Hormonal regulation of estrogen receptor messenger ribonucleic acid in T47Dco and MCF-7 breast cancer cells

Using a combination of hormone-binding assays, immunologic techniques, and mRNA hybridizations we have measured the estrogen receptor (ER) content and studied the hormonal regulation of ER mRNA in one estrogen responsive and one estrogen unresponsive breast cancer cell line, MCF-7 and T47Dco, respectively. Estradiol binding could be detected in cytosol from MCF-7 cells but not in T47Dco cells. However, when measured by an enzyme-linked immunosorbent assay, T47Dco cells were found to contain approximately 15 fmol ER/mg cytosolic protein or 10% of the ER content in MCF-7 cells. Immunologically reactive ER in T47Dco cells was indistinguishable in size (approximately equal to 68 KD) from the ER in MCF-7 cells, as shown by Western blotting using a monoclonal antihuman ER antibody. Quantification of ER mRNA in MCF-7 and T47Dco cells indicated that T47Dco cells contained approximately 50% of the ER mRNA levels found in MCF-7 cells. This basal level of ER mRNA in T47Dco cells was not decreased by estradiol treatment, as opposed to in MCF-7 cells where estradiol caused 40-60% decrease in the ER mRNA expression. Also, estradiol did not increase the progesterone receptor (PR) mRNA levels in T47Dco cells whereas in MCF-7 cells an approximately 5-fold increase of the PR mRNA levels occurred after estradiol treatment. However, incubation of the cells with the synthetic progestin R5020 decreased the ER mRNA levels to approximately the same degree in both cell lines. In conclusion, we have shown that estrogen down-regulates ER mRNA and up-regulates PR mRNA in MCF-7 cells. Neither of these estrogenic effects were seen in T47Dco cells. It appears that the steroid-resistance in T47Dco cells does not occur as a consequence of a complete absence of ER mRNA or protein.(ABSTRACT TRUNCATED AT 250 WORDS)     

In Vivo Substitution of Choline for Sodium Evokes a Selective Osmoinsensitive Increase of Extracellular Taurine in the Rat Hippocampus

Recent investigations have demonstrated that taurine and phosphoethanolamine (PEA) are the amino acids most sensitive to microdialysis-perfusion with reduced concentrations of NaCl. The aim of the present work was to assess the importance of Na+ deficiency in evoking this response. Further, the previously described selectivity of replacement of Cl- with acetate with respect to amino acid release was reinvestigated. The hippocampus of urethane-anesthetized rats was dialyzed with Krebs-Ringer bicarbonate buffer, and amino acid concentrations of the perfusate were determined. Choline chloride was then stepwise substituted for NaCl, and, in some cases, mannitol (122 mM) was included in low sodium-containing media. In other experiments, NaCl was replaced with sodium acetate. The dialysate levels of taurine increased selectively in response to Na+ substitution. The elevation of taurine was linearly related to the increase in choline chloride, and maximal levels amounted to 335% of basal levels. The increase in extracellular taurine was not inhibited by perfusion with medium made hyperosmotic with mannitol. Replacement of Cl- with acetate stimulated the release of taurine to 652% of resting levels. In addition, PEA levels increased to 250% of control concentration. Other amino acids were unaffected by Cl- substitution. The results show that taurine transport is considerably more sensitive to Na+ depletion than glutamate transport, which also is known to be Na+ dependent. The taurine increase evoked by low Na+ is not caused by cellular swelling as it was unaffected by hyperosmolar medium. Finally, substitution of acetate for Cl- causes a specific elevation of extracellular taurine and PEA, possibly as a result of cytotoxic edema.     

Glial fibrillary acidic protein and desmin in salivary neoplasms. Expression of four different types of intermediate filament proteins within the same cell type

The presence of intermediate filament proteins (IFP) in normal salivary gland tissue and investigated by immunohistochemical techniques on frozen sections. Cytokeratins (CKs) were seen in almost all normal epithelial cells. In the parotid gland and in palatal gland tissue, a co-expression of cytokeratin and glial fibrillary acidic protein (GFAP) was seen in some myoepithelial cells, but this was not apparent in the submandibular gland. In some pleomorphic adenomas, carcinomas in pleomorphic adenomas, one mucoepidermoid carcinoma, one mucus-producing adenopapillary carcinoma and one adenoid cystic carcinoma, cells expressing three different IFP classes were found (CKs, vimentin, GFAP). These cells were most often situated peripherally in the tumour cords or ducts. The cytokeratin pattern in these cells, as revealed by mAbs PKK1-3, was similar to that in normal myoepithelial cells. Furthermore, reactivity for a fourth class of IFP, desmin, could be seen in this cell type in two carcinomas in pleomorphic adenomas, and also in a few cells in a pleomorphic adenoma and an adenoid cystic carcinoma. Thus the pattern of IFP expression in salivary gland neoplasms, is very complex, and cannot always be related to the normal tissue.     

