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ARTICLE |
and ERß, in Estrogen Target Tissues in Vivo through the Use of an ER-Selective Ligand
Women’s Health Research Institute (H.H.), Wyeth Research, Collegeville, Pennsylvania 19426; and Departments of Chemistry (J.A.K.) and Molecular and Integrative Physiology (B.S.K.), University of Illinois, Urbana, Illinois 61801
Address all correspondence and requests for reprints to: Dr. Benita S. Katzenellenbogen, Department of Molecular and Integrative Physiology, 524 Burrill Hall, 407 South Goodwin Avenue, University of Illinois, Urbana, Illinois 61801-3704. E-mail: katzenel{at}life.uiuc.edu.
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Abstract |
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and ERß, in mediating these bioactivities is incompletely understood. In this study, we investigated the activity of an ER-selective agonist ligand, propyl pyrazole triol (PPT), in several rat animal models to define the involvement of ER in these biological responses. In a short-term (4 d) uterotrophic assay, PPT was found to be as efficacious as 17-ethinyl-17ß-estradiol in stimulating uterine weight gain and up-regulating complement 3 gene expression. In a 6-wk chronic model, PPT completely prevented the ovariectomy-induced body weight increase and loss of bone mineral density. It also increased uterine weight and markedly reduced plasma cholesterol levels in these mature animals. PPT was also effective in the brain. It increased progesterone receptor mRNA in the arcuate and ventromedial nuclei of the hypothalamus and prevented experimentally induced hot flushes. Our findings indicate that several physiologically relevant estrogen-induced tissue responses can be effectively evoked via ER alone. By providing an approach that is complementary to that of analyzing the phenotype and response of ER knockout animals, our findings also demonstrate that ER subtype-selective ligands can play a valuable role in enhancing our understanding of how estrogens work through the two ER subtypes. |
Introduction |
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and ERß (1, 2, 3). The tissue distribution of these subtypes is not completely coincident, as some tissues (e.g. uterus, vagina, and hypothalamic arcuate nucleus of the brain) express predominantly ER, whereas others (e.g. lung, prostate, ovarian granulosa cells, and hypothalamic paraventricular nucleus of the brain) express predominantly ERß. Other tissues, such as bone and the pituitary, express both ER and ERß (3, 4, 5).
The respective roles of ER and ERß have been investigated through the use of ER, ERß, or ER/ERß knockout mice, where the deletion of one or both of the ERs can be correlated with changes in the effects of estrogen in different tissues and on different responses (6, 7, 8). Although analysis of these knockout mice has been key to expanding our understanding of the physiological roles of estrogens, this approach is not without its drawbacks. For example, one cannot separate the developmental effects of estrogens from those of adulthood because the ER knockout mice produced to date are not conditional knockouts. In addition, some biological responses may still occur as a consequence of the small quantities of variant ER transcripts in some ERKO mice (7).
Synthetic estrogens that are ER subtype-selective also have the potential to be very useful tools for elucidating the biological roles of ER and ERß (9, 10, 11). These compounds can be used in normal animals at any stage of development. Results obtained from this approach should complement studies with knockout mice and help to further expand our appreciation of the complexities of estrogen action.
We have developed a synthetic estrogen agonist, propyl pyrazole triol (PPT), which is highly selective for ER. Previously, we have shown that PPT binds to ER with an affinity that is 400-fold higher than for ERß (11). In gene transcription assays in cells, PPT is a potent agonist on ER and is devoid of activity on ERß (10, 11, 12). In this report, we have used PPT to define the physiological roles played by ER in mediating several well-known estrogenic responses in vivo.
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Materials and Methods |
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coding region (13) using 2% stripped FBS phenol red-free medium with antibiotics and GlutaMAX-1 for 2 h at 37 C. Medium containing virus was aspirated, and the cells were washed with medium. Fresh medium was added and the cells were allowed to recover overnight at 37 C. Cells were then treated with various compounds for 24 h. For single doses, test compounds are usually administered at 1 µM, and 17ß-estradiol (E2) is used at 10 nM. To measure antagonist activity, test compounds are coadministered with E2.
