Simultaneous plasma radioimmunoassay of estradiol, progesterone, 17-hydroxy-progesterone, testosterone and androstenedione in farm animals.
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Androstenedione↗
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The absorption of oral micronized progesterone: the effect of food, dose proportionality, and comparison with intramuscular progesterone.
OBJECTIVES: To examine the effects of food ingestion and administered dose on the absorption of oral micronized P (Utrogestan; Besins-Iscovesco, Paris, France) and to compare the bioavailability of intramuscular versus oral routes of administration. DESIGN: Prospective, randomized, open label crossover protocol with 7 days between dosages. SETTING: Academic institution. PARTICIPANTS: Fifteen normal postmenopausal women. INTERVENTIONS: All subjects participated in three separate protocols: [1] micronized P (200 mg) or placebo under fasting or nonfasting conditions once daily for 5 days; [2] micronized P (100, 200, or 300 mg) once daily under fasting conditions for 5 days; and [3] micronized P (200 mg) or intramuscular P (50 mg in oil) administered once daily for 2 days. MAIN OUTCOME MEASURES: Serum P concentrations were measured in all groups. RESULTS: Concomitant food ingestion increased the area under the serum P concentration versus time curve (AUC0 to 24) and the maximum serum P concentration (Cmax) without affecting time to maximum serum concentration (Tmax) (P < 0.05). Micronized P absorption and elimination were first-order processes and exhibited dose-independent pharmacokinetics between 100 and 300 mg. After intramuscular P, Cmax was higher and Tmax occurred later compared with the oral P preparation. Oral P had lower relative bioavailability (8.6%) than intramuscular P. CONCLUSIONS: Absorption of micronized P was enhanced twofold in the presence of food. Both absorption and elimination were dose-independent, dose proportionality being confirmed. Bioavailability of the oral P was approximately 10% compared with intramuscular P.
Administration, Oral↗
Progesterone in uterus and plasma. IV. Progesterone receptor(s) in guinea pig uterus cytosol.
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Animals↗
Review: progesterone or progesterone lead to a marginal reduction in premenstrual syndrome symptoms.
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Comment↗
[Steroid alkaloids. 53. Synthesis of dihydro-4,5 hydroxy-18 progesterone and nor-18 hydroxy-13 dihydro-4,5 progesterone starting with conessine].
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Alkaloids↗
Specific non-genomic, membrane-localized binding sites for progesterone in the bovine corpus luteum.
Fractionation of bovine corpus luteum (CL) homogenates on continuous sucrose density gradients with and without preincubation with 3H-progesterone demonstrated high levels of tracer binding and high content of endogenous progesterone associated with particulate membrane fractions. Analysis of gradient fractions for a range of luteal plasma membrane and intracellular organelle marker enzyme activities indicated that endogenous progesterone content and 3H-progesterone-binding activity were associated with fractions enriched in luteal plasma membrane markers. This was confirmed by pretreatment of homogenates with the saponin, digitonin, prior to fractionation. Digitonin perturbed the buoyant density of luteal surface membrane markers and 3H-progesterone binding to a similar extent, but did not perturb the buoyant densities of other intracellular markers to the same degree. Interestingly, digitonin pretreatment also increased the proportion of progesterone tracer that entered the gradients. We consistently failed to demonstrate significant binding of 3H-progesterone to membrane fractions incubated with progesterone tracer in vitro. However, when digitonin was included in the in vitro binding assay, we observed a dramatic, dose-dependent stimulation of 3H-progesterone binding by digitonin. Other radiolabeled steroids tested (3H-cortisol, 3H-testosterone) bound poorly in the presence or absence of digitonin. 3H-Progesterone binding in the presence of optimal digitonin concentrations increased linearly with increasing luteal membrane concentration; was dependent on the pH, duration, and temperature of incubation; and low levels of progesterone (68 nM) competed for tracer binding. A range of other steroids tested (androgens, estrogens, corticosteroids, steroid precursors) competed at higher concentrations (10- to 100-fold) or did not compete at all for 3H-progesterone binding. There was no correlation between the hydrophobicity of various steroids and their ability to compete for binding. Moreover, a number of agonists and antagonists specific for the genomic progesterone receptor, agonists of peripheral benzodiazepine receptors, and inhibitors of a range of steroidogenic enzymes did not compete for 3H-progesterone binding. Western blots confirmed that detergent-solubilized progesterone-binding sites could be resolved from cytochrome P450 side-chain cleavage and 3beta-hydroxysteroid dehydrogenase. Moreover, extraction of bound steroid from the binding site and HPLC demonstrated identity to progesterone, suggesting that no metabolism of the progesterone tracer had occurred during incubation. Progesterone binding to membranes of large luteal cells was higher compared with binding to small luteal cells, and levels were similar in membranes prepared from CL at all stages of the luteal phase. We suggest that bovine luteal progesterone-binding sites may play a role either in sequestration of newly synthesized progesterone or in the mediation of autocrine and/or paracrine actions of progesterone in the CL.
