The development of ovarian tissue cryopreservation in Edinburgh: Translation from a rodent model through validation in a large mammal and then into clinical practice.

Ovarian tissue cryopreservation has developed rapidly from its origins in experiments in sheep in the 1990s, and is now becoming recognized as a standard, rather than experimental, procedure. This review summarizes the origins of the technique, and key milestones in its development since the first reported sheep pregnancy in 1994 to the first successful human pregnancy in 2004, and now to the first baby born following cryopreservation of prepubertal ovarian tissue. Many challenges remain to optimize this technique, to improve the survival of follicles within the reimplanted ovarian tissue, to improve its reproducibility and hence the success rate and the lifespan of the graft. The other key area remains the possibility of the grafted tissue containing malignant cells, most importantly in leukemia.

Can midlevel health-care providers administer early medical abortion as safely and effectively as doctors? A randomised controlled equivalence trial in Nepal.

BACKGROUND: Medical abortion is under-used in developing countries. We assessed whether early fi rst-trimester medical abortion provided by midlevel providers (government-trained, certified nurses and auxiliary nurse midwives) was as safe and effective as that provided by doctors in Nepal. METHODS: This multicentre randomised controlled equivalence trial was done in fi ve rural district hospitals in Nepal. Women were eligible for medical abortion if their pregnancy was of less than 9 weeks (63 days) and if they resided less than 90 min journey away from the study clinic. Women were ineligible if they had any contraindication to medical abortion. We used a computer-generated randomisation scheme stratified by study centre with a block size of six. Women were randomly assigned to a doctor or a midlevel provider for oral administration of 200 mg mifepristone followed by 800 µg misoprostol vaginally 2 days later, and followed up 10-4 days later. The primary endpoint was complete abortion without manual vacuum aspiration within 30 days of treatment. The study was not masked. Abortions were recorded as complete, incomplete, or failed (continuing pregnancy). Analyses for primary and secondary endpoints were by intention to treat, supplemented by per-protocol analysis of the primary endpoint. This trial is registered with ClinicalTrials.gov, NCT01186302. FINDINGS: Of 1295 women screened, 535 were randomly assigned to a doctor and 542 to a midlevel provider. 514 and 518, respectively, were included in the analyses of the primary endpoint. Abortions were judged complete in 504 (97.3%) women assigned to midlevel providers and in 494 (96.1%) assigned to physicians. The risk difference for complete abortion was 1.24% (95% CI -0.53 to 3.02), which falls within the predefined equivalence range (-5% to 5%). Five cases (1%) were recorded as failed abortion in the doctor cohort and none in the midlevel provider cohort; the remaining cases were recorded as incomplete abortions. No serious complications were noted. INTERPRETATION: The provision of medical abortion up to 9 weeks' gestation by midlevel providers and doctors was similar in safety and effectiveness. Where permitted by law, appropriately trained midlevel health-care providers can provide safe, low-technology medical abortion services for women independently from doctors. FUNDING: UNDP/UNFPA/WHO/World Bank Special Programme of Research, Development and Research Training in Human Reproduction (HRP), Department of Reproductive Health and Research (RHR), World Health Organization.

Emergency contraception: how does it work?

Emergency (or post-coital) contraception is any substance or device that is used to prevent pregnancy after unprotected intercourse. Currently used hormonal methods of emergency contraception (high-dose combined oral contraceptive pill or levonorgestrel) prevent about 50-80% of pregnancies. Research has demonstrated that these methods inhibit the midcycle surge of LH from the pituitary and, if given at least 2 days before ovulation, ovulation is delayed or prevented. Ovulation still occurs if administration is delayed until ovulation is imminent. Biological data that suggest that the most likely mode of action is by preventing fertilization are supported by the clinical observation that the greater the interval between coitus and administration the greater the chance of pregnancy. There are no data supporting the view that levonorgestrel can impair the development of the embryo or prevent implantation. In contrast, other very effective methods of emergency contraception, such as mifepristone and intrauterine devices, can also inhibit implantation.

