Experience improves performance of hysterosalpingo-contrast sonography (HyCoSy): a comprehensive and well-tolerated screening modality for the subfertile patient.

PURPOSE: To investigate the clinical observations, provider experience, safety, and tolerance of the hysterosalpingo-contrast sonography (HyCoSy) procedure. MATERIALS AND METHODS: A retrospective study design in which data was collected from ninety-six subfertile women who underwent the HyCoSy procedure at the University of Louisville over a 16-month interval. RESULTS: Ninety-six HyCoSy procedures were performed by a single investigator and contained complete records for review. The authors observed significant decreases in the quantities of saline and air utilized per procedure over time (p < 0.0001 and p = 0.0001). Results from the HyCoSy studies were more often non-diagnostic or non-patent in women with a body mass index (BMI) > 30. Reported pain scores did not statistically differ over the course of the study interval. There were no procedure-related complications noted. CONCLUSION: The HyCoSy procedure is a timely and minimally invasive study that can be implemented in an office setting with minimal prior operator experience that improves over time.

Dysregulation of leptin and testosterone production and their receptor expression in the human placenta with gestational diabetes mellitus.

Whether the placenta contributes to some of the abnormal hormonal profiles in gestational diabetes mellitus (GDM) pregnancies and whether GDM affects placental endocrine signaling pathways are yet to be established. The objective of this study was to investigate differences in the expression of the placental steroid and peptide hormone synthesis-related factors, enzymes and their receptors between normal and GDM pregnancies. Nine term placentae from GDM pregnancies and twelve from healthy pregnancies were collected. The results of immunohistochemistry, Western blotting and semiquantitative RT-PCR indicated that mRNA and protein levels of leptin, leptin receptors, androgen receptor and FGF2 were significantly higher in the GDM placentae than non-GDM placentae; while NRIH3, NRIH2, StARD3, CYP11A1, HSD3B, HSD11B, HSD17B, ERalpha, ERbeta, progesterone receptor, FGF receptor-2, insulin receptor-alpha and -beta showed no differences. Interestingly, Western blotting and immunohistochemistry revealed that aromatase protein concentrations in the GDM placentae were significantly reduced without a change in mRNA levels. Moreover, androgen upregulated FGF2 expression in the placental villous explants. These findings suggest that the placentae of GDM pregnancies contribute to elevated testosterone and leptin levels due to a decrease in the conversion of testosterone to estrogens and to an increase in leptin production. The androgen and leptin signaling pathways may be over-activated by the presence of excessive ligands and overexpressed receptors in GDM placentae. Dysregulation of these two endocrine networks may contribute to placental abnormalities eventually increasing the frequency of maternal and fetal complications associated with GDM.

Human fetal nongonadal tissues contain human chorionic gonadotropin/luteinizing hormone receptors.

Human chorionic gonadotropin (hCG), a heterodimeric glycoprotein hormone produced in abundance by placental syncytiotrophoblasts, is preferentially secreted into maternal circulation. Fetal circulation also contains low levels of hCG that are probably derived from fetal kidney, liver, anterior pituitary gland, etc. In addition, the fetus has access to hCG present in exocoelomic and amniotic fluids. hCG has been found in a number of fetal tissues known to stimulate fetal adrenal and testicular steroidogenesis and is also thought to play a role in growth and differentiation of fetal tissues. This led us to test the hypothesis that fetal nongonadal tissues, as in the adult, may also contain hCG/LH receptors. This hypothesis was tested by immunocytochemistry, Western blotting, in situ hybridization, and RT-PCR. The results demonstrate that kidney, liver, pancreas, lung, small and large intestines, and adrenals contained hCG/LH receptors. Although the role of fetal nongonadal hCG/LH receptors is not known, they may mediate the pleiotropic actions of hCG in the growing human fetus.

The presence of functional luteinizing hormone/chorionic gonadotropin receptors in human sperm.

