| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Regular Article |
a Center for Reproductive Medicine, Departments of Physiology and of Obstetrics and Gynecology, Sahlgrenska University Hospital, SE-413 45 Göteborg, Sweden
b Institute of Anatomy and Cell Biology, Medical Faculty, Göteborg University, SE-405 30 Göteborg, Sweden
ABSTRACT
Almost all ovarian follicles undergo atresia during follicular development. However, the number of corpora lutea roughly equals the number of preovulatory follicles in the ovary. Because apoptosis is the cellular mechanism behind follicle and luteal cell demise, this suggests a change in apoptosis susceptibility during the periovulatory period. Sex steroids are important regulators of follicular cell survival and apoptosis. The aim of the present work was to study the role of progesterone receptor-mediated effects in the regulation of granulosa cell apoptosis. The levels of internucleosomal DNA fragmentation were evaluated in rat granulosa cells before and after induction of the nuclear progesterone receptor, using hCG treatment to eCG-primed rats to mimic the naturally occurring LH surge. Granulosa cells isolated from hCG-treated rats showed a several-fold increase in the expression of progesterone receptor mRNA and a 47% decrease (P < 0.01) in DNA fragmentation after 24 h incubation in serum-free medium compared to granulosa cells isolated from rats treated with eCG only. The effect of hCG treatment in vivo was dose-dependently reversed in vitro by addition of antiprogestins (Org 31710 or RU 486) to the culture medium, demonstrated by increased DNA fragmentation as well as increased caspase-3 activity. Addition of antiprogestins to granulosa cells isolated from immature or eCG-treated rats did not result in increased DNA fragmentation. The results suggest that progesterone receptor-mediated effects are involved in regulating the susceptibility to apoptosis in LH receptor-stimulated preovulatory rat granulosa cells.
apoptosis, granulosa cells, hCG, hormone action, LH, ovary, progesterone, progesterone receptor
INTRODUCTION
The majority of ovarian follicles undergo atresia, a hormonally controlled apoptotic process [1]. Atresia occurs throughout follicle development, with an extensive reduction in the number of follicles present at birth [2]. This drastic reduction is, however, not present among preovulatory follicles responding to the ovulatory LH surge, as the number of corpora lutea roughly equals the number of preovulatory follicles.
One of the major characteristics of granulosa cells after LH receptor stimulation is the ability to produce large amounts of progesterone. Before the LH surge, the level of circulating progesterone is low. Progesterone within the ovary is critical for regulating specific events such as ovulation and luteinization [3–5]. Effects of progesterone are mediated via the nuclear progesterone receptor, a typical three-domain steroid receptor, existing in a long form (B) and two truncated forms (A and C) and acting as a transcription factor upon ligand activation [6, 7].
In the rat, the LH surge acts directly to induce expression of progesterone receptor mRNA and protein in differentiated rat granulosa cells, expressing elevated levels of LH receptor and aromatase cytochrome P450 [8]. The induction of progesterone receptor mRNA is rapid but transient, with detectable amounts observed 2 h after the LH surge, increasing further after 4 h, and declining after 8 h. The appearance of receptor protein follows the same pattern with an increase seen 3–5 h after the LH surge and a further increase seen after 7–9 h. No progesterone receptor mRNA is observed in rat corpus luteum [8]. However, progesterone has been shown to mediate effects in the rat corpus luteum, using the glucocorticoid receptor [9]. In the brain, progesterone may mediate effects using the 
-aminobutyric acid (GABA) receptor [10]. Indeed, the existence of GABA receptors in the rat ovary has been reported [11, 12]. Ovarian effects mediated by progesterone using classic GABA receptors have not been demonstrated, though ovarian effects of progesterone using a GABA-like receptor have been reported [13]. Progesterone binding to a membrane-bound protein in immature rat and bovine granulosa cells has recently been reported [14, 15]. In addition, an orphan nuclear receptor, activated by pregnenolone and progesterone, was recently characterized in mouse liver, intestine, kidney, and stomach [16]. Thus, effects of progesterone seen before the LH surge or after ovulation may be transferred via other signalling pathways than the classical nuclear receptor pathway.
