Case report: an identical twin with Sertoli-Leydig cell tumor.

Our report details the workup and management of a 43-year-old woman with an identical twin who presented with 2 years of virilization and secondary amenorrhea. Serum total testosterone was elevated. An MRI did not identify adnexal or adrenal pathology. Subsequent ovarian vein sampling demonstrated unilateral testosterone elevation. The patient underwent laparoscopic unilateral oophorectomy resulting in the diagnosis of Sertoli-Leydig cell tumor (SLCT). Although SLCT is a rare sex-cord ovarian tumor, it is associated with endometrial hyperplasia and malignancy. Our goals are to review the workup of androgen-secreting tumors and discuss the clinical importance of the DICER1 mutation in the context of SLCT. In this case, an identical twin underwent DICER1 testing which was one of the essential steps in her clinical management.

Adverse perinatal outcomes associated with crown-rump length discrepancy in in vitro fertilization pregnancies.

OBJECTIVE(S): To determine whether an association exists between small crown-rump length (CRL) and adverse obstetrical outcomes in pregnancies conceived by IVF and to compare a CRL reference based on IVF pregnancies to a reference based on spontaneous pregnancies. DESIGN: Retrospective cohort study. CRL was classified as small by comparing it with the local university hospital maternal fetal medicine standard and the Monash IVF reference chart. SETTING: University-affiliated fertility center. PATIENT(S): Singleton pregnancies conceived by IVF with ultrasounds performed between 7+0 and 8+6 weeks of gestational age. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Pregnancy loss, preterm birth, and low birth weight. RESULT(S): Included were 940 clinical pregnancies. The overall and CRL-discrepant miscarriage rates were 12.7% and 41%, respectively. When CRL was small, the maternal age-adjusted odds of miscarriage were 13.8 times higher (95% confidence interval [CI], 8.9-21.6). At age 30, small CRL was associated with a 30% risk of miscarriage, versus 61% at age 45. There was no association between small CRL and preterm birth or low birth weight. The sensitivity and specificity for predicting miscarriage from the optimal Monash cut point were 0.69 (95% CI, 0.61-0.77) and 0.84 (95% CI, 0.82-0.87), which were similar to those of the CRL reference based on spontaneous pregnancies. CONCLUSION(S): Small CRL in IVF pregnancy was strongly associated with miscarriage, especially in the context of advanced maternal age. Small CRL was not associated with preterm birth or low birth weight. A CRL reference based on IVF pregnancies was equivalent to the standard reference for predicting miscarriage.

Peak Serum Estradiol Level During Controlled Ovarian Stimulation Is not Associated with Lower Levels of Pregnancy-Associated Plasma Protein-A or Small for Gestational Age Infants: A Cohort Study.

OBJECTIVE: During controlled ovarian stimulation in IVF, supraphysiologic levels of estradiol (E2) have been associated with poor placentation and adverse pregnancy outcomes. This study aimed to investigate whether high peak E2 on the day of human chorionic gonadotropin trigger is associated with low pregnancy-associated plasma protein-A (PAPP-A) and adverse perinatal outcomes. METHODS: We performed a retrospective cohort study at a private, university-affiliated fertility centre in Vancouver, BC. We enrolled 216 patients with a singleton pregnancy after fresh embryo transfer who also underwent first trimester screening. Adverse perinatal outcomes were collected from a local registry and included preterm birth, hypertension in pregnancy, antepartum hemorrhage, intrauterine growth restriction, SGA, stillbirth, admission to the NICU, and neonatal death. RESULTS: High serum E2 (≥13 035 pmol/L) at controlled ovarian stimulation was not correlated with low PAPP-A (<0.4 multiples of the median) at first trimester screening (P = 0.46). When each adverse outcome was analysed separately, there was no association between high E2 and any of the outcomes (P > 0.05 for all). High peak E2 was not associated with a total composite of maternal and neonatal adverse birth outcomes (P = 0.30). CONCLUSION: Our results do not support the theory that high E2 at fresh embryo transfer impedes placentation. We found no association between peak E2 and low PAPP-A levels or adverse pregnancy outcomes.

