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Biomedical subjects

E B Thompson

Publications and source records attributed to E B Thompson.

At least 145 records · Page 8Linked to original sources
PubMed1981

Dexamethasone 21-mesylate: an affinity label of glucocorticoid receptors from rat hepatoma tissue culture cells.

We recently described the biological properties of an alpha-keto mesylate derivative of cortisol, cortisol-Mes. Cortisol-Mes exhibited long-term antiglucocorticoid activity, but there was no firm evidence that this activity was irreversible or receptor-mediated. Here we report that dexamethasone mesylate (Dex-Mes), which is the alpha-keto mesylate derivative of the more active glucocorticoid dexamethasone, is a candidate for a steroid-specific affinity label of glucocorticoid receptors. Dex-Mes is relatively stable, like cortisol-Mes, but possesses greater whole-cell antiglucocorticoid activity. However, Dex-Mes also possesses partial agonist activity, which is expressed at somewhat higher concentrations of Dex-Mes than the antagonist activity. Dex-Mes is more efficient than cortisol-Mes in competing for dexamethasone binding to glucocorticoid receptors. Furthermore, Dex-Mes is effective at lower concentrations than cortisol-Mes in causing long-term apparently irreversible antiglucocorticoid effects in whole and broken cells. The cell-free effect of Dex-Mes is specifically prevented by coincubation with an excess of cortisol. These facts argue that the apparently irreversible effects of Dex-Mes are steroid mediated. [3H]Dex-Mes has been used to identify a glucocorticoid-specific, covalently labeled fraction on sodium dodecyl sulfate/polyacrylamide gels with a molecular weight of approximately 85,000. Thus Dex-Mes appears to have been established as an affinity label for glucocorticoid receptors.

Affinity Labels↗
PubMed1981

Isolation and characterization of dexamethasone-resistant mutants from human lymphoid cell line CEM-C7.

Fifty-four independent dexamethasone-resistant clones were isolated from the clonal, glucocorticoid-sensitive human leukemic T-cell line CEM-C7. Resistance to 1 microM dexamethasone was acquired spontaneously at a rate of 2.6 X 10(-5) per cell per generation as determined by fluctuation analysis. After mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), the phenotypic expression time for dexamethasone resistance was determined to be 3 days. Spontaneous acquisition of resistance to 0.1 mM 6-thioguanine appeared to occur at a much slower rate, 1.6 X 10(-6) per cell per generation. However, the expression time after MNNG mutagenesis for this resistant phenotype was greater than 11 days, suggesting that the different rates of acquisition for the two phenotypes measured by fluctuation analysis were the results of the disparate expression times. The mutagens ICR 191 and MNNG were effective in increasing the dexamethasone-resistant fraction of cells in mutagenized cultures; ICR 191 produced a 35.6-fold increase, and MNNG produced an 8.5-fold increase. All the spontaneous dexamethasone-resistant clones contained glucocorticoid receptors, usually less than half of the amount found in the parental clone. They are therefore strikingly different from dexamethasone-resistant clones derived from the mouse cell lines S49 and W7. Dexamethasone-resistant clones isolated after mutagenesis of CEM-C7 contained, on the average, lower concentrations of receptor than did those isolated spontaneously, and one clone contained no detectable receptor. These results are consistent with a mutational origin for dexamethasone resistance in these human cells at a haploid or functionally hemizygous locus. They also suggest that this is a useful system for mutation assay.

Cell Division↗
PubMed1981

Factors affecting adrenocortical hormone function.

This paper will briefly outline those elements which must be considered in assessing the effects of any given compound which might impinge on the actions of adrenocortical steroids. Considerations that ought to be taken into account include the following: the delivery to the affected cells of corticosteroids, the uptake of the hormone by the cells, the metabolism of the hormone by the cells, the intracellular actions of the hormone, the possible secondary interactions between cells, and multihormonal actions on the final target cell.

Adrenal Cortex↗
PubMed1981

Some pharmacological properties of a novel series of 2-substituted 2-phenylcyclohexyl N,N-diethylaminoethyl ethers. II.