Molecular profiling of the human testis reveals stringent pathway-specific regulation of RNA expression following gonadotropin suppression and progestogen treatment.

Gonadotropin withdrawal induces changes in gene expression in all 3 major cell types of the testis. Knowledge of the genes affected, in both the presence and absence of additional progestogen, will give insight into the regulation of human testicular function and aid development of novel contraceptive methods. We have undertaken a whole-genome analysis of RNA expression in testicular biopsies from normal men and after 4 weeks of gonadotropin suppression induced by gonadotropin-releasing hormone antagonist plus testosterone administration sufficient to cause marked suppression of spermatogenesis. Microarray analysis shows that interindividual variability is markedly low, and the response to treatment is focused on a small subset of genes particularly related to pathways in steroidogenesis and cholesterol biosynthesis or metabolism, the Leydig cell gene INSL3, and genes involved in early meiosis or Sertoli-germ cell junctions. These changes in expression were confirmed by quantitative reverse transcriptase polymerase chain reaction. No major changes in gene expression were identified in men additionally treated with a progestogen, although FLJ35767, an expressed sequence tag that is expressed in the germ cell compartment, did show a small but significant additional effect of progestogen. Overall, the results of this investigation disclose a remarkably stringent regulation of testicular gene expression, revealing the genes most sensitive to gonadotropin withdrawal, and might reflect the most labile pathways in the regulation of testicular function.

7alpha-methyl-19-nortestosterone (MENT) vs testosterone in combination with etonogestrel implants for spermatogenic suppression in healthy men.

Testosterone with a progestogen can suppress spermatogenesis for contraception. The synthetic androgen 7alpha-methyl-19-nortestosterone (MENT) may offer advantages because it is resistant to 5alpha-reduction and is therefore less active at the prostate. This study aimed to investigate MENT implants in combination with etonogestrel on spermatogenesis, gonadotropins, and androgen-dependent tissues in comparison with a testosterone/etonogestrel regimen. Healthy men (n = 29) were recruited and randomized to receive 2 etonogestrel implants with either 600-mg testosterone pellets repeated every 12 weeks or 2 MENT implants for up to 48 weeks. Testosterone concentrations in the testosterone group remained in the normal range. Subjects with 2 MENT implants showed peak MENT levels at 4 weeks with testosterone concentrations of 2 nmol/L. Sperm concentrations fell rapidly to less than 1 x 10(6)/mL at 12 weeks in 8 of 10 subjects in the MENT group and 13 of 16 subjects in the testosterone group with equally suppressed gonadotropins. Thereafter, suppression was not maintained in the MENT group, and 6 men noted loss of libido. Fourteen men completed 48 weeks of testosterone treatment, and all became azoospermic. Hemoglobin concentrations rose, and high density lipoprotein-cholesterol (HDL-C) fell in both groups. The MENT group showed a fall in prostate-specific antigen with no change in bone mass. MENT with a progestogen can achieve rapid suppression of spermatogenesis similar to testosterone, but this promising result was not sustained due to a decline in MENT release from the implants. This dose of testosterone, compared with previous studies using a lower dose with a higher dose of etonogestrel, had nonreproductive side effects without any increase in spermatogenic suppression. These data indicate the importance of the doses of progestogen and testosterone for optimum spermatogenic suppression while minimizing side effects.

Changes in vaginal morphology, steroid receptor and natural antimicrobial content following treatment with low-dose mifepristone.