Longer ovarian stimulation reduces embryo number and clinical pregnancy rate in long GnRH agonist cycles.

BACKGROUND: The aim of this study was to analyze whether the length of controlled ovarian stimulation affects in vitro fertilization (IVF) cycle outcomes. METHODS: This retrospective cohort study was performed at a private, university-affiliated fertility centre. We reviewed 1522 IVF cycles, comprising 979 long gonadotropin-releasing hormone (GnRH) agonist and 543 GnRH antagonist protocols. All subjects underwent controlled ovarian stimulation followed by fresh embryo transfer. Logistic regression analysis was used to examine the relationship between trigger day and the following cycle outcomes: normal fertilization rate (FR), proportion of mature oocytes, proportion of cycles with embryos for cryopreservation, and clinical pregnancy rate (CPR). RESULTS: In long agonist cycles, having more days of stimulation was associated with a lower clinical pregnancy rate (OR=0.87, 95% CI=0.80-0.96, P=0.01). Longer stimulation also resulted in fewer cycles with supernumerary embryos for cryopreservation (OR=0.84, 95% CI=0.77-0.92, P=0.0005), despite a having greater number of mature oocytes retreived (OR=1.05, 95% CI=1.01-1.10, P=0.04). For each additional day of stimulation in a long agonist protocol, the odds of achieving a clinical pregnancy were reduced by 13% and of achieving cryopreservation by 16%. In the antagonist protocol group, the length of ovarian stimulation did not have an effect on the clinical pregnancy and cryopreservation rates. CONCLUSIONS: Longer duration of ovarian stimulation appears to reduce clinical pregnancy and embryo cryopreservation rates in subjects undergoing long GnRH agonist cycles. The number of days of stimulation does not appear to affect those using the GnRH antagonist protocol.

Malignant ovarian Sertoli-Leydig cell tumor localized with selective ovarian vein sampling.

Sertoli-Leydig cell tumors (SLCT) are rare, comprising less than 0.5% of ovarian neoplasms. They are most often diagnosed in premenopausal women and may produce androgens, resulting in hirsuitism, voice deepening, frontal balding, terminal hair growth, and clitoromegaly. SLCT are malignant in 15%-20% of cases. We discuss a 25-year-old patient with persistent hyperandrogenemia. Noninvasive imaging cannot conclusively differentiate between SCLT and other diagnoses such as polycystic ovary syndrome, ovarian hyperthecosis, idiopathic hyperandrogenism, idiopathic hirsuitism, and 21-hydroxylase-deficient nonclassic adrenal hyperplasia. Selective ovarian vein sampling revealed a 15-fold greater testosterone production from the right ovary compared with the left, which guided appropriate surgical management.

The mechanism for protein kinase C inhibition of androgen production and 17alpha-hydroxylase expression in a theca cell tumor model.