TaqMan quantitative RT-PCR
RNA was isolated using RNeasy columns (QIAGEN, Valencia, CA). Each sample was deoxyribonuclease treated directly on the column. For IGF binding protein (IGFBP)-4, one-step TaqMan RT-PCR was performed on a set of six standard RNAs with amounts of 200, 40, 8, 1.6, 0.32, and 0.016 ng and on each Saos-2 RNA sample of 40 ng in a 50-µl reaction. The reaction contained 1x TaqMan One Step RT-PCR Master Mix (Perkin-Elmer, Foster City, CA), 0.5 µM IGFBP-4-specific forward and reverse primers (forward: 5'-TTTTCAGCCTTGGGAGGTTTTAT-3' and reverse: 5'-AGGCTTGAACTCTCCTTATAGGAGTG-3'), 0.2 µM IGFBP-4-specific TaqMan probe (5'-6-FAM-CTCCACATGCCAAAATCAGAGGAAGTCAG-TAMRA-3'), 1x MultiScribe/Ribonuclease Inhibitor Mix (Perkin-Elmer). Reactions were incubated at 48 C for 30 min followed by 10 min at 95 C then 40 cycles of PCR as follows: 95 C for 15 sec then 60 C for 1 min in an ABI 7700.
For metallothionein-II, one-step TaqMan RT-PCR was performed as described above except that 0.9 µM of metallothionein-II-specific primers was used. The primer sequences were forward: 5'-AAAGAACGCGACTTCCACAAA-3' and reverse: 5'-TTCAAGTCAAAGCTGTTTTATTATCATTC-3'. The probe sequence was 5'-6-FAM-CCCTGACCGTTTGCTATATTCCTTT TTCTATGAA-TAMRA-3'.
Real-time quantitation of PCRs was done using Sequence Detector Software (Applied Biosystems Inc., Foster City, CA). Amount of IGFBP-4 and metallothionein-II RNA in each sample was calculated based on the standard curve RNA and expressed as nanograms.
Quantitation of plasma levels of PPT following sc administration
All studies involving animals were conducted in accordance with the NIH Guide for the Care and Use of Laboratory Animals, with all protocols approved by Wyeth’s Institutional Committee on Animal Care and Use.
Young male Sprague Dawley rats (165 g) were dosed sc with 10 mg/kg of PPT in a vehicle of 50% dimethylsulfoxide (DMSO)/50% 1x Dulbecco’s PBS. Rats were euthanized at various times after dosing (three per group), and blood was collected via cardiac puncture. Coagulation was prevented by combining 1ml of blood with 50 µl of 0.5 M EDTA. Plasma was prepared by centrifuging blood at 10,000 x g for 5 min and collecting the supernatant solution. Plasma was stored frozen at -80 C until use.
The plasma samples were prepared for analysis by combining 100 µl of each plasma sample with 100 µl of a 200 ng/ml aqueous solution of Biochanin A (purchased from Sigma, St. Louis, MO). After dilution, the samples were loaded onto the autosampler of a 2790 HPLC system (Waters Corp., Milford, MA) for injection. The extraction of PPT from the plasma was performed by the injection of 10 µl of the plasma/internal standard solution onto an Oasis extraction column (2.1 x 20 mm; 25 µm; Waters Corp.) and washing at 4 ml/min for 0.3 min using an aqueous 0.02% triethylamine solution (approximately 20-column volumes). After the extraction was complete, a 2690 HPLC system (Waters Corp.) was used to reverse elute the trapped PPT from the Oasis column using a fast water/acetonitrile gradient. To improve resolution and peak shape of the compound of interest, PPT was eluted from the Oasis extraction column onto a Waters XTerra MS C18 column (3 x 30 mm; 3.5 µm) using 0.02% triethylamine in each mobile phase gradient mobile phase. The quantity of PPT in the sample was quantified using a Micromass Quattro Ultima triple quadrupole mass spectrometer. Multiple reaction monitoring was used as the detection mechanism. In general, standard curves from 1–1000 ng/ml were created by dissolving known amounts of pure PPT in rat plasma purchased from Pel-Freez Biologicals, Inc. (Rogers, AR).
Evaluation of uterotrophic activity and complement 3 (C3) gene expression
Sexually immature (18 d of age) Sprague Dawley rats were obtained from Taconic Farms, Inc. (Germantown, NY) and provided unrestricted access to a casein-based diet (Purina Mills 5K96C, St. Louis, MO) and water. On d 19, 20, and 21, the rats were dosed sc with EE2 (0.06 µg/rat·d), PPT, or vehicle (50% DMSO/50% Dulbecco’s PBS). Doses are expressed as micrograms/rat because the rats grow significantly during the study from about 35 g to about 50 g. Rats (n = 6 per group) were euthanized approximately 24 h after the last injection by CO2 asphyxiation and pneumothorax. Uteri were removed and weighed after trimming associated fat and expressing any internal fluid.
Complement component C3 gene expression was assessed by Northern blot analysis as previously described (14).