BACKGROUND: We have previously shown that the antigestagen mifepristone is contraceptive when given in a daily dose of 5 mg, po. Epidemiological studies suggest that gestagen-only contraceptives may increase the risk of transmission of human immunodeficiency virus (HIV) due to effects on the vaginal defenses to infection. We investigate the effects of mifepristone on vaginal thickness, steroid receptor and natural antimicrobial content and pharmacokinetics of mifepristone. METHODS: In a pilot study, eight women were given mifepristone 5 mg/day for an average of 33 days. Ovarian function was assessed by measurement of estradiol and progesterone in blood and their metabolites in urine and by serial ultrasound of their ovaries. Vaginal biopsies were collected before (late proliferative) and after taking mifepristone. RESULTS: All subjects showed a similar pattern of descending serum concentrations of mifepristone. The elimination phase half-life was 18+/-5.1 h (mean+/-SD). Mean Cmax measured at 1 h was 641.7 nmol/L (range, 502-740 nmol/L). All eight women reported amenorrhea for the duration of treatment and seven of eight women showed biochemical and ultrasound evidence of anovulation. There was no significant change in vaginal thickness following treatment [342+/-40 microm pretreatment, 303+/-69 microm posttreatment (mean+/-SEM); p>.05]. Estrogen (ERalpha, ERbeta) and androgen receptor were expressed in both vaginal epithelium and subepithelial stroma, whereas progesterone receptor was expressed predominantly in the subepithelial stroma. There was no change in receptor content and distribution following mifepristone treatment. Natural antimicrobial mRNA [secretory leukocyte protease inhibitor, human beta defensins mRNA (HBD1, HBD2, HBD3, HBD5), granulysin and elafin] was extracted from the vaginal tissues, and the content was unaffected by mifepristone treatment. CONCLUSION: The absence of changes in vaginal thickness, steroid receptor and natural antimicrobial content and its distribution in this preliminary study suggests that in contrast to other estrogen-free contraceptives, mifepristone is unlikely to be associated with the increased risk of transmission of HIV and other sexually transmitted infections.

A diurnal variation in testicular hormone production is maintained following gonadotrophin suppression in normal men.

OBJECTIVE: A diurnal variation in serum testosterone in adult men is well recognized, but whether this occurs during exogenous testosterone administration and the degree to which it is endogenous to the testis is unclear. DESIGN: A clinical research centre investigation of testicular function in normal men. PATIENTS: Twenty normal men were recruited, 10 of whom were investigated during administration of testosterone with etonogestrel to suppress gonadotrophin secretion. MEASUREMENTS: Hourly blood samples were taken over 24 h for measurement of testosterone, inhibin B, LH, FSH and cortisol. Urinary excretion of testosterone and the testicular steroid epitestosterone was also measured. RESULTS: In the controls, a diurnal variation in serum testosterone and LH but not FSH was detected. The treated group had similar testosterone concentrations but showed no diurnal variation. Periodicity was also detected in inhibin B concentrations in 5 of the controls and in 9 of the treated group, who also showed synchrony not seen in the controls. Both groups showed diurnal variation in cortisol. Urinary testosterone excretion did not show a diurnal variation in either group, but this was apparent for epitestosterone with a morning peak in both groups despite the markedly lower excretion in the treated men. CONCLUSIONS: The diurnal variation of testosterone in normal men is due to a change in secretion rather than in clearance and is largely LH driven. An endogenous rhythm in both testicular steroidogenesis (epitestosterone) and Sertoli cell function (inhibin B) is also present.

Regulation of human endometrial function: mechanisms relevant to uterine bleeding.

This review focuses on the complex events that occur in the endometrium after progesterone is withdrawn (or blocked) and menstrual bleeding ensues. A detailed understanding of these local mechanisms will enhance our knowledge of disturbed endometrial/uterine function--including problems with excessively heavy menstrual bleeding, endometriosis and breakthrough bleeding with progestin only contraception. The development of novel strategies to manage these clinically significant problems depends on such new understanding as does the development of new contraceptives which avoid the endometrial side effect of breakthrough bleeding.

Toward developing a once-a-month pill: a double-blind, randomized, controlled trial of the effect of three single doses of mifepristone given at midcycle on the pattern of menstrual bleeding.