INTRODUCTION: In polycystic ovary syndrome (PCOS), there is increased formation of androgens by thecal cells. Moreover, PCOS ovaries have been shown to have decreased levels of c-fos transcription factor. We hypothesize that c-fos expression inhibits 17alpha-hydroxylase 17,20 lyase (CYP17) activity in the human ovary, and its decreased expression seen in PCOS may lead to elevated CYP17 transcription, resulting in increased androgen production. OBJECTIVE: Our objective was to define the role of the activator protein-1 transcription factors, namely c-fos, in the regulation of CYP17 expression in theca cells. METHODS: Human ovarian thecal-like tumor cells were used for all experiments. The following techniques were used: steroid quantification, mRNA extraction, microarray analysis, transfection, small interfering RNA, and immunohistochemistry. RESULTS: Stimulation of human ovarian thecal-like tumor cells with the protein kinase A pathway activator forskolin resulted in stimulation of C19 androgen production. In contrast, treatment with the protein kinase C pathway activator tetradecanoylphorbol acetate (TPA) resulted in decreased androgen production with a shift toward C21 progesterone production. TPA also led to complete inhibition of CYP17. Microarray data showed a 37-fold increase in c-fos after treatment with TPA. Transfection with steroidogenic factor 1 resulted in an increase in CYP17 promoter activity, which was significantly inhibited in the presence of c-fos. c-fos gene silencing led to an increase in CYP17 mRNA levels. Immunohistochemical staining for c-fos in ovaries demonstrated strong staining in granulosa cells, but not theca. CONCLUSIONS: The activator protein-1 transcription factor c-fos plays a role in the inhibition of CYP17 expression. The decreased levels of c-fos expression in polycystic ovaries may be responsible for increased CYP17 levels in PCOS.

Effect of vitamin E supplementation with and without hormone therapy on circulatory inflammatory markers in postmenopausal women.

OBJECTIVE: To investigate the effect of vitamin E with and without estrogen replacement therapy or hormone therapy (HT) on inflammatory markers in postmenopausal women. DESIGN: Prospective, observational study, followed by a randomized, prospective, double-blind study. SETTING: Healthy volunteers in an academic medical referral center in Dallas, Texas. PATIENT(S): Seventy-five postmenopausal healthy women, ages 40 to 65 years, with follicle-stimulating hormone (FSH) levels greater > or =30 mIU/mL and a serum estradiol level < or =30 pg/mL. INTERVENTION(S): After enrollment, all women were studied at baseline and received vitamin E for 4 weeks. They were then randomized from week 4 to week 12 to receive vitamin E in conjunction with conjugated equine estrogen (CEE) (0.625 mg), CEE (0.625 mg) plus medroxyprogesterone acetate (MPA) (2.5 mg), or placebo. MAIN OUTCOME MEASURE(S): Change from baseline and between groups effects of vitamin E with and without estrogen or hormone therapy on seven circulatory inflammatory markers in postmenopausal women. RESULT(S): Vitamin E levels increased to a similar extent in all three groups compared with baseline at weeks 4 and 12. Vitamin E increased serum interleukin-6 levels. Combination CEE plus MPA significantly increased C-reactive protein levels. However, there were no consistent statistically significant effects on six other inflammatory markers. CONCLUSION(S): Vitamin E attenuated C-reactive protein increases in postmenopausal women treated with estrogen replacement therapy but not with HT. Because there was no other persistent effect on six additional inflammatory markers, it can be concluded that vitamin E and HT do not play a major role in promoting a change in cardiovascular inflammatory markers.

The post-menopausal ovary displays a unique pattern of steroidogenic enzyme expression.

BACKGROUND: While menopause results in the loss of cyclic steroid production, evidence exists for persistent, albeit reduced, ovarian androgen production. In order to continue to synthesize ovarian androgens, the steroidogenic enzymes necessary for androgen biosynthesis must be present. Few studies have selectively analysed some of the steroidogenic enzymes present in the post-menopausal ovary (PMO), and a comprehensive study of this matter has never been undertaken. METHODS: RNA and protein were obtained from PMO, pre-menopausal ovarian stroma, corpora lutea (CL), ovarian follicles, placenta, and myometrium. Oligonucleotide microarray analysis was performed to compare the gene expression profiles of PMO with pre-menopausal ovarian stroma. Real-time RT-PCR was performed for LH/HCG receptor (LHCGR), steroidogenic acute regulatory (StAR), cholesterol side-chain cleavage (CYP11A), 3beta-hydroxysteroid dehydrogenase type I (HSD3B1) and type II (HSD3B2, 3betaHSD), 17a-hydroxylase (CYP17), cytochrome b5 (CytB5), and aromatase (CYP19). Western blot analysis was performed for StAR, CYP11A, CYP17,and 3betaHSD. RESULTS: The PMO and pre-menopausal ovarian stroma had a similar pattern of steroidogenic enzyme expression. The PMO had persistent, but reduced, levels of LHCGR and most steroidogenic enzymes. CYP19 and HSD3B2 mRNA were greatly reduced in PMO in comparison with CL (50-fold and 2000-fold less respectively). HSD3B2 was not detectable in PMO by western analysis. CONCLUSIONS: This study supports the idea that the PMO retains some steroidogenic capacity. However, based on steroidogenic enzyme expression, the PMO has a unique pattern of steroidogenic enzyme expression that favors Delta5 steroid formation over Delta4 steroid formation.

