Estrogen-Independent Ovary Formation in the Medaka Fish, Oryzias latipes

To investigate whether a female sex steroid, estrogen, acts as a natural inducer of female gonadal sex determination (or ovary formation) in the medaka fish, Oryzias latipes, the effects of an aromatase inhibitor and anti-estrogens on sexual differentiation of gonads were examined. We found that both drugs did not show any discernible effects on the genetically determined sex differentiation in both sexes. However, the aromatase inhibitor impaired the paradoxical effects of androgen (a male sex steroid), and the anti-estrogens inhibited the male-to-female sex reversal caused by estrogen. Treatments of the fertilized eggs with androgen disturbed the gonadal sex developments in both sexes, suggesting that sex steroid synthesis is detrimental to the gonadal sex developments in the medaka embryos. These results are consistent with the previous observation that sex steroids are not synthesized before the onset of gonadal sex differentiation, and suggest that ovary formation in the genetic females of the medaka fish is not dependent on estrogen.     

Assay and Characterization of Ovarian Testosterone Receptor

Abstract

Ovarian cytosol from immature, hypophysectomized rats was used to characterize the biochemical properties of ovarian androgen receptor and to develop a reliable and convenient assay procedure for its measurement in small quantities of tissue. The results show that Sephadex G-25 columns provide a rapid and reliable method to separate bound from free testosterone for assay of androgen receptor in the ovarian tissue. The binding affinity (K_D=0.6 nM) and sedimentation value (ES) for the ovarian androgen receptor are similar to other steroid receptors and is saturable. It is inactivated by heat and protease digestion and the binding is specific for potent androgens. The ovary contains approximately equal quantities of androgen and estrogen receptor. A physiological role for the presence of androgen receptor in the ovarian tissue is proposed.     

Absence of mutations in the coding regions of follicle-stimulating hormone receptor gene in Singapore Chinese women with premature ovarian failure and polycystic ovary syndrome.

Normal gonadal function is critically dependent on the integrity of pituitary-gonadal axis, where follicle-stimulating hormone (FSH) plays a key role. In the female, FSH is required for follicular growth, estrogen production and oocyte maturation. Its function is mediated by its specific receptor (FSHR), and defective FSHR has been shown to affect folliculogenesis and ovarian function. In this study, we screened the entire coding region of FSHR gene for pathogenic mutations in women with premature ovarian failure (POF) (n = 16) and polycystic ovary syndrome (PCOS) (n = 124) and found no mutations in these patients. Two known polymorphisms, Thr307Ala and Ser680Asn showed similar distributions of the allelic variations and protein isoforms in PCOS and normal control subjects (n = 236). It appears from this study that mutations in the coding regions of FSHR gene are not a causative factor of the above clinical manifestations in Chinese Singapore women.     

Ovarian theca cell

Theca cells are the endocrine cells associated with ovarian follicles that play an essential role in fertility by producing the androgen substrate required for ovarian estrogen biosynthesis. Theca cells differentiate from the interfollicular stroma in response to proteins secreted from growing follicles. The most common endocrine cause of infertility is associated with excessive proliferation of theca cells and ovarian hyperandrogenism. Cell facts: -ovarian androgen-producing cells; -are associated only with developing follicles; -over-activity of theca cells causes infertility due to hyperandrogenism; -under-activity of theca cells causes infertility due to lack of estrogen. Theca cells: androgen-producing cells in the ovary.     

Effects of methoxychlor on IGF-I signaling pathway in rat ovary

Follicular development and other ovarian functions are regulated by growth factors that can be affected by exogenous agents. Methoxychlor (MXC) is an organochloride pesticide that causes female infertility. We investigated how MXC affects the distribution of developing ovarian follicles in adult rats after treatment between embryonic day (E) 18 and postnatal day (PND) 7. We also measured insulin-like growth factor-I (IGF-I) and its receptor, IGF-IR, expressions in ovarian follicles and investigated whether MXC changed the levels of IGF-I and IGF-IR in the ovary. Using immunohistochemical (IHC) staining, we detected IGF-I expression in oocytes and granulosa cells of the follicles, luteal cells, interstitial cells, theca externa and theca interna, and the smooth muscle of ovarian vessels. IGF-IR was co-localized with IGF-I in the ovary except for the theca externa. IGF-I expression was decreased in granulosa cells of preantral and antral follicles after treatment with MXC compared to granulosa cells of preantral and antral follicles of the control group. We also observed that oocytes of secondary follicles and granulosa cells of secondary and preantral follicles of the MXC treated groups showed increased IGF-IR expression compared to oocytes of secondary follicles and granulosa cells of secondary and preantral follicles of the control group. We also detected more secondary and preantral follicles, and fewer primordial and antral follicles after MXC administration compared to controls. Therefore, the IGF signaling pathway may participate in MXC induced ovary dysfunction and female infertility.     