Evaluation of effects of PPT on bone mineral density, body weight, uterine weight, and plasma cholesterol in a 6-wk rat model of osteopenia
Female Sprague Dawley rats, ovariectomized or sham operated, were obtained 1 d after surgery from Taconic Farms, Inc. (weight range 240–275 g). They were housed three or four rats per cage in a room on a 12-h light, 12-h dark schedule and provided with food (Purina 5K96C rat chow) and water ad libitum. Treatment for all studies began 1 d after arrival and rats were dosed 7 d per week as indicated for 6 wk. A group of age-matched sham operated rats not receiving any treatment served as an intact, estrogen-replete control group for each study.
PPT was prepared in a vehicle of 50% DMSO/50% 1x Dulbecco’s PBS at defined concentrations so that the treatment volume was 0.1 ml/100 g body weight. E2 was dissolved in corn oil (20 µg/ml) and delivered sc, 0.1 ml/rat. All dosages were adjusted at 3-wk intervals according to group mean body weight measurements, and given sc.
Five weeks after the initiation of treatment and 1 wk before the termination of the study, each rat was evaluated for bone mineral density. The total and trabecular density of the proximal tibia were evaluated in anesthetized rats using an XCT-960M [peripheral quantitative computer tomography (pQCT); Stratec Medizintechnik, Pforzheim, Germany]. The measurements were performed as follows: 15 min before scanning, each rat was anesthetized with an ip injection of 45 mg/kg ketamine, 8.5 mg/kg xylazine, and 1.5 mg/kg acepromazine. The right hind limb was passed through a polycarbonate tube with a diameter of 25 mm and taped to an acrylic frame with the ankle joint at a 90° angle and the knee joint at 180°. The polycarbonate tube was affixed to a sliding platform that maintained it perpendicular to the aperture of the pQCT. The platform was adjusted so that the distal end of the femur and the proximal end of the tibia was in the scanning field. A two-dimensional scout view was run for a length of 10 mm and a line resolution of 0.2 mm. After the scout view was displayed on the monitor, the proximal end of the tibia was located. The pQCT scan was initiated 3.4 mm distal from this point. The pQCT scan was 1 mm thick, has a voxel (three-dimensional pixel) size of 0.140 mm, and consisted of 145 projections through the slice.
After the pQCT scan was completed, the image was displayed on the monitor. A region of interest including the tibia, but excluding the fibula, was outlined. The soft tissue was mathematically removed using an iterative algorithm. The density of the remaining bone (total density) is reported in mg/cm3. The outer 55% of the bone was mathematically peeled away in a concentric spiral. The density of the remaining bone (trabecular density) is reported in mg/cm3.
One week after bone mineral density evaluation, the rats were euthanized by CO2 asphyxiation and pneumothorax, and blood was collected for cholesterol determination. The uteri were also removed and weighed after trimming associated fat and expressing any luminal fluid. Total cholesterol was determined using a Roche Molecular Biochemicals (Indianapolis, IN) Hitachi 911 clinical analyzer and the Cholesterol/HP kit (15). Statistics were compared using one-way ANOVA with Dunnett’s test.
Induction of progesterone receptor (PR) mRNA in the brain
This study was similar to previous studies where PR mRNA levels were examined (16), except that the arcuate and ventromedial hypothalamic nuclei were analyzed here rather than the preoptic area. Briefly, ovariectomized Sprague Dawley rats were treated with a single injection of either E2 (50 µg/kg, sc) or PPT 6 h (both in 50% DMSO/50% 1x Dulbecco’s PBS) before euthanasia. The brains were frozen, cryosectioned (20 µm) and sections through the ventromedial and arcuate hypothalamic nuclei were processed for PR mRNA by in situ hybridization as previously described (16). The slides were then exposed to autoradiographic film for 5 d at room temperature and the films were analyzed for levels of PR mRNA by measuring optical density using image analysis software (C-Imaging, Pittsburgh, PA). OD readings were obtained from a defined window that included both nuclei in six sections/animal (both sides of the brain). The sections were approximately 60 µm apart, and the six individual measurements were averaged to obtain a single optical density value for each animal.
Analysis of hot flush in a rat model
The effects of PPT and 17-ethinyl-17ß-estradiol (EE2) on prevention of hot flushes were evaluated in mature ovariectomized rats using a protocol previously described (17).
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or ERß selective using several estrogen-responsive promoter-reporter gene constructs transfected together with either ER or ERß in a variety of cell lines (11, 12). To further investigate the ER selectivity of PPT in cells, we examined its ability to regulate expression of two endogenous genes, one of which is regulated by both ER and ERß (IGFBP-4), and the other only by ERß (metallothionein-II) (13). Saos-2 cells expressing either human ER or ERß were treated with 10 nM E2 or 1 µM PPT for 24 h, RNA was extracted, and quantitative RT-PCR (TaqMan) was used to measure the levels of these two mRNAs.