OBJECTIVE: To test the feasibility of timing the administration of mifepristone as a once-a-month contraceptive pill on the 12th day before the next menses, as calculated from the length of the previous menstrual cycles. DESIGN: Double-blind, randomized, controlled trial. SETTING: Five family planning centers across the world. PATIENT(S): Three hundred ninety-nine women attending family planning clinics. INTERVENTION(S): Randomized to receive 10, 25, or 200 mg of mifepristone or a placebo. MAIN OUTCOME MEASURE(S): Lengthening or shortening of the normal menstrual cycle length following administration of the drug by at least 5 days. RESULT(S): The menstrual period came within 5 days of the predicted date in 88% of women receiving the placebo, 84% of women receiving 10 mg, 72% of women receiving 25 mg of mifepristone, and only 48% of women treated with 200 mg of mifepristone. Increasing the dose of mifepristone was associated with an increased chance of having a delayed period (P<.001). Only 45% of women were in the peri-ovulatory phase of the cycle according to LH and P measurements on the day of drug administration. Women treated before ovulation were more likely to have delayed menses with all three doses of mifepristone. CONCLUSION(S): Because of the disruption in cycle length, it appears unlikely that mifepristone administered once a month, at a calendar-based time, would provide a reliable method of contraception.

Long-term suppression of reproductive function by a single dose of gonadotropin-releasing hormone antagonists in a sheep model.

OBJECTIVE: To determine the effect of single long-acting doses of GnRH antagonists on reproductive function in a sheep model. DESIGN: Observational, model study. SETTING: University-affiliated research unit. ANIMAL(S): Nine intact mature Merino sheep in experiment 1 and 12 mature Merino-crossed ewes with the ovary autotransplanted to the neck in experiment 2. INTERVENTION(S): Synchronization of estrous cycle either with intravaginal progestins or prostaglandin F2alpha analogues and treatment with a single dose of GnRH antagonist; evaluation of reproductive activity, plasma sampling, and ovarian ultrasonography. MAIN OUTCOME MEASURE(S): Determination of estrus behavior; plasma concentrations of P, FSH, LH, and inhibin A; and number and size of ovarian follicles. RESULT(S): In both experiments, the concentrations of FSH and LH were suppressed when compared with those in control ewes. In experiment 1, the ovulatory cycles were suppressed for > or = 55 days in treated sheep. In experiment 2, there were no follicles sized > or = 5 mm in treated ewes for 50 days. CONCLUSION(S): The suppression of the development of large follicles for > or = 30 days after a single injection of a long-acting GnRH antagonist provides a novel convenient method of pretreatment before COS.

Mifepristone-induced amenorrhoea is associated with an increase in microvessel density and glucocorticoid receptor and a decrease in stromal vascular endothelial growth factor.

BACKGROUND: We have previously shown that the progesterone antagonist mifepristone is a contraceptive when given in a dose of 2 or 5 mg per day. The majority of women experience amenorrhoea rather than the irregular break through bleeding usually occurring with other estrogen-free contraceptive pills, such as progestogen-only pill (POP). We investigated the effects of low-dose mifepristone on endometrial parameters which may be associated with changes in endometrial function, such as microvasculature, vascular endothelial growth factor (VEGF) and glucocorticoid receptor (GR) content. METHODS AND RESULTS: Endometrial biopsies were collected from 16 women before (late proliferative phase) and 60 and 120 days after taking 2 or 5 mg mifepristone daily for 120 days. Seven of the eight women who received 2 mg mifepristone and all eight women who received 5 mg were amenorrhoeic during the study. Mean estradiol (E(2)) concentrations remained in the mid-proliferative range, and the majority (9/16) of women showed proliferative endometrial histology at 60 and 120 days following treatment. There was a significant increase in the density of the endometrial stroma (P < 0.05) and microvessels (P < 0.01) following 120 days of treatment. Immunocytochemistry showed that GR, hitherto localized specifically in endometrial stroma, was up-regulated in the nuclei of glands (P < 0.05) and surface (luminal) epithelium (P < 0.01) by 60 days and maintained at 120 days. There was a significant reduction in stromal VEGF protein expression by day 120 of treatment (P < or = 0.01). CONCLUSION: The high incidence of amenorrhoea in women taking mifepristone may be related to changes in the regulation of vascular function.

Direct effect of progestogen on gene expression in the testis during gonadotropin withdrawal and early suppression of spermatogenesis.

