A critical analysis of the accuracy, reproducibility, and clinical utility of histologic endometrial dating in fertile women.

OBJECTIVE: To refine or redefine the traditional histologic criteria used to date the secretory phase endometrium. DESIGN: Randomized, observational study. SETTING: Academic clinical research center. PATIENT(S): One hundred and thirty healthy, regularly cycling, fertile volunteers, aged 18 to 35 years. INTERVENTION(S): Patients were randomized to undergo endometrial sampling and measurement of serum estradiol and progesterone 1 to 14 days after the midcycle urinary luteinizing hormone surge. Three gynecologic histopathologists objectively scored each tissue specimen for 32 distinct histologic features and dated the endometrium using traditional histologic criteria. MAIN OUTCOME MEASURE(S): The 32 features were evaluated for [1] temporally dependent variation, [2] the amplitude of variations in score observed across the secretory phase, and [3] interobserver variability. Additionally, traditional dating criteria were analyzed. RESULT(S): The traditional endometrial histologic dating criteria are much less temporally distinct and discriminating than originally described, due to considerable intersubject, intrasubject, and interobserver variability. Neither traditional dating criteria nor any combination of the best performing histologic features identified by our objective and systematic analyses could reliably distinguish any specific cycle day or narrow interval of days. CONCLUSION(S): Histologic endometrial dating does not have the accuracy or the precision necessary to provide a valid method for the diagnosis of luteal phase deficiency or to otherwise guide the clinical management of women with reproductive failure.

Soy protein isolate with isoflavones does not prevent estradiol-induced endometrial hyperplasia in postmenopausal women: a pilot trial.

OBJECTIVE: To test the hypothesis that soy protein isolate (SPI) with isoflavones opposes the proliferative effects of exogenous estradiol (E2) on the endometrium after menopause. DESIGN: Thirty-nine postmenopausal women were randomized to receive daily for 6 months either 0.5 mg E2 + placebo, 1.0 mg E2 + placebo, 0.5 mg E2 + 25 g SPI with 120 mg isoflavones, or 1.0 mg E2 + 25 g SPI with 120 mg isoflavones. Primary outcome measures were endometrial histology, ultrasound endometrial thickness, and Ki67 staining quantification, a marker of cellular proliferation. Secondary outcome measures were serum lipids and markers of bone resorption. RESULTS: Endometrial hyperplasia, endometrial stromal and epithelial cellular proliferation, and sonographically measured endometrial thickness were similarly affected in all groups. SPI did not lessen the beneficial effects of E2 on lipids and markers of bone resorption. CONCLUSION: In this pilot study, SPI with isoflavones did not protect the endometrium from E2-induced hyperplasia in postmenopausal women. If higher, long-term doses of isoflavone supplementation are found to be safe for postmenopausal women, then future studies combining E2 with isoflavones may be feasible as an alternative to traditional hormone replacement therapy.

Effect of stress on pregnancy outcome among women undergoing assisted reproduction procedures.

BACKGROUND: This study was performed to examine the effect of stress on pregnancy outcome in women who underwent assisted reproductive technology (ART) procedures. METHODS: In a controlled clinical study of healthy volunteers in an academic research environment, stress was measured subjectively by administering patient questionnaires and biochemically by examining urinary excretion of cortisol and 6-sulfatoxy-melatonin (6-SM), the primary metabolite of melatonin and a marker of peripheral stress response. A total of 42 women who underwent ART procedures during an 18-month period agreed to participate in the study and were enrolled consecutively. The women collected 24-hour urine specimens on the day after human chorionic gonadotropin administration and concurrently completed the State-Trait Anxiety Inventory questionnaire. We measured the cortisol and 6-SM levels in the urine collection for each of the 42 women and for 10 oocyte donors who served as controls. Pregnancy tests were performed 14 days after embryo transfer. Analysis of covariance was used to compare the relationship of stress ratings to urinary cortisol and 6-SM levels among the women who became pregnant, the women who did not, and the women who served as controls. Other variables were explored by performing chi2 analysis and Fisher's exact test. RESULTS: Neither self-ratings of acute anxiety, nor total daily 6-SM value, nor cortisol levels were associated with pregnancy outcome in the ART procedures. CONCLUSION: Biochemical markers of stress failed to support a deleterious effect of stress on pregnancy outcome in women who underwent ART procedures. Subjective measurement of stress levels did not differ between women who became pregnant and those who did not.