Effects of a single administration of prostaglandin F2alpha, or a combination of prostaglandin F2alpha and prostaglandin E2, or placebo on fertility variables in dairy cows 3–5 weeks post partum, a randomized, double-blind clinical trial

In female mammals, including humans, deviations from normal androgenic or estrogenic exposure during fetal development are detrimental to subsequent adult ovarian function. Androgen deficiency, without accompanying estrogen deficit, has little apparent impact on ovarian development. Fetal estrogen deficiency, on the other hand, results in impaired oocyte and follicle development, immature and abnormal adult ovaries, and excessive ovarian stimulation from endogenous gonadotropins ultimately generating hemorrhagic follicles. Complete estrogen deficiency lasting into adulthood results in partial ovarian masculinization. Fetal androgen excess, on the other hand, mediated either by direct androgen action or following androgen aromatization to estrogen, reprograms ovarian development and reproductive neuroendocrinology to mimic that found in women with polycystic ovary syndrome: enlarged, polyfollicular, hyperandrogenic, anovulatory ovaries with accompanying LH hypersecretion. Oocyte developmental competence is also compromised. Insulin is implicated in the mechanism of both anovulation and deficient oocyte development. Fetal estrogen excess induces somewhat similar disruption of adult ovarian function to fetal androgen excess. Understanding the quality of the fetal female sex steroid hormone environment is thus becoming increasingly important in improving our knowledge of mechanisms underlying a variety of female reproductive pathologies.     

No effects of estrogen receptor overexpression on gonadal sex differentiation and reversal in medaka fish

In order to elucidate a possible role of estrogen receptor in the gonadal sex differentiation and the sex reversal with sex steroids, we examined for the formation of testis or ovary in transgenic medaka fish overexpressing the medaka estrogen receptor under the constitutive medaka beta-actin promoter. The transgenic fish underwent the genetically determined gonadal differentiation and showed the same sex-reversal rates as those of wild-type non-transgenic fish after treatments with estrogen and androgen. These results present invaluable data to reconsider the role of estrogen receptor in the gonadal sex determination.     

重复制动应激对雌性大鼠下丘脑-垂体-卵巢轴的影响

目的: 探索重复制动应激对雌性大鼠下丘脑-垂体-卵巢轴的影响。方法: 40只SD雌鼠随机分为两组(n=20),对照组和实验组,一组正常饲养,一组采取递增负荷束缚应激,每天置于束缚器内制动应激一次(从上午9:00开始),第1日制动2 h,以后采用递增负荷,每日增加0.5 h,持续两周,通过检测体重、脏器系数、动情周期、性激素、病理和相关基因的表达探索其对下丘脑-垂体-卵巢轴的危害。结果: 重复制动应激使雌性大鼠体重下降、动情周期延长,卵巢和子宫的脏器系数和形态发生改变,利用qPCR技术对其相关基因检测,发现下丘脑促性腺激素释放激素、垂体促性腺激素释放激素受体、促卵泡生成素和促黄体生成素mRNA的表达显著下降,卵巢促卵泡生成素和黄体生成素受体 mRNA的表达显著上升,卵巢和子宫雌激素受体mRNA的表达显著下降。结论: 重复制动应激可能通过干扰下丘脑-垂体-卵巢轴的内分泌调节作用,使动情周期紊乱,从而损伤雌性动物的性腺和生殖内分泌功能。     