The second of a series of 2-phenylcyclohexyl N,N-diethylaminoethyl ethers, (trans-compound), was investigated for its pharmacological actions in mice and rats. The trans-compound shortened the duration of methacholine (20 mg/kg)-induced salivation in mice, which was observed to be inversely proportional to the intraperitoneally administered dose of the compound. In anesthetized rats, a time-course study revealed the effects of the compound to be brief (5-15 minutes). When administered alone, it decreased heart rate, systolic, diastolic and mean arterial pressures and increased pulse-pressure. It produced transient apnea followed by slight increase in respiration. In other anesthetized rats, pretreatment with 5 mg/kg trans-compound reversed the depression response and bradycardia induced by injection of increasing doses of methacholine. Respiratory rate was similarly decreased due to initial transient apnea followed by a slight increase. There was a non-significant decrease in rectal temperature when methacholine was injected before or after the compound. These experiments suggest that the trans-compound possesses a weaker anticholinergic activity compared to the cis-compound.

Animals↗
PubMed1980

Cytosolic proteins that bind oxygenated sterols. Cellular distribution, specificity, and some properties.

The differential binding of cholesterol and 25-hydroxycholesterol to cytosolic proteins in various types of cells was investigated. 25-Hydroxycholesterol taken up by six different established cell culture lines and by mouse spleen cells in primary culture was bound to cytosolic components which, during velocity gradient centrifugation, displayed sedimentation coefficients of approximately 5 S and 8 S. In contrast, cholesterol taken up by the cells was concentrated near the bottom (approximately 20 S) of the sucrose density gradient but was distributed throughout. Results with primary cultures of mouse fetal liver differed from thos obtained with other cell cultures in that both sterols appeared principally in a 5 S band. Further characterization of the binding components from intact L cells indicated that binding of 25-hydroxycholesterol to the 8 S fraction was saturable and reversible, whereas binding to the 5 S band was not saturable. The 8 S 25-hydroxycholesterol complex involved a protein with a relatively long half-life. The complex was essentially stable at 0 degrees C but dissociated slowly at 25 degrees C. Sulfhydryl functions were not required for sterol binding, and formation of the complex was not dependent upon cAMP. Competition studies with intact cells and with isolated cytosol indicated that a number of other oxygenated sterols bind to 8 S sites occupated by 25-hydroxycholesterol. Those sterols which are potent suppressors of 3-hydroxy-3-methylglutaryl coenzyme A reductase (EC 1.1.1.34) activity competed for these binding sites, while those which do not suppress the reductase did not compete for them. These studies suggest that the binding of oxygenated sterols to the 8 S cytosolic component may be in some sense specific, while that to the 5 S component is nonspecific. The kinetics of formation and dissociation of the 8 S 25-hydroxy-cholesterol.protein complex and of the suppression of 3-hydroxy-3-methylglutaryl-CoA reductase in the presence of the diol was also consistent with a postulated role for the complex in the regulation of the enzyme.

Animals↗
PubMed1980

'Activation-labile' glucocorticoid-receptor complexes of a steroid-resistant variant of CEM-C7 human lymphoid cells.

For cytoplasmic glucocorticoid-receptor complexes to enter and accumulate in the nucleus a temperature-dependent event, 'activation' is required. Activation can be achieved in vitro by increased ionic strength, dilution or gel filtration and is manifested by an increased affinity of steroid-receptor complex for DNA and an altered elution profile from ion-exchange resins. Munck and Foley have shown that activated complexes isolated from thymocytes elute from DEAE-cellulose in a manner identical to complexes activated in vitro. We report here that DEAE-cellulose chromatography of steroid-receptor complexes from CEM-C7, a cloned human leukaemic T-cell line sensitive to the cytolytic action of glucocorticoids, and its steroid-resistant subclone 4R4 demonstrated that steroid receptors of clone 4R4 cannot form stable activated complexes. This defines a new defect in receptor action, activation lability (r+act1), which is unlike either the r-, r+nt-, or r+nti phenotypes previously described for mouse lymphoid variants.

Animals↗
PubMed1980

Genomic organization of rat prolactin and growth hormone genes.

Five overlapping cloned DNAs containing the rat prolactin gene and its flanking sequences, as well as one cloned DNA containing the rat growth hormone gene and its flanking sequences, were isolated from a chromosomal DNA library. They were characterized by restriction enzyme mapping and electron microscopy. In each gene, the structural gene sequence coding for mature mRNA of a length of about 1 kilobase is split into at least five segments by a minimum of four intervening sequences. The two genes are similar in the length and organization of their coding regions, consistent with the suggestion that they are derived from a common ancestral gene. However, the two genes differ greatly in the lengths of their intervening sequences. That leads to a total gene length of 10 kilobase pairs for the prolactin and 2.1 kilobase pairs for the growth hormone gene. At least one intervening sequence appears to be in the 5' nontranslated regions of the prolactin and growth hormone mRNA coding sequences.

