Unoccupied Nuclear Receptors for Estrogen in Human Endometrial Tissue

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Abstract
Extracts of cell nuclei from normal, nonmalignant, and cancerous human endometrial tissues were found to contain material capable of binding estradiol at 0 C in vitro. Measurements at the same temperature imply that only binding sites unoccupied by endogenous estrogen are determined. The estrogenic receptor character of this binding was demonstrated by: 1) high affinity of binding to estradiol (Kd = ∼0.5 nM as assayed by the Scatchard method); 2) specificity of binding, competition by diethylstilbestrol and estriol for estradiol binding, and the absence of competition by cortisol, progesterone, and 5α-dihydrotestosterone; and 3) sedimentation constant at about 4S in a sucrose density gradient. Both unoccupied and occupied nuclear receptors were determined in normal, nonmalignant, and cancerous endometrial tissues. Unoccupied receptors were measured at 0 C for 2 h, while the occupied receptors were calculated from the total binding measured at 30 C for 2 h. Receptors were measured by a single saturating dose of 7.5 nM [3H]estradiol with or without a 100-fold excess of diethylstilbestrol to estimate the amount of nonspecific binding. Unoccupied nuclear receptors were found in all specimens assayed. The unoccupied nuclear receptor comprises 9–37% of the total estradiol receptors (cytoplasmic plus nuclear). The finding that a substantial number of receptors in the nuclei of normal and pathological endometrial tissue are unoccupied may indicate that the unoccupied receptor is a necessary product in the normal mechanism of estradiol action.