| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Regular Article |
a Departments of Medical Nutrition and
b Biosciences, Karolinska Institute, NOVUM, S-14186 Huddinge, Sweden
c Department of Obstetrics and Gynaecology, Huddinge University Hospital, Karolinska Institute, S-14186 Huddinge, Sweden
d Department of Anatomy, Institute of Biomedicine, University of Turku, FIN-20520 Turku, Finland
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
is not detectable in granulosa cells in the mouse ovary. In ERß knockout (BERKO) mice, there is abnormal follicular development and very reduced fertility. At 3 wk of age, no significant morphologic differences were discernable between wild type (WT) and BERKO mouse ovaries, but by 5 mo of age, atretic follicles were abundant in BERKO mice and there were very few healthy late antral follicles or corpora lutea. At 2 yr of age, unlike the ovaries of their WT littermates, BERKO mouse ovaries were devoid of healthy follicles but had numerous large, foamy lipid-filled stromal cells. The late antral and atretic follicles in BERKO mice were characterized by a high level of expression of the androgen receptor (AR) and IGF-1 receptor. These proteins were abundantly expressed in granulosa cells of preantral and early antral follicles in both genotypes, but their expression was extinguished in late antral follicles of WT mice. Healthy late antral follicles and corpora lutea were restored in BERKO ovaries after 15 days of treatment of mice with the antiandrogen flutamide. The results suggest that in the absence of ERß there was a loss of regulation of AR. Because androgens enhance recruitment of primordial follicles into the growth pool and cause atresia of late antral follicles, the inappropriately high level of AR probably is related to the follicular atresia and to the early exhaustion of follicles in BERKO mice.
androgen receptor, courpus luteum, estradiol receptor, follicular development, granulosa cells, ovary, ovulation
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Follicular atresia is hormonally controlled apoptosis [2–4]. FSH and estrogen are essential for follicles to escape atresia and reach the preovulatory follicle stage [2, 6–8]. The major role of FSH in this process is to elevate estrogen levels through stimulation of aromatase expression in granulosa cells [4]. In addition, several ovarian growth factors such as insulinlike growth factor-1 (IGF-1), epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), and the cytokine interleukin-1ß participate in prevention of follicular apoptosis [2–4].
In the rodent ovary, estrogen receptor (ER)- and ERß are expressed in distinctly different cell types. ERß is expressed in the nuclei of granulosa cells of primary, secondary, and mature follicles but not in the germinal epithelium or in thecal, luteal, or interstitial cells. ER protein is not detectable in granulosa cells but is found in the germinal epithelium and in interstitial and thecal cells [9–13]. The distinct patterns of distribution of ER and ERß in the ovary suggest that the 2 ERs mediate different aspects of estrogen action in this organ. ER knockout (ERKO) mice are infertile, with follicular development arrested at the preovulatory stage [14, 15]. The primary cause of this defect is an elevated level of serum LH [16]. ERß knockout (BERKO) mice exhibit abnormal follicular maturation and have very few corpora lutea. Superovulation experiments indicate that the reduction in fertility in BERKO mice is the result of reduced ovarian efficiency [9]. The ovarian phenotype of aromatase knockout (ARKO) mice, in which no estrogen is produced [17], is different from that of either ERKO or BERKO mice, indicating that ER and ERß have distinct roles in follicular maturation.
Androgens serve 2 distinct functions in the ovary. As substrates of P450arom, they are precursors of estrogen, but they also act via androgen receptors (ARs) at 2 stages in the development of follicles. During initial recruitment, androgen is a stimulating factor increasing the recruitment of primordial follicles into the growing pool of developing follicles. Treatment of female mice with androgens increases the number of primary follicles in the ovary and the expression of IGF-1 receptor (IGF-1R) mRNA in primordial follicle oocytes [18, 19]. During cyclic recruitment, androgens promote atresia [1–3]. In the present study, we investigated the ovarian phenotypes of BERKO mice from 3 wk to 2 yr of age and gathered evidence for a relationship between the extensive follicular atresia in these mice and a defect in regulation of AR expression.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
The ovaries of wild-type (WT) mice (5 mice 3 wk old, 28 mice 5 mo old, 13 mice 2 yr old) and BERKO mice (5 mice 3 wk old, 25 mice 5 mo old, 15 mice 2 yr old) were removed for morphologic observation. Immunohistochemical studies were performed on 14 WT and 15 BERKO mice 5 mo of age. Animals were used under the Guidelines for Care and Use of Experimental Animals issued by Stockholm Södra Djurförsöksetiska Nämnd. Animals were maintained under standard environmental conditions, with free access to food and water. WT and BERKO mice were bred from heterozygous male and female mice. Genotyping by polymerase chain reaction was performed on DNA isolated from tails of 2-wk-old mice [20]. Mice were killed by CO2 asphyxiation. The ovaries were immediately removed, and 1 ovary of each mouse was frozen in liquid nitrogen and the other was fixed in 4% paraformaldehyde overnight and routinely embedded in paraffin.