Biogenesis of the somatogenic receptor in rat liver

Certain structural characteristics, in particular the type of oligosaccharide chains associated with the rat liver somatogenic (GH) receptors, were studied in different isolated organelles involved in receptor biosynthesis, maturation, and binding, with the use of ligand-affinity cross-linking, incubation with various oligosaccharide chain-cleaving enzymes, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In an endoplasmic reticulum-enriched fraction, a somatogenic receptor with Mr 33,000, after correction for bound ligand (assuming a 1:1 binding ratio of ligand to receptor) was found to contain N-linked high mannose oligosaccharide chain(s). In an intermediate density fraction, enriched in cis-Golgi, a major receptor of Mr 43,000 was found to contain N-linked complex type of oligosaccharide chains. In a low density membrane fraction, containing trans-Golgi complex membranes and endocytic vesicles, three receptors of Mr 95,000, 55,000, and 43,000 were found. These three receptors contain N-linked complex-type oligosaccharide chains. Neuraminidase treatment resulted in a decrease of the Mr 95,000 and 43,000 receptors to Mr 81,000 and 39,000, respectively. Two specific somatogenic receptors of Mr 95,000 and 43,000 containing N-linked complex type of oligosaccharides were found in an isolated plasma membrane-enriched fraction. When isolated hepatocytes were analyzed, the Mr 95,000 receptor was found to be the major labeled species. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis (first dimension nonreducing and the second dimension reducing conditions), showed that the Mr 43,000 receptor is contained within the Mr 95,000 receptor. The data suggest that the Mr 33,000 receptor found in endoplasmic reticulum constitutes a precursor to the Mr 43,000 receptor and that the Mr 43,000 receptor is complexed with an unknown subunit during transport through the Golgi complex to form an Mr 95,000 receptor present on the cell surface.     

Activation of rat liver microsomal glutathione S-transferase by hydrogen peroxide: role for protein-dimer formation.

The mechanism of oxygen radical-dependent activation of hepatic microsomal glutathione S-transferase by hydrogen peroxide was studied. Glutathione S-transferase activity in liver microsomes was increased 1.5-fold by incubation with 0.75 mM hydrogen peroxide at 37 degrees C for 10 min, and the increase in activity was reversed by incubation with dithiothreitol. Purified glutathione S-transferase was also activated by hydrogen peroxide after incubation at room temperature, and the increase in the activity was also reversed by dithiothreitol. Immunoblotting with anti-microsomal glutathione S-transferase antibodies after sodium dodecyl sulfate-polyacrylamide gel electrophoresis of hydrogen peroxide-treated microsomes or purified glutathione S-transferase revealed the presence of a glutathione S-transferase dimer. These results indicate that the hydrogen peroxide-dependent activation of the microsomal glutathione S-transferase is associated with the formation of a protein dimer.     

Armaments and ornaments: an evolutionary explanation of traits of dual Utility

Secondary sexual characters in many species function both in male-male competition and as cues for female choice. Based on a literature compilation of existing knowledge of traits with this dual function, we propose that they commonly arise through intersexual selection processes and serve as honest signals to other males regarding fighting ability or dominance. Faking these traits, here called armaments, (i.e. weapons and status badges) is difficult, as they are constantly put to trial in male-male contests. Females that subsequently utilize them as indicators of male phenotypic quality when selecting a partner will benefit by acquiring males of higher quality to father their offspring. Thus, evolution of armaments through male-male competition is seen as a usually initiating process, whereas female choice later may assume a role as an additional selective factor. The reverse, that males use information from traits evolved through female choice, is, however, also possible. The traditional view of independently evolved and temporarily unordered intra- and intersexual selection processes fails to explain dual trait functions. Moreover, our model may more satisfyingly than traditional ones explain how trait honesty and trait genetic variance are maintained: theoretical and empirical evidence suggests that such honesty and variation are more easily maintained under male-male competition than under female choice.