CONTEXT: Testicular production of steroids and gametes is under gonadotropin support, but there is little information as to the molecular mechanisms by which these are regulated in the human. The testicular response to gonadotropin withdrawal is important for the development of effective contraceptive methods. OBJECTIVE: Our objective was investigation of expression of genes in the normal human testis reflecting steroidogenesis, Sertoli cell function, and spermatogenesis after short-term gonadotropin withdrawal and the effects of activating testicular progesterone receptors. DESIGN AND SETTING: We conducted a randomized controlled trial at a research institute. PATIENTS: Thirty healthy men participated. INTERVENTIONS: Subjects were randomized to no treatment or gonadotropin suppression by GnRH antagonist (cetrorelix) with testosterone (CT group) or with additional administration of the gestogen desogestrel (CTD group) for 4 wk before testicular biopsy. Gene expression was quantified by RT-PCR. RESULTS: Both treatment groups showed similar suppression of gonadotropins and sperm production and markedly reduced expression of steroidogenic enzymes. Addition of progestogen in the CTD group resulted in reduced expression of 5alpha-reductase type 1 compared with both controls and the CT group. Inhibin-alpha and the spermatocyte marker acrosin-binding protein were significantly lower in the CTD but not CT groups, compared with controls, but did not differ between treated groups. Men who showed greater falls in sperm production also showed reduced expression of these three genes but not of the spermatid marker protamine 1. CONCLUSIONS: These data provide evidence for direct progestogenic effects on the testis and highlight steroid 5alpha-reduction and disruption of spermiation as important components of the testicular response to gonadotropin withdrawal.

Comparison of a transdermal continuous combined and an interrupted progestogen HRT.

OBJECTIVES: Pilot study to compare the effects of a continuous combined hormone replacement therapy (HRT) regimen with an interrupted progestogen regimen administered transdermally, upon the endometrium of postmenopausal women, the incidence of amenorrhoea and relief of menopausal symptoms. METHODS: Fifty-nine postmenopausal women aged 50-63 years were randomised to either (i) continuous combined regimen: combined oestrogen/progestogen skin patches (releasing continuous 50 microg estradiol and 20 microg levonorgestrel/day) or (ii) interrupted regimen: oestrogen-only patches (releasing 80 microg estradiol/day) for 4 days followed by combined oestrogen/progestogen patches (releasing continuous 50 microg estradiol and 20 microg levonorgestrel/day) for 3 days, for 6 months. An endometrial biopsy was performed at end of treatment for histological analysis. RESULTS: Thirty-three women (56%) completed the study. Significantly higher rates of amenorrhoea were observed with the interrupted than continuous combined regimen (P<0.0001; 25% versus 7% at 6 months). The interrupted regimen was also associated with fewer days of bleeding overall (total 20 versus 44 days during months 4-6; P=0.001). Both regimens improved vasomotor symptoms. No endometrial hyperplasia or atypical changes were observed in endometrial biopsies. CONCLUSIONS: Although significantly less bleeding was observed with the interrupted regimen, it did not have a sufficiently high incidence of amenorrhoea to render it clinically useful.

Leukocyte populations and steroid receptor expression in human first-trimester decidua; regulation by antiprogestin and prostaglandin E analog.

CONTEXT: Progesterone acting via its cognate receptor is critical to maintaining a viable endometrial environment for implantation and pregnancy. During medical termination of pregnancy, the biological effect of progesterone is pharmacologically withdrawn and prostaglandins administered exogenously. Leukocytes within the uterus are the effector cells of an inflammatory response and play important roles in both tissue breakdown and remodeling. OBJECTIVE: The aim of this study was to identify the separate and combined effects of the antiprogestin Mifepristone (single dose, 200 mg) and the prostaglandin E (PGE) analog (gemeprost) on leukocyte populations and steroid receptor expression in human first-trimester decidua. PATIENTS: Eighty women were recruited from the termination of pregnancy service with a gestational age of between 35 and 65 d at the time of surgical termination of pregnancy. MAIN OUTCOME MEASURES: Immunohistochemistry was used to measure macrophage (CD68 +ve), neutrophil (neutrophil elastase +ve), and uterine natural killer cell (CD56 +ve) populations and progesterone (PR(A) and PR(B)), estrogen (ERalpha and ERbeta), and androgen receptor (AR) expression. RESULTS: After administration of both antiprogestin and the PGE analog, macrophage and neutrophil numbers were significantly increased, whereas natural killer cell numbers were unchanged. Antiprogestin and PGE analog coadministration also significantly decreased PR and ERalpha immunoreactivity but had no effect on androgen receptor or ERbeta receptor expression. PGE analog alone was also capable of reducing PR expression. CONCLUSIONS: In this study, we demonstrate that the inflammatory response induced by antiprogestin in combination with PGE analog is accompanied by both increases in macrophages and neutrophils numbers and decreases in PR and ERalpha expression in human first-trimester decidua.

Multiple factors affecting the efficiency of multiple ovulation and embryo transfer in sheep and goats.

This review offers an overview of the basic characteristics of in vivo embryo technologies, their current status, the main findings and the advances gained in recent years, and the outstanding subjects for increasing their efficiency. The use of superovulation and embryo transfer procedures remains affected by a high variability in the ovulatory response to hormonal treatment and by a low and variable number of transferable embryos and offspring obtained. This variability has been classically identified with both extrinsic (source, purity of gonadotrophins and protocol of administration) and intrinsic factors (breed, age, nutrition and reproductive status), which are reviewed in this paper. However, emerging data indicate that the main causes of variability are related to endocrine and ovarian factors, and so the number of studies and procedures addressing a better understanding and control of these factors may be increased in the future. The accomplishment of this objective, the improvement of procedures for embryo conservation and for the selection and management of recipient females, will allow further development and application of this technology.

Low-dose mifepristone inhibits endometrial proliferation and up-regulates androgen receptor.

Mifepristone in daily low doses has contraceptive potential by inhibiting ovulation. Follicular development is maintained, and although the endometrium is exposed to unopposed estrogen, there are no signs of hyperplasia or atypia. The mechanism of this antiestrogenic action is unknown. We have investigated the effect of daily low-dose mifepristone on proliferation markers and steroid receptors in surface epithelium, glands, and stroma of the endometrium. Endometrial biopsies were collected from 16 women before (late proliferative) and 60 and 120 d after taking 2 or 5 mg mifepristone daily for 120 d. Endometrial proliferation (H3 mitosis marker) and steroid (estrogen, progesterone, and androgen) receptor content were studied using standard immunocyotchemistry techniques. There was a significant decrease in the expression of H3 mitosis marker (P

Long-term ovarian function in sheep after ovariectomy and autotransplantation of cryopreserved cortical strips.

Transplantation of cryopreserved ovarian tissue offers a means of preserving reproductive function in girls/women whose ovarian function is compromised by radio- and/or chemotherapy for cancer. Studies in small laboratory rodents have demonstrated that fertility can be restored by transplanting whole cryopreserved ovaries. In large animals, it is not possible to freeze or transplant ovaries without vascular anastomosis. However, primordial oocytes in cortical strips survive freezing, thawing and culture in vitro. We have used sheep as an experimental model for the human because the size of the ovary and length of folliculogenesis (4-6 months) are similar. Following orthotopic transplantation the grafts, become vascularised within a few days and ovulatory cycles become established by 4 months. Several lambs have been born in ewes after autotransplantation of the grafts which continue to function for up to 2 years. The levels of FSH remain consistently raised after transplantation due to a reduced secretion of inhibin A, reflecting the reduction in the size of the ovarian pool of follicles.

Effect of ageing on hormone secretion and follicular dynamics in sheep with and without the Booroola gene.