二甲双胍对多囊卵巢不孕症患者卵巢功能、雌激素水平及胰岛素抵抗的影响

目的:探讨二甲双胍对多囊卵巢不孕患者卵巢功能、雌激素水平及胰岛素抵抗的影响。方法:选取我院收治的多囊卵巢不孕症患者72例,根据用药不同分为对照组与实验组,每组36例。对照组患者给予来曲唑促排卵治疗,实验组患者在对照组基础上予以二甲双胍治疗。观察并比较治疗前后两组患者的卵巢功能、雌激素水平、胰岛素抵抗及受孕情况。结果:与治疗前比较,两组患者治疗后卵巢功能、雌激素水平、胰岛素抵抗及受孕情况均得到改善,差异具有统计学意义(P0.05);与对照组比较,实验组患者治疗后卵巢功能显著改善,雌激素水平显著提高,胰岛素抵抗情况明显好转,受孕情况显著改善,差异均具有统计学意义(P0.05)。结论:二甲双胍能够调节多囊卵巢不孕症患者的卵巢功能,提高雌激素水平,改善胰岛素抵抗,提高受孕率,值得临床推广应用。     

Effect of p-chlorophenylalanine on the reproduction of the rat (author's transl)]

The effect of p-chlorophenylalanie (p-CPA) --300 mg/kg-- on reproduction has been studied in the female rat. Groups of animals were injected with a dose of 300 mg/kg of p-CPA 48 hours before proper copulation conditions at different moments along the ovarian cycle. Presence of spermatozoa in the vaginal frotis was negative in treated rats; in the control groups however, positivity was found in variable proportions according to the phase of the ovarian cycle: 30 ad 90% in diestrus and proestrus respectively. Treated animals showed continuous diestrus phases and diffuse luteinitation of the ovary. The results may indicate that a decrease of cerebral 5-HT, caused by p-CPA, lessens the reproductive behaviour of the female rat through mechanisms depending probably on the liberation of gonadrotrophins.     

The Role of Gonadal Steroid Receptor Activation in the Restoration of Sociosexual Behavior in Adult Male Rats

This work tested the hypothesis that gonadal steroid receptor activation was necessary for the restoration of several sociosexual behaviors (such as copulatory behavior, partner preference, 50-kHz vocalizations, and scent marking) in testosterone-treated gonadectomized male rats. Gonadal steroid receptors were blocked by systemic administration of the antiandrogen hydroxyflutamide, the antiestrogen RU 58668, or both antagonists simultaneously in a restoration paradigm. Inhibiting androgen receptors with hydroxyflutamide blocked the restoration of male copulatory behavior, partner preference (time spent with a sexually receptive female over a nonreceptive female), 50-kHz ultrasonic vocalizations, and scent marking. On the other hand, we did not find that blocking estrogen receptors with RU 58668 inhibited the restoration of copulatory behavior or partner preference in testosterone-treated gonadectomized male rats, even though the level of brain nuclear estrogen receptor occupation was significantly reduced to the level found in gonadectomized males. However, the restoration of scent marking and 50-kHz vocalizations were impaired by RU 58668. Blocking both nuclear androgen and estrogen receptors with the two antagonists simultaneously did not have a greater inhibitory effect than treatment with each antagonist alone. Therefore, the activation of nuclear estrogen receptors is necessary for the restoration of some, but not all, sociosexual behaviors, which are also androgen receptor-dependent. Besides nuclear estrogen receptors, there are additional, but unknown, targets of estradiol that play a role in mediating copulatory behavior in adult male rats. Moreover, the signals from multiple gonadal steroid signaling pathways converge in the regulation of some sociosexual behaviors in adult male rats.     

雄激素受体缺乏抑制肝脏Vtg合成导致卵巢发育障碍

为研究雄激素受体(Androgen receptor, Ar)在卵巢发育中的作用机制, 文章以雄性激素受体敲除的雌性斑马鱼(Danio rerio; ar–/–)为研究对象, 利用ELISA和蛋白质免疫印迹等方法分析Ar对斑马鱼肝脏中卵黄蛋白原(Vitellogenin, Vtg)产生、母体营养通过Vtg运输、卵黄形成和卵巢成熟的影响。研究发现在ar–/–雌性斑马鱼的肝脏中, Vtg的产生和雌激素受体的表达水平显著降低。在ar–/–雌性斑马鱼中的卵巢质量、脂质含量和类胡萝卜素含量均显著下降, 表明Ar的缺失可导致雌性斑马鱼通过Vtg转运到卵巢的脂质和类胡萝卜素等营养物质供应减少。此外, ar–/–雌性斑马鱼中与卵巢发育相关的几个基因转录表达水平显著下调, 这与卵巢发育异常存在相关性。研究结果表明, Ar的缺乏可通过对肝脏中的雌激素受体表达的影响, 降低肝脏Vtg蛋白的合成, 从而损害由Vtg向卵巢的营养物质运输和卵黄的正常形成, 影响卵巢发育。研究阐明了雄激素信号通路与体内肝脏Vtg产生之间的联系。     