As shown in Table 1
, the ER and ERß agonist, E2, up-regulated expression of IGFBP-4 mRNA in cells when either ER or ERß was expressed. PPT, however, only up-regulated IGFBP-4 mRNA in Saos-2 cells expressing ER.
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in these cells (13), and these observations were confirmed in this study (Table 1
). Of note, PPT did not up-regulate metallothionein-II mRNA when cells expressed either ER or ERß.
With both genes, PPT had no influence on the stimulation by E2, indicating that it does not act as an ER antagonist. These results indicate that PPT retains its specificity as an ER agonist on these endogenously expressed genes.
PPT pharmacokinetics
A preliminary assessment was made of PPT plasma levels after treatment. Rats received 10 mg/kg PPT sc, and the levels of PPT in the plasma were monitored by HPLC-mass spectroscopy analysis. The concentration of PPT in plasma was 705 ng/ml at 0.5 h and remained above 450 ng/ml for at least 4 h after injection.
Dose-response effects of PPT and E2 on uterine weight and complement 3 gene expression
Sexually immature Sprague Dawley rats were treated with various doses of PPT (5–1000 µg/rat·d) or with EE2 (0.06 µg/rat·d) for 3 d, and uterine weights were determined approximately 24 h after the last dose. As shown in Fig. 1, PPT stimulated uterine weight gain in a dose-dependent manner and was as efficacious as EE2 but was less potent, so much higher doses of PPT were needed.
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, at the two higher doses, PPT stimulated C3 gene expression as effectively as EE2.
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Ovariectomy is known to cause a marked increase in body weight in this animal model. As seen in Fig. 3A, both PPT and E2 completely prevented the ovariectomy-induced weight gain.
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). These uterine weight results are consistent with those seen in immature rats (Fig. 1) in which PPT was as efficacious as estrogen in increasing uterine weight. PPT reduced total plasma cholesterol by approximately 50%. The amount of E2 used in this study was below that required to reduce cholesterol, so no change was seen (15). A comparison of the effectiveness of PPT vs. E2 in the uterine weight and body weight vs. cholesterol endpoints highlights the fact that different estrogens can have very different potencies in different tissue responses.
As shown in Fig. 3D, PPT was as efficacious as E2 in preventing the ovariectomy-induced loss of bone mineral density in the proximal tibia. Preservation of bone density was seen in both the total and trabecular compartments.
Comparison of the effects of PPT and E2 in the brain
PR induction by PPT and E2.
Estrogen is well known to dramatically increase PR mRNA in several hypothalamic brain regions including the medial preoptic area, the ventromedial nucleus, and arcuate nucleus (16, 18). Because in rats the anterior ventrolateral ventromedial and arcuate hypothalamic nuclei contain almost exclusively ER, we examined the effectiveness of E2 and PPT to induce PR mRNA specifically in these nuclei. Using in situ hybridization, we found that ovariectomized rats treated with PPT for 6 h had a dose-dependent induction of PR mRNA in both brain regions (Fig. 4, A and B). These data indicate that PPT is able to cross the blood-brain barrier and induce gene expression in neurons known to contain ER.
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, PPT (15 mg/kg) and EE2 (0.3 mg/kg) were equally effective at preventing the tail skin temperature increase in this animal model. These hormone doses were also equally and maximally effective in increasing uterine weight (Fig. 5C). A lower dose of PPT (5 mg/kg) was ineffective at preventing hot flush but was fully uterotrophic (data not shown), again indicating a potency separation between different estrogenic endpoints in vivo. |
Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-selective ligand, PPT, to delineate physiological functions mediated by ER vs. ERß. We find that several biologically important estrogen-induced tissue responses can be effectively evoked by activation of ER only.
In the uterus, PPT was as efficacious as the ER subtype nonselective ligand EE2 at increasing tissue weight and up-regulating C3 mRNA. These data indicate that stimulation of ER is sufficient to elicit a full uterotrophic response and that ERß is not required for these estrogenic effects.
The uterine results are consistent with the predominant expression of ER in this tissue, and with the findings from ERKO mice in which E2 was unable to stimulate significant uterine weight gain (7). However, in other tissues such as bone, where the two ERs are expressed at more comparable levels (1, 3), it is of interest that our study clearly shows that activation of ER is sufficient to mediate the bone sparing effects of estrogens in a clinically relevant model of osteopenia.
Because estrogens exert important effects in the brain (16, 18), we were particularly interested in using PPT to assess the contribution of ER in mediating estrogenic effects in the central nervous system. We assessed two end points known to be estrogen regulated, induction of PR mRNA, and suppression of hot flushes. PPT was able to up-regulate PR in the anterior ventrolateral ventromedial and arcuate regions of the hypothalamus, which are known to express ER almost exclusively (16).