Antibodies
AR rabbit polyclonal IgG (N20), IGF-1R rabbit polyclonal IgG (C20), and cyclin D2 rabbit polyclonal IgG (M20) were obtained from Santa Cruz Biotechnology (Santa Cruz, CA). ERß rabbit polyclonal IgG (C-terminal) was purchased from Zymed Laboratories (South San Francisco, CA). Biotinylated goat anti-rabbit IgG and avidin-biotin complex (ABC) kits were obtained from Vector Laboratories (Burlingame, CA).
Morphologic Classification of Growing Follicles
Sections were taken at intervals of 50 µm, and 5-µm paraffin-embedded sections were mounted on slides (SuperFrost, Braunschweig, Germany). Routine hematoxylin and eosin staining was performed for histologic examination under a light microscope. The number of follicles in 10 sections from each mouse ovary was evaluated by 2 independent investigators. If the assessments were similar, the values were averaged. If there were big differences between evaluations, the sections were examined again. The area of ovary was measured with Easy Image Mating (version 1.66; Bergstr
m Instrument AB, Solna, Sweden). The abundance of each type of follicle was normalized by the area of ovary in the sections [21]. Follicle types in ovarian cross-sections were defined as follows. Primary follicles comprised an oocyte surrounded by a single layer of cuboidal granulosa cells. Preantral follicles comprised an oocyte surrounded by 2 or more layers of granulosa cells with no antrum. Antral follicles were distinguished by the presence of an antrum within the granulosa cell layers enclosing the oocyte. Follicles were determined to be atretic if they displayed 2 or more of the following criteria within a single cross-section: more than 2 pyknotic nuclei, granulosa cells within the antral cavity, granulosa cells pulling away from the basement membrane, or uneven layers of granulosa cells [22].
Detection of Lipid Droplet by Oil Red O Staining
Oil red O (500 mg; Sigma Chemical Co., St. Louis, MO) was dissolved in 100 ml of 99% isopropanol (Merck, Darmstadt, Germany) as storage solution. The working solution was prepared by filtration after mixing 6 ml of storage solution with 4 ml of double-distilled H2O and was used within 1 h. The 8-µm-thick cryostat sections were placed on poly-L-lysine (Sigma)-coated slides, stained with oil red O for 2 min, and lightly counterstained with hematoxylin [23].
Immunohistochemistry
Sections were subjected to a microwave antigen retrieval technique by boiling in 10 mM citrate buffer (pH 6.0) in a microwave oven for 30 min. The cooled sections were incubated in 1% H2O2 for 30 min to quench endogenous peroxidase and then incubated with 1% Triton X-100 in PBS for 10 min. To block the nonspecific binding of secondary antibodies, sections were incubated in normal serum prepared from the host of secondary antibodies (goat serum, Sigma) for 1 h at 4°C. Sections were then incubated with the following specific rabbit antisera: anti-AR (1:100), anti-IGF-1R (1:500), anti-cyclin D2 (1:500), or anti-ERß (1:500) in 3% BSA overnight at 4°C. Negative controls were incubated with only 3% BSA without primary antibody. The ABC method was used to visualize the signal according to the manual provided by the manufacturer (Vector). The sections were incubated in biotinylated goat anti-rabbit immunoglobulin (1:200 dilution) for 2 h at room temperature, followed by washing with PBS and incubation in avidin-biotin-horseradish peroxidase for 1 h. After thorough washing in PBS, sections were developed with 3,3'-diaminobenzidine tetra-hydrochloride substrate (DAKO, Carpinteria, CA) and slightly counterstained with Mayer hematoxylin, dehydrated through an ethanol series and xylene, and mounted. Staining for AR and IGF-1 was evaluated by 3 persons according to the intensity of the brown color: +, weak staining; ++, moderate staining; and +++, strong staining. The scores for the 3 observations for each sample were averaged before analysis.