It has been suggested that ewes carrying the Booroola gene (Fec(B)) consistently ovulate more follicles because they recruit more primordial follicles and/or have a lower rate of atresia. If the former is correct, the pool of follicles would be depleted sooner in Fec(B) animals. We have studied follicular dynamics and endocrine function during follicular and early luteal phases of the estrous cycle of older ewes with or without the fecundity gene and compared this data with data obtained 6 yr previously in the same animals. Older sheep carrying the Booroola gene maintained a significantly higher ovulation rate than noncarrier ewes [4.2 +/- 0.8 vs. 2.2 +/- 0.6 corpora lutea (CL), respectively; P < 0.05], and in keeping with data from young animals, both ovulatory follicles and CL (4.7 +/- 0.3 vs. 6.9 +/- 0.7 mm and 12.8 +/- 0.5 vs. 16.7 +/- 0.8 mm, respectively) were smaller than those of noncarrier ewes (P < 0.05). The interval from luteolysis to the onset of the LH surge increased with age in all the animals (from 52.0 +/- 8.0 to 67.0 +/- 7.5 h in gene carrier sheep and from 56.0 +/- 2.0 to 79.5 +/- 9.6 h in noncarrier sheep, P < 0.05). The concentration of estradiol and inhibin A in the early luteal phase was lower in older noncarrier ewes (P = 0.08 and P < 0.05, respectively), and the level of inhibin A was inversely related to the level of FSH in aged sheep of both genotypes (P < 0.0001). In contrast, the number of developing follicles in older ewes of both genotypes was similar to the number found in younger ewes, suggesting that increased ovulation rate in sheep carrying the Fec(B) mutation is related to a reduced rate of atresia.

Evidence of a role for follicle-stimulating hormone in controlling the rate of preantral follicle development in sheep.

Autografting ovarian cortex results in the loss of growing follicles and elevated gonadotropins. This paradigm was employed to examine the effect of gonadotropins on preantral follicle development in sheep. Ovarian tissue was recovered at 1, 2, 3, and 4 months after grafting from ewes that were either hyper- (n = 12; untreated) or hypogonadotropic (n = 12; GnRH-agonist and estradiol implants). Compared with the Hypo group, Hyper ewes had higher (P < 0.001) gonadotropins, had greatly enlarged grafts, had reestablished a normal follicular hierarchy 2 months earlier (P < 0.05), had higher (P < 0.05) levels of proliferating cell nuclear antigen expression in tertiary, preantral, and antral follicles, and had higher (P < 0.01) concentrations of inhibin A and estradiol. Compared with time zero controls, increases in the number of primary follicles and the rate of proliferation in primary and secondary follicles in both groups of autografts (P < 0.05) were also observed. In conclusion, the results of this experiment provide the first evidence that gonadotropins can affect the rate of development of preantral follicles in vivo in a large monovulatory species. Furthermore data are presented to support the existence of a gonadotropin-independent intraovarian feedback loop regulating both the rate of primordial follicle initiation and primary and secondary follicle development.

A role for the androgen receptor in the endometrial antiproliferative effects of progesterone antagonists.

In women and nonhuman primates, treatment with progesterone antagonists suppresses estrogen-dependent mitotic activity in the endometrial glands. This antiproliferative effect is paradoxical, because progesterone antagonists do not bind to the estrogen receptor (ER). While this phenomenon has been termed a "functional noncompetitive antiestrogenic effect," it does not occur in all species or in all regions of the primate reproductive tract, so is best referred to as an "endometrial antiproliferative effect." Recent studies of ours in both women and macaques revealed that the endometrial androgen receptor (AR) was increased by progesterone antagonist treatment. Because androgens are known to suppress estrogen-dependent endometrial proliferation, we hypothesized that the AR was involved in the antiproliferative effects induced by progesterone antagonists. In a test of this hypothesis, we administered the antiandrogen, flutamide, along with progesterone antagonists to ovariectomized, estrogen-treated macaques. Flutamide counteracted the suppressive effects of the progesterone antagonists on endometrial wet weight, thickness, stromal compaction, and mitotic index. Hyaline degeneration of the spiral arteries was also blocked by flutamide. These data implicate the AR as a functional component of the mechanism through which progesterone antagonists induce endometrial antiproliferative effects in the presence of estrogens.