Neonatal exposure to estradiol valerate programs ovarian sympathetic innervation and follicular development in the adult rat

A single injection of estradiol valerate (EV) to 14-day-old rats (when the ovarian follicle population has been already established) disrupts cyclicity, increases the activity of key enzymes of androgen biosynthesis, and develops polycystic ovary by a causally related increase in ovarian noradrenaline (NA). The current study examined an early window of ovarian development to look for a specific stage of development at which estradiol can induce such changes in sympathetic activity and follicular development. A single dose of EV applied to rats before the first 12 h of life rapidly increases (after 24 h) the ovarian expression of nerve growth factor (Ngfb) and p75 low-affinity neurotrophic receptor (Ngfr) mRNAs. When adults, rats presented early vaginal opening, disrupted cyclicity, appearance of follicular cyst, absence of corpus luteum, and infertility. Total follicles decreased, mainly due to a reduced number of primordial follicles, suggesting that estradiol acts in the first stages of folliculogenesis, when primordial follicles are organizing. These changes paralleled a 6-fold increase in NA concentration. No changes in NA content were found in the celiac ganglia, suggesting a local, non-centrally mediated effect of estradiol. Surgical section of the superior ovarian nerve (the main source of sympathetic nerves to the ovary) to rats neonatally treated with EV decreased intraovarian NA, delayed vaginal opening, and blocked the development of follicular cyst and that of preovulatory follicles. Therefore, we can conclude that early exposure to estradiol permanently modifies ovarian sympathetic activity and causes profound changes in follicular development, leading to the polycystic ovary condition.     

Steroid hormone receptors in the uterus and ovary of immature rats treated with gonadotropins

We administered either saline (group A) or 10 IU of pregnant mare serum gonadotropin (PMS; groups B and C) to female immature rats. Fifty-three hours later, the rats were injected with saline (groups A and B) or 30 IU of human chorionic gonadotropin (hCG; group C). The rats were decapitated 17 h after the last treatment, and the serum levels of progesterone (P4) and estradiol (E2) were measured by specific radioimmunoassays (RIA). The receptor levels of progesterone (PR) and estrogen (ER) in the uterus and ovaries were measured and the dissociation constant (Kd) of PR was obtained. The highest serum level of P4 was found in group C and that of E2 in group B. Cytosol levels of PR and ER in the uterus and ovary of the group B were the highest. It was indicated that the PMS treated-group (B), which had developing follicles in the ovary and the high serum level of E2, showed the highest concentration of ER and PR in both the ovary and the uterus. In the PMS and hCG-treated group (C), the uterine and ovarian steroid receptors decreased probably because of the luteinization and the high serum level of P4. The Kd uterine PR value was less than that of ovarian PR.     

Feminine sexual behavior in rats enhanced by prenatal inhibition of androgen aromatization

Male and female rats were exposed to the aromatization inhibitor 1,4,6-androstatriene-3, 17-dione (ATD) in utero via prenatal injections to the pregnant mother. In adulthood, lordosis behavior was measured in response to ovarian hormones. Males and females exposed prenatally to ATD showed enhanced lordosis behavior in response to estrogen alone and in response to estrogen plus progesterone when compared to controls. These data lend further support to the idea of a prenatal, androgen-sensitive phase of sexual differentiation in which defeminization normally occurs in both male and female rats. Further, these data support the concept that androgen aromatization is an important process in this defeminization.     

Aromatase inhibitors in ovarian stimulation

The selective estrogen receptor modulator, clomiphene citrate (CC), has been the principal drug used for induction of ovulation in women with polycystic ovarian syndrome (PCOS). CC is associated with adverse side effects and low pregnancy rates attributed to long-lasting estrogen receptor depletion. Anastrozole and letrozole are potent, non-steroidal, reversible aromatase inhibitors, developed for postmenopausal breast cancer therapy. We hypothesized that aromatase inhibitors could mimic the action of CC in reducing estrogen negative feedback on follicle stimulating hormone (FSH) secretion, without depleting estrogen receptors. In a series of preliminary studies, we reported the success of aromatase inhibition in inducing ovulation in anovulatory women with PCOS. Moreover, we showed that concomitant use of aromatase inhibitors resulted in a significant reduction of the FSH dose needed for controlled ovarian hyperstimulation. We suggest that aromatase inhibitors act through an increase in endogenous gonadotropin secretion as well as through increased intraovarian androgen levels that may increase ovarian FSH receptors. Recently, we demonstrated the safety of aromatase inhibitors in pregnancy outcome studies examining spontaneous pregnancy loss, multiple pregnancy rates and congenital anomalies compared to a control group of infertility patients treated with CC.     