Treatment with Flutamide
WT and BERKO mice (8 in each group: 3 mice 8 wk old and 5 mice 5 mo old) that were matched littermates were treated with s.c. injections of flutamide (Sigma), 3 mg kg-1 day-1, for 15 consecutive days (4 estrous cycles). Flutamide was dissolved in Intralipid (Pharmacia & Upjohn, Stockholm, Sweden). The WT and BERKO age-matched control animals (8 mice in each group) received the same amount of vehicle. Both ovaries were taken from each mouse and were fixed in 4% paraformaldehyde overnight and routinely embedded in paraffin. Sections were cut at 50 µm and stained with haematoxylin and eosin. The total number of corpora lutea in each ovary was evaluated by counting corpora lutea in each section.
Statistical Analysis
The statistical differences between WT and BERKO mice were analyzed using the Student t-test and the chi-square test with SPSS 8.0 software (SPSS, Chicago, IL). Data are presented as means ± SD. Differences were considered significant at P < 0.05.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Ovaries of 5 WT and 5 BERKO mice at 3 wk of age were compared. There was no significant difference between the WT and BERKO ovaries in abundance of the follicles. In both genotypes, there were 16–21 preantral follicles and 1 or 2 early antral follicles per section. No atretic follicles could be identified at this time (Fig. 1, A and D).
|
Morphology of 5-Mo-Old Mouse Ovaries
At 5 mo of age, the most obvious difference was the almost complete lack of corpora lutea in BERKO mice. Of the 28 ovaries examined from WT mice, there were 1–7 corpora lutea in every section (Fig. 1B). However, of the 25 BERKO mice examined, corpora lutea were seen in only 7 ovaries and in these, there were only 1 or 2 corpora lutea in each section (Fig. 1E). Quantitative analysis of these differences is illustrated in Table 1. The mean (±SD) number of corpora lutea was 2.8 ± 1.0/mm2 in WT mice and 0.2 ± 0.2/mm2 in BERKO mice (P < 0.001).
|
The second striking feature of the BERKO ovaries was the abundance of atretic follicles and the paucity of Graafian follicles (Fig. 1, E and F). There was no significant difference in number of primary or preantral follicles between WT and BERKO mice. The number of healthy antral follicles (Fig. 1C) per mm2 in the sections was 2.7 ± 0.7 in WT and 1.5 ± 0.4 in BERKO mice (P < 0.01), and the number of atretic follicles (Fig. 1F) was 1.5 ± 1.3 in WT and 5.2 ± 2.2 in BERKO mice (P < 0.001). Of the population of follicles at the antral stage, 72.9% were atretic in BERKO mice and 38.0% were atretic in WT mice (P < 0.01).
Morphology of 2-Yr-Old Mouse Ovaries
At 2 yr of age, BERKO mice ovaries (Fig. 1J) were smaller than those of their WT littermates, had fewer growing follicles, and showed a marked absence of corpora lutea (Fig. 1G). Of the 15 BERKO mice examined, no corpora lutea were found in any ovaries, whereas in WT mice, corpora lutea were present in the ovaries of 10 of the 13 mice examined (P < 0.05). Follicles at various stages were present in all 13 of the WT mice, but follicles were seen in only 6 of the 15 BERKO mice, and none of these 6 BERKO mice had healthy antral follicles (P < 0.001). Atretic follicles accounted for 83% of total follicles in BERKO mice and 31% of follicles in WT mice (P < 0.001).
In the stroma of 2-yr-old BERKO ovaries, large round foamy stromal cells were abundant (Fig. 1K). These cells were only occasionally found in WT ovaries (Fig. 1H). At 2 yr of age, foamy cells accounted for more than 50% of the stromal volume in 13 of 15 BERKO mice compared with 4 of 13 WT mice (P < 0.05). At 5 mo of age, foamy stromal cells were present in 9 of the 25 BERKO ovaries but none were detectable in any of the 28 WT ovaries (P < 0.01).