Postmenopausal women often suffer from hot flushes, and indeed, this is the primary reason that many choose to begin hormone replacement therapy. Because both ER and ERß are expressed throughout the brain, we examined the efficacy of PPT on ameliorating hot flushes in a rat model of vasomotor instability. As was observed in the bone osteopenia model, PPT was as efficacious as EE2 in blunting experimentally induced hot flushes. Again, we conclude that activation of ER is sufficient to produce this effect.
We observed that the doses of PPT required to obtain activity on brain endpoints (PR induction and hot flushes) were markedly higher than those required to elicit uterine effects. In our experience, this is not unusual and probably results from several factors. First, the exposure and metabolism of compounds may differ between the brain and the rest of the body. In addition, biological endpoints can exhibit differential sensitivities. For example, when EE2 is administered sc, the EC50 for uterine weight gain in adult ovariectomized rats is 0.3 µg/kg, whereas the IC50 for cholesterol reduction is 22 µg/kg—a difference of two orders of magnitude (15). The high doses of PPT required for in vivo effectiveness, despite the high affinity of PPT for ER, may reflect high protein binding of this compound, which would affect its pharmacodynamic behavior.
It should be noted that an observation that PPT acting through ER alone is capable of evoking the full response in a particular bioactivity is evidence only that the response can be evoked by ER. It does not demonstrate that the in vivo response might not also be capable of being elicited by estrogen action through ERß. Further studies will also be needed to investigate whether estrogen action through ERß might enhance or suppress the activity or potency of estrogens acting through ER, as suggested in model cell culture systems (19).
The approach of using ER-selective ligands serves as a valuable complement to the use of knockout animals in understanding the respective biological roles of ER and ERß. It is worth noting that there are some limitations in the knockout approach, in that the animals are genetically devoid of one or both of the ERs from the earliest embryonic stages, and thus, there is the potential for complexities that might arise through developmental compensation over these long periods of time. The development of conditional knockout animals, including tissue-specific knockouts, would avoid some of these potential problems. However, knockouts may not always be complete, leaving residual expression of variant receptor forms that can result in a low level of responsiveness (20, 21). In addition, deletion of only one ER subtype eliminates the modulation of responses that might arise from synergistic or antagonistic interactions between the two proteins, and may also alter possible up- or down-regulation of one receptor subtype by the other. Hence, the use of ER subtype-specific ligands provides an alternate approach for probing the biological roles of the two ERs, although with limitations based on the degree to which such ligands are selective only for the intended receptor target. Work in progress in our laboratories, as well as in others, is directed at obtaining agonist and antagonist ligands that are highly selective for each ER subtype. These compounds could be used in combinatorial studies to elucidate such ER subtype interactions and resultant bioactivities.
In summary, our findings document that several physiologically relevant estrogen-mediated tissue responses can be effectively and exclusively evoked through ER. They also demonstrate that ER subtype selective ligands can play a valuable role in enhancing our understanding of how estrogens work through ER and ERß to regulate the numerous important processes that are under estrogen control.
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Acknowledgments |
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Footnotes |
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Abbreviations: DMSO, Dimethylsulfoxide; E2, 17ß-estradiol; EE2, 17-ethinyl-17ß-estradiol; ER, estrogen receptor; FBS, fetal bovine serum; IGFBP, IGF binding protein; PPT, propyl pyrazole triol; pQCT, peripheral quantitative computer tomography; PR, progesterone receptor.
Received April 15, 2002.
Accepted for publication July 5, 2002.
