The interaction between TRH and GH3 pituitary tumor cells was studied in monolayer cultures or membrane-containing fractions. In intact cells, the apparent dissociation constant (Kd) was approximately 10 nM, and the total number of binding sites was approximately 1.4 pmol/mg protein at temperatures of 0-37 C. In broken cell preparations, the number of sites occupied at saturating TRH concentrations was reduced by half when the temperature was increased from 0 to 30 C. Linear Scatchard plots were obtained under all conditions. The rate of dissociation of TRH was temperature dependent, and first order plots were nonlinear. The half-times for dissociation at 0 C were over 240 min in cells and membranes, while at 37 C, the half-time values were 24 min (cells) and less than 0.5 min (membranes). Identical dissociation kinetics were obtained by dilution alone or dilution with excess unlabeled hormone. When cultures which had been incubated with TRH were lightly fixed with glutaraldehyde, dissociation at 37 C became immeasurably slow. However, TRH dissociated immediately when sodium dodecyl sulfate, ethanol, or acetone was added, indicating that the tripeptide was not covalently coupled to cell proteins. The data indicate that binding of TRH to high affinity GH3 receptors is not cooperative, and that the majority of TRH bound after short incubations is dissociable.
The interrelationships between thyroid hormone and cortisol actions were investigated in GH3 pituitary tumor cells. When GH3 cells were grown in thyroid hormone-deficient medium, cortisol did not affect the concentration of TRH receptors. Both thyroid hormones and TRH normally decrease the number of TRH receptors, and cortisol inhibited down-regulation by both hormones. TRH caused a greater increase in PRL synthesis when TRH receptors were high in the presence of cortisol and T3 than when TRH receptors were low (T3 alone). In the presence of cortisol, higher concentrations of T3 were required to decrease TRH receptors, while lower concentrations were necessary to stimulate GH synthesis. Cortisol and T3 alone stimulated GH synthesis 6- and 10-fold, respectively, while together they caused an 830-fold increase. In contrast, T3 did not alter the inhibition of PRL synthesis by the glucocorticoid. Cortisol did not significantly affect the amount of [125I]T3 bound to nuclei from cells incubated in thyroid hormone-deficient or T3-supplemented medium (approximately 100 and approximately 25 fmol/mg cell protein). The data suggest that cortisol modifies thyroid hormone action at a step subsequent to T3 receptor binding.
The use of Thorotrast as a contrast medium is now of historical interest. Thorotrast-induced angiosarcoma, though rare, still generates considerable clinical interest because of the characteristic opacification of the liver, spleen, and lymph nodes, and the long latency period between exposure and the onset of the tumor. We present a case of hepatic angiosarcoma which developed 37 years after the administration of Thorotrast.
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