To determine whether these foamy cells represent a population in which there was cholesteryl ester accumulation, cryostat sections were stained with oil red O, a dye that detects cholesteryl ester droplets in stromal cells [23]. A cross-section through a WT ovary (Fig. 1I) showed oil red O staining as expected in the thecal cells and stromal cells but none in the granulosa cells. Lipid droplets were large and abundant and filled the cytoplasm in the stromal cells of the BERKO ovaries (Fig. 1L). There was very little oil red O staining in the epithelium.
ERß Localization
In ovaries of WT mice, nuclear ERß staining was observed in granulosa cells of follicles at different stages. The intensity of ERß immunoreactivity was stronger in antral follicles than in primary follicles (Fig. 2A). Thecal cells, interstitial cells, oocytes, and germinal epithelium showed no nuclear ERß immunoreactivity. In BERKO mice (Fig. 2B), no ERß expression was detected.
|
AR Localization
In WT mice (Fig. 2, C and E), AR nuclear staining was most abundant in granulosa cells of healthy preantral and early antral follicles. AR staining was markedly reduced or completely extinguished in healthy late antral and preovulatory follicles. Atretic follicles maintained a high level of AR expression. As in WT mice, AR nuclear staining in preantral and early antral follicles was stronger than that in primary follicles in BERKO mice (Fig. 2, D and F). However, unlike the situation in the WT mice, there was no reduction in the level of AR in late antral follicles, and most of these follicles were atretic (Table 2). Thecal cells, stromal cells, and corpora lutea in both WT (Fig. 2, C and E) and BERKO (Fig. 2, D and F) mice had weak to moderate AR expression without significant differences between the 2 genotypes.
|
IGF-1R Expression
IGF-1R was detected in granulosa, thecal, stromal, and germinal epithelial cells in both WT (Fig. 2G) and BERKO (Fig. 2H) mouse ovaries, with strongest staining in granulosa cells. There was no obvious difference in the staining in granulosa cells of primary and preantral follicles, thecal cells, and stromal cells between the 2 genotypes. In atretic follicles, the staining of IGF-1R was reduced in WT mice, but it remained strong in BERKO mouse ovaries. Semiquantitative analysis of the IGF-1R staining revealed that the difference was significant. Of the atretic follicles examined in 12 WT mice, 44% were scored as +, 48% as ++, and 7% as +++. Of the atretic follicles examined in 10 BERKO mice, 5% were +, 46% were ++, and 48% were +++ (P < 0.05).
Flutamide Reversal of BERKO Mouse Ovary Phenotype
After 15 consecutive days of treatment, there were no marked changes in follicle abundance or in the appearance of corpora lutea between age-matched, vehicle-treated, and flutamide-treated WT mice (Fig. 3, A and B). In BERKO mice treated with flutamide, late antral follicles and 1–5 corpora lutea were found in each ovary in all of 3 BERKO mice 8 wk of age and in 3 of 5 BERKO mice 5 mo of age (Fig. 3D). In 2 of the 5-mo-old BERKO mice, no corpora lutea were detected after flutamide treatment. No corpora lutea were seen in any of the 8 BERKO mice that received vehicle treatment only (Fig. 3C).
|
The proliferation marker cyclin D2 [5] was used to evaluate the proliferation pattern of granulosa cells after flutamide treatment. In both WT and BERKO mice that received vehicle only, cyclin D2 was expressed in healthy growing follicles but not in the atretic follicles, as expected (Fig. 3, E and G). The cumulus oophorus of the late antral follicles in WT mice still expressed high levels of cyclin D2 (Fig. 3E). After flutamide treatment, most of the granulosa cells in healthy late antral follicles in both WT and BERKO mice expressed high levels of cyclin D2 (Fig. 3, F and H).
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Experimental evidence [2–4] suggests that initial recruitment of follicles is not stimulated by FSH or estrogen. In both WT and BERKO prepubertal mice, follicles developed normally to the preantral stage. At the age of 5 mo, there was no significant difference in the number of primary or preantral follicles between WT and BERKO mice. However, there was an increase in the total number of follicles (healthy and atretic) in the ovaries of BERKO mice at 5 mo of age and an earlier exhaustion of follicles in 2-yr-old BERKO ovaries. This finding suggests that there was enhanced follicular recruitment in BERKO mice.
There was no reduction in recruitment of follicles into the growth pool and there were less healthy antral follicles, more atretic follicles, and very few corpora lutea in mature BERKO ovaries, indicating that the late antral follicle was the stage at which arrest of maturation occured. Cyclic recruitment and survival of follicles depend on pituitary hormones, estrogen, IGF-1, EGF, bFGF, interleukin-1ß, etc. [1–4, 24]. Because FSH and LH levels are normal in BERKO females [13], it is unlikely that FSH deficiency is responsible for the increased follicular atresia in BERKO females. Superovulation of BERKO mice with gonadotropins did not completely restore normal ovulation, further evidence that FSH deficiency is not the cause of the defect [9].
IGF-1, via its receptor on the granulosa cells, enhances the gonadotropin-induced stimulation of aromatase activity, promotes granulosa cell differentiation, and suppresses follicular apoptosis [2, 25]. In IGF-1 knockout mice, there is arrest of follicular development at the antral stage, absence of corpora lutea, low serum estradiol concentration, and failure of gonadotropin administration to reverse the phenotype [22]. In the present study, both WT and BERKO follicles expressed IGF-1R. In BERKO mice, there was an even higher expression of IGF-1R in atretic follicles. IGF-1R, therefore, does not appear to be a limiting factor in follicular development and lack of this factor cannot explain the increased follicular atresia in BERKO mouse ovaries.
Androgens are also important for follicular development. In addition to being precursors of estrogen, androgens via AR enhance FSH-stimulated follicular differentiation and inhibit follicular maturation. Androgens, therefore, either enhance or inhibit granulosa cell steroidogenesis depending on the stage of follicular development. In primary and preantral follicles, there is low P450arom activity but high levels of AR expression. Androstenedione diffuses from thecal cells into granulosa cells, where it is converted to testosterone. Testosterone then activates AR, leading to increased expression of IGF-1R, stimulation of granulosa cell proliferation, and enhancement of the effects of FSH on induction of P450arom [18]. As the environment of the oocyte changes from androgen dominant to estrogen dominant, follicles develop from preantral to antral and preovulatory stages. An increase in circulating estradiol produced by the growing follicles exerts a positive feedback regulation on the hypothalamus-pituitary axis and, prior to ovulation, induces the LH surge, a paradoxical estrogen-induced increase in LH secretion that is critical for initiation of ovulation [26, 27]. Increased plasma levels of estrogen also cause a reduction in FSH secretion from the pituitary gland, a fall in the circulating level of FSH, and consequently a reduction in FSH-dependent aromatase gene expression. In less well-developed follicles, this reduction in aromatase activity results in a reconversion to an androgenic microenvironment and to initiation of the process of atresia [26, 27].
In granulosa cells, FSH induces AR in primary but not mature follicles [28]. As follicles mature, there is a concomitant increase in estrogen levels in follicular fluid and a decrease in AR levels in granulosa cells. Because granulosa cells express ERß but not ER [29], we hypothesize that AR expression in the granulosa cells of late antral follicles is repressed by the activation of ERß, and this repression changes the microenvironment of the oocyte from androgen dominant to estrogen dominant. This change is critical for survival of the follicle [2, 22]. If AR expression remains high, antral follicles cannot undergo conversion to estrogen dominance and their fate is atresia [27]. Such a regulatory mechanism would be compatible with the overexpression of AR in atretic follicles in BERKO mice. A working hypothesis of the role of ERß in follicular maturation, based on current concepts and the present data, is presented in Figure 4.
|
Flutamide is a nonsteroidal antiandrogen of known efficacy in hirsutism in polycystic ovarian syndrome (PCOS) patients who have hyperandrogenism [30, 31]. In young women with PCOS, ovulation was restored after 6 mo of treatment with flutamide [32]. In the present study, after 15 days (4 estrous cycles) of treatment with flutamide, healthy late antral follicles appeared in BERKO mice. In addition, all 3 of the 8-wk-old mice and three of five 5-mo-old BERKO mice had corpora lutea after treatment. Healthy late antral follicles and corpora lutea were very rare in the ovaries of untreated BERKO mice. Cyclin D2, a proliferation marker for granulosa cells [5], was expressed in the healthy growing follicles in both WT and BERKO mice. The cumulus oophorus of the preovulatory follicles in WT mice still expressed cyclin D2. However, cyclin D2 was not expressed in atretic follicles in either WT or BERKO mice. In BERKO mouse ovaries, most of the late antral follicles did not express cyclin D2. This finding is consistent with the morphologic observation of an increased number of atretic follicles seen at late antral stage. Treatment with flutamide increased the number of healthy follicles in BERKO mice, with a concomitant increase in the expression of cyclin D2 in antral follicles. These results show that upon administration of flutamide, proliferation of granulosa cells was stimulated and follicles developed to the late antral stage. These results support our notion that it is the lack of downregulation of AR in BERKO mouse ovary that results in the increased follicular atresia. Previous studies [33] have shown that flutamide does not change uterine or ovarian weight, alter serum estradiol or progesterone levels, or have progesterone effects. We cannot, however, rule out the possibility that the effect of flutamide in the BERKO ovary was mediated by its action outside of the ovary.