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References |
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and ß. Endocrinology 138:863–870 (ER) and ß (ERß) on mouse reproductive phenotypes. Development 127:4277–4291[Abstract]
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transcriptional activity and is a key regulator of the cellular response to estrogens and antiestrogens. Endocrinology 140:5566–5578This article has been cited by other articles:
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 R. W. Hsieh, S. S. Rajan, S. K. Sharma, Y. Guo, E. R. DeSombre, M. Mrksich, and G. L. Greene Identification of Ligands with Bicyclic Scaffolds Provides Insights into Mechanisms of Estrogen Receptor Subtype Selectivity J. Biol. Chem., June 30, 2006; 281(26): 17909 - 17919. [Abstract] [Full Text] [PDF]
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 L. B. J. Morales, K. K. Loo, H.-b. Liu, C. Peterson, S. Tiwari-Woodruff, and R. R. Voskuhl Treatment with an estrogen receptor alpha ligand is neuroprotective in experimental autoimmune encephalomyelitis. J. Neurosci., June 21, 2006; 26(25): 6823 - 6833. [Abstract] [Full Text] [PDF]
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 S. J. Warden, L. K. Saxon, A. B. Castillo, and C. H. Turner Knee ligament mechanical properties are not influenced by estrogen or its receptors Am J Physiol Endocrinol Metab, May 1, 2006; 290(5): E1034 - E1040. [Abstract] [Full Text] [PDF]
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J Christoffel, G Rimoldi, and W Wuttke Effects of 8-prenylnaringenin on the hypothalamo-pituitary-uterine axis in rats after 3-month treatment. J. Endocrinol., March 1, 2006; 188(3): 397 - 405. [Abstract] [Full Text] [PDF]
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T. D. Lund, L. R. Hinds, and R. J. Handa The Androgen 5{alpha}-Dihydrotestosterone and Its Metabolite 5{alpha}-Androstan-3beta, 17beta-Diol Inhibit the Hypothalamo-Pituitary-Adrenal Response to Stress by Acting through Estrogen Receptor beta-Expressing Neurons in the Hypothalamus J. Neurosci., February 1, 2006; 26(5): 1448 - 1456. [Abstract] [Full Text] [PDF]
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M. T. Follettie, M. Pinard, J. C. Keith Jr., L. Wang, D. Chelsky, C. Hayward, P. Kearney, P. Thibault, E. Paramithiotis, A. J. Dorner, et al. Organ Messenger Ribonucleic Acid and Plasma Proteome Changes in the Adjuvant-Induced Arthritis Model: Responses to Disease Induction and Therapy with the Estrogen Receptor-{beta} Selective Agonist ERB-041 Endocrinology, February 1, 2006; 147(2): 714 - 723. [Abstract] [Full Text] [PDF]
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C. Bodo, A. E. Kudwa, and E. F. Rissman Both Estrogen Receptor-{alpha} and -{beta} Are Required for Sexual Differentiation of the Anteroventral Periventricular Area in Mice Endocrinology, January 1, 2006; 147(1): 415 - 420. [Abstract] [Full Text] [PDF]
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 K. Kublickiene, E. Svedas, B.-M. Landgren, M. Crisby, N. Nahar, H. Nisell, and L. Poston Small Artery Endothelial Dysfunction in Postmenopausal Women: In Vitro Function, Morphology, and Modification by Estrogen and Selective Estrogen Receptor Modulators J. Clin. Endocrinol. Metab., November 1, 2005; 90(11): 6113 - 6122. [Abstract] [Full Text] [PDF]
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B. S. Komm, Y. P. Kharode, P. V. N. Bodine, H. A. Harris, C. P. Miller, and C. R. Lyttle Bazedoxifene Acetate: A Selective Estrogen Receptor Modulator with Improved Selectivity Endocrinology, September 1, 2005; 146(9): 3999 - 4008. [Abstract] [Full Text] [PDF]
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D. A. Schreihofer, K. D. Do, and A. M. Schreihofer High-soy diet decreases infarct size after permanent middle cerebral artery occlusion in female rats Am J Physiol Regulatory Integrative Comp Physiol, July 1, 2005; 289(1): R103 - R108. [Abstract] [Full Text] [PDF]
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S. Gingerich and T. L. Krukoff Estrogen Modulates Endothelial and Neuronal Nitric Oxide Synthase Expression via an Estrogen Receptor {beta}-Dependent Mechanism in Hypothalamic Slice Cultures Endocrinology, July 1, 2005; 146(7): 2933 - 2941. [Abstract] [Full Text] [PDF]
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N. R. Miller, T. Jover, H. W. Cohen, R. S. Zukin, and A. M. Etgen Estrogen Can Act via Estrogen Receptor {alpha} and {beta} to Protect Hippocampal Neurons against Global Ischemia-Induced Cell Death Endocrinology, July 1, 2005; 146(7): 3070 - 3079. [Abstract] [Full Text] [PDF]