The functional receptors that bind human CG (hCG) and LH have recently been identified in a number of nongonadal human tissues. The current experiments tested the hypothesis that human ejaculated sperm may also contain them. The data revealed that they, indeed, do as determined by the presence of receptor messenger RNA and receptor protein that can bind (125)I-hCG. The receptors were functional, as indicated by an increase in cyclic AMP levels and activation of sperm protein kinase A following treatment with hCG or LH. Treatment with these hormones, on the other hand, had no effect on sperm protein kinase C activity. Now that the functional LH/hCG receptors are found in human sperm, it is important to determine whether hCG treatment could improve the outcome of infertility procedures.

Differences in hormonal characteristics of conceptive versus nonconceptive menstrual cycles.

OBJECTIVE: To determine the hormonal characteristics of a fecund menstrual cycle. DESIGN: Prospective observational study. SETTING: Clinical research center. PATIENT(S): Fourteen patients having artificial insemination with donor semen provided daily blood and urine samples in a nonconceptive cycle and the consecutive conceptive cycle. INTERVENTION(S): Concentrations of E2, luteinizing hormone, follicle-stimulating hormone (FSH), and P4 were measured in serum. Urine samples were analyzed for metabolites of E2, P4, and FSH. MAIN OUTCOME MEASURE(S): The serum and urinary hormone profiles of the periovulatory period were compared in conceptive and nonconceptive cycles. RESULT(S): The mean peak value of periovulatory urinary FSH was significantly higher in conceptive cycles than in nonconceptive cycles. The mean serum E2 concentration was significantly higher on day 0 (day of peak urinary FSH concentration) in conceptive cycles than in nonconceptive cycles, but mean peak values of serum E2 did not differ significantly. No other significant differences were observed in serum and urinary hormone profiles between conceptive and nonconceptive cycles. CONCLUSION(S): A lower, broader peak of FSH in urine was observed in nonconceptive cycles compared with conceptive cycles. Urinary FSH measurements may be useful in predicting less fecund ovulatory cycles and may identify some types of reduced female fertility.

Sperm morphology, motility, and concentration in fertile and infertile men.

BACKGROUND: Although semen analysis is routinely used to evaluate the male partner in infertile couples, sperm measurements that discriminate between fertile and infertile men are not well defined. METHODS: We evaluated two semen specimens from each of the male partners in 765 infertile couples and 696 fertile couples at nine sites. The female partners in the infertile couples had normal results on fertility evaluation. The sperm concentration and motility were determined at the sites; semen smears were stained at the sites and shipped to a central laboratory for an assessment of morphologic features of sperm with the use of strict criteria. We used classification-and-regression-tree analysis to estimate threshold values for subfertility and fertility with respect to the sperm concentration, motility, and morphology. We also used an analysis of receiver-operating-characteristic curves to assess the relative value of these sperm measurements in discriminating between fertile and infertile men. RESULTS: The subfertile ranges were a sperm concentration of less than 13.5 x 10(6) per milliliter, less than 32 percent of sperm with motility, and less than 9 percent with normal morphologic features. The fertile ranges were a concentration of more than 48.0 x 10(6) per milliliter, greater than 63 percent motility, and greater than 12 percent normal morphologic features. Values between these ranges indicated indeterminate fertility. There was extensive overlap between the fertile and the infertile men within both the subfertile and the fertile ranges for all three measurements. Although each of the sperm measurements helped to distinguish between fertile and infertile men, none was a powerful discriminator. The percentage of sperm with normal morphologic features had the greatest discriminatory power. CONCLUSIONS: Threshold values for sperm concentration, motility, and morphology can be used to classify men as subfertile, of indeterminate fertility, or fertile. None of the measures, however, are diagnostic of infertility.

Initial experience with a modification of the follicle aspiration, sperm injection, and assisted rupture technique.