The aim of this study was to investigate the role of nuclear progesterone receptor-mediated effects in regulation of apoptosis in rat granulosa cells during the period between LH receptor stimulation and ovulation. Internucleosomal DNA fragmentation and caspase-3 activity, late and early events in the apoptotic process [17], were used as endpoints.
MATERIALS AND METHODS
Animals
Immature, female Sprague-Dawley rats were purchased from B&K Universal AB, Sollentuna, Sweden, and treated in accordance with the requirements of the local ethical committee.
Materials, Tissue Isolation, and Cell Culture
Twenty six-day-old animals were treated with s.c. injections of 10 IU eCG (Sigma, St. Louis, MO) to induce follicle maturation. Some animals were also given an i.p. injection with 50 IU hCG (N.V. Organon, Oss, Holland) 48 h after eCG treatment to mimic the endogenous LH surge. A third group of animals did not receive any treatment. Isolated ovaries were used to isolate granulosa cells from preovulatory follicles. Granulosa cells were either snap frozen or incubated (500 000/0.5 ml) in tubes (Falcon 12 by 75 mm; Becton Dickinson) with culture medium (Eagle MEM with glutamax-I [L-alanyl-L-glutamine], Earle salts, and Hepes; Life Technologies, Täby, Sweden), supplemented with penicillin (100 U/ml), streptomycin (100 µg/ml, Life Technologies, Paisley, Scotland), and BSA, fraction V (0.1%, Sigma-Aldrich Chemie, Gmbh, Steinheim, Germany), with or without addition of the antiprogestins Org 31710 (0.1–10 µM, N.V. Organon, Oss, the Netherlands) or RU 486 (1–50 µM, Exelgyn, Paris, France), the GABA receptor antagonists picrotoxin (100 µM, Sigma-Aldrich Chemie) or bicuculline (100 µM, Sigma-Aldrich Chemie), the GABA receptor agonist muscimol (100 µM, Sigma-Aldrich Chemie) or progesterone (1–100 µM, Sigma). Org 31710 and bicuculline were dissolved in dimethyl sulfoxide (DMSO), RU 486 in ethanol, and picrotoxin and muscimol in culture medium. The final concentration of ethanol or DMSO in the incubation medium was less than 0.1% in all experiments. The incubations were performed at 37°C in 5% CO2 and 95% humidified air for 24 h.
Gel Electrophoretic Analysis of Internucleosomal DNA Fragmentation
To estimate the degree of internucleosomal DNA fragmentation, granulosa cells were gently homogenized and incubated (30 min, 65°C) in a buffer containing 0.1 M NaCl, 0.01 M EDTA, 0.3 M Tris-HCl, 0.2 M sucrose, and 10% SDS, followed by addition of 8 M potassium acetate and incubation on ice for 60 min. After phenol:chloroform extraction and ethanol precipitation of the DNA, RNase A (Boerhinger Mannheim, Bromma, Sweden) was added, followed by another phenol:chloroform extraction and ethanol precipitation. The DNA was finally dissolved in distilled water. Aliquots of DNA from each sample were labeled with 35S-dATP (Amersham Sweden AB, Stockholm, Sweden), using Klenow enzyme (Promega, Sweden) [18]. Equal amounts of labeled DNA (either 10 or 30 ng/lane) were fractionated through 1.8% agarose gels and dried in a slab gel dryer before exposure to x-ray films (Amersham Hyperfilm, Stockholm, Sweden) at room temperature. After autoradiography, the samples were excised with a scalpel, each lane being separated into one high (>2 kb) and one low (<2 kb) molecular fraction and then counted in a ß-counter to provide a quantitative estimate of the degree of internucleosomal DNA fragmentation among samples [19]. The degree of apoptosis is defined as the level of low molecular weight DNA radioactivity in each sample compared to control and is expressed as apoptotic index. The value for the control group was set as 1.0.
