [3H]Flu binding and weaker pentobarbital potentiation. This is consistent with other observations of an earlier phylogenetic, as well as ontogenetic, emergence in mammals of the BZ2 receptor subtype than the BZ1. Codfish GABA receptor is postulated to be a homo-oligomer in which the conformation of GABA and BZ recognition sites is very similar to that in the mammalian hetero-oligomeric GABAA receptor. The codfish receptor appears to be encoded by an ancestral gene and indicates an early development of BZ-GABA coupling.

Animals↗
PubMed1990

Inhibitory and regulatory binding sites on the rat brain serotonin transporter: molecular weight of the [3H]paroxetine and [3H]citalopram binding proteins.

Citalopram binds with high affinity to a specific binding site located on the serotonin (5-HT) transporter in 5-HT neurons. The binding affinity of [3H]citalopram was found to increase with increasing concentration of citalopram. This may be a homotropic positive allosteric effect, thus indicating the presence of an allosteric binding site for citalopram. The molecular weight of the proteins containing the high-affinity binding sites for citalopram and paroxetine, as well as the allosteric binding site for citalopram were determined by the irradiation method. The molecular weights of the three binding site proteins were found to be the same, suggesting that all three binding sites are located on the same protein molecule in the 5-HT transporter. The results support a hypothesis that the binding area for [3H]citalopram is located deeper in the transport channel than the [3H]paroxetine binding area. Thus the two high-affinity binding sites probably cover different, but overlapping, parts of the protein molecule. The allosteric binding site may be located elsewhere on the protein where it induces conformational changes of the 5-HT transporter with the result that high-affinity bound ligands get trapped in the transport channel, thereby explaining the increase in affinity.

Animals↗
PubMed1990

Target size of 5-HT3 receptors in N1E-115 neuroblastoma cells and rat brain.

Radiation inactivation was used to determine the molecular target size of the binding site for [3H]GR65630, a specific 5-HT3 receptor ligand, in two different neuronal tissues. Using a calibration curve of known molecular weight enzymes, the target sizes of [3H]GR65630 binding sites in N1E-115 neuroblastoma cells and rat brain were 98,600 +/- 11,300 and 49,100 +/- 8,500 Da, respectively. The results suggest 5-HT3 receptors may be present as dimers in N1E-115 neuroblastoma cells.

Animals↗