Human myometrial gene expression before and during parturition.

Identification of temporal and spatial changes in myometrial gene expression during parturition may further the understanding of the coordinated regulation of myometrial contractions during parturition. The objective of this study was to compare the gene expression profiles of human fundal myometrium from pregnant women before and after the onset of labor using a functional genomics approach, and to further characterize the spatial and temporal expression patterns of three genes believed to be important in parturition. Fundal myometrial mRNA was isolated from five women in labor and five women not in labor, and analyzed using human UniGEM-V microarrays with 9182 cDNA elements. Real-time polymerase chain reaction using myometrial RNA from pregnant women in labor or not in labor was used to examine mRNA levels for three of the genes; namely, prostaglandin-endoperoxide synthase 2 (PTGS2), calgranulin B (S100A9), and oxytocin receptor (OXTR). The spatial expression pattern of these genes throughout the pregnant uterus before and after labor was also determined. Immunolocalization of cyclooxygenase-2 (also known as PTGS2) and S100A9 within the uterine cervix and myometrium were analyzed by immunohistochemistry. Few genes were differentially expressed in fundal myometrial tissues at term with the onset of labor. However, there appears to be a subset of genes important in the parturition cascade. The cellular properties of S100A9, its spatial localization, and dramatic increase in cervix and myometrium of women in labor suggest that this protein may be very important in the initiation or propagation of human labor.

The NGFI-B family of transcription factors regulates expression of 3beta-hydroxysteroid dehydrogenase type 2 in the human ovary.

The nerve growth factor-induced clone B (NGFI-B) family of transcription factors are orphan members of the steroid hormone receptor superfamily. The NGFI-B expression was recently shown in the rat ovarian tissue and appears to be regulated by gonadotrophins. The purpose of our study was to investigate the role of the three members of this family [NGFI-B, Nur-related factor 1 (NURR1) and neuron derived orphan receptor 1 (NOR-1)] in the transcription of genes that encode key steroidogenic enzymes and examine expression in the human ovary. Real-time RT-PCR was used to quantify mRNA expression levels of the NGFI-B family members in human ovarian follicles, corpora lutea and in human granulosa cells after FSH, phorbol ester (TPA) and forskolin treatment. NGFI-B was expressed at higher levels than both NURR1 and NOR-1 in both ovarian follicles and corpora lutea. In human granulosa tumour (HGT) cells, the NGFI-B expression increased after TPA, and to a lesser extent, after forskolin treatment. Treatment of primary cultures of human granulosa cells with forskolin and FSH rapidly increased the NGFI-B mRNA levels followed by an increase in 3beta-hydroxysteroid dehydrogenase type 2 (HSD3B2). Transcription of HSD3B2 was studied by transfecting NGFI-B and steroidogenic factor 1 (SF1) expression vectors with reporter constructs prepared with human steroidogenic acute regulatory protein, cholesterol side-chain cleavage, and HSD3B2 genes. NGFI-B increased the transcription of HSD3B2 in HGT cells which is significantly more than SF1. Mutation or deletion of the NGFI-B response element in the HSD3B2 promoter significantly reduced the NGFI-B-mediated transcription of HSD3B2. Therefore, our data suggest that the NGFI-B may play a significant role in up-regulation of HSD3B2 that leads to the increase in progesterone production that is seen in granulosa cells at ovulation.