Animals↗
Studies on the influence of the structural modifications in the tracer on the immunoassay of progesterone.
The main objective of the present study was to examine the influence of different bridges in radioiodinated tracers on the assay performance of progesterone using antibodies. Three homologous and two heterologous immunoassay systems for the measurement of progesterone in human serum are described. Using an antiserum raised against progesterone-11alpha-hemisuccinate-bovine serum albumin (BSA), assays with homologous radioligands, namely progesterone-11alpha-hemisuccinate-125I-tyrosine methyl ester (TME) and progesterone-11alpha-hemisuccinate-125I-histamine, heterologous bridge radioligand, namely progesterone-11alpha-hemiphthalate-125I-TME, and a heterologous site radioligand namely progesterone-3-(O-carboxymethyl) oxime (CMO)-125I-histamine were optimized. A homologous assay system, using antiserum raised against progesterone-3-carboxymethyl oxime-BSA and progesterone-3-CMO-125I-histamine as the radioligand was also optimized to develop a radio-immunoassay (RIA) for serum progesterone. Amongst the two homologous radioligands, viz., progesterone-11alpha-hemisuccinate-125I-histamine and the corresponding TME conjugate tracer, the former yielded a standard curve with a higher slope (-0.6) as compared to the latter (-0.5). The heterologous bridge system with progesterone-11alpha-hemiphthalate-125I-TME resulted in a more sensitive assay (slope of -0.8) than the homologous tracers, whilst the heterologous site radioligand, viz., progesterone-3-CMO-125I-histamine gave the most sensitive assay (slope of -1.2). The homologous assay with antiserum against progesterone-3-CMO-BSA and progesterone-3-CMO-125I-histamine tracer gave a standard curve having a slope of -0.97. The two antibodies developed against progesterone, viz., progesterone-11alpha-hemisuccinate-BSA and progesterone-3-CMO-BSA were characterized for their titre, sensitivity, and specificity. Considering the slope, sensitivity, cross-reactivity, and the quality of tracer, the assay system using antiserum against progesterone-11alpha-hemisuccinate-BSA and progesterone-3-CMO-125I-histamine was found to be suitable for the development of RIA for serum progesterone. The bridges used in an immunogen for production of antibodies, as well as in the preparation of tracer, have a great influence on the assay characteristics.
Animals↗
Stimulatory and inhibitory effects of progesterone on FSH secretion by the anterior pituitary.
The purpose of this study was to investigate whether progesterone exerted progesterone receptor mediated direct effects on the anterior pituitary in the secretion of FSH and whether such effects were mediated through the 5 alpha-reduction of progesterone. Treatment of anterior pituitary dispersed cells for 48 h with 0.5 nM estradiol reduced the ED50 for gonadotropin releasing hormone (GnRH)-stimulated FSH release from 0.58 to 0.36 ng/ml and the ED50 for GnRH-induced LH release from 0.54 to 0.19 ng/ml. When dispersed pituitary cells were treated with 0.5 nM estradiol and exposed to various doses of progesterone for 1 to 6 h, the most consistent rise in basal and GnRH-stimulated FSH release was observed with the 50 nM dose of progesterone with a 3-h exposure period. All three doses of progesterone elevated basal LH and GnRH-stimulated LH was increased by the 50 and 100 nM doses of progesterone during the 3-h period of treatment. Using the 50 nM dose of progesterone, basal and GnRH-stimulated LH was increased after 2, 3 and 6 h of progesterone treatment. When the period of exposure of progesterone was extended to 12, 36 or 48 h, there was a significant inhibition of GnRH-stimulated FSH release. GnRH-stimulated LH release was inhibited at 36 and 48 but not 12 h after progesterone treatment. These studies showed that the effect of progesterone administered for periods of 1 to 6 h enhanced the secretion of LH and FSH whereas progesterone administered for periods beyond 12 h inhibited FSH and LH release by dispersed pituitary cells in culture. These results are similar to those observed in vivo after progesterone treatment. Furthermore estrogen priming of the dispersed pituitary cells was necessary to observe the effects of progesterone. The progesterone antagonist RU486 prevented the progesterone-induced rise in GnRH-stimulated FSH release. Furthermore the 5 alpha-reductase inhibitor N,N-diethyl-4-methyl-3-oxo-4-aza-5 alpha-androstane- 17 beta-carboxamide also prevented the progesterone-induced rise in GnRH-stimulated FSH release in estrogen-treated dispersed pituitary cells. These results indicate that the anterior pituitary is a major site of action of progesterone in the release of FSH and that 5 alpha-reduction of progesterone plays an important role in FSH release.
5-alpha Reductase Inhibitors↗
Progesterone receptor in non-small cell lung cancer--a potent prognostic factor and possible target for endocrine therapy.
A possible involvement of gender-dependent factors has been postulated in development of human non-small-cell lung cancers (NSCLC), but its details remain unclear. In this study, we examined biological significance of progesterone receptor in NSCLCs. Progesterone receptor immunoreactivity was detected in 106 of 228 NSCLCs (46.5%). Progesterone receptor-positive NSCLC was frequently detected in female and adenocarcinoma, and was inversely associated with tumor-node-metastasis stage and histologic differentiation. Progesterone receptor status was also associated with better clinical outcome of the patients, and a multivariate analysis revealed progesterone receptor status as an independent prognostic factor. Progesterone-synthesizing enzymes were detected in NSCLCs, and tissue concentration of progesterone was higher in these cases (n = 42). Immunoblotting analyses showed the presence of progesterone receptor in three NSCLC cell lines (A549, LCSC#2, and 1-87), but not in RERF-LC-OK or PC3. Transcriptional activities of progesterone receptor were increased by progesterone in these three progesterone receptor-positive NSCLC cells by luciferase assays. Cell proliferation was inhibited by progesterone in these progesterone receptor-positive NSCLC cells in a dose-dependent manner, which was inhibited by progesterone receptor blocker. Proliferation of these tumor cells injected into nude mice was also dose-dependently inhibited by progesterone, with a concomitant increase of p21 and p27 and a decrease of cyclin A, cyclin E, and Ki67. Results of our present study suggested that progesterone receptor was a potent prognostic factor in NSCLCs and progesterone inhibited growth of progesterone receptor-positive NSCLC cells. Therefore, progesterone therapy may be clinically effective in suppressing development of progesterone receptor-positive NSCLC patients.
Adult↗
Progesterone inhibits the induction of aromatase activity in rat granulosa cells in vitro.
The development of ovulatory follicles is suppressed during reproductive states with high circulating levels of progesterone. We have investigated the effects of progesterone on estradiol secretion by rat granulosa cells in vitro to determine if progesterone has direct effects on follicular aromatization. Relatively undifferentiated granulosa cells were obtained from hypophysectomized (HPX), diethylstilbestrol-treated immature rats and differentiated granulosa cells were obtained from preovulatory follicles on the morning of proestrus. Progesterone suppressed, in a dose-dependent fashion, the accumulation of estradiol when HPX or proestrous cells were cultured with follicle-stimulating hormone (FSH) (300 ng/ml) and testosterone (0.5 microM), but granulosa cells from proestrous rats were less sensitive to the inhibitory effects of progesterone. When aromatase was first induced in cells from HPX rats by culturing them for 1, 2, or 3 days with FSH, the cells were less susceptible to the inhibitory effects of progesterone during the subsequent 3 days of culture. The inhibitory effects of progesterone on estradiol secretion appeared irreversible when cells were exposed to progesterone for longer than 1 day. The effects of progesterone on HPX granulosa cells were specific since the progesterone precursor, pregnenolone, was as inhibitory as progesterone, while the progesterone metabolite, 17 alpha-hydroxyprogesterone, had no effect. When granulosa cells from proestrous follicles were cultured with testosterone and graded doses of progesterone, a slight inhibition of estradiol production occurred. This inhibition was much less than that observed when FSH was present. Therefore, the inhibitory effects of progesterone are exerted primarily on the induction of aromatase by FSH, rather than on the functioning of existing aromatase. Whole proestrous follicles were also cultured with graded doses of progesterone in the presence or absence of FSH and/or testosterone. In those experiments progesterone increased estradiol on the first day of culture, but inhibited the effects of FSH in maintaining estradiol secretion later in culture. These experiments have shown that progesterone exerts a specific and irreversible inhibitory effect on estradiol production by granulosa cells. Less differentiated granulosa cells appear to be more sensitive to the inhibitory effects of progesterone. These results suggest that in vivo progesterone may directly suppress development of ovulatory follicles by inhibiting aromatase in granulosa cells of developing follicles.
Animals↗
Physiological profiles of episodic progesterone release during the midluteal phase of the human menstrual cycle: analysis of circadian and ultradian rhythms, discrete pulse properties, and correlations with simultaneous luteinizing hormone release.
To define the physiological relationships between episodic progesterone and LH release, we measured serum progesterone and LH concentrations in blood sampled at 10-min intervals for 24 h in seven young women in the midluteal phase of the menstrual cycle. The resultant time series were assessed further by Fourier transformation, Cluster analysis, and cross-correlation analysis with autoregressive modeling. These techniques permitted an examination of circadian rhythms, discrete (ultradian) pulse properties, and simultaneous or lagged correlations between progesterone and LH release. We found the following. 1) Both serum LH and progesterone concentrations had significant circadian periodicities, with similar acrophases (times of maximal nyctohemeral values). LH and progesterone also manifested multiple ultradian rhythms of similar periodicities (range, 48-241 min). 2) Discrete serum progesterone peaks occurred at a mean interpulse interval of 118 +/- 12 (+/- SE) min, had durations of 92 +/- 12 min, and had incremental amplitudes of 4.3 +/- 0.9 ng/mL (14 +/- 3 nmol/L). The frequency and duration characteristics of the progesterone and LH peaks were not significantly different, but progesterone fractional peak amplitudes were one quarter those of LH pulses. 3) Fractional progesterone peak amplitudes in the seven women correlated inversely (r = -0.811) with 24-h mean LH concentrations, suggesting a negative feedback relationship between progesterone and LH release. 4) LH and progesterone interpulse intervals both exhibited significant nyctohemeral variations, with diurnal amplitudes of 73 +/- 12 min for LH and 43 +/- 8.9 min for progesterone (P less than 0.01). 5) Significant positive cross-correlations existed in all seven women between serum LH and progesterone concentrations considered simultaneously and at progesterone time lags of 10-50 min. By autoregressive modeling, the later (20-50 min) cross-correlations could be accounted for by sustained autocorrelations in the individual progesterone and LH time series and significant cross-correlations between LH and simultaneous progesterone concentrations and between LH and 10-min lagged progesterone concentrations. We conclude that progesterone release occurs in a periodic (circadian and ultradian) fashion as well as in a discrete (episodic or pulsatile) mode. Moreover, both positive and negative feedback relationships operate to coordinate LH and progesterone secretion in the midluteal phase of the human menstrual cycle.
Adult↗
[Studies on progesterone receptor in rabbit uterine cytosol -- nuclear translocation and chromatin binding-- (author's transl)].
Estrogen priming increases uterine 8S macromolecule which binds progesterone specifically. Progesterone-8S complex in the cytoplasm enters into nucleus and is bound to chromatin finally. In this paper, the mode of nuclear translocation of steroid in exchange assay of receptor introduced by Anderson et al., and the mode of binding to chromatin were studied on the progesterone-receptor complex in the uterus of estrogen primed female rabbit. 1. After intravenous administration of 200 mug progesterone into the estrogen primed immature rabbit, uterine nuclei were prepared by the method in Table 1. These nuclei were incubated with 3H-progesterone and cold steroids at 4 degrees C for 30 minutes, and then washed with buffer A. The radioactivity of the nuclei was counted. This experiment was performed at 4 degrees C because progesterone receptor and chromatin were observed to be degraded at 37 degrees C for 20 minutes. The effect of cold steroids in vitro on the incorporation of 3H-progesterone into the uterine nuclei of rabbit pretreated with progesterone was found to be similar to their effect on progesterone-receptor binding in cytosol or chromatin (Fig. 1). 2. The effect of cold steroids on 3H-progesterone-receptor-chromatin triplex (Table 2 and Fig. 2) was examined. Once 3H-progesterone-receptor-chromatin triplex was formed, it was difficult to exchange 3H-progesterone to other steroids at 4 degrees C. These results (1 & 2) indicate that progesterone-receptor complex enters into nucleus and is bound to chromatin. Exchange of steroid may occur in the nuclear progesterone-receptor complex, which is free from the binding with chromatin. And thus exchange assay cannot represent quantitative data on receptor content. 3. 3H-progesterone-8S or 5S complexes were obtained by 5 approximately 20% sucrose linear gradient centrifugation (Fig. 3). The same molar concentration of these complexes from estrogen primed or castrated rabbit uterus were incubated with primed uterine chromatin for 30 minutes. Then the chromatin was washed with buffer A and the radioactivity was counted. It was shown in Fig. 4 that 3H-progesterone-8S complex was bound to chromatin much more tightly than 3H-progesterone 5-S complex in preparations obtained from both castrated or primed uterine cytosols. All these results indicate that 8S may be the biologically active form of the receptor. 4.3H-progesterone uptake into uterine nuclei was observed in very limited amount following the injection into uterine artery. The radioactivity in nuclei decreased easily by washing with buffer A as in Fig. 5. The small amount of residual radioactivity after washing, that is, very limited number of binding sites with high affinity is considered to be indicative of biologically active binding.
Animals↗
Control of endometrial oxytocin receptor and uterine response to oxytocin by progesterone and oestradiol in the ewe.
The effects of administration of progesterone and oestradiol on ovine endometrial oxytocin receptor concentrations and plasma concentrations of 13,14-dihydro-15-keto prostaglandin F-2 alpha (PGFM) after oxytocin treatment were determined in ovariectomized ewes. Ewes received progestagen pre-treatment, progesterone and/or oestradiol in 11 different treatment schedules. Progestagen pre-treatment decreased oxytocin receptor concentrations in endometrium from ewes treated subsequently with either progesterone for 5 days or progesterone for 5 days plus oestradiol on Days 4 and 5 of progesterone treatment. Oestradiol increased endometrial oxytocin receptor concentrations when administered on Days 4 and 5 of 5 days progesterone treatment. Progestagen pre-treatment followed by progesterone treatment for 12 days caused a large increase in oxytocin receptors and no further increase occurred when ewes were given oestradiol on Days 11 and 12, or when progesterone was withdrawn on Days 11 and 12, or these two treatments were combined. Oxytocin administration caused an increase in plasma PGFM concentrations in ewes which did not receive progestagen pre-treatment, and subsequently received progesterone treatment for 5 days and oestradiol treatment on Days 4 and 5 of progesterone treatment. Similarly treated ewes which received progestagen pre-treatment did not respond to oxytocin. Oxytocin administration also increased plasma PGFM concentrations in ewes which received progestagen pre-treatment followed by progesterone treatment for 12 days, progesterone treatment for 12 days plus oestradiol on Day 11 and 12 of progesterone treatment, progesterone withdrawal on Day 11 and 12, or progesterone withdrawal and oestradiol treatment combined. The results indicate that (1) progesterone pre-treatment affects oxytocin receptor concentrations in the endometrium and uterine responsiveness to oxytocin and (2) progesterone treatment alone for 12 days after a treatment which mimics a previous luteal phase and oestrus is sufficient to induce oxytocin receptors and increase oxytocin-induced PGF release. These results emphasize the importance of progesterone and provide information which can be used to form an hypothesis for control of luteolysis and oestrous cycle length in the ewe.
Animals↗
Hormonal regulation of oestrogen and progesterone receptors in cultured bovine endometrial cells.
Changes in the number of progesterone and oestradiol receptors in the endometrium are thought to play a role in the induction of luteolysis. The effect of oestradiol and progesterone on the regulation of their receptors in cultured bovine uterine epithelial and stromal cells was examined to determine the mechanisms involved in this process. Cells were obtained from cows at days 1-3 of the oestrous cycle and were cultured for 4 or 8 days in medium alone (RPMI medium + 5% (v/v) charcoal-dextran stripped newborn calf serum) or with oestradiol, progesterone or oestradiol and progesterone. At the end of culture, receptor binding was measured by saturation analysis. Specific binding of both [3H]ORG 2058 (16 alpha-ethyl-21-hydroxy-19-nor (6,7-3H) pregn-4-ene-3,20-dione) and [3H]oestradiol to epithelial and stromal cells showed high affinities (Kd = 1.1 x 10(-9) and 6 x 10(-10) mol l-1, respectively, for progesterone receptors; Kd = 5.5 x 10(-9) and 7 x 10(-10) mol l-1, respectively, for oestradiol receptors). In the stromal cells, oestradiol (0.1-10 nmol l-1) increased the number of oestradiol receptors from 0.21 +/- 0.06 to 0.70 +/- 0.058 fmol microgram-1 DNA and the number of progesterone receptors from 1.4 +/- 0.83 to 6.6 +/- 0.70 fmol microgram-1 DNA in a dose-dependent manner after 4 days of culture (P < 0.01). After culture for 8 days, the stimulatory effect of oestradiol increased. Progesterone (50 nmol l-1) had no effect on the number of oestradiol or progesterone receptors (P > 0.05). However, progesterone inhibited the stimulatory effect of oestradiol. In epithelial cells, the lower concentrations of oestradiol (0.1 and 1 nmol l-1) stimulated the number of progesterone receptors (P = 0.05) after 4 days culture, whereas the highest concentration of oestradiol (10 nmol l-1), progesterone (50 nmol l-1) and progesterone (50 nmol l-1) plus oestradiol (1 nmol l-1) had no effect. After culture for 8 days, the stimulatory effect of oestradiol decreased. In contrast to progesterone receptors, the number of oestradiol receptors increased with oestradiol concentration (P < 0.01). These data show that the number of progesterone receptors was higher in the stromal cells than in epithelial cells, whereas the number of oestradiol receptors was higher in the epithelial cells than in stromal cells. Oestradiol upregulates its own receptor and increases the number of progesterone receptors in both cell types in vitro, whereas progesterone has little effect, but inhibits the effects of oestradiol on progesterone receptors.
Analysis of Variance↗
Failure of protein synthesis inhibition to block progesterone desensitization of lordosis in female rats.