In addition to abnormal follicular maturation, there were also marked morphologic changes in the stroma of adult BERKO mouse ovaries. With increasing age, numerous foamy cells appeared in the stroma of BERKO ovaries. At 2 yr of age, most of the stromal cells in BERKO mice were replaced by these foamy cells but these cells were only occasionally seen in WT mice. Oil red O staining revealed an abnormal accumulation of lipid droplets in the cytoplasm of the stromal cells in BERKO mouse ovaries. The reason for the abundance of these cells in BERKO mouse ovaries remains to be determined.
Ovaries from BERKO mice are characterized by abnormal follicular maturation, increased atresia, paucity of corpora lutea, and early follicle exhaustion. Although it might not be the only cause, the dysregulation of AR appears to play an important role in the BERKO ovarian phenotype.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
1 Supported by grants from KaroBio AB, the Swedish Cancer Fund, and the Swedish Medical Research Council. S.S. has a fellowship from the Wenner-Gren Foundation and a research grant from the Scandinavia-Japan Sasakawa Foundation. 
2 Correspondence: Jan-Åke Gustafsson, Department of Medical Nutrition, Karolinska Institute, NOVUM, S-14186, Huddinge, Sweden. FAX: 46 8 779 87 95; jan-ake.gustafsson{at}mednut.ki.se
Accepted: August 16, 2001.
Received: April 11, 2001.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
and ß mRNA in the female reproductive organ of rats as revealed by in situ hybridization. J Endocrinol 2000; 165:59-66[Abstract]-reductase inhibitors on estrus duration in the cycling female rat. Physiol Behav 1983; 30::519-524[CrossRef][Medline]This article has been cited by other articles:
|
|
 |
|
  B. R. Bhavnani, S.-P. Tam, and X. Lu Structure Activity Relationships and Differential Interactions and Functional Activity of Various Equine Estrogens Mediated via Estrogen Receptors (ERs) ER{alpha} and ER{beta} Endocrinology, October 1, 2008; 149(10): 4857 - 4870. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. da Silva Faria, F. de Bittencourt Brasil, F. J B Sampaio, and C. da Fonte Ramos Maternal malnutrition during lactation alters the folliculogenesis and gonadotropins and estrogen isoforms ovarian receptors in the offspring at puberty J. Endocrinol., September 1, 2008; 198(3): 625 - 634. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L.-M. Chen, R.-S. Wang, Y.-F. Lee, N.-C. Liu, Y.-J. Chang, C.-C. Wu, S. Xie, Y.-C. Hung, and C. Chang Subfertility with Defective Folliculogenesis in Female Mice Lacking Testicular Orphan Nuclear Receptor 4 Mol. Endocrinol., April 1, 2008; 22(4): 858 - 867. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K.A. Walters, C.M. Allan, and D.J. Handelsman Androgen Actions and the Ovary Biol Reprod, March 1, 2008; 78(3): 380 - 389. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. A. Pangas, C. J. Jorgez, M. Tran, J. Agno, X. Li, C. W. Brown, T. R. Kumar, and M. M. Matzuk Intraovarian Activins Are Required for Female Fertility Mol. Endocrinol., October 1, 2007; 21(10): 2458 - 2471. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Stettner, S. Kaulfuss, P. Burfeind, S. Schweyer, A. Strauss, R.-H. Ringert, and P. Thelen The relevance of estrogen receptor-{beta} expression to the antiproliferative effects observed with histone deacetylase inhibitors and phytoestrogens in prostate cancer treatment Mol. Cancer Ther., October 1, 2007; 6(10): 2626 - 2633. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. A. Walters, C. M. Allan, M. Jimenez, P. R. Lim, R. A. Davey, J. D. Zajac, P. Illingworth, and D. J. Handelsman Female Mice Haploinsufficient for an Inactivated Androgen Receptor (AR) Exhibit Age-Dependent Defects That Resemble the AR Null Phenotype of Dysfunctional Late Follicle Development, Ovulation, and Fertility Endocrinology, August 1, 2007; 148(8): 3674 - 3684. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Tiwari-Pandey, Y. Yang, J. Aravindakshan, and M.R. Sairam Normalization of hormonal imbalances, ovarian follicular dynamics and metabolic effects in follitrophin receptor knockout mice Mol. Hum. Reprod., May 1, 2007; 13(5): 287 - 297. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Smitz, A.N. Andersen, P. Devroey, J.-C. Arce, and for the MERIT Group Endocrine profile in serum and follicular fluid differs after ovarian stimulation with HP-hMG or recombinant FSH in IVF patients Hum. Reprod., March 1, 2007; 22(3): 676 - 687. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Inzunza, A. Morani, G. Cheng, M. Warner, J. Hreinsson, J.-A. Gustafsson, and O. Hovatta Ovarian wedge resection restores fertility in estrogen receptor beta knockout (ERbeta-/-) mice PNAS, January 9, 2007; 104(2): 600 - 605. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Poutanen Toward understanding the endocrine regulation of gonadal somatic cells. Endocrinology, August 1, 2006; 147(8): 3662 - 3665. [Full Text] [PDF]
|
|
|
|
|
|
S. A. Pangas, X. Li, E. J. Robertson, and M. M. Matzuk Premature Luteinization and Cumulus Cell Defects in Ovarian-Specific Smad4 Knockout Mice Mol. Endocrinol., June 1, 2006; 20(6): 1406 - 1422. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Morani, R. P. A. Barros, O. Imamov, K. Hultenby, A. Arner, M. Warner, and J.-A. Gustafsson Lung dysfunction causes systemic hypoxia in estrogen receptor beta knockout (ERbeta-/-) mice PNAS, May 2, 2006; 103(18): 7165 - 7169. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. Imamov, G.-J. Shim, M. Warner, and J.-A. Gustafsson Estrogen Receptor beta in Health and Disease Biol Reprod, November 1, 2005; 73(5): 866 - 871. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. F. Couse, M. M. Yates, B. J. Deroo, and K. S. Korach Estrogen Receptor-{beta} Is Critical to Granulosa Cell Differentiation and the Ovulatory Response to Gonadotropins Endocrinology, August 1, 2005; 146(8): 3247 - 3262. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. M. A. Emmen, J. F. Couse, S. A. Elmore, M. M. Yates, G. E. Kissling, and K. S. Korach In Vitro Growth and Ovulation of Follicles from Ovaries of Estrogen Receptor (ER){alpha} and ER{beta} Null Mice Indicate a Role for ER{beta} in Follicular Maturation Endocrinology, June 1, 2005; 146(6): 2817 - 2826. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. F. Koehler, L. A. Helguero, L.-A. Haldosen, M. Warner, and J.-A. Gustafsson Reflections on the Discovery and Significance of Estrogen Receptor {beta} Endocr. Rev., May 1, 2005; 26(3): 465 - 478. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Omoto, R. Lathe, M. Warner, and J.-A. Gustafsson Early onset of puberty and early ovarian failure in CYP7B1 knockout mice PNAS, February 22, 2005; 102(8): 2814 - 2819. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y.-C. Hu, P.-H. Wang, S. Yeh, R.-S. Wang, C. Xie, Q. Xu, X. Zhou, H.-T. Chao, M.-Y. Tsai, and C. Chang Subfertility and defective folliculogenesis in female mice lacking androgen receptor PNAS, August 3, 2004; 101(31): 11209 - 11214. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T.E. Hickey, D.L. Marrocco, R.B. Gilchrist, R.J. Norman, and D.T. Armstrong Interactions Between Androgen and Growth Factors in Granulosa Cell Subtypes of Porcine Antral Follicles Biol Reprod, July 1, 2004; 71(1): 45 - 52. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. Imamov, A. Morani, G.-J. Shim, Y. Omoto, C. Thulin-Andersson, M. Warner, and J.-A. Gustafsson Estrogen receptor {beta} regulates epithelial cellular differentiation in the mouse ventral prostate PNAS, June 22, 2004; 101(25): 9375 - 9380. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