The orphan nuclear receptor, liver receptor homolog-1, regulates cholesterol side-chain cleavage cytochrome p450 enzyme in human granulosa cells.

After ovulation, there is a shift in ovarian steroidogenesis from an estrogen-producing ovarian follicle to a progesterone-producing corpus luteum. The first step in human ovarian steroidogenesis is catalyzed by cholesterol side-chain cleavage cytochrome P450 (CYP11A1) enzyme. Steroidogenic factor-1 is an orphan nuclear receptor that regulates several steroidogenic enzymes, including CYP11A1. Liver receptor homolog-1 (LRH-1) is another orphan nuclear receptor that is expressed in the human ovary. After ovulation there is a down-regulation in steroidogenic factor-1, which is associated with an up-regulation of LRH-1 expression. These changes coincide with increased level of CYP11A1 expression in human corpus luteum. In this study, we examined the role of LRH-1 in the regulation of human granulosa cell CYP11A1 expression. Cotransfection of human granulosa cell tumor cells with CYP11A1 promoter and LRH-1 expression vector resulted in a significant increase in CYP11A1 expression. Deletion analysis revealed two putative LRH-1 binding sites at -1580 and -40, which was confirmed by EMSA. Dosage-sensitive sex-reversal-adrenal hypoplasia congenita critical region on the X chromosome, gene-1 inhibited LRH-1 stimulated CYP11A1 expression, and that was not overcome by the presence of PKA agonist. We conclude that CYP11A1 expression in human granulosa cells is regulated by LRH-1. We propose that LRH-1 could be the major transcription factor for the post-ovulatory surge in human ovarian steroidogenesis.

Ovarian granulosa cell lines.

The ovary is a complex endocrine gland responsible for production of sex steroids and is the source of fertilizable ova for reproduction. It also produces various growth factors, transcription factors and cytokines that assist in the complex signaling pathways of folliculogenesis. The ovary possesses two primary steroidogenic cell types. The theca cells (and to a lesser extent, the stroma) are responsible for androgen synthesis, and the granulosa cells are responsible for conversion of androgens to estrogens, as well as progesterone synthesis. These cells undergo a transformation in the luteal phase of the menstrual cycle, converting them from estrogen producing, to predominantly progesterone producing cells. Understanding the molecular mechanisms regulating these cells is essential in understanding the regulation of steroidogenesis and reproduction. Creation of appropriate in vitro cell model systems can provide important tools for the study of ovarian function. This has led to the development of ovarian steroidogenic cell lines in several laboratories. Developing theca cell lines has met with limited success. Conversely, numerous human and animal granulosa cell lines have been developed. This review will discuss the existing granulosa cell lines and their characteristics.

The rise in adrenal androgen biosynthesis: adrenarche.

Adrenarche is characterized by the increase in adrenal androgen production, namely dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulfate (DHEAS) that occurs around 6 years of age. These steroids are secreted by the zona reticularis (ZR) of the adrenal gland. This is associated with pubarche or the increase in androgen-dependent hair growth at the time of puberty. The increase in adrenal androgen production can be explained by the increase in the expression of DHEA-synthesizing steroidogenic enzymes in the ZR. Adrenarche is an event independent of gonadarche and is found only in humans and select nonhuman primates. Although numerous prenatal and postnatal factors are important in the onset of adrenarche, a specific adrenal cortical androgen-stimulating hormone has not been identified. Evidence also exists for a role for adrenarche in behavior, skeletal maturation, and postpubertal well-being. Adrenarche is influenced by sex and race, and some of this variation may be related to the insulin and insulin-like growth factor (IGF) signaling pathways. In addition, children with premature and exaggerated adrenarche may be predisposed to certain diseases later in life.