OBJECTIVE: To establish cycle fecundity with a modification of the follicle aspiration, sperm injection, and assisted rupture (FASIAR) technique. DESIGN: Prospective, observational study. SETTING: University and health maintenance organization-based infertility centers. PATIENT(S): Infertile couples were enrolled from our professional practices. All patients were 16-18 mm in diameter by transvaginal ultrasonography. A modified FASIAR procedure was performed 22 to 28 hours after hCG injection. MAIN OUTCOME MEASURE(S): Clinical pregnancy rate. RESULT(S): No clinical pregnancies were observed with the modified FASIAR technique. CONCLUSION(S): The FASIAR technique is still an attractive and economical technique. Our modification of the FASIAR technique, however, resulted in a suboptimal cycle fecundity.

Alterations in humoral immune responses associated with recurrent pregnancy loss.

OBJECTIVE: To investigate the reactivity of maternal antibodies with endometrium-derived antigens and to correlate their association with recurrent pregnancy loss (RPL). DESIGN: Prevalence study. SETTING: Academic research center. PATIENT(S): Nulliparous women (n = 10), women with RPL (n = 15), pregnant women (n = 8), and multiparous women with a normal obstetric history (n = 20). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Reactive antibodies were analyzed by Western immunoblot techniques and quantitated by densitometry. RESULT(S): Antibodies from women with RPL and multiparous women recognized antigens ranging from 10-120 kd on normal endometrium and endometrial tumors. Antibodies from most women with RPL (10/15) and from multiparous women (15/20) recognized 65-kd and 80-kd proteins in normal endometrium. Antibodies from women with RPL recognized 21-kd and 28-kd antigens (12/15 and 13/15, respectively) in endometrial tumors at a significantly greater rate (than did antibodies from multiparous women (5/20 and 8/20, respectively). Women with RPL had significantly lower levels of asymmetric IgG compared with controls. CONCLUSION(S): Recurrent pregnancy loss may be linked with the failure to elicit asymmetric IgG and a unique immunologic recognition of endometrial antigens.

Efficacy of superovulation and intrauterine insemination in the treatment of infertility. National Cooperative Reproductive Medicine Network.

BACKGROUND: Induction of superovulation with gonadotropins and intrauterine insemination are frequently used to treat infertility. We conducted a large, randomized, controlled clinical trial of these treatments. METHODS: We studied 932 couples in which the woman had no identifiable infertility factor and the man had motile sperm. The couples were randomly assigned to receive intracervical insemination, intrauterine insemination, superovulation and intracervical insemination, or superovulation and intrauterine insemination. Treatment continued for four cycles unless pregnancy was achieved. RESULTS: The 231 couples in the group treated with superovulation and intrauterine insemination had a higher rate of pregnancy (33 percent) than the 234 couples in the intrauterine-insemination group (18 percent), the 234 couples in the group treated with superovulation and intracervical insemination (19 percent), or the 233 couples in the intracervical-insemination group (10 percent). Stratified, discrete-time Cox proportional-hazards analysis showed that the couples in the group treated with superovulation and intrauterine insemination were 3.2 times as likely to become pregnant as those in the intracervical-insemination group (95 percent confidence interval, 2.0 to 5.3) and 1.7 times as likely as those in the intrauterine-insemination group (95 percent confidence interval, 1.2 to 2.6). The couples in the intrauterine-insemination group and in the group treated with superovulation and intracervical insemination were nearly twice as likely to conceive as those in the intracervical-insemination group. CONCLUSIONS: Among infertile couples, treatment with induction of superovulation and intrauterine insemination is three times as likely to result in pregnancy as is intracervical insemination and twice as likely to result in pregnancy as is treatment with either superovulation and intracervical insemination or intrauterine insemination alone.

Total urinary follicle stimulating hormone as a biomarker for detection of early pregnancy and periimplantation spontaneous abortion.