Temporal and morphologic characteristics of pinopod expression across the secretory phase of the endometrial cycle in normally cycling women with proven fertility.

OBJECTIVE: To assess the temporal and morphologic characteristics of pinopod expression on the surface of endometrium across the secretory phase, in LH-timed endometrial samples in normal, healthy women. DESIGN: Prospective, randomized study. SETTING: Academic teaching hospital. PATIENT(S): Sixty-eight healthy volunteers with proven fertility. INTERVENTION(S): Urinary LH-timed endometrial and blood sampling was performed on each subject on a randomly selected day of the secretory phase. MAIN OUTCOME MEASURE(S): Histologic dating, assessment of pinopods using scanning electron microscopy, and comparison with serum P levels. RESULT(S): Eighty-six endometrial tissue samples obtained from 68 subjects were evaluated under scanning electron microscopy. Pinopods were first observed on luteal day 5, corresponding with the onset of the midluteal phase increase in serum P levels. Pinopods persisted for the entire duration of the secretory phase, but their morphology changed as the cycle advanced. CONCLUSION(S): The present findings demonstrate that pinopods are a characteristic feature of the mid to late secretory phase endometrial epithelium, exhibit cycle-dependent changes in morphology, and are most prominent during the putative implantation interval.

Oil-soluble contrast during hysterosalpingography in women with proven tubal patency.

To determine if there are therapeutic advantages to oil-soluble contrast medium compared with water-soluble medium during hysterosalpingography.A randomized, controlled trial including 56 infertile patients undergoing hysterosalpingography was performed. After a hysterosalpingogram with water-soluble contrast demonstrated tubal patency, 30 patients were randomized to receive oil-soluble contrast medium (oil group) and 26 patients received no additional contrast medium (control group). The outcome was pregnancy and timing of pregnancy in relation to hysterosalpingography. There were 18 (64%) pregnancies in the oil group and 14 (56%) pregnancies in the control group. Mean time to achieve pregnancy was shorter in the oil group: 3.8 months in the oil group compared with 6.1 months in the control group (P =.06) There was a clinically meaningful improvement in pregnancy rates between the oil group and the control group at 1 month postprocedure (relative risk [RR] 2.1, 95% confidence interval [CI] 0.6, 7.2). However, at 12 months postprocedure, the advantage was diminished. (RR 1.3, CI 0.8, 2.1)Eighteen months after hysterosalpingography, contrast does not appear to influence cumulative pregnancy rates; however, the addition of oil-soluble contrast medium to water-soluble contrast medium may have the potential to reduce the time to conception.

Steroid receptor coactivator expression throughout the menstrual cycle in normal and abnormal endometrium.

The endometrium of reproductive aged women undergoes cyclic developmental changes in preparation for implantation in response to estrogen and progesterone. These steroids and their receptors are tightly regulated throughout the menstrual cycle, and their actions are facilitated by the presence of steroid receptor coactivators of the p160 family. In this study using immunohistochemistry and Western blot analysis, we characterize the expression patterns of three coactivators, steroid receptor coactivator-1, amplified in breast cancer-1 (AIB1), and transcriptional intermediary factor-2 in human endometrium obtained prospectively from normal fertile women throughout the menstrual cycle. With the exception of glandular AIB1, which increased in the late secretory phase, none of the coactivators changed significantly during the menstrual cycle. We compared coactivator expression patterns in fertile endometrium to the endometrium of anovulatory (proliferative; n = 3) and clomiphene-induced ovulatory (secretory; n = 13) women with polycystic ovarian syndrome (PCOS), a group that have a higher likelihood of developing estrogen-induced endometrial hyperplasia and cancer. To control for the effect of clomiphene citrate, an additional group was included consisting of ovulatory women treated with clomiphene citrate for "male factor" infertility. Compared with both fertile and infertile controls, PCOS women exhibited elevated levels of AIB1 and transcriptional intermediary factor-2 expression in both epithelial and stromal cells. We postulate that increased coactivator expression may render the endometrium more sensitive to estrogen. In support of this, we describe an increased expression of ERalpha (an estrogen-induced gene product) during the menstrual cycle in PCOS endometrium compared with fertile controls. In summary, we demonstrate that the expression of p160 coactivators are regulated in endometrium during the menstrual cycle in normal fertile women but are overexpressed in the endometrium of women with PCOS. Based on these findings, we suggest a possible mechanism to explain the poor reproductive performance observed in PCOS and the increased incidence of endometrial hyperplasia and cancer noted in this group of women.

Effects of the route of estrogen administration and exercise on hormonal levels in postmenopausal women.

OBJECTIVE: To examine the effects of exercise on serum estrogens, growth hormone, insulin, cortisol, lactate, and glucose levels in postmenopausal women receiving two routes of administration of estrogen replacement therapy (ERT). DESIGN: Prospective, randomized, crossover study. SETTING: The general clinical research center of an academic medical center. PATIENT(S): Eleven active, postmenopausal women. INTERVENTION(S): The patients were screened with exercise stress testing, then oral micronized estradiol or transdermal estradiol was administered, followed by two 45-minute submaximal exercise tests. Dietary intake before the tests was standardized. MAIN OUTCOME MEASURE(S): The study measured maximal heart rate and aerobic power (VO2max), and serum levels of estradiol (E2), estrone (E1), cortisol, growth hormone (GH), insulin, glucose, and lactate. RESULT(S): Growth hormone, cortisol, and insulin all changed significantly in response to the 45-minute exercise bouts, but no differences were observed between the oral micronized estradiol and transdermal estradiol responses. E2 levels increased significantly during the transdermal estradiol 45-minute exercise bout; this change did not occur during the oral estradiol exercise bout. In the transdermal estradiol treatment group, the E2 levels at +30 and +45 minutes of exercise were elevated compared to the post-exercise levels at -15, 0, and 30 minutes. E1 was not significantly changed during the 45-minute exercise bouts in either group. CONCLUSION(S): During exercise, serum E2 levels rise significantly higher with transdermal but not oral routes of E2 administration. However, the elevated levels are not prolonged and normalize by 30 minutes after exercise.

In vivo and in vitro evidence suggest that HB-EGF regulates endometrial expression of human decay-accelerating factor.

Human endometrium expresses the critical complement component C3 in a cyclic fashion, with the highest expression in the secretory phase. As activated complement can kill cells, self or foreign, the secretory endometrial epithelium protects itself by concomitant expression of complement-protective proteins. The objectives of our present study were to describe the spatial and temporal regulation of the complement-protective protein decay-accelerating factor (DAF) in human endometrium and to identify local regulators of its expression. To describe the cyclic regulation of DAF, immunohistochemistry was performed using the IH4 monoclonal antibody on secretory phase endometrial biopsies taken from normal fertile volunteers in LH-timed cycles (n = 114). DAF expression in human endometrium was predominantly localized to the apical membrane of glandular and luminal epithelium. DAF expression, as assessed by histological scoring analysis, was minimal in the proliferative and early secretory phases and increased markedly on approximately day LH +7 (lumen) and LH +8 (glands). Maximal expression was seen in both glands and lumen by LH +8, and this persisted into menses. Using the RL95-2 endometrial epithelial cancer cell line as a model system, we next examined the cellular regulation of DAF. Treatment with E2 and progesterone, alone or in combination, had little effect on DAF expression. Heparin-binding epidermal growth factor-like growth factor (HB-EGF) treatment increased cell surface and total DAF protein, increasing the signal by 260% on flow cytometry and by 200% on Western blot analysis. Stimulation of DAF protein expression was dose dependent, with maximal expression seen at 1 ng/ml. The stimulatory effects of HB-EGF were also observed at the mRNA level. EGF had effects similar to those of HB-EGF on DAF mRNA and protein expression, suggesting that the HB-EGF effect was mediated at least in part by the Her1 EGF receptor subunit. These studies suggest that DAF expression in the midsecretory phase is stimulated by HB-EGF or other members of the EGF family and may function to protect the epithelial integrity of human endometrium in the face of increased complement expression.

