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Ovary |
-Inhibin Expression in the Fetal Ovary of Estrogen-Suppressed Baboons Is Associated with Impaired Fetal Ovarian Folliculogenesis1
Department of Physiological Sciences,3 Eastern Virginia Medical School, Norfolk, Virginia 23501-1980
Departments of Obstetrics/Gynecology/Reproductive Sciences and Physiology, The Center for Studies in Reproduction,4 University of Maryland School of Medicine, Baltimore, Maryland 21201
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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and ß, the mechanism(s) of estrogen action remains to be determined. It is well established that inhibin and activins function as autocrine/paracrine factors that impact adult ovarian function. However, our understanding of the expression of these factors in the primate fetal ovary is incomplete. Therefore, we determined the expression of -inhibin, activin ßA, activin ßB, and activin receptors in fetal ovaries obtained at mid and late gestation from untreated baboons and at late gestation from animals in which fetal estrogen levels were reduced by >95% by maternal administration of the aromatase inhibitor CGS 20267 or restored to 30% of normal by treatment with CGS 20267 and estradiol benzoate. Immunocytochemical expression of -inhibin was minimal to nondetectable in fetal ovaries from untreated baboons. In contrast, in baboons depleted of estrogen, -inhibin was abundantly expressed in pregranulosa cells of interfollicular nests and granulosa cells of primordial follicles. Thus, the number (mean ± SEM) per 0.08 mm2 of fetal ovarian cells expressing -inhibin, determined by image analysis, was similar at mid and late gestation and increased approximately 8-fold (P < 0.01) near term in baboons treated with CGS 20267 and was restored (P < 0.01) to normal in baboons treated with CGS 20267 plus estradiol. Activin ßA was detected in oocytes and pregranulosa cells at midgestation and in oocytes and granulosa cells of primordial follicles at late gestation. Activin ßB was also expressed in pregranulosa cells and granulosa cells at mid and late gestation, respectively, but was not detected in oocytes. Neither the pattern nor the apparent level of expression of activin ßA or ßB were altered in fetal ovaries of baboons treated with CGS 20267 or CGS 20267 and estrogen. Activin receptors IA, IB, IIA, and IIB were detected by Western blot analysis in fetal ovaries at mid and late gestation, and expression was not altered by treatment with CGS 20267 or CGS 20267 and estrogen. Activin receptors IB and IIA were localized to oocytes and pregranulosa cells at midgestation and to granulosa cells and oocytes of primordial follicles at late gestation. Thus, the decrease in the number of follicles in the primate fetal ovary of baboons deprived of estrogen in utero was associated with increased expression of -inhibin. Therefore, we propose that estrogen regulates fetal ovarian follicular development by controlling -inhibin expression and, thus, the intraovarian inhibin:activin ratio.
activin, estradiol, follicular development, inhibin, ovary
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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and ß [2] and placental estrogen secretion into the fetus increased with advancing gestation [3], we have proposed that estrogen regulates fetal ovarian folliculogenesis in the primate [1]. However, the mechanism(s) by which estrogen regulates development of the primate fetal ovary remains to be determined.
It is well established that inhibin and activin function as intragonadal autocrine/paracrine factors that regulate adult ovarian function [4, 5]. Inhibins are glycoproteins that belong to the transforming growth factor ß superfamily and are composed of an subunit and one of two ß subunits giving rise to two functional glycoproteins, inhibin A (ßA) and inhibin B (ßB). Activins are homodimers or heterodimers of either of the two ß subunits (ßA:ßA, activin A; ßB:ßB, activin B; or ßA:ßB; activin AB) [6, 7]. At the cell membrane, activins interact with serine/threonine kinase receptors classed as type I or type II. Because a specific receptor(s) for inhibin remains to be characterized [8], it would appear that inhibins act either by binding the ß subunit to produce an inactive dimer (i.e., inhibin) and/or by binding to the activin receptor, thereby preventing the action of activin [6, 7].
In mice, activin subunit gene disruption resulted in females with impaired reproductive ability, whereas mice deficient in the -inhibin subunit gene developed gonadal stromal tumors (i.e., unopposed action on activin-stimulated proliferation) and were infertile [9, 10]. Moreover, overexpression of the -inhibin subunit gene in mice caused several ovarian pathologies, including development of ovarian cysts [11, 12]. Based on these observations and the findings that activin promoted in vitro development of theca-free granulosa-oocyte complexes isolated from immature rat ovaries [13], stimulated growth of rat preantral follicles and oocyte maturation [14], and induced proliferation of human luteinized granulosa cells [15], it has been suggested that ovarian function is modulated by the relative levels of inhibin and activin [11].
It has also been demonstrated that the antioncogenic transcription factor Wilms tumor-1 (Wt-1) is expressed in the human fetal ovary and kidney [16–18] and is critical for postnatal ovarian development in the rodent [6, 9, 19]. In adult murine, monkey, and human ovaries, Wt-1 is expressed in granulosa cells of preantral follicles and is regulated in a maturation- and gonadotroph-dependent manner [20]. Mutations of Wt-1 have been linked to sex cord-stromal tumors of the ovary in humans [18], and Wt-1 has been shown to suppress the expression of several growth factors (e.g., -inhibin, transforming growth factor ß) [21]. Therefore, it has been suggested that Wt-1 may be a key factor in maintaining a balance between factors promoting follicular development and regression as well as promoting the preservation of ovarian follicles in a quiescent state throughout development and reproductive life [17].
In contrast to the extensive literature on the expression and roles of inhibins, activins, and Wt-1 on adult ovarian function, our understanding of the developmental pattern and site of expression of these factors as well as the activin receptors in the primate fetal ovary is incomplete [15], and to our knowledge, no studies have elucidated regulation in utero [18, 22]. Therefore, in the present study, we determined the temporal pattern and site of expression of activins, the activin receptors, -inhibin, and Wt-1 in the baboon fetal ovary and whether expression was altered in animals deprived of estrogen during the second half of gestation.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Fetal ovaries were obtained on Days 100 (n = 6) and 165–170 (n = 6) of gestation from untreated baboons (Papio anubis) and on Days 165–170 from animals treated daily beginning on Day 100 of gestation (term = Day 184) with the aromatase inhibitor CGS 20267 (Letrozole; 4,4-[1,2,3-triazol-1yl-methylene] bis-benzonitrite; Norvartis Pharma AG, Basel, Switzerland) administered s.c. (115 µg kg body weight [BW]-1 day- per 0.05 ml sesame oil, n = 6) to the mother or treated with CGS 20267 (115 µg kg BW-1 day-1) plus estradiol benzoate at doses (50–175 µg kg BW-1 day-1 per 0.1 ml sesame oil, n = 5) designed to replicate the normal pattern of serum estradiol essentially as described previously [1]. Briefly, baboons were sedated with ketamine, anesthetized with isoflurane, the placenta and the fetus delivered by cesarean section, and the fetus killed with an overdose of sodium pentobarbital. Fetal ovaries were excised, trimmed of fat, and weighed. One ovary was then fixed in 10% buffered formalin and paraffin-embedded for subsequent histologic/immunocytochemical analyses as previously described [1, 2], and the other ovary was stored in liquid nitrogen for Western blot analysis. Blood samples (3–5 ml) were also obtained at 1- to 4-day intervals between Days 85 and 165 of gestation via a maternal saphenous vein after sedation with an i.m. injection of ketamine-HCl (10 mg/kg BW; Parke-Davis, Detroit, MI) and from the umbilical vein before delivery of the fetus between Days 165 and 170 of gestation.
RIA of Estradiol
Maternal and umbilical venous serum estradiol levels have been reported previously [1, 23], and they have been recompiled for presentation in the present study. Estradiol was determined by RIA using an automated chemiluminescent immunoassay system (Immulite; Diagnostic Products Corp., Los Angeles, CA) as described previously [24].
Histology and Immunocytochemistry
Fetal ovarian histology and immunocytochemical expression of Wt-1, -inhibin, activin ßA, activin ßB, and activin receptors IB and IIA were determined essentially as described previously [2, 25]. Briefly, representative sections (thickness, 4 µm) of paraffin-embedded fetal ovaries were mounted onto Superfrost microscope slides (Fisher Scientific Co., Arlington, VA) and then heat-fixed and endogenous peroxidase-blocked following incubation for 10 or 30 min with 0.4% or 1.0% (activin ßB) H2O2 in methanol, respectively. Except for analysis of -inhibin, sections were microwaved in 10 mM sodium citrate buffer (pH 6; Sigma Chemical Co., St. Louis, MO) at a power-level setting of 9 (General Electric Model JE 1540 oven; maximum power, 900 W) for 1–2 min (activins), 5 min (activin receptors), or 20 min (Wt-1) and then cooled for 30 min before analysis. All sections were washed in PBS and preblocked with 5% normal goat serum (NGS) or 5% normal horse serum (NHS) in PBS for 30 min at room temperature. Sections (n = 5–10/animal) were then incubated (4°C) for 48 h with mouse monoclonal antibody to the human subunit of inhibin (generously supplied by Dr. Nigel Groome, Oxford, U.K.), goat polyclonal antibody to the ßA subunit of human activin A (Research and Development Systems, Minneapolis, MN), mouse monoclonal antibody to the ßB subunit of human activin B (Serotec, Inc., Raleigh, NC), goat polyclonal antibodies to the human activin receptors IB and IIA (Research and Development Systems), or a mouse anti-human monoclonal antibody to Wt-1 (Santa Cruz Biotechnology, Inc., Santa Cruz, CA). Antibodies were diluted in 5% NGS-PBS or 5% NHS-PBS and used at concentrations of 5 µg/ml (-inhibin, Wt-1, and activin ßB), 3 µg/ml (activin ßA), or 0.5 and 2 µg/ml (activin receptors). Except for analysis of activin ßB, ovarian sections were then washed twice in PBS (10 min) and incubated (40 min) with biotinylated goat anti-mouse or mouse anti-goat immunoglobulin (Ig) G (Vector Laboratories, Burlingame, CA) and then with the VectaStain Elite Kit (Vector Laboratories). After rinsing in PBS, sections were stained with diaminobenzidine (DAB)-imidazole-H2O2 and, for Wt-1, with DAB-nickel sulfate (0.250 g of nickel sulfate, 2 mg of DAB, and 8.3 µl of 3% H2O2 in 10 ml of 0.175 M acetate buffer, pH 6.0) as described by Hoffman et al [26]. For activin ßB, sections were washed in PBS and incubated (100 min) with F(ab)2 rabbit anti-mouse IgG conjugated to horseradish peroxidase (Serotec), washed twice in PBS, and then incubated with DAB, imidazole, and H2O. All tissue sections except for Wt-1 were then counterstained with Gill hematoxylin (diluted 1:5 in H2O), mounted in Cytoseal XYL (Richard Allan Scientific, Kalamazoo, MI), and examined by light microscopy. Studies were also performed without primary antibody or with primary antibodies preabsorbed with immunizing peptide (-inhibin and activin receptor IIA) or recombinant proteins to activin A (Research and Development Systems) before application to tissue sections.
Image Analysis of -Inhibin Expression
A minimum of 10 areas (0.08 mm2) of the fetal ovarian cortex from five to eight randomly selected sections of each fetal ovary were examined by light microscopy using an Optiphot-2 microscope attached to a video-based Image-1 analysis system (Universal Imaging Corp., West Chester, PA) essentially as described previously [1]. The average number of cells expressing -inhibin per 0.08 mm2 were calculated for each animal, and the data are expressed as the overall mean ± SEM.
Western Immunoblot Analysis
Western blot analyses of the activin receptors IA, IB, IIA, and IIB as well as Wt-1 were performed essentially as described previously [2, 27]. Briefly, samples of frozen fetal ovaries were suspended in 1% cholic acid (Sigma), 0.1% SDS (Sigma), and 1 mM EDTA in PBS (pH 7.4) containing 0.1 mg/ml of PMSF, 10 µg/ml of aprotinin, and 0.1 mg/ml of soybean inhibitor (Sigma) and then homogenized on ice. After addition of Laemmli buffer [28], samples were heated (100°C for 5 min), cooled, and then loaded (25 µg protein/lane) onto discontinuous 12% SDS-polyacrylamide minigels in electrophoresis chambers containing chilled 0.025 M Tris, 0.192 M glycine (Bio-Rad Laboratories, Inc., Richmond, CA), and 0.1% SDS buffer (pH 8.3). Samples were electrophoresed and wet-transferred to Immobilon P (Millipore Corp., Bedford, MA), and membranes were then blocked and incubated with antibodies described above as well as with polyclonal antibodies to the human activin receptors (Research and Development Systems) diluted to 0.2 µg/ml (IA, IB, and IIB) or 0.5 µg/ml (IIA) in buffer I (50 mM Tris [pH 7.5], 150 mM NaCl, 0.05% Tween-20, and 0.05% IGEPAL CA-630) containing 1.5% BSA. Membranes were washed and then incubated with donkey anti-goat IgG horseradish peroxidase-conjugated second antibody (Amersham Life Sciences, Inc., Arlington Heights, IL) in buffer I containing 1.5% BSA. After washing and application of enhanced chemiluminescent reagent (Amersham Life Sciences), membranes were placed in x-ray film cassettes containing Fugi Medical x-ray film (Fugi Medical Systems, Stamford, CT) and exposed in a dark room for 30–120 sec. The second antibody contributed no nonspecific bands at the concentrations employed.
Statistics
Serum estradiol levels and the number of fetal ovarian cells expressing -inhibin were analyzed by analysis of variance with post hoc comparisons of the means by the Newman-Keuls multiple-comparison test.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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We have previously demonstrated [1, 23] that maternal serum estradiol levels increased from approximately 1 ng/ml on Days 85–120 of gestation to 2.5–3.0 ng/ml by Day 165 and, within 48–72 h of the onset of CGS 20267 treatment on Day 100, decreased to and remained at approximately 0.1 ng/ml. The pattern of serum estradiol in baboons treated with CGS 20267 plus estradiol benzoate was similar to that in the control, although absolute levels were slightly greater than normal. Thus, as seen in Table 1, mean ± SEM estradiol levels in maternal serum on Day 165 of gestation in untreated baboons were significantly (P < 0.01) reduced by administration of CGS 20267 and restored to normal in baboons treated with CGS 20267 and estradiol. Umbilical serum estradiol levels in untreated baboons (0.59 ± 0.13 ng/ml) were also reduced (P < 0.01) by administration of CGS 20267 (0.04 ± 0.01 ng/ml) and restored (0.19 ± 0.08 ng/ml) to 30% of normal (P < 0.05) in baboons treated with CGS 20267 and estrogen. The restoration of maternal, but not of fetal, serum estradiol to normal in baboons treated with CGS 20267 and estradiol benzoate presumably reflects the fact that placental estradiol is preferentially secreted into the maternal circulation during primate pregnancy [3] and that estradiol benzoate was injected into and, thus, was distributed to all maternal tissues rather than originating exclusively in the placenta.
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Expression of -Inhibin
The immunocytochemical expression of -inhibin was minimal to nondetectable in virtually all sections of fetal ovaries from all untreated baboons (Fig. 1, A and B). When detected, -inhibin was lightly expressed in pregranulosa cells of germ cell cords at midgestation (not shown) and in the pregranulosa cells in interfollicular nests at Days 165–170 of gestation. The subunit of inhibin, however, was never detected in granulosa cells of primordial follicles. In contrast, in all sections of late-gestation fetal ovaries from six of six baboons depleted of estrogen, -inhibin expression was abundant and detected in a significant number of pregranulosa cells of the interfollicular nests and in granulosa cells of most primordial follicles (Fig. 1, C and D). This abundant up-regulation of -inhibin in estrogen-suppressed baboons was either prevented (three of five animals) or markedly reduced (two of five animals) in fetal ovaries of baboons treated with CGS 20267 and estradiol (Fig. 1, E and F). Thus, in fetal ovaries of these animals, -inhibin expression was minimal and only occasionally detected in pregranulosa cells of interfollicular nests. We previously showed that these interfollicular nests are a discrete group of cells and not components of follicles resulting from the cutting of sections adjacent to or just into a follicle [1]. Finally, specificity of antibody for detection of -inhibin was confirmed in previous studies that demonstrated absence of staining in sections of fetal baboon adrenal glands incubated with antibody preabsorbed with excess recombinant -inhibin peptide subunit [27].
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The effects of estrogen deprivation on fetal ovarian -inhibin immunocytochemical expression were quantified by image analysis. Thus, as shown in Figure 2, the number (mean ± SEM) per 0.08 mm2 of fetal ovarian cells of untreated baboons expressing -inhibin was similar at midgestation (8.5 ± 2.1) and at late gestation (2.3 ± 0.7), increased approximately 8-fold (P < 0.01) near term in baboons treated with CGS 20267 (70.9 ± 7.5), and restored (P < 0.01) to normal in baboons treated with CGS 20267 plus estradiol (5.6 ± 1.4).
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Expression of Activins ßA and ßB
Both activin ßA and ßB subunits were abundantly expressed in the baboon fetal ovary at mid and late gestation (Fig. 3). Thus, cytoplasmic expression of activin ßA was detected in oocytes and pregranulosa cells at midgestation (Fig. 3A) and in oocytes and granulosa cells of primordial follicles at late gestation (Fig. 3B). Activin ßB was also expressed in the cytoplasm of pregranulosa cells and granulosa cells at midgestation (Fig. 3D) and late gestation (Fig. 3E), respectively, but was not detected in oocytes. Neither the pattern nor the apparent level of expression of activin ßA (Fig. 3C) or activin ßB (Fig. 3F) appeared to be markedly altered at term in fetal ovaries of baboons treated with CGS 20267 or CGS 20267 and estrogen. Specificity was confirmed by absence of staining in sections incubated without primary antibody (not shown).
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Expression of Activin Receptors
Western blot analyses demonstrated that the activin receptors IA and IB were detected as 60-kDa proteins and the type IIA and II B receptors as 68-kDa proteins in fetal ovaries of untreated baboons at both mid (not shown) and late gestation (Fig. 4A). The level of expression of all four receptors at late gestation did not appear to be altered by in vivo treatment with CGS 20267 or CGS 20267 and estrogen (Fig. 4A). Immunocytochemistry showed that activin receptors IB (Fig. 4B) and IIA (Fig. 4C) were localized to oocytes and pregranulosa cells at midgestation and to granulosa cells and/or oocytes of primordial follicles at late gestation.
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Expression of Wt-1
The Wt-1 protein was expressed in the baboon fetal ovary and localized exclusively to the nuclei of pregranulosa cells at midgestation (Fig. 5A) and to the nuclei of granulosa cells of primordial follicles at late gestation (Fig. 5B). Although Wt-1 was also detected in pregranulosa cells of the interfollicular nests in the late-gestation fetal ovary (Fig. 5B), this transcription factor was not expressed in oocytes at either mid or late gestation. The site of expression of Wt-1 was not altered in the near-term fetal ovary of baboons treated with CGS 20267 (Fig. 5C) or CGS 20267 and estrogen (Fig. 5D). Specificity was confirmed by absence of staining in sections of fetal ovary incubated without primary antibody and detection of Wt-1 in sections of fetal kidney, but not of other fetal tissues, incubated with primary antibody as well as by Western blot analyses (results not shown).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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-inhibin expression. Moreover, expression of -inhibin was prominent in pregranulosa cells but not in oocytes, a population of cells that we previously showed were present within interfollicular nests which comprise the primate fetal ovary at midgestation [1, 2]. It is generally believed [29] that the onset of follicle formation, which is initiated at this time during gestation in the human [30] and nonhuman primate [31, 32], involves encapsulation of oocytes by surrounding pregranulosa, a process that results by late gestation in formation of the pool of primordial follicles available for reproductive/ovarian function during adulthood. In our previous studies [1], we showed that the 50% decrease in the number of follicles in fetal ovaries of estrogen-deprived baboons was associated with a comparable increase in the number of interfollicular nests (i.e., oocytes and pregranulosa). We proposed that the decrease in follicle formation reflected, at least in part, a decrease in pregranulosa envelopment of oocytes in the interfollicular nests. Therefore, based on the results of the present study, we further propose that this decrease in follicle formation may result from increased formation of the subunit of inhibin secondary to a loss of estrogen.