Fluorospectrophotometric Analysis of DNA Fragmentation
Fragmentation of DNA in isolated granulosa cells was determined essentially as described earlier [20] with the following modifications: cells were pelleted (200 x g, 5 min) and treated with a lysis buffer (5 mM Tris, 20 mM EDTA, and 0.5% Triton, pH 8). After incubation on ice for 20 min and sixfold dilution with a buffer containing Tris (5 mM) and EDTA (20 mM), the samples were centrifuged for 20 min at 11 000 x g to separate fragmented DNA (supernatant) from intact DNA (pellet). The pellets were incubated overnight with proteinase-K (1 mg/ml; Merck, Darmstadt, Germany) at 45°C in a buffer containing Tris (10 mM) and EDTA (1 mM). The DNA content in the supernatant and pellet, respectively, was measured with a fluorescence spectrophotometer (356 nm excitation and 458 nm emission; F-2000, Hitachi, KEBO, Göteborg, Sweden), after addition of Hoechst dye H33258 (0.2 µg/ml in 2 M NaCl, 1 mM EDTA, and 10 mM Tris, pH 7.4) [21]. The apoptotic index reflects the ratio between low molecular weight DNA (isolated in the supernatant) and total DNA content (supernatant DNA and pellet DNA). The value for the control group was set as 1.0.
Reverse Transcriptase-Polymerase Chain Reaction
Total RNA was isolated from granulosa cells, small intestine, and uterus by acid guanidinium thiocynate:phenol:chloroform extraction as described by Chomczynski and Sacchi [22]. Complementary DNA was synthesized using 2 µg total RNA, 10 nmol dNTPs, and 1 µg oligo-dT15 primers (Promega) that were denatured at 70°C for 3 min and placed on ice. Two hundred units of mouse mammary leukemia virus reverse transcriptase (RT; Promega), and corresponding buffer was added and cDNA synthesis was carried out at 42°C for 60 min in a total volume of 20 µl.
To detect the expression of the mRNA encoding the progesterone receptor, 1 µl of the RT reaction was subjected to polymerase chain reaction (PCR) amplification using gene specific primers for the progesterone receptor hormone-binding domain, common to the A and B isoforms, and ribosomal protein L19 (RPL19) as internal standard [23]. The primer sequences were 5'-CTGCTGGATGAGCCTGATGGTG-3' and 5'-CACCATCCCTGCCAGGATCTTG-3' for the progesterone receptor sense and antisense primers, respectively, and 5'-CTGAAGGTCAAAGGGAATGTG-3' and 5'-GGACAGAGTCTTGATGATCTC-3' for sense and antisense primers for RPL19. The PCR amplification was carried out in the presence of 2.5 µCi 
-32P-dCTP for 25 cycles with an annealing temperature of 60°C, using a Perkin Elmer GeneAmp PCR system 9700 (Perkin Elmer, Stockholm, Sweden). All PCR products were electrophoretically separated on a 1% 0.5x Tris-borate-EDTA agarose gel. Following autoradiography, the gel was analyzed using a Molecular Dynamics Storm 820 Phosphoimager and Image Quant 5.0 software (Molecular Dynamics, Uppsala, Sweden).
Progesterone Assay
Progesterone in spent medium was analyzed by time-resolved immunofluorometric assay (DELFIA, Wallac Oy, Turku, Finland).
Caspase-3 Activity Assay
Granulosa cells were isolated from rats treated with hCG for 12 h and incubated for 24 h in serum-free medium with or without addition of Org 31710 (10 µM) or RU 486 (25 µM). After incubation, medium was removed by centrifugation and granulosa cells were washed in ice-cold 1x PBS before freezing at -70°C. The frozen cell pellets were lysed by addition of 3-[(3-cholamidopropyl)-dimethyl-ammonio]-1-propanesulfonate (CHAPS)-containing buffer (50 mM Tris-HCl, 100 mM NaCl, 5 mM EDTA, 1 mM EGTA, 3 mM NaN3, 0.2% CHAPS, pH 7.3). Trypsin inhibitor (final concentration 5 µg/ml), pepstatin (0.5 µg/ml), leupeptin (1.25 µg/ml), and PMSF (0.5 mM) (all from Sigma) were added to the buffer to minimize activity of other proteases than caspase-3. Cell lysate from three tubes in each group were pooled and statistically calculated as n = 1. Preincubation with inhibitor-containing CHAPS buffer was made for at least 30 min at room temperature, after which 20 µl was removed for protein determination (see below). The synthetic substrate acetyl-Asp-Glu-Val-Asp-7-amido-4-methyl-coumarin (Ac-DEVD-AMC; Calbiochem, La Jolla, CA) was dissolved in water as a 1 mM stock and diluted to a final concentration of 28 µM in 50 mM Tris-HCl, 100 mM NaCl, 5 mM EDTA, 1 mM EGTA, 3 mM NaN3,pH 7.3, with the addition of 2 mM dithiothreitol (final concentration in the assay).
At the start of the proteolytic assay, 50 µl of cell lysate and 50 µl of substrate solution were added to a 96-well plate (Dynex Technologies). Fluorescence of the cleavage product was measured over time at 37°C in a microplate spectrofluorometer (SPECTRAmax GEMINI, Molecular Devices, Sunnyvale, CA), excitation wavelength 380 nm, emission wavelength 440 nm. Activity was normally measured during 3 to 5 h and Vmax determined using SOFTmax PRO Version 2.6 as software (Molecular Devices). Proteolytic activity was calculated as relative fluorescence units per second and gram of protein (U/sec g).
For protein determination, Pierce bicinchononic acid protein assay (Pierce, Rockford, IL) with BSA as standard was used. Absorbance was measured at 570 nm in a microplate reader (Emax, Molecular Devices) using SOFTmax version 2.01 as software.
Statistical Analysis
Experiments were done at least three times, if not stated otherwise. All statistical analyses were performed by ANOVA one-way analysis, followed by the Student-Newman-Keuls multiple range test. Numerical data are presented as mean ± SEM. A P value less than 0.05 was considered significant.
RESULTS
Effect of In Vivo Differentiation on Internucleosomal DNA Fragmentation and Expression of Progesterone Receptor mRNA in Rat Granulosa Cells
The degree of granulosa cell internucleosomal DNA fragmentation was decreased in a time-dependent manner after eCG treatment in vivo (0, 24, and 48 h). Internucleosomal DNA fragmentation was further decreased after LH receptor stimulation with hCG in vivo (Fig. 1A). Quantitative analysis of the 35S-labeled low molecular granulosa cell DNA separated by gel electrophoresis demonstrated that the degree of internucleosomal DNA fragmentation in cells isolated from hCG-treated rats was reduced to less than one fifth compared to granulosa cells isolated from untreated controls. To exclude that the effect of hCG seen in granulosa cells was due to selection of follicles during granulosa cell isolation, DNA from whole ovaries was isolated and electrophoretically separated. Similar results were achieved as for isolated granulosa cells (data not shown).
|
To investigate the expression of nuclear progesterone receptor mRNA in rat granulosa cells, RT-PCR was performed. Six hours after hCG treatment in vivo, the expression was dramatically increased, compared to the expression in granulosa cells isolated from immature rats and from rats treated with eCG for 24 and 48 h, respectively (Fig. 1B).
Effect of In Vivo Differentiation on DNA Fragmentation and Progesterone Production in Rat Granulosa Cells In Vitro
Granulosa cells isolated from hCG-treated rats and incubated for 24 h showed a 47% decrease in DNA fragmentation compared to controls (Fig. 2). Analysis of spent medium showed a dramatic increase in progesterone production by granulosa cells isolated from hCG-treated rats (33.03 ± 0.295 nmol/L, n = 3) compared to controls (0.409 ± 0.034 nmol/L, n = 3). Supplementation of the culture medium with progesterone (1–100 µM) only accentuated the inhibition of DNA fragmentation in granulosa cells from hCG-treated rats in the 100 µM dose (three magnitudes higher than the concentration in spent medium from these cells), which conferred an additional decrease of 20% (P < 0.01).
|
Induction of DNA Fragmentation and Caspase-3 Activity by Org 31710 or RU 486
Granulosa cells were incubated for 24 h with or without addition of the nuclear progesterone receptor antagonist Org 31710 (10 µM). Internucleosomal DNA fragmentation was visualized with gel electrophoresis. The degree of internucleosomal DNA fragmentation in granulosa cells isolated from hCG-treated rats was one fourth of the internucleosomal DNA fragmentation in granulosa cells isolated from eCG-treated rats. Treatment with Org 31710 more than doubled the internucleosomal DNA fragmentation in granulosa cells isolated from hCG-treated rats (Fig. 3). Internucleosomal DNA fragmentation induced by Org 31710 could not be reversed by adding more progesterone (100 µM) to the levels of progesterone produced by granulosa cells during incubation (data not shown).
|
The decrease in internucleosomal DNA fragmentation induced by hCG treatment in vivo for 12 h was dose-dependently reversed after addition of increasing concentrations of Org 31710 (0.1–10 µM) (Fig. 4). The RU 486, another progesterone receptor antagonist, also showed a dose-dependent increase in DNA fragmentation when added (1–50 µM) to the medium during 24 h incubation of granulosa cells isolated from hCG-treated rats (Fig. 5). The increase in DNA fragmentation in these cells after treatment with Org 31710 or with RU 486 was parallelled by an increase in caspase-3 activity (Fig. 6, A and B). No effect on DNA fragmentation or caspase-3 activity could be demonstrated when Org 31710 or RU 486 was added to the culture medium of granulosa cells isolated from immature rats treated with eCG for 48 h (data not shown).
|
|
|
No Effect of GABA Receptor Agonist or Antagonists on DNA Fragmentation in Granulosa Cells In Vitro
No effect on DNA fragmentation, measured fluorospectrophotometrically, was seen in granulosa cells isolated from rats either treated with eCG for 48 h or additionally treated with hCG for 12 h, when the GABA receptor antagonists picrotoxin (100 µM) or bicuculline (100 µM) or the GABA receptor agonist muscimol (100 µM) were added to the culture medium during 24 h incubation (Table 1).
|
DISCUSSION
In this study, nuclear progesterone receptor-mediated effects on rat preovulatory granulosa cell susceptibility to apoptosis in vitro, measured as internucleosomal DNA fragmentation and caspase activity, was investigated. An increase in progesterone receptor mRNA and a decrease in internucleosomal DNA fragmentation were seen after granulosa cell LH receptor stimulation in vivo. The decrease in internucleosomal DNA fragmentation was also seen after incubation for 24 h, compared to granulosa cells isolated from rats treated with eCG only. The decrease in DNA fragmentation was dose-dependently inhibited in vitro by addition of the antiprogestins Org 31710 or RU 486 to the culture medium.
Gonadotropins are required for the growth and development of ovarian follicles [24, 25]. Antral follicles, expressing the FSH receptor, are dependent upon FSH stimulation for survival. However, studies using cultured rat granulosa cells have shown that treatments with FSH, LH/hCG, or insulin-like growth factor-I are ineffective in the prevention of spontaneous apoptosis, despite their apoptosis-suppressing action in cultured rat follicles [26]. This implies the importance of neighboring theca cells and local factors produced in the ovary for regulation of follicle growth and atresia.
Follicle survival factors, e.g., epidermal growth factor/transforming growth factor, basic fibroblast growth factor [27], interleukin-1ß [28], as well as atretogenic factors, e.g., tumor necrosis factor- [29], Fas ligand [30], GnRH [31], have been characterized. Other important regulators of follicle survival and atresia are sex steroids. In the rat ovary, estrogens act as survival factors, whereas androgens promote apoptosis [32]. In contrast to their action in the ovary, androgens can act as survival factors in the testis and in the prostate gland [33, 34]. Progesterone has also been reported to affect cell survival [14, 35, 36]. In the female, the circulating level of progesterone is low until the preovulatory LH surge. A characteristic feature of the luteinization process of granulosa cells is the ability of the cells to produce large amounts of progesterone. Interestingly, this stage of differentiation differs from preceding ones concerning follicular demise, as the number of postovulatory corpora lutea roughly equals the number of preovulatory follicles. Thus, the apoptotic program, operating at a high rate at all other stages of follicular development, seems to be changed drastically during the transformation of preovulatory follicles into corpora lutea.