Animals↗
PubMed1980

Unique long-acting antiglucocorticoid in whole and broken cell systems.

The biological properties of cortisol 21-mesylate (CM), an alkylating derivative of cortisol, were investigated in a line of rat hepatoma tissue culture (HTC) cells. CM appears to bind to glucocoticoid receptors in cell-free extracts because CM inhibits the specific binding of [3H]dexamethasone. However, in whole cells CM not only fails to induce the enzyme tyrosine aminotransferase (TyrATase) but also inhibits the induction of TyrATase by dexamethasone. Thus CM is an antiglucocorticoid. This is not caused by cell death, because CM is relatively nontoxic up to concentrations of 10 microM. The concentration of CM needed for half maximal inhibition of TyrATase induction is an order of magnitude lower than that predicted from the apparent cell-free affinity of CM for the glucocorticoid receptors of HTC cells, which suggests that the cell-free binding data does not reflect an equilibrium situation. In fact, the reactive alpha-keto mesylate group was intentionally incorporated into cortisol in hopes of obtaining a steroid capable of undergoing irreversible reactions. When HTC cells were preincubated with either CM or the reversible antiglucocorticoid progesterone and then washed to remove free steroid, only the CM-treated cells failed to show subsequent induction of TyrATase by dexamethasone. Furthermore, preincubation of HTC-cell cytosol with CM blocked approximately 75% of the subsequent exchange binding of [3H]dexamethasone to glucocorticoid receptor sites. Thus, the actions of CM in whole and broken cells either require an exceptionally long time for reversal or are not reversible. Together, these results indicate that CM is a unique antagonist and could be an irreversible antiglucocorticoid in vitro.

Animals↗
PubMed1980

Clinical correlations of steroid receptors and male breast cancer.

Estrogen receptors (ER) were present in tumor specimens from 29 of 34 cases of male breast cancer. There was a significant negative correlation of ER concentration with age. The quantity of ER tended to correlate directly with progesterone receptor levels, disease-free interval, and response duration among responders, but not to a statistically significant extent. In 13 patients for whom response data were available, no significant correlation was observed between ER levels and either frequency or duration of orchiectomy response. Among the six patients with tumor ER levels of less than 30 fmol per mg of protein, however, only two brief responses to orchiectomy occurred that were of little clinical benefit, while three of seven patients with higher ER responded more favorably. Thus, although this suggests that a relationship between low ER and unfavorable orchiectomy response may emerge as more patients are studied, currently available data do not justify basing therapeutic intervention on ER status of a biopsy in a manner analogous to that used for female breast cancer. Nine of 14 male breast cancer patients had positive progesterone receptor assays and several had androgen or glucocorticoid receptors. Tissue from only three of ten men with gynecomastia had measurable ER, and these were limited to the 4S component on sucrose gradients.

Adrenalectomy↗
PubMed1980

Some pharmacological properties of a novel series of 2-substituted 2-phenylcyclohexyl N,N-diethylaminoethyl ethers I.

Two 2-phenylcyclohexyl N,N-diethylaminoethyl ethers were studied for their pharmacological actions in vitro-the cis- and trans- isomers; and in vivo,-the cis- isomer. These compounds (5 mg/ml) significantly depressed acetylcholine-induced contractions in the isolated guineapig ileum. The trans- isomer produced a greater depression of contraction. The cis- isomer shortened the duration of methacholine-induced salivation in mice when administered intraperitoneally in increasing doses. The duration was found to be inversely proportional to the dose of the compound. In anesthetized rats, pretreatment with either a dose of 5 mg/Kg of the cis- compound or 0.25 mg/Kg of atropine changed the methacholine depressor response to a pressor response. Heart rate was increased in both groups of pretreated rats. Respiratory rate was further augmented by methacholine in the presence of either atropine or the cis- compound, while rectal temperature was slightly decreased. Effects of the cis- isomer when administered alone, on heart rate and blood pressure responses usggest cholinergic activity. However, its ability to decrease the duration of methacholine-induced salivation in mice and its reversal of methacholine-evoked cardiovascular effects are indicative of anticholinergic activity. These experiments suggest that the cis- isomer has a mixed cholinergic and anticholinergic action and thus may be classed as a partial antagonist of the parasympathetic system.

Animals↗