As previously demonstrated in the human and rhesus monkey fetus [15], the results of the present study show that the baboon fetal ovary also expressed the ßA and ßB subunits of activin and that these factors were localized to oocytes and/or pregranulosa cells at midgestation and oocytes and/or granulosa cells of primordial follicles at late gestation. However, our studies are the first, to our knowledge, to document that the primate fetal ovary also expressed the receptors for activin and that these are localized to oocytes and granulosa cells at both mid and late gestation. Moreover, because the subunit of inhibin was minimally expressed in the fetal ovary of untreated (i.e., estrogen-replete) baboons, it would appear that activin, and not inhibin, is produced at these sites. Based on these observations, plus the fact that activins can act as paracrine factors to regulate oocyte maturation and granulosa cell function in the adult ovary [11, 14, 15], we suggest that activin also has the potential to act as an autocrine/paracrine factor in the primate fetal ovary to regulate envelopment of oocytes by pregranulosa and granulosa proliferation. A role for activin is, however, suggested by the finding that in estrogen-deprived baboons of the present study, -inhibin was up-regulated and, in these animals, fetal ovarian folliculogenesis was significantly impaired [1] and oocyte atresia markedly increased (unpublished observations). Presumably, up-regulation of -inhibin could result in either depletion of activin (i.e., formation of inhibin) or interference of activin signaling at the receptor level. Studies are currently in progress to examine these possibilities.
In our previous studies, we showed that pregranulosa and granulosa cells of the baboon fetal ovary expressed both ER and ß at mid and late gestation [2]. It appears, therefore, that -inhibin expression, which was minimal in fetal ovaries of baboons either untreated (i.e., estrogen levels normally elevated) or treated with CGS 20267 and estradiol benzoate, is inhibited during fetal development directly and/or indirectly by estrogen. Therefore, we suggest that estrogen regulates fetal ovarian folliculogenesis by controlling, in a cell-specific manner, the ratio of activin to inhibin. Indeed, McMullen et al. [12] have shown that overexpression of -inhibin in the mouse ovary independent of pituitary FSH resulted in ovarian pathologies. Although the sites and mechanisms of estrogen action in the baboon fetal ovary remain to be defined, this proposed action of estrogen is absolutely essential for normal fetal ovarian maturation. Based on studies of the expression of -inhibin, activin subunits, the activin receptors, and the intracellular Smad signaling proteins in adult human ovary, it has been suggested that the activin signal transduction system in granulosa and theca cells undergoes significant changes during follicular maturation and atresia [32]. Whether these critical events in adult ovarian activin signaling and function are dependent on estrogen-dependent programming of the fetal ovary in utero remains to be determined.
Results of the present study also show that the Wt-1 protein was exclusively expressed in somatic cells, but not in oocytes, of the baboon fetal ovary at mid and late gestation as well as in granulosa cells of primordial follicles at late gestation. These observations are consistent with the generally accepted theory, originally put forth by Ohno and Smith [29] in the human and by Sawyer et al. [33] in the sheep, that the somatic cells in germ cell cords are pregranulosa and, thus, the cells that ultimately surround oocytes to form primordial follicles. Interestingly, in baboons deprived of estrogen, the Wt-1 protein continued to be expressed and localized to somatic cells in the interfollicular nests. Thus, the fact that the number of these interfollicular nests of cells was minimal in the fetal ovary of normal baboons as well as those treated with aromatase inhibitor and estrogen further supports the suggestion that the decrease in the number of primordial follicles in estrogen-suppressed fetal baboons reflects, in part, a decrease in encapsulation of oocytes by the pregranulosa. Finally, although Wt-1 has been shown to bind to the promoter region and, thus, can inhibit the expression of -inhibin [21], the results of the present study suggest that changes in the distribution and apparent expression of this protein cannot explain the marked increase in -inhibin in fetal ovaries of baboons deprived of estrogen. Whether other factors that have been shown to modulate -inhibin gene expression (e.g., GATA-1, cAMP) [34, 35] have been modified by estrogen deprivation, however, remains to be determined.
In summary, the results of the present study show that -inhibin expression was markedly up-regulated in pregranulosa and granulosa cells of baboon fetuses in which the number of ovarian follicles was significantly reduced by estrogen deprivation. Moreover, activin ßA was detected in oocytes and activins ßA and ßB were detected and activin receptors were expressed in pregranulosa and granulosa cells of the baboon fetal ovary at mid and late gestation. However, the site and/or level of expression of activins and the activin receptors were not altered by estrogen deprivation. In contrast, the baboon fetal ovary does not express subunit throughout the second half of gestation; thus, the primate fetal ovary presumably does not produce inhibin. Therefore, we propose that estrogen regulates envelopment of oocytes by pregranulosa (i.e., folliculogenesis) by controlling -inhibin expression and, thus, the fetal ovarian activin:inhibin ratio.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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2 Correspondence: Gerald J. Pepe, Department of Physiological Sciences, Eastern Virginia Medical School, P.O. Box 1980, Norfolk, VA 23501-1980. FAX: 757 624 2269; pepegj{at}evms.edu
Received: 1 October 2002.
First decision: 22 October 2002.
Accepted: 26 December 2002.
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and ß in the baboon fetal ovary. Biol Reprod 2002 66:1054-1060-Inhibin is a tumour-suppressor gene with gonadal specificity in mice. Nature 1992 360:313-319[CrossRef][Medline]-subunit. Endocrinology 2001 142:5005-5014-subunit gene. Endocrinology 2001 142:4994-5004 gene promoter. Mol Endocrinol 1995 9:1356-1366-inhibin. Biol Reprod 1999 61:142-146-subunit(s). J Clin Endocrinol Metab 1995 80:3173-3179[Abstract]-subunit gene in MA-10 Leydig tumor cells. Mol Endocrinol 1998 12:378-390-, ßA- and ßB-inhibin/activin genes. Identification of transcription factor AP-2-binding sites in the 5'-flanking regions by DNase I footprinting. Eur J Biochem 1994 226:751-764[Medline]This article has been cited by other articles:
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  C. Pelusi, Y. Ikeda, M. Zubair, and K. L. Parker Impaired Follicle Development and Infertility in Female Mice Lacking Steroidogenic Factor 1 in Ovarian Granulosa Cells Biol Reprod, December 1, 2008; 79(6): 1074 - 1083. [Abstract] [Full Text] [PDF]
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 M. R. Hussein Apoptosis in the ovary: molecular mechanisms Hum. Reprod. Update, March 1, 2005; 11(2): 162 - 178. [Abstract] [Full Text] [PDF]
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E. D. Albrecht, R. B. Billiar, G. W. Aberdeen, J. S. Babischkin, and G. J. Pepe Expression of Estrogen Receptors {alpha} and {beta} in the Fetal Baboon Testisand Epididymis Biol Reprod, April 1, 2004; 70(4): 1106 - 1113. [Abstract] [Full Text] [PDF]
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R. B. Billiar, J. B. St. Clair, N. C. Zachos, M. G. Burch, E. D. Albrecht, and G. J. Pepe Localization and Developmental Expression of the Activin Signal Transduction Proteins Smads 2, 3, and 4 in the Baboon Fetal Ovary Biol Reprod, March 1, 2004; 70(3): 586 - 592. [Abstract] [Full Text] [PDF]
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 N. C. Zachos, R. B. Billiar, E. D. Albrecht, and G. J. Pepe Regulation of Oocyte Microvilli Development in the Baboon Fetal Ovary by Estrogen Endocrinology, February 1, 2004; 145(2): 959 - 966. [Abstract] [Full Text] [PDF]
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