Seasonal shifts in the provisioning behavior of chinstrap penguins, Pygoscelis antarctica.

Whether parents are able to adapt food gathering to rising offspring demands, or if they are controlled largely by extrinsic factors, is important for understanding key limits on fitness. Over seven breeding seasons, we studied the provisioning behavior of chinstrap penguins, Pygoscelis antarctica, at Seal Island, Antarctica, during parents' transition to leave broods of one or two chicks unguarded. By measuring the frequency, duration, and diel timing of foraging trips and the quantity of prey brought to chicks, we examined the extent to which variation in parents' feeding behavior could be attributed to provisioning costs which increase with chick growth. Estimates of the energy content of food loads were combined with foraging patterns and brood requirements to model parents' seasonal provisioning budget. The frequency of foraging trips increased from the guard to post-guard phase and was higher, and the seasonal effect larger, in parents with two chicks. The duration of overnight trips (~16 h) increased with seasonally increasing night length; diurnal trip duration (~8 h) showed no seasonal pattern. Birds exhibited a seasonal shift to diurnal foraging, a trend that was generally weaker in parents of smaller broods. Food loads increased with chick mass only in parents of one chick; parents of two chicks had larger but more constant food loads. Based on per trip calculations, parents foraging overnight could not have delivered to two-chick broods enough food to meet their demands unless chicks were small. Diurnal foragers (regardless of brood size) and overnight foragers with one chick could meet brood demands at chicks' peak mass. The combined daily effort of parents indicated that mated pairs on average had ample resources to meet chick demands through most of rearing. A brief period when demands could not be met was predicted in two-chick broods just before chicks were left unguarded and again as they neared fledging. Our findings suggest that penguins both increased provisioning frequency and favored foraging under higher light intensity in conjunction with increasing chick demands, tactics which required parents to leave chicks unattended. The ability to maintain intrinsic control over provisioning has bearing on how penguins may be limited by extrinsic constraints. Prey surveys conducted annually near colonies show abundant resources 10-20 km offshore with no consistent seasonal shifts in abundance. These findings support a prominent role for intrinsic factors in the foraging decisions of chinstrap penguins.

Effect of sex steroids on beta-endorphin levels at rest and during submaximal treadmill exercise in anovulatory and ovulatory runners.

OBJECTIVE: To examine the interaction between circulating beta-endorphin levels and sex steroids during sustained submaximal exercise in runners who are either anovulatory and oligomenorrheic (AO) or ovulatory and eumenorrheic (EO). DESIGN: Controlled clinical study. SETTING: General clinical research center at an academic medical center. PATIENT(S): Three AO and four EO runners. INTERVENTION(S): The athletes underwent 60 minutes of submaximal treadmill exercise on three separate occasions. Anovulatory and oligomenorrheic runners underwent exercise at baseline and after physiologic estrogen and combined estrogen and progesterone replacement. Ovulatory and eumenorrheic runners underwent exercise in the follicular and luteal phases and after GnRH agonist desensitization. MAIN OUTCOME MEASURE(S): Serum cortisol, beta-endorphin, progesterone, estrogen, and gonadotropin levels at rest and during exercise. RESULT(S): Serum levels of E2 increased in response to exercise in both EO and AO runners during sex steroid replacement. Baseline peripheral beta-endorphin and cortisol levels were not different between the EO and AO groups. A significant increase in beta-endorphin levels in response to exercise occurred only in the EO group after GnRH agonist desensitization. CONCLUSION(S): Alterations in menstrual cyclicity and ovulation in conditioned runners probably are not due to an increase in opioid tone. The hypothalamic-gonadotropic axis appears to be intact in AO runners, as measured by the gonadotropic response to exogenous exposure to estrogen and progesterone. Sex steroid administration had no effect on basal beta-endorphin levels, but this probably was not due to preexisting increased opioid tone.