Total concentrations of follicle stimulating hormone (FSH) were evaluated in daily urine samples from conceptive and nonconceptive menstrual cycles by measurement of the FSH beta subunit following treatment of the samples to dissociate the FSH heterodimer. Samples were self-collected by normal subjects during cycles in which daily blood samples also were obtained. Daily blood and urine specimens were collected prospectively from 10 subject in conceptive cycles, which led to normal pregnancies, and from 10 subjects with bilateral tubal ligations to provide control samples form nonconceptive cycles. Mean serum and urinary FSH concentration profiles wer parallel in both groups following ovulation and during he first 9 days of the luteal phase. Mean values for both serum and urinary FSH rose significantly above the postovulatory baseline by 10-12 days following the midcycle luteinizing hormone (LH) peak in nonconceptive cycles, but did not rise at any time following ovulation during conceptive cycles. Following regression analysis of the changing FSH concentration between days 9-14 post-LH surge in conceptive cycles, a slope of

Analysis of pre-ovulatory changes in cervical mucus hydration and sperm penetrability.

Changes in cervical mucus occur during the proliferative phase of the menstrual cycle and are known to correlate with receptivity to sperm and to the endocrine milieu. Prior studies, however, have often lacked biological incisiveness and technical objectivity and precision. This study analyzed daily changes in mucus water content (hydration) prior to the LH surge (LH+0) in normal women, in relation to daily levels of serum LH, FSH, estradiol and progesterone, and to daily tests of sperm penetration of the mucus. Cervical mucus was studied for 12 cycles in 10 ovulating women. Three to ten mucus specimens were collected per cycle, over the days LH-8 to LH+0. Each specimen was subjected to measurement of both water content (hydration) and penetration by spermatozoa from fresh specimens of normal human semen. For the latter, a new microscale assay was developed and applied, which was amenable to very small volumes of mucus. The new technique determines objective measures of both the numbers of penetrating sperm (motile and non-motile) and the distance penetrated by the forward most vanguard sperm. In these experiments, variations in semen quality were controlled by performing a companion penetration assay in an artificial 1.5% polyacrylamide gel. The patterns of change in mucus hydration varied quantitatively among women, with preovulatory baseline levels ranging from 93.8-96.5%. All normal cycles (as defined by endocrine profiles) displayed a significant increase in hydration over a one-day period occurring 3-4 days before the LH peak. The magnitude of this shift varied among women between 2 and 3% (absolute hydration), a distinction well within the precision of the hydration assay. This quantum increase in hydration was more pronounced than the corresponding increase in serum estradiol on the same day. The change in mucus hydration, and the associated increase in sperm penetrability, were more consistent among cycles than the changes in reproductive hormones. There was a strong but non-linear correlation between mucus hydration and sperm penetrability. Once the value of hydration rose above approximately 97.5%, there was a substantial increase in penetrability. This 'cut-off point' in sperm penetrability was in the middle of the range of hydration values (across women) which preceded the quantum jump in hydration-which, itself, preceded the surge of LH. Hydration began to increase approximately 2 days before measurable increases in sperm penetration of the mucus in vitro. These results demonstrate that mucus hydration may be a valuable marker of the approach to ovulation and delineation of the fertile period. They also provide new methods for assessing sperm penetration into both large peri-ovulatory and very small samples of collected mucus.

The role of relaxin in glycodelin secretion.

Glycodelin is a glycoprotein named for its unique carbohydrate structure. Glycodelin is produced by the secretory endometrium during the late luteal phase and returns to baseline during menses of the ensuing cycle, whereas in conceptive cycles it rapidly increases. Although progesterone and possibly estradiol are required for glycodelin production, they are not directly involved in the synthesis and release of this protein. Their role may be development of the endometrial secretory glandular elements, whereas other factors are required to initiate and maintain glycodelin secretion. The pattern of relaxin secretion during the luteal phase and early pregnancy is similar to that of glycodelin, but their profiles have not been determined simultaneously. To investigate the relationship of relaxin and glycodelin, two studies were conducted. In the first study, relaxin, glycodelin, and ovