Association of raised serum progesterone level with ovulation trigger and histology of endometrium in stimulated cycles

Abstract This was a forthcoming study of those patients, who undergo in-vitro fertilization (IVF) and freeze-all embryo, who acquiesce for the study. The number of participated patients (n=350) in this study, underwent for IVF. The blood sample was collected from patients to evaluate the level of serum progesterone in vacuum vials on the day of ovulation trigger. After 36 hrs of ovulation trigger, ovum picked up was done. Quantitative methods were used to estimate the level of serum progesterone through the electrochemiluminescence immunoassay and correlation of serum progesterone with embryo transfer (ET) outcomes. Main outcome of this current study was to evaluate the value of mean serum progesterone level i.e.0.868± 0.712 ng/ml and 0.88±0.723 ng/ml was found in case of pregnancy positive and negative respectively, at p=0.216 value. In antagonist (n=40) and agonist (n=310) cases, it was 8(20%) and 37(11.94%) PL occurrence was noted at p=0.143 respectively. An overall value of the premature lutenization (PL) occurrences was 13.63% and 15.25% observed in both positive and negative cases of pregnancy at p=0.216 respectively. This study concluded that 12.66% of PL occurrences were recorded in the case of IVF. Study results proved, there were no significant effect of PL on pregnancy outcomes.

Is it time for a paradigm shift in drug research and development in endometriosis/adenomyosis?

BACKGROUND: The drug research and development (R&D) for endometriosis/adenomyosis has been painfully slow. Most completed clinical trials on endometriosis did not publish their results, and presumably failed. While few published trials did report how they foundered, the reasons why they failed are often completely unclear. Surprisingly, there has been no open discussion on why these trials failed. If the causes for these failed trials remain unelucidated, mistakes made in these failed trials may be repeated in the future. Since failure can be infinitely more instructive and educational than success, elucidating the causes for failed clinical trials may yield a treasure trove for future drug R&D. Given our growing understanding of the natural history of ectopic endometrium, it is also important to make an inventory of biologicals/compounds that are currently under development to see where we stand and whether they would stand a better chance of gaining regulatory approval than their predecessors. OBJECTIVE AND RATIONALE: We provide an overview of all compounds under clinical investigation and in development in order to assess the evolution of R&D since the last inventory, reported in 2013. We also have attempted to analyse selected failed clinical trials in the context of published translational/preclinical research and our growing understanding of the natural history of endometriotic/adenomyotic lesions, in the hope that the lessons learned will steer investigators toward the right track in future drug R&D. SEARCH METHODS: We searched ClinicalTrials.gov and a database containing information on drugs gathered daily by Thomson Reuters from a wide range of sources (e.g. patent offices, biomedical literature, congresses, symposia, meetings, company information, regulatory information) for all therapeutic compounds that have undergone or are under clinical trials, or in the developmental stage, and then searched PubMed and Google to determine their publication status using trial identifiers. For trials that were completed at least 2 years ago and have, or have not, published their results, a PubMed search was performed using the name of the therapeutic that has been tested and 'endometriosis' or 'adenomyosis' to identify published preclinical studies prior to the launch of the trial. For those published trials, the cited preclinical studies were also retrieved and scrutinized. OUTCOMES: Despite repeated calls for more transparency, only a small fraction of completed trials on endometriosis has been published. A large number of 'novel' compounds under development are simply repurposed drugs, which seem to be ill-prepared to combat the fibroproliferative nature of endometriosis/adenomyosis. This sobering picture indicates an alarming innovation 'drought' in the drug R&D front, resulting in trickling drug pipelines. Some trials foundered owing to unanticipated serious side-effects, or because attempts were made to suppress a target that can be compensated for by redundant pathways, but many failed in efficacy, indicating that the translational value of the current models is seriously questionable. All existing animal models of endometriosis do not recapitulate the key features of human conditions. WIDER IMPLICATIONS: The glaring innovation drought in drug R&D for endometriosis/adenomyosis should sound alarms to all stake-holders. The failed clinical trials in endometriosis also indicate that some past research had serious deficiencies. In light of the recent understanding of the natural history of ectopic endometrium, it is perhaps time to shift the research paradigm and revamp our research focus and priorities.

Effects of gonadotropin-releasing hormone agonist on human chorionic gonadotropin activity in granulosa cells of immature female rats.

Although the expression of gonadotropin-releasing hormone (GnRH) in the ovaries is well established, its physiological role remains unknown. The aim of this study was to determine whether ovarian GnRH mediates the actions of human chorionic gonadotropin (hCG) in the granulosa cells of immature female rats. Follicular growth was induced by administration of pregnant mare serum gonadotropin (PMSG, 15 IU/0.15 ml) on day 25 after birth, and hCG (20 IU/0.2 ml) was administered on day 27 revealing the increase of plasma progesterone level. Primary cultures of granulosa cells were established from large follicles 2 days after PMSG treatment. Progesterone synthesis was augmented by hCG in a dose-dependent manner. Annexin A5 (ANXA5), a biomarker of GnRH, was expressed in the granulosa-luteal cells after hCG treatment, as shown by immunohistochemistry, suggesting that hCG treatment induced GnRH action. The GnRH mRNA level was increased by hCG, and treatment with GnRH agonist (GnRHa) increased ANXA5 mRNA levels in the primary cultures of granulosa cells. Concomitant incubation of GnRH (10-7 M) or GnRHa (fertirelin acetate, 10-8 M) with hCG suppressed progesterone synthesis during a 3 h incubation period. The mRNA expression of luteinizing hormone receptor (LHR) and follicle-stimulating hormone receptor (FSHR) was synergistically stimulated and suppressed by hCG and GnRHa, respectively. GnRHa stimulated p21 expression, and GnRHa and hCG synergistically reduced the mRNA expression levels of p27 and FOXO1. These data suggest that GnRH induced by LH may have a role for the LH-mediated luteinization of granulosa cells. In addition, ANXA5 may be involved in GnRH action. GnRH-ANXA5 would be an important mechanism in cell differentiation.

A regulatory role of androgen in ovarian steroidogenesis by rat granulosa cells.

Excess androgen and insulin-like growth factor (IGF)-I in the ovarian follicle has been suggested to be involved in the pathophysiology of polycystic ovary syndrome (PCOS). Here we investigated the impact of androgen and IGF-I on the regulatory mechanism of ovarian steroidogenesis using rat primary granulosa cells. It was revealed that androgen treatment with dihydrotestosterone (DHT) amplified progesterone synthesis in the presence of FSH and IGF-I, whereas it had no significant effect on estrogen synthesis by rat granulosa cells. In accordance with the effects of androgen on steroidogenesis, DHT enhanced the expression of progesterogenic factors and enzymes, including StAR, P450scc and 3ßHSD, and cellular cAMP synthesis induced by FSH and IGF-I. Of note, treatment with DHT and IGF-I suppressed Smad1/5/8 phosphorylation and transcription of the BMP target gene Id-1, suggesting that androgen and IGF-I counteract BMP signaling that inhibits FSH-induced progesterone synthesis in rat granulosa cells. DHT was revealed to suppress the expression of BMP-6 receptors, consisting of ALK-2, ALK-6 and ActRII, while it increased the expression of inhibitory Smads in rat granulosa cells. In addition, IGF-I treatment upregulated androgen receptor (AR) expression and DHT treatment suppressed IGF-I receptor expression on rat granulosa cells. Collectively, the results indicate that androgen and IGF-I mutually interact and accelerate progesterone production, at least in part, by regulating endogenous BMP signaling in rat granulosa cells. Cooperative effects of androgen and IGF-I counteract endogenous BMP-6 activity in rat granulosa cells, which is likely to be functionally linked to the steroidogenic property shown in the PCOS ovary.

Butyric acid regulates progesterone and estradiol secretion via cAMP signaling pathway in porcine granulosa cells.

Butyric acid (BA), one of the short chain fatty acids (SCFAs), has positive actions on the metabolism, inflammation, etc. However, whether it influences the reproductive physiology and if so the detail mechanism involved has not yet been determined. In this study, the porcine granulosa cells (PGCs) were treated with gradient concentrations of BA. After 24h culture, 0.05mM BA significantly stimulated the progesterone (P4) secretion (P<0.05), 5mM and 10mM BA significantly inhibited the P4 secretion (P<0.05). Simultaneously, BA up-regulated the estradiol (E2) secretion in a dose dependent manner, 5mM and 10mM BA significantly promoted the E2 level (P<0.05). In addition, 10mM BA significantly promoted the G-protein-coupled receptor 41/43 mRNA (P<0.05). Interestingly, 5mM BA treatment significantly down-regulated cyclic adenosine monophosphate (cAMP) content (P<0.05), steroidogenic acute regulatory (StAR), steroidogenic factor 1 (SF1), P450scc in the mRNA and/or protein level (P<0.05), and these actions were reversed by cAMP activator forskolin (FK). Moreover, the co-treatment of 5mM BA and bupivacaine (BPC, the cAMP inhibitor) significantly accumulated the inhibition action of BPC on cAMP, the secretion of P4, and the abundance of StAR mRNA (P<0.05), inhibited the up-regulation of 5mM BA on the E2 secretion (P<0.05). Further, the Global Proteome and KEGG pathway analysis found that 5mM BA significantly up-regulated the I3LM80 proteins (P<0.05), which is involved in the steroid biosynthesis signaling pathway. 5mM BA significantly decreased the F2Z5G3 protein level (P<0.05), and the cAMP signaling pathway. In conclusion, present findings for the first time demonstrated that BA could regulate the P4 and E2 hormone synthesis in PGCs via the cAMP signaling pathway.

The Rising Phoenix-Progesterone as the Main Target of the Medical Therapy for Leiomyoma.

Leiomyomas, also known as uterine fibroids, are a common benign tumor in women of reproductive age. These lesions disrupt the function of the uterus causing menorrhagia and pelvic pressure as well as reproductive disorders. These women pose a true challenge for clinicians in the attempt of choosing the suitable treatment for each patient. Patient's age, interest in fertility preservation, and leiomyoma location and size are all factors to be taken into account when deciding upon the preferable therapeutic option. For the past few decades, surgical treatment was the only reliable long-term treatment available. A variety of surgical approaches have been developed over the years but these developments have come at the expense of other treatment options. The classical medical treatment includes gonadotropin-releasing hormone (GnRH) agonists and antagonists. These agents are well known for their limited clinical effect as well as their broad spectrum of side effects, inspiring a need for new pharmacological treatments. In recent years, promising results have been reported with the use of selective progesterone receptor modulators (SPRM). Long-term clinical trials have shown a reduction in bleeding and shrinkage of leiomyoma mass. These results instill hope for women suffering from symptomatic leiomyomas seeking an effective, long-term medical option for their condition.

Toxicological effects of fumonisin B1 alone and in combination with other fusariotoxins on bovine granulosa cells.

There is now overwhelming evidence of global contamination of commodities with Fusarium mycotoxins. Fumonisin B1 (FB1) is a Fusarium mycotoxin frequently occurring in corn in combination with deoxynivalenol (DON), α-zearalenol (α-ZEA) and ß-zearalenol (ß-ZEA). The aim of this study was to determine if FB1, alone and combined with DON or α-ZEA or ß-ZEA, can affect cell proliferation and steroid production of bovine granulosa cells (GC). A species-specific model with bovine granulosa cells (GC) was used to study the potential endocrine disruptor effects of FB1 alone and in co-exposure. In the presence of ß-ZEA (30 ng/mL), FB1 at 30 ng/mL showed a stimulatory effect on GC numbers. Insulin-like growth factor-1 (IGF1)-stimulated cell proliferation was decreased after exposure to ß-ZEA alone at 5.0 µg/mL and FB1 with α-ZEA and ß-ZEA at the same concentration. Regarding steroid production, FB1 at 30 ng/mL and 100 ng/mL amplified the inhibitory effect of ß-ZEA (30 ng/mL) on estradiol (E2) production, while FB1 alone increased (P < 0.05) IGF1-induced E2 production. α-ZEA alone decreased (P < 0.05) E2 production, whereas ß-ZEA alone and in combination with FB1 decreased (P < 0.05) E2 production. These studies indicate for the first time that the Fusarium mycotoxin FB1 along with other mycotoxins can affect GC proliferation and steroid production, which ultimately could influence reproductive function in cattle.

Organotin compounds cause structure-dependent induction of progesterone in human choriocarcinoma Jar cells.

Organotin compounds, such as tributyltin (TBT) and triphenyltin (TPT), are typical environmental contaminants and suspected endocrine-disrupting chemicals because they cause masculinization in female mollusks. In addition, previous studies have suggested that the endocrine disruption by organotin compounds leads to activation of peroxisome proliferator-activated receptor (PPAR)γ and retinoid X receptor (RXR). However, whether organotin compounds cause crucial toxicities in human development and reproduction is unclear. We here investigated the structure-dependent effect of 12 tin compounds on mRNA transcription of 3ß-hydroxysteroid dehydrogenase type I (3ß-HSD I) and progesterone production in human choriocarcinoma Jar cells. TBT, TPT, dibutyltin, monophenyltin, tripropyltin, and tricyclohexyltin enhanced progesterone production in a dose-dependent fashion. Although tetraalkyltin compounds such as tetrabutyltin increased progesterone production, the concentrations necessary for activation were 30-100 times greater than those for trialkyltins. All tested active organotins increased 3ß-HSD I mRNA transcription. We further investigated the correlation between the agonistic activity of organotin compounds on PPARγ and their ability to promote progesterone production. Except for DBTCl2, the active organotins significantly induced the transactivation function of PPARγ. In addition, PPARγ knockdown significantly suppressed the induction of mRNA transcription of 3ß-HSD I by all active organotins except DBTCl2. These results suggest that some organotin compounds promote progesterone biosynthesis in vitro by inducing 3ß-HSD I mRNA transcription via the PPARγ signaling pathway. The placenta represents a potential target organ for these compounds, whose endocrine-disrupting effects might cause local changes in progesterone concentration in pregnant women.

Effects of sulpiride and ethylene glycol monomethyl ether on endometrial carcinogenicity in Donryu rats.

Sulpiride and ethylene glycol monomethyl ether (EGME) are known ovarian toxicants that stimulate prolactin (PRL) secretion, resulting in hypertrophy of the corpora lutea and increased progesterone (P4) production. The purpose of the present study was to investigate how the PRL stimulatory agents affected uterine carcinogenesis and to clarify the effects of PRL on endometrial adenocarcinoma progression in rats. Ten-week-old female Donryu rats were treated once with N-ethyl-N'-nitro-N-nitrosoguanidine (20 mg kg(-1) ), followed by treatment with sulpiride (200 ppm) or EGME (1250 ppm) from 11 weeks of age to 12 months of age. Sulpiride treatment inhibited the incidence of uterine adenocarcinoma and precancerous lesions of atypical endometrial hyperplasia, whereas EGME had no effect on uterine carcinogenesis. Sulpiride markedly prevented the onset of persistent estrus throughout the study period, and EGME delayed and inhibited the onset of persistent estrus. Moreover, sulpiride-treated animals showed high PRL and P4 serum levels without changes in the levels of estradiol-17ß, low uterine weights and histological luteal cell hypertrophy. EGME did not affect serum PRL and P4 levels. These results suggest that the prolonged low estradiol-17ß to P4 ratio accompanied by persistent estrous cycle abnormalities secondary to the luteal stimulatory effects of PRL may explain the inhibitory effects of sulpiride on uterine carcinogenesis in rats. Copyright © 2015 John Wiley & Sons, Ltd.

Aniline Is Rapidly Converted Into Paracetamol Impairing Male Reproductive Development.

Industrial use of aniline is increasing worldwide with production estimated to surpass 5.6 million metric tons in 2016. Exposure to aniline occurs via air, diet, and water augmenting the risk of exposing a large number of individuals. Early observations suggest that aniline is metabolized to paracetamol/acetaminophen, likely explaining the omnipresence of low concentrations of paracetamol in European populations. This is of concern as recent studies implicate paracetamol as a disrupter of reproduction. Here, we show through steroidogenic profiling that exposure to aniline led to increased levels of the Δ4 steroids, suggesting that the activity of CYP21 was decreased. By contrast, paracetamol decreased levels of androgens likely through inhibition of CYP17A1 activity. We confirm that aniline in vivo is rapidly converted to paracetamol by the liver. Intrauterine exposure to aniline and paracetamol in environmental and pharmaceutical relevant doses resulted in shortening of the anogenital distance in mice, a sensitive marker of fetal androgen levels that in humans is associated with reproductive malformations and later life reproductive disorders. In conclusion, our results provide evidence for a scenario where aniline, through its conversion into antiandrogenic paracetamol, impairs male reproductive development.

The non-benzodiazepine anxiolytic drug etifoxine causes a rapid, receptor-independent stimulation of neurosteroid biosynthesis.

Neurosteroids can modulate the activity of the GABAA receptors, and thus affect anxiety-like behaviors. The non-benzodiazepine anxiolytic compound etifoxine has been shown to increase neurosteroid concentrations in brain tissue but the mode of action of etifoxine on neurosteroid formation has not yet been elucidated. In the present study, we have thus investigated the effect and the mechanism of action of etifoxine on neurosteroid biosynthesis using the frog hypothalamus as an experimental model. Exposure of frog hypothalamic explants to graded concentrations of etifoxine produced a dose-dependent increase in the biosynthesis of 17-hydroxypregnenolone, dehydroepiandrosterone, progesterone and tetrahydroprogesterone, associated with a decrease in the production of dihydroprogesterone. Time-course experiments revealed that a 15-min incubation of hypothalamic explants with etifoxine was sufficient to induce a robust increase in neurosteroid synthesis, suggesting that etifoxine activates steroidogenic enzymes at a post-translational level. Etifoxine-evoked neurosteroid biosynthesis was not affected by the central-type benzodiazepine (CBR) receptor antagonist flumazenil, the translocator protein (TSPO) antagonist PK11195 or the GABAA receptor antagonist bicuculline. In addition, the stimulatory effects of etifoxine and the triakontatetraneuropeptide TTN, a TSPO agonist, were additive, indicating that these two compounds act through distinct mechanisms. Etifoxine also induced a rapid stimulation of neurosteroid biosynthesis from frog hypothalamus homogenates, a preparation in which membrane receptor signalling is disrupted. In conclusion, the present study demonstrates that etifoxine stimulates neurosteroid production through a membrane receptor-independent mechanism.

Human steroidogenesis: implications for controlled ovarian stimulation with exogenous gonadotropins.

In the menstrual cycle, the mid-cycle surge of gonadotropins (both luteinising hormone [LH] and follicle-stimulating hormone [FSH]) signals the initiation of the periovulatory interval, during which the follicle augments progesterone production and begins to luteinise, ultimately leading to the rupture of the follicle wall and the release of an oocyte. The administration of gonadotropins in controlled ovarian stimulation (COS) leads to supraphysiological steroid concentrations of a very different profile compared with those seen during natural cycles. It has been suggested that these high steroid concentrations cause alterations in endometrial development, affecting oocyte viability in assisted reproductive technology. Furthermore, it has been proposed that elevated progesterone levels have a negative effect on the reproductive outcome of COS. This may arise from an asynchrony between embryo stage and endometrium status at the window of implantation. The regulation of progesterone production by the developing follicles during COS is a complicated interplay of hormonal systems involving the theca and granulosa cells, and the effect of the actions of both LH and FSH. The present paper reviews current knowledge of the regulation of progesterone in the human ovary during the follicular phase and highlights areas where knowledge remains limited. In this review, we provide in-depth information outlining the regulation and function of gonadotropins in the complicated area of steroidogenesis. Based on current evidence, it is not clear whether the high levels of progesterone produced during COS have detrimental effects on fertility.

[Selective progesterone receptor modulators and their therapeutical use]. / Selektivní modulátory progesteronového receptoru a jejich terapeutické vyuzití

Currently developed selective progesterone receptor modulators (SPRMs) are steroid derived compounds with a bulky radical substitution at carbon 11. They interact with progesterone receptor and exert antagonistic or/and agonistic effects. Mifepristone was approved for pregnancy termination and ulipristal acetate as emergency contraception and pharmacological therapy of uterine fibroids. SPRMs inhibit endometrial proliferation and myoma growth, this suggests a therapeutical effect in cases of endometriosis and other estrogen-dependent diseases.

An early single dose of progesterone agonist attenuates endogenous progesterone surge and reduces ovulation rate in immature rat model of induced ovulation.

Inhibition of preovulatory synthesis and action of progesterone impairs ovulation in rodents. We evaluated effects of supplementation of exogenous progesterone on human chorionic gonadotropin (hCG)-induced ovulatory response in immature rats. Equine CG-primed mature follicles responded to hCG with induction of immunoreactive steroidogenic acute regulatory protein (StAR) mainly in thecal layers and a transient enhancement in progesterone synthesis peaking at 6h after hCG (hCG6h). A single dose of natural progesterone or a synthetic agonist (MP) at hCG0h both decreased ovulation rates in dose-dependent manners. MP was still effective when treated at hCG4h. Treatment with these agents at hCG0h reduced circulating progesterone and thecal expression of StAR at hCG6h. The treatments further attenuated induction of cyclooxygenase (COX)-2 in mural granulosa cells and ovarian prostaglandin (PG) E(2) level at hCG8h. We also found a significant reduction in bromo-deoxyuridine incorporation by mural granulosa cells. Obtained results show that the early treatment with exogenous progesterone agonist caused attenuated amplitude of endogenous progesterone surge, reduced COX-2/PGE(2) system, dysregulated mitosis of granulosa cells, and decreased oocytes release. We suggest that optimal progesterone synthesis and action are an early critical component of hCG-initiated ovulatory cascade that regulates biochemical function of granulosa cells.

Cyclin D1 enhances the response to estrogen and progesterone by regulating progesterone receptor expression.

Estrogen and progesterone are the defining hormones of normal female development, and both play critical roles in breast carcinogenesis. Cyclin D1 is a breast cancer oncogene whose amplification is linked to poor prognosis in estrogen and progesterone receptor-positive breast cancers. Here we report that cyclin D1 regulates progesterone receptor expression, consequently enhancing responses to estrogen and progesterone. Estrogen treatment of cyclin D1 transgenic mice increased progesterone receptor expression and induced mammary hyperplasias that were stimulated by progesterone and blocked by a progesterone antagonist. Progesterone receptor levels decreased in cyclin D1 knockout mice. Cyclin D1 regulated progesterone receptor expression through a novel estrogen- and cyclin D1-responsive enhancer in DNA encoding part of the 3' untranslated region of the progesterone receptor gene. Small inhibitory RNAs for cyclin D1 decreased progesterone receptor expression and estrogen receptor binding to the 3' enhancer region in human breast cancer cells. Since estrogen and progesterone regulate cyclin D1, our results suggest that cyclin D1's participation in a feed-forward loop could contribute to increased breast cancer risks associated with estrogen and progesterone combinations. Additionally, its regulation of the progesterone receptor identifies a novel role for cyclin D1 in ovarian hormone control of breast development and breast carcinogenesis.

Seasonality, estrous cycle characterization, estrus synchronization, semen cryopreservation, and artificial insemination in the Pacific white-sided dolphin (Lagenorhynchus obliquidens).

The reproductive physiology of the Pacific white-sided dolphin, Lagenorhynchus obliquidens, was characterized to facilitate the development of artificial insemination (AI) using cryopreserved spermatozoa. Specific objectives were to: 1) describe reproductive seasonality of the Pacific white sided dolphins; 2) describe urinary LH and ovarian steroid metabolites during the estrous cycle; 3) correlate LH and ovarian steroidal metabolite patterns to ultrasound-monitored follicular growth and ovulation; and 4) assess the efficacy of synchronizing estrus, sperm collection/cryopreservation, and intrauterine insemination. Ovulations (64%, n=37) and conceptions (83%, n=18) occurred from August to October. Peak mean serum testosterone (24 ng/ml), cross-sectional testicular area (41.6 cm(2)), and sperm concentration (144.3 x 10(7) sperm/ml) occurred in July, August, and September respectively. Spermatozoa were only found in ejaculates from July to October. Estrous cycles (n=22) were 31 d long and were comprised of a 10 d follicular and 21 d luteal phase. Ovulation occurred 31.2 h after the onset of the LH surge and 19.3 h after the LH peak. Follicular diameter and circumference within 12 h of ovulation were 1.52 and 4.66 cm respectively. Estrus synchronization attempts with altrenogest resulted in 17 (22%) ovulatory cycles with ovulation occurring 21 d post-altrenogest. Ten AI attempts using cryopreserved semen resulted in five pregnancies (50%). The mean gestation length was 356 days (range 348-367). These data provide new information on the Pacific white-sided dolphin's reproductive physiology and collectively enabled the first application of AI in this species.

Danazol regulates the functions of normal human endometrial stromal cell subpopulations by modifying endometrial cytokine networks.