The effect of destruction and stimulation of the lateral nucleus of the septum pellucidum on the structure and functions of the ovaries and the mechanism of such influences]

The experiments on the immature female rats were carried out to show the effects of damage or stimulation of the lateral septal nucleus (LSN) on the ovary function and its response to chorionic gonadotrophin. The damage of the LSN decreases the ovary weight, but enlarges mature ovarian follicles and increases the estrogen production. LSN lesions in hypophysectomized rats have no effect on the ovary and uterus weights as compared to hypophysectomized ones, but decrease the size of mature ovarian follicles against a background of hypophysectomy or LSN lesions. Damage of the LSN increases ovary and uterus sensitivity to the chorionic gonadotrophin injections. A conclusion is drawn on the existence of parahypophysial way to transfer the influence of LSN on the rat ovaries. The possible mechanisms of LSN influence on the reproduction system of female rats is under discussion.     

Reproductive activities of female albino rats treated with quassin, a bioactive triterpenoid from stem bark extract of Quassia amara

To evaluate the effect of quassin on female reproductive functions, 42 albino rats (35 females and 7 males) were used. The female albino rats were divided into seven groups of five rats each. Group I served as the control group and received distilled water while Groups II, III and IV rats were treatedorally with 0.1mg/kg, 1.0 mg/kg and 2.0 mg/kg body weight of quassin for 60 days respectively. Groups V, VI and VII rats were also treated orally with 0.1 mg/kg, 1.0mg/kg and 2.0 mg/kg body weight of quassin for 60 days but were left untreated for another 30 days, to serve as the recovery groups. At the end of each experimental period, blood samples were collected from each rat. Fertility study was done by cohabiting one untreated male with the five female rats in each group for 10 days. Quassin did not adversely affect the weight of the kidney, heart, liver and the body of the rats. However there was a significant decrease in the weight of the ovary and uterus in all the groups relative to the control. There was also a significant decrease in serum estrogen levels in quassin treated rats. The quassin treated rats had a significantly decreased mean litter number and weight. Histological studies show a disorganization and degeneration in the ovary while the uterus showed signs of vacuolation and disorganization. However, these effects were ameliorated after quassin was withdrawn from the rats. The results suggest that quassin has female anti-fertility properties, possibly acting via inhibition of estrogen secretion. Keyword: Quassin, Female rat, Reproduction, Estrogen.     

Testosterone environment of splenocytes modifies the steroidogenesis of polycystic ovary in rats.

The functional relationship between the ovary and immune cells is well known. The modulation of ovarian steroidogenesis in adult rats with polycystic ovary (PCO) by secretions of cultured splenocytes treated with 10 (-6) M testosterone or 10 (-6) M testosterone plus 10 (-4) M flutamide, an androgen receptor antagonist, was investigated. Polycystic ovary was induced by estradiol valerate (2 mg/rat). Polycystic ovary splenocyte secretions decreased the release of androstenedione from PCO ovaries in contrast to the effect of non-PCO splenocyte secretions. This decrease was associated with a significant decrease in androgen receptor and IL-12 mRNA expression in PCO splenocytes. When splenocytes were treated with testosterone, their conditioned media further decreased androstenedione release from the ovary and had a greater inhibitory effect on PCO ovary compared with non-PCO ovary. This effect was reversed by flutamide. Polycystic ovary splenocytes showed a decrease in IL-1 beta mRNA expression. Their secretions scarcely affected progesterone release from non-PCO ovaries but significantly stimulated progesterone release from PCO ovary by an androgen-independent mechanism. The differential steroidogenic ability of splenocyte secretions from PCO rats is associated with the IN VITRO testosterone environment. Polycystic ovary splenocytes might exert a protective action against PCO effects through their secretions by inducing a low androstenedione response from the ovary.