Progesterone's desensitization effect on lordosis has been shown to correlate with a decreased concentration of hypothalamic progestin receptors after progesterone injection. In a recent study, one group of investigators found that the protein synthesis inhibitor anisomycin appeared to block progesterone's desensitization effect. Despite decreased levels of cytoplasmic progestin receptors, progesterone + anisomycin-treated rats exhibited a high level of lordosis four hr after a second progesterone injection. Because this finding conflicts with a progestin receptor model of progesterone's desensitization effect, we investigated it further. In the first experiments, ovariectomized rats were injected with estradiol benzoate followed 24 hr later by either progesterone or vehicle. Anisomycin injected 3 hr after progesterone, at a dose that causes inhibition of hypothalamic protein synthesis for at least 4 hr, was without effect on progesterone desensitization a day later. In other experiments silastic implants containing estradiol were inserted into ovariectomized rats. Forty-five hr later, rats received progesterone or vehicle, followed by injections of anisomycin or saline. Rats receiving anisomycin + progesterone were still highly receptive at 30 hr while saline + progesterone controls were not. Furthermore, the results were similar 4 hr after a second injection of progesterone at 30 hr. In a related experiment, we confirmed that anisomycin delayed dramatically termination of the period of sexual receptivity. In this laboratory anisomycin does not seem to block progesterone's desensitization effect. However, with certain procedures anisomycin delays the termination of sexual receptivity. Thus it is important in investigations of the mechanism of progesterone's desensitization effect that animals be tested prior to the second progesterone injection to determine if they are actually responding to the progesterone.
Animals↗
Antiprogestin ZK-98.299 and progesterone display differential binding characteristics in the human myometrial cytosol.
To investigate whether the synthetic progesterone antagonist ZK-98.299 binds to progesterone receptor or also has distinct binding sites, the binding characteristics of ZK-98.299 were compared with those of progesterone in the human myometrial cytosol. [3H]ZK-98.299 and [3H]progesterone showed specific binding in the myometrial cytosol and the binding of each radiolabelled ligand could be displaced with the respective ligand in a dose-response manner. However, while the binding of [3H]progesterone could be completely blocked with progesterone or ZK 98.299, the binding of [3H]ZK-98.299 could not be displaced more than 50%. The non-specific binding of [3H]ZK-98.299 was very high as compared to that of [3H]progesterone. Using [3H]progesterone, the relative binding affinity (RBA) of progesterone was more than that of ZK 98.299, whereas using [3H]ZK-98.299 the RBA of ZK 98.299 exceeded that of progesterone. Treatment of myometrial cytosol with increasing concentrations of -SH-modifying agents (iodoacetamide (IA) 0-10 mM or N-ethylmaleimide (NEM) 0-1000 nM) decreased the binding of progesterone by over 80%, whereas similar treatment did not have appreciable effect on the binding of [3H]ZK-98.299. Although both preformed ligand-receptor complexes were relatively stable in the presence of IA and NEM, the [3H]progesterone-receptor complex was more sensitive as compared to the [3H]ZK-98.299-receptor complex. The addition of 20 mM molybdate in the cytosol had a protective effect against the -SH-modifying agents. [3H]ZK-98.299 and [3H]progesterone-receptor complexes also showed differential stability when incubated at elevated temperatures (25 degrees C and 37 degrees C), [3H]ZK-98.299-binding sites being more thermolabile as compared to [3H]progesterone binding sites. Prior occupation of the receptor by the two ligands gave the complexes the ability to resist an elevated temperature of 25 degrees C. Moreover, molybdate stabilized both the liganded and unoccupied receptors at 25 degrees C. When the ligand-receptor complexes were applied onto a prefocused polyacrylamide gel, the progesterone and ZK-98.299-receptor complexes were resolved and focused at pH 7.2 and 8.4, respectively. The results of this study suggest that although progesterone and ZK-98.299 are mutually competitive for binding to progesterone receptor, ZK-98.299 also has distinct binding sites.