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D. Rai, A. Frolova, J. Frasor, A. E. Carpenter, and B. S. Katzenellenbogen Distinctive Actions of Membrane-Targeted Versus Nuclear Localized Estrogen Receptors in Breast Cancer Cells Mol. Endocrinol., June 1, 2005; 19(6): 1606 - 1617. [Abstract] [Full Text] [PDF]
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M M. Elloso, K. Phiel, R. A Henderson, H. A Harris, and S. J Adelman Suppression of experimental autoimmune encephalomyelitis using estrogen receptor-selective ligands J. Endocrinol., May 1, 2005; 185(2): 243 - 252. [Abstract] [Full Text] [PDF]
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 G.-S. Lee, H.-J. Kim, Y.-W. Jung, K.-C. Choi, and E.-B. Jeung Estrogen Receptor {alpha} Pathway Is Involved in the Regulation of Calbindin-D9k in the Uterus of Immature Rats Toxicol. Sci., April 1, 2005; 84(2): 270 - 277. [Abstract] [Full Text] [PDF]
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 V. Krishnan, H. A. Bullock, B. C. Yaden, M. Liu, R. J. Barr, C. Montrose-Rafizadeh, K. Chen, J. A. Dodge, and H. U. Bryant The Nongenotropic Synthetic Ligand 4-Estren-3{alpha}17{beta}-diol Is a High-Affinity Genotropic Androgen Receptor Agonist Mol. Pharmacol., March 1, 2005; 67(3): 744 - 748. [Abstract] [Full Text] [PDF]
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C. C. Chadwick, S. Chippari, E. Matelan, L. Borges-Marcucci, A. M. Eckert, J. C. Keith Jr., L. M. Albert, Y. Leathurby, H. A. Harris, R. A. Bhat, et al. Identification of pathway-selective estrogen receptor ligands that inhibit NF-{kappa}B transcriptional activity PNAS, February 15, 2005; 102(7): 2543 - 2548. [Abstract] [Full Text] [PDF]
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T. D. Lund, T. Rovis, W. C. J. Chung, and R. J. Handa Novel Actions of Estrogen Receptor-{beta} on Anxiety-Related Behaviors Endocrinology, February 1, 2005; 146(2): 797 - 807. [Abstract] [Full Text] [PDF]
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 K. M. Egan, J. A. Lawson, S. Fries, B. Koller, D. J. Rader, E. M. Smyth, and G. A. FitzGerald COX-2-Derived Prostacyclin Confers Atheroprotection on Female Mice Science, December 10, 2004; 306(5703): 1954 - 1957. [Abstract] [Full Text] [PDF]
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 M. Zaitseva, D. S. Yue, J. A. Katzenellenbogen, P. A. W. Rogers, and C. E. Gargett Estrogen Receptor-{alpha} Agonists Promote Angiogenesis in Human Myometrial Microvascular Endothelial Cells Reproductive Sciences, December 1, 2004; 11(8): 529 - 535. [Abstract] [PDF]
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V. M. Navarro, J. M. Castellano, R. Fernandez-Fernandez, M. L. Barreiro, J. Roa, J. E. Sanchez-Criado, E. Aguilar, C. Dieguez, L. Pinilla, and M. Tena-Sempere Developmental and Hormonally Regulated Messenger Ribonucleic Acid Expression of KiSS-1 and Its Putative Receptor, GPR54, in Rat Hypothalamus and Potent Luteinizing Hormone-Releasing Activity of KiSS-1 Peptide Endocrinology, October 1, 2004; 145(10): 4565 - 4574. [Abstract] [Full Text] [PDF]
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A.G.B. Hurst, D.W. Goad, M. Mohan, and J.R. Malayer Independent Downstream Gene Expression Profiles in the Presence of Estrogen Receptor {alpha} or {beta} Biol Reprod, October 1, 2004; 71(4): 1252 - 1261. [Abstract] [Full Text] [PDF]
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 H.V.O. Carswell, I.M. Macrae, L. Gallagher, E. Harrop, and K.J. Horsburgh Neuroprotection by a selective estrogen receptor {beta} agonist in a mouse model of global ischemia Am J Physiol Heart Circ Physiol, October 1, 2004; 287(4): H1501 - H1504. [Abstract] [Full Text] [PDF]
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 A. S. Lovegrove, J. Sun, K. A. Gould, D. B. Lubahn, K. S. Korach, and P. H. Lane Estrogen receptor {alpha}-mediated events promote sex-specific diabetic glomerular hypertrophy Am J Physiol Renal Physiol, September 1, 2004; 287(3): F586 - F591. [Abstract] [Full Text] [PDF]
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L. Carta and D. Sassoon Wnt7a Is a Suppressor of Cell Death in the Female Reproductive Tract and Is Required for Postnatal and Estrogen-Mediated Growth Biol Reprod, August 1, 2004; 71(2): 444 - 454. [Abstract] [Full Text] [PDF]
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J.-x. Zhang, D. C. Labaree, G. Mor, and R. B. Hochberg Estrogen to Antiestrogen with a Single Methylene Group Resulting in an Unusual Steroidal Selective Estrogen Receptor Modulator J. Clin. Endocrinol. Metab., July 1, 2004; 89(7): 3527 - 3535. [Abstract] [Full Text] [PDF]
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V. Unfer, M. L. Casini, L. Costabile, M. Mignosa, S. Gerli, and G. C. Di Renzo High Dose of Phytoestrogens Can Reverse the Antiestrogenic Effects of Clomiphene Citrate on the Endometrium in Patients Undergoing Intrauterine Insemination: A Randomized Trial Reproductive Sciences, July 1, 2004; 11(5): 323 - 328. [Abstract] [PDF]
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A. Hillisch, O. Peters, D. Kosemund, G. Muller, A. Walter, B. Schneider, G. Reddersen, W. Elger, and K.-H. Fritzemeier Dissecting Physiological Roles of Estrogen Receptor {alpha} and {beta} with Potent Selective Ligands from Structure-Based Design Mol. Endocrinol., July 1, 2004; 18(7): 1599 - 1609. [Abstract] [Full Text] [PDF]
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F. Stossi, D. H. Barnett, J. Frasor, B. Komm, C. R. Lyttle, and B. S. Katzenellenbogen Transcriptional Profiling of Estrogen-Regulated Gene Expression via Estrogen Receptor (ER) {alpha} or ER{beta} in Human Osteosarcoma Cells: Distinct and Common Target Genes for These Receptors Endocrinology, July 1, 2004; 145(7): 3473 - 3486. [Abstract] [Full Text] [PDF]
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M. B. Hawkins and P. Thomas The Unusual Binding Properties of the Third Distinct Teleost Estrogen Receptor Subtype ER{beta}a Are Accompanied by Highly Conserved Amino Acid Changes in the Ligand Binding Domain Endocrinology, June 1, 2004; 145(6): 2968 - 2977. [Abstract] [Full Text] [PDF]
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F. Barchiesi, E. K. Jackson, B. Imthurn, J. Fingerle, D. G. Gillespie, and R. K. Dubey Differential Regulation of Estrogen Receptor Subtypes {alpha} and {beta} in Human Aortic Smooth Muscle Cells by Oligonucleotides and Estradiol J. Clin. Endocrinol. Metab., May 1, 2004; 89(5): 2373 - 2381. [Abstract] [Full Text] [PDF]
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M. A. Iannone, C. A. Simmons, S. H. Kadwell, D. L. Svoboda, D. E. Vanderwall, S.-J. Deng, T. G. Consler, J. Shearin, J. G. Gray, and K. H. Pearce Correlation between in Vitro Peptide Binding Profiles and Cellular Activities for Estrogen Receptor-Modulating Compounds Mol. Endocrinol., May 1, 2004; 18(5): 1064 - 1081. [Abstract] [Full Text] [PDF]
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C. Hegele-Hartung, P. Siebel, O. Peters, D. Kosemund, G. Muller, A. Hillisch, A. Walter, J. Kraetzschmar, and K.-H. Fritzemeier Impact of isotype-selective estrogen receptor agonists on ovarian function PNAS, April 6, 2004; 101(14): 5129 - 5134. [Abstract] [Full Text] [PDF]
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M. Tena-Sempere, V.M. Navarro, A. Mayen, C. Bellido, and J.E. Sanchez-Criado Regulation of Estrogen Receptor (ER) Isoform Messenger RNA Expression by Different ER Ligands in Female Rat Pituitary Biol Reprod, March 1, 2004; 70(3): 671 - 678. [Abstract] [Full Text] [PDF]
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Y. Ikeda, A. Nagai, M.-A. Ikeda, and S. Hayashi Sexually Dimorphic and Estrogen-Dependent Expression of Estrogen Receptor {beta} in the Ventromedial Hypothalamus during Rat Postnatal Development Endocrinology, November 1, 2003; 144(11): 5098 - 5104. [Abstract] [Full Text] [PDF]
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D. P. McDonnell Mining the Complexities of the Estrogen Signaling Pathways for Novel Therapeutics Endocrinology, October 1, 2003; 144(10): 4237 - 4240. [Full Text] [PDF]
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H. A. Harris, L. M. Albert, Y. Leathurby, M. S. Malamas, R. E. Mewshaw, C. P. Miller, Y. P. Kharode, J. Marzolf, B. S. Komm, R. C. Winneker, et al. Evaluation of an Estrogen Receptor-{beta} Agonist in Animal Models of Human Disease Endocrinology, October 1, 2003; 144(10): 4241 - 4249. [Abstract] [Full Text] [PDF]
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J. Frasor, D. H. Barnett, J. M. Danes, R. Hess, A. F. Parlow, and B. S. Katzenellenbogen Response-Specific and Ligand Dose-Dependent Modulation of Estrogen Receptor (ER) {alpha} Activity by ER{beta} in the Uterus Endocrinology, July 1, 2003; 144(7): 3159 - 3166. [Abstract] [Full Text] [PDF]
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J. Sun, J. Baudry, J. A. Katzenellenbogen, and B. S. Katzenellenbogen Molecular Basis for the Subtype Discrimination of the Estrogen Receptor-{beta}-Selective Ligand, Diarylpropionitrile Mol. Endocrinol., February 1, 2003; 17(2): 247 - 258. [Abstract] [Full Text] [PDF]
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