Local danazol therapy can improve endometriotic signs and symptoms without causing any menstrual disorders. As a consequence, certain direct actions of danazol on endometriotic tissues have been proposed, but the mechanisms of these actions have not been clarified. In the present study, the direct effects of danazol on normal human endometrial stromal cells (ESCs) were examined using in vitro decidualization assays. Danazol did not affect the viable cell numbers of unstimulated ESCs or 8Br-cAMP-stimulated decidualized ESCs, but significantly enhanced the viable cell numbers of 8-Br-cAMP-stimulated ESCs during decidualization in a dose-dependent manner. Danazol had no effect on PRL secretion by 8-Br-cAMP-stimulated decidualized ESCs. Danazol, as well as progesterone and medroxyprogesterone acetate (MPA), induced ESC decidualization. Danazol synergistically enhanced the differentiation process of 8-Br-cAMP-stimulated ESCs during decidualization. Although progesterone and MPA increased G-CSF and IL-8 secretion by ESCs in similar manner to 8-Br-cAMP, danazol had no such effects. Moreover, remarkable increases in G-CSF and IL-8 secretions by 8-Br-cAMP-stimulated ESCs during decidualization were completely inhibited by cotreatment with danazol. These results indicate that danazol has specific pharmacological effects on ESCs, rather than progesterone-like effects or similar effects to those reported for endometrial cytokines. According to the results, normal human ESCs can be classified into at least four functional subpopulations. Therefore, under certain circumstances, danazol has similar or opposite effects on ESCs to certain endometrial cytokines, suggesting that it regulates functional cellular subpopulation ratios of normal human ESCs by modifying the endometrial cytokine network in endometrial stromal tissues.

Spontaneous regression of an asymptomatic meningioma associated with discontinuation of progesterone agonist administration.

An 80-year-old male visited the hospital as an outpatient with a head injury sustained in a traffic accident. Brain computed tomography incidentally revealed a left frontal lobe tumor measuring 5 cm in a diameter. The patient had a history of taking chlormadinone acetate (a progesterone agonist) prescribed several years previously as treatment for benign prostatic hypertrophy. The tumor was seen as an isointense lesion on T(1)-weighted magnetic resonance (MR) images with enhancement by gadolinium, and as a heterogeneously hyperintense mass on T(2)-weighted MR images. The tentative diagnosis was left frontal meningioma attached to the sphenoid ridge or sphenoid plane. The patient was managed conservatively because of his advanced age and no symptoms or progression were observed during a 9-month follow-up period. The medication for benign prostatic hypertrophy was changed from chlormadinone acetate to naftopidil (an alpha-2-blocker) about 9 months after his first presentation. The patient presented again 2 years later complaining of dizziness. Computed tomography and MR imaging performed at this time revealed remarkable regression of the tumor. The signal intensity change with regression of the tumor on T(2)-weighted images was observed as a hypointense lesion. Thus, we wish to emphasize that treatment of meningiomas, especially those diagnosed incidentally, must be based on a thorough consideration of any history of hormonal therapy with prostate disease.

Screening of some anti-progestin endocrine disruptors using a recombinant yeast based in vitro bioassay.

The present study was aimed to develop a sensitive, fast and user friendly progesterone receptor transactivation assay using recombinant yeast cells, Saccharomyces cerevisiae, modified to express human progesterone receptor (PR) and progesterone response element (PRE) driving the expression of green fluorescent protein. Stimulation of cells with increasing concentrations of progesterone resulted in significant elevation in fluorescence activity, with the minimum effective dose of progesterone being 0.1 nM. RU486, significantly inhibited progesterone induced transactivation and non-progesterogenic steroids failed to transactivate PR till 10 microM concentrations. About 7 different chemicals (mostly pesticides or their metabolites) like DDT and its metabolites, nonylphenol, endosulfan were screened in this assay system for their role in transactivation and they were all found to be anti-progestative and IC50 values within the range of 3-20 microM. Further, the assay was used to analyze the endocrine disrupting activity of extracted water samples from leather industries known for their high content of various chemicals and it was found to be rich in anti-progestative compounds. It resulted in about 30% reduction in transactivation. In conclusion, we demonstrated that this yeast based bioassay provides a rapid and robust assay for high throughput screening of (anti)progestative compounds from various sources.

11-(pyridinylphenyl)steroids--a new class of mixed-profile progesterone agonists/antagonists.

Recently, a new class (often referred to as SPRMs: selective progesterone receptor modulators) of progesterone receptor ligands with mixed agonist/antagonist properties has been described. Such compounds are envisaged, for example, as treatment for endometriosis, uterine fibroids, and leiomyomas. Existing SPRMs include Asoprisnil 1 and Uliprisnil acetate 2. In our hands, however, these compounds proved to have a predominantly or exclusively antagonistic in vitro profile, which may make this type of compound less attractive, for example, as contraceptives. We therefore aimed at a class of mixed-profile compounds that would show a more evenly balanced agonist/antagonist profile. A novel class of 11beta-[4-(heteroaryl)phenyl]-substituted pregnanes was identified that displayed the desired balance. Contrary to known SPRMs, this novel class of MPP (mixed-profile progestagen) was found to have a truly mixed activity, including a sizeable agonist component.