Binding Sites↗
Effects of progesterone administration on N-nitrosomethylurea-induced rat mammary carcinogenesis.
N-Nitrosomethylurea (NMU), 4 mg/100 g body wt, was given to female Sprague-Dawley rats by i.v. injection on 2 occasions, 4 weeks apart. One group of 20 animals also received 5 mg of progesterone s.c. on the morning before, of and after each NMU dose (acute progesterone treatment). A second group of 21 rats was given progesterone 2.5 mg twice a day throughout the experiment commencing 7 days before the first dose of NMU (chronic progesterone treatment). The third group of 20 animals comprised the NMU-exposed controls. The latent period for mammary tumor development was reduced and the number of tumors per rat was increased by the acute progesterone treatment. The final mammary tumor incidence for the chronic progesterone treatment group (62%) was lower than that of the controls (85%) and the acute progesterone-treated rats (80%), and tumor multiplicity was less. Estrogen receptor levels were significantly higher in tumors from the chronic progesterone group than in those from the acute progesterone-treated animals (P less than 0.01), and progesterone receptor levels were lower in comparison to either of the other 2 groups. Serum progesterone concentrations were subnormal in the NMU-exposed controls but the estrogens were unaffected. The acute progesterone-treated rats also had reduced serum progesterone levels when compared with normal animals, although they were significantly higher than those of the NMU-exposed controls (P less than 0.01). Extremely high serum progesterone levels in rats treated chronically with progesterone were accompanied by reduced estrogen concentrations. Serum prolactin levels were elevated in the NMU-exposed controls and chronic progesterone-treated groups compared with non-NMU-exposed normal rats, while growth hormone concentrations were reduced by progesterone administration. All 3 NMU-exposed groups had elevated serum TSH levels.
Animals↗
Myometrial progesterone responsiveness and the control of human parturition.
OBJECTIVE: The goal of this review is to assess the body of literature addressing the mechanism of progesterone withdrawal in the control of human parturition and in particular the recent advances in testing the hypothesis that human parturition is initiated by decreased myometrial responsiveness to progesterone, ie, functional progesterone withdrawal. METHODS: Published studies of progesterone responsiveness of the pregnant human myometrium in the context of parturition control were reviewed. RESULTS: Advances in understanding the molecular basis for progesterone receptor (PR)-mediated control of progesterone responsiveness has led to the hypothesis that functional progesterone withdrawal in human parturition is mediated by specific changes in myometrial PR expression, function, or both. The human PR exists as two major subtypes, PR-A and PR-B. As PR-A represses progesterone actions mediated by PR-B, the extent of progesterone responsiveness is inversely related to the PR-A/PR-B expression ratio. In women, the onset of term labor is associated with a significant increase in the myometrial PR-A/PR-B expression ratio that may facilitate functional progesterone withdrawal. Interestingly, expression of the estrogen receptor-alpha (ERalpha) increases concordantly with the PR-A/PR-B expression ratio in nonlaboring myometrium. This finding indicates that functional estrogen activation and functional progesterone withdrawal are linked. CONCLUSION: Functional progesterone withdrawal in human parturition is likely mediated by an increase in the myometrial PR-A/PR-B expression ratio and possibly by modulation of coactivator and corepressor proteins. Functional progesterone withdrawal appears to induce functional estrogen activation. Thus, for most of pregnancy, progesterone may decrease myometrial estrogen responsiveness by inhibiting ERalpha expression. Such an interaction would explain why the human myometrium is refractory to the high levels of circulating estrogens for most of pregnancy. At term, functional progesterone withdrawal removes the suppression of ERalpha expression leading to an increase in ERalpha and a concomitant increase in myometrial estrogen responsiveness. Estrogen can then act to transform the myometrium to a contractile phenotype. This model explains why disruption of progesterone action alone triggers the full parturition cascade. The link between functional progesterone withdrawal and functional estrogen activation may be a critical mechanism for the endocrine control of human parturition.
Estrogens↗