Estrogen Receptors In Nuclei Research Articles

Estrogen receptor and aromatic hydrocarbon receptor in the primate ovary

We have previously shown by immunocytochemistry and autoradiography the presence of estrogen receptors (ER) in rhesus monkey ovary. Intense chromogen staining showed specific binding for ER in nuclei of germinal epithelium and granulosa cells of antral follicles; and radiolabeled ligand bound specifically to functional corpora lutea (CL). Although it is accepted that the germinal epithelium of the primate ovary contains ER, some controversy still persists regarding the intraovarian localization of this molecule. In addition, no data exist that localize the aromatic hydrocarbon (dioxin) receptor (AHR), which is known to modulate ER, to the primate ovary. In the present study, we show the presence of ER using Western blot analysis, and ER capable of binding DNA within intraovarian compartments in two species of the genusMacaca (rhesus macaque,Macaca mulatta and stumptail macaque,Macaca arctoides); extend these findings to human ovarian granulosa cells (GC) using Western blot, reverse transcription-polymerase chain reaction (RT-PCR), and gel mobility-shift analysis; and localize the AHR to intraovarian compartments of the macaque ovary by Western blots and gel-shift assays. These experiments strongly suggest that estrogens can exert effects on follicle development directly at the ovary, and provide the first direct evidence that AHR-mediated toxicity may be manifested at the ovary to induce possible antifertility effects.

Estrogen and androgen receptors in Sertoli, Leydig, myoid, and epithelial cells: effects of time in culture and cell density.

Estrogen and androgen receptors were measured in the cytosol prepared from established cell lines of Sertoli, Leydig, myoid, and endothelial cells as well as in primary Sertoli cell-enriched cultures. Estradiol-binding sites with characteristics of estrogen receptors were identified in established lines of Sertoli cells from rat [43 +/- 7 (mean +/- SEM) fmol/mg protein; Kd = 1.1 nM] and mouse (34 +/- 4 fmol/mg protein; Kd = 0.9 nM). The binding sites were estrogen specific, since only estradiol and diethylstilbestrol, but not testosterone, progesterone, or dexamethasone, competed with [3H]estradiol for binding sites in the cytosol prepared from the cell lines. Exposure of the cells to estradiol (10 nM) resulted in accumulation of estrogen receptors in nuclei, with maximal uptake by 30 min. The estrogen receptor concentration was very low or undetectable (less than 10 fmol/mg protein) in primary cultures of rat Sertoli cells that were cultured for 3 days. However, after 15 days in culture, the estrogen receptor concentration increased and reached levels similar to those in the established Sertoli cell lines. No estrogen receptors were measurable in myoid or endothelial cells. By contrast, androgen receptors were identified in all five cell lines and in primary Sertoli cells cultured for 3 and 15 days. The content of both estrogen and androgen receptors in the mouse Sertoli cell line increased as a function of cell density. We conclude from these studies that androgen receptors are present in all of the testicular somatic cell lines examined and in primary Sertoli cells; estrogen receptors are present in Sertoli cell and Leydig cell lines, but not in myoid and endothelial cell lines; the low estrogen receptor concentration in Sertoli cells cultured for 3 days increases 4-fold after 15 days in culture; and cell density is a major regulator of the concentrations of estrogen and androgen receptors in the Sertoli cell line.

Hepatic nuclear estrogen receptor concentrations in the rat—influence of age, sex, gestation, lactation and estrous cycle

Nuclear estrogen receptor concentrations in rat liver were determined by exchange assay. The concentration of estrogen receptors in nuclei from vehicle-treated male and female rats show agedependent, but not sex-dependent variations in the course of life. Levels are highest during the perinatal period (~ 1500 binding sites/nucleus), whereafter they decrease towards the onset of puberty (~ 300 binding sites/nucleus) before rising again to reach the postpuberal maximum (~ 800 binding sites/nucleus). Pregnancy further raised receptor concentrations in the last week of gestation when they reach ~ 1200 binding sites/nucleus. Studies with ethynylestradiol-treated rats demonstrated that virtually no translocation can be detected before the onset of puberty; thereafter the number of translocated receptors increases dramatically reaching a maximum (9000 binding sites/nucleus) between day 80 and 87 of life. The extractability of the nuclear receptors with 0.4 M KC1 varies during the course of life. Extractability is very high (~90%) up to about day 12 of life, but then decreases markedly (to ~70% in vehicle-treated and to ~ 50% in ethynylestradiol-treated rats) before the onset of puberty.