To mimic the period between LH receptor stimulation and ovulation, immature Sprague-Dawley rats were treated with eCG and hCG. A decrease in internucleosomal DNA fragmentation in granulosa cells was seen after hCG treatment in vivo as well as after incubation for 24 h, suggesting that stimulation of the LH receptor confers resistance to apoptosis in rat granulosa cells in vivo as well as in vitro. One possible mediator of the observed LH receptor-mediated effect is progesterone, as the midcycle LH surge induces production of high levels of progesterone in the follicle. Progesterone mediates its effects via the nuclear progesterone receptor, but other pathways have also been demonstrated [10–16].
To evaluate the hypothesis that stimulation of the LH receptor confers resistance to apoptosis in rat granulosa cells, Org 31710, a highly selective antiprogestin, was added in vitro. Org 31710 interacts with the progesterone receptor at the ligand-binding domain. In contrast to RU 486, Org 31710 has a highly selective spectrum, with little antiglucocorticoid activity and no other hormonal activities, except for weak interactions with the androgen receptor [37, 38]. Org 31710 dose-dependently reversed the DNA fragmentation protecting effect induced by stimulation of the LH receptor with hCG in vivo. Org 31710 did not show any effect on DNA fragmentation during 24 h incubation of granulosa cells isolated from immature rats or from rats treated with eCG for 48 h. In these cells only minute levels of progesterone receptor were detected using RT-PCR, compared to high levels in granulosa cells isolated from hCG-treated rats. Indeed, similar expression patterns of progesterone receptor mRNA and protein have previously been reported by others [8, 39]. Thus, there is no evidence for expression of functional nuclear progesterone receptor in rat granulosa cells until after the midcycle LH surge. The results support the theory of progesterone receptor involvement in regulation of granulosa cell susceptibility to apoptosis after LH receptor stimulation.
Progesterone mediating its effects via the nonclassical progesterone receptor GABA [10], via the recently characterized pregnenolone receptor [16], or via other signalling pathways [13–15] has been reported. However, we have not been able to confirm progesterone signalling via the GABA receptor in our system, using the GABA receptor antagonists picrotoxin and bicuculline or the GABA-agonist muscimol. To our knowledge, no interactions of Org 31710 with the GABA receptor or with the recently characterized pregnenolone receptor have been reported.
In studies regarding progesterone interaction with granulosa cell apoptosis prior to the LH surge [13–15], the antiprogestin RU 486 has been used. Beside its antagonistic effects on the progesterone receptor, this compound also has antiglucocorticoid activity that may affect the pituitary-adrenal axis, making it difficult to interpret in vivo data. In addition, in vitro studies have shown that RU 486 may interfere with progesterone production in both rat and human granulosa cells [40]. It may also, through inhibition of 17 OH-activity, influence androgen as well as estrogen levels [41]. In our experiments, RU 486, similar to Org 31710, showed a dose-dependent increase in DNA fragmentation when added in vitro to granulosa cells isolated from rats treated with hCG but not when added to granulosa cells isolated from 28-day-old immature rats or from rats treated with eCG for 48 h. The increase in DNA fragmentation induced by Org 31710 or by RU 486 was parallelled by an increase in caspase-3 activity, an early event in the apoptotic process [17]. This suggests a similar apoptosis-inducing action of RU 486 and Org 31710 via the nuclear progesterone receptor. No effect on caspase-3 activity was detected after in vitro treatment with Org 31710 or RU 486 to rat granulosa cells isolated from immature rats treated with eCG for 48 h.
The LH-induced expression of the nuclear progesterone receptor and increase in progesterone production, as a result of luteinization of the follicular cells, may induce a progesterone receptor-mediated differentiation of the cells to become dependent on progesterone as a survival factor. Indeed, in the progesterone receptor knockout mouse, the follicles do not ovulate or luteinize but persist, giving the ovaries a polycystic appearance [5]. It was recently demonstrated that decreased progesterone levels and progesterone receptor antagonists promote apoptotic cell death in bovine luteal cells [42].
In conclusion, the results presented here suggest that progesterone receptor expression and activation are involved in regulating rat granulosa cell susceptibility to apoptosis after LH receptor stimulation.
ACKNOWLEDGMENTS
The Org 31710 was generously provided by N.V. Organon, Oss, the Netherlands; RU 486 was a gift from Exelgyn, Paris, France. The authors thank Ann Wallin for technical assistance with progesterone measurements.
FOOTNOTES
First decision: 10 March 2000.
1 This work was supported by grants 10380 and 11134 from The Swedish MRC, by grants from the Assar Gabrielsson Foundation, The Research Foundation of Hjalmar Svensson, The Göteborg Medical Society, The Foundation of Clas Groschinskys Memorial Fund, and by grants from the State under the LUA agreement. 
2 Correspondence: Håkan Billig, Department of Physiology, P.O. Box 434, SE-405 30, Göteborg, Sweden. FAX: 46 31 7733531; hakan.billig{at}fysiologi.gu.se
Accepted: June 21, 2000.
Received: February 7, 2000.
REFERENCES
-hydroxysteroid dehydrogenase expression in the rat corpus luteum through the glucocorticoid receptor. Endocrinology 1997; 138:4497–4500. and its second messenger, ceramide, stimulate apoptosis in cultured ovarian follicles. Endocrinology 1996; 137:4864–4870.[Abstract]This article has been cited by other articles:
|
|
 |
|
  J. J. Peluso, X. Liu, A. Gawkowska, and E. Johnston-MacAnanny Progesterone Activates a Progesterone Receptor Membrane Component 1-Dependent Mechanism That Promotes Human Granulosa/Luteal Cell Survival But Not Progesterone Secretion J. Clin. Endocrinol. Metab., July 1, 2009; 94(7): 2644 - 2649. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. A. Friberg, D.G. J. Larsson, and H. Billig Dominant Role of Nuclear Progesterone Receptor in the Control of Rat Periovulatory Granulosa Cell Apoptosis Biol Reprod, June 1, 2009; 80(6): 1160 - 1167. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. J. Peluso, J. Romak, and X. Liu Progesterone Receptor Membrane Component-1 (PGRMC1) Is the Mediator of Progesterone's Antiapoptotic Action in Spontaneously Immortalized Granulosa Cells As Revealed by PGRMC1 Small Interfering Ribonucleic Acid Treatment and Functional Analysis of PGRMC1 Mutations Endocrinology, February 1, 2008; 149(2): 534 - 543. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. J. Peluso, X. Liu, and J. Romak Progesterone Maintains Basal Intracellular Adenosine Triphosphate Levels and Viability of Spontaneously Immortalized Granulosa Cells by Promoting an Interaction between 14-3-3{sigma} and ATP Synthase{beta}/Precursor through a Protein Kinase G-Dependent Mechanism Endocrinology, May 1, 2007; 148(5): 2037 - 2044. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Yacobi, A. Tsafriri, and A. Gross Luteinizing Hormone-Induced Caspase Activation in Rat Preovulatory Follicles Is Coupled to Mitochondrial Steroidogenesis Endocrinology, April 1, 2007; 148(4): 1717 - 1726. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Engmann, R. Losel, M. Wehling, and J. J. Peluso Progesterone Regulation of Human Granulosa/Luteal Cell Viability by an RU486-Independent Mechanism J. Clin. Endocrinol. Metab., December 1, 2006; 91(12): 4962 - 4968. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. J. Peluso Multiplicity of Progesterone's Actions and Receptors in the Mammalian Ovary Biol Reprod, July 1, 2006; 75(1): 2 - 8. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. J. Peluso, A. Pappalardo, R. Losel, and M. Wehling Progesterone Membrane Receptor Component 1 Expression in the Immature Rat Ovary and Its Role in Mediating Progesterone's Antiapoptotic Action Endocrinology, June 1, 2006; 147(6): 3133 - 3140. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Z. Cai and C. Stocco Expression and Regulation of Progestin Membrane Receptors in the Rat Corpus Luteum Endocrinology, December 1, 2005; 146(12): 5522 - 5532. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. J. Peluso, A. Pappalardo, R. Losel, and M. Wehling Expression and Function of PAIRBP1 Within Gonadotropin-Primed Immature Rat Ovaries: PAIRBP1 Regulation of Granulosa and Luteal Cell Viability Biol Reprod, August 1, 2005; 73(2): 261 - 270. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Rung, P. A. Friberg, R. Shao, D.G. J. Larsson, E. Ch. Nielsen, P.-A. Svensson, B. Carlsson, L. M.S. Carlsson, and H. Billig Progesterone-Receptor Antagonists and Statins Decrease De Novo Cholesterol Synthesis and Increase Apoptosis in Rat and Human Periovulatory Granulosa Cells In Vitro Biol Reprod, March 1, 2005; 72(3): 538 - 545. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. M. Quirk, R. G. Cowan, and R. M. Harman Progesterone Receptor and the Cell Cycle Modulate Apoptosis in Granulosa Cells Endocrinology, November 1, 2004; 145(11): 5033 - 5043. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. J. Peluso, A. Pappalardo, G. Fernandez, and C. A. Wu Involvement of an Unnamed Protein, RDA288, in the Mechanism through which Progesterone Mediates Its Antiapoptotic Action in Spontaneously Immortalized Granulosa Cells Endocrinology, June 1, 2004; 145(6): 3014 - 3022. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Yacobi, A. Wojtowicz, A. Tsafriri, and A. Gross Gonadotropins Enhance Caspase-3 and -7 Activity and Apoptosis in the Theca-Interstitial Cells of Rat Preovulatory Follicles in Culture Endocrinology, April 1, 2004; 145(4): 1943 - 1951. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. M. Quirk, R. G. Cowan, R. M. Harman, C.-L. Hu, and D. A. Porter Ovarian follicular growth and atresia: The relationship between cell proliferation and survival J Anim Sci, January 1, 2004; 82(13_suppl): E40 - 52. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Shao, E. Markstrom, P. A. Friberg, M. Johansson, and H. Billig Expression of Progesterone Receptor (PR) A and B Isoforms in Mouse Granulosa Cells: Stage-Dependent PR-Mediated Regulation of Apoptosis and Cell Proliferation Biol Reprod, March 1, 2003; 68(3): 914 - 921. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Wang, C. Xiao, and A. K. Goff Progesterone-Modulated Induction of Apoptosis by Interferon-Tau in Cultured Epithelial Cells of Bovine Endometrium Biol Reprod, February 1, 2003; 68(2): 673 - 679. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J.J. Peluso, T. Bremner, G. Fernandez, A. Pappalardo, and B.A. White Expression Pattern and Role of a 60-Kilodalton Progesterone Binding Protein in Regulating Granulosa Cell Apoptosis: Involvement of the Mitogen-Activated Protein Kinase Cascade Biol Reprod, January 1, 2003; 68(1): 122 - 128. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. A. Goyeneche, R. P. Deis, G. Gibori, and C. M. Telleria Progesterone Promotes Survival of the Rat Corpus Luteum in the Absence of Cognate Receptors Biol Reprod, January 1, 2003; 68(1): 151 - 158. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. S. Quadros, J. L. Pfau, A. Y. N. Goldstein, G. J. De Vries, and C. K. Wagner Sex Differences in Progesterone Receptor Expression: A Potential Mechanism for Estradiol-Mediated Sexual Differentiation Endocrinology, October 1, 2002; 143(10): 3727 - 3739. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Sasson and A. Amsterdam Stimulation of Apoptosis in Human Granulosa Cells from in Vitro Fertilization Patients and Its Prevention by Dexamethasone: Involvement of Cell Contact and Bcl-2 Expression J. Clin. Endocrinol. Metab., July 1, 2002; 87(7): 3441 - 3451. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J.J. Peluso, G. Fernandez, A. Pappalardo, and B.A. White Characterization of a Putative Membrane Receptor for Progesterone in Rat Granulosa Cells Biol Reprod, July 1, 2001; 65(1): 94 - 101. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