Alterations induced by clomiphene in the concentrations of oestrogen receptors in the uterus, pituitary gland and hypothalamus of female rats.

Alterations in the concentrations of oestrogen receptors in the uterus, pituitary gland and hypothalamus during the 2 weeks following a single administration of clomiphene citrate (Clomid) to immature, bilaterally ovariectomized rats were investigated. Examination of the uterine wet weight at 1, 7 and 14 days following a single injection of Clomid (100 micrograms, 250 micrograms or 10 mg) indicated significant time- and dose-related increments from a control value of 45 +/- 2 (S.E.M.) mg to a maximum of 123 +/- 3 mg (250 micrograms dose at 14 days). In contrast, a single injection of oestradiol led to a transient increase in the uterine weight on day 1 to 94 +/- 6 mg, but was without effect by days 7 and 14. Analysis of the uterine DNA content 7 and 14 days after treatment with Clomid revealed significant increments from control values of 390 +/- 10 micrograms to a high level of 558 +/- 9 micrograms (10 mg dose at 7 days). There was a transient retention of nuclear oestrogen receptors and rapid replenishment of cytoplasmic oestrogen receptors in less than 24 h in the uteri of animals treated with oestradiol (25 micrograms), but determinations of receptor content in Clomid-treated animals revealed prolonged retention of nuclear receptors and delayed replenishment of cytoplasmic receptors. The duration and extent of retention of nuclear receptors and depletion of cytoplasmic receptors after treatment with Clomid were found to be dose-dependent. Fourteen days after Clomid treatment, levels of oestrogen receptors in nuclei from the uterus were still raised in all treatment groups, whereas replenishment of cytoplasmic receptors was complete in animals treated with the lower doses (100 and 250 micrograms) of Clomid. A single injection of Clomid (250 micrograms) induced similar prolonged retention of nuclear receptors and delayed depletion of cytoplasmic receptors in pituitary tissue. In contrast, changes in the content of oestrogen receptors in the hypothalamus following Clomid treatment were minimal. The limited effect of Clomid on hypothalamic tissue may mean that the pituitary gland is a more important target for this compound than is the hypothalamus. The findings have confirmed earlier reports on the long-term uterotrophic effect of Clomid and have suggested that under these long-term, in-vivo conditions, Clomid acts in the uterus and pituitary gland as a long-acting oestrogen characterized by prolonged retention of oestrogen receptors in the nucleus and delayed, but otherwise effective, replenishment of the oestrogen receptors in the cytoplasm.

Androgen-induced nuclear accumulation of the estrogen receptor

The effect of androgens on the nuclear uptake of both tritiated estradiol ( 3H-E 2) and the estrogen receptor was studied in immature rat uteri. It was demonstrated that in vitro preincubation of immature rat uteri with various androgens (1 μM to 50μM) followed by incubation with 3H-E 2 (20nM) resulted in a greatly decreased specific nuclear uptake of 3H-E 2. Non-androgenic steroids had no effect. It was also confirmed that 5a-dihydrotestosterone (DHT) causes the accumulation of the estrogen receptor in the nuclei of uterine tissue. In vitro incubations of rat uteri with DHT (1μM and 50μM) were found to cause maximal nuclear estrogen receptor accumulation to the same degree as caused by estradiol, i.e. the nuclear uptake of approximately 100% of the estrogen receptor. Antiandrogens, which block the binding of androgens to the testosterone receptor in various tissues, did not inhibit the DHT -induced decrease in the 3H-E 2uptake by the uterine nuclei or the DHT — caused accumulation of the estrogen receptor in nuclei. These results seem to indicate that the uterine testosterone receptor has no role in the androgen — induced nuclear uptake of the estrogen receptor. However, the non-steroidal antiestrogens inhibited the DHT — induced nuclear accumulation of the estrogen receptor. This would seem to indicate that the estrogen — and androgen — induced nuclear accumulation of the estrogen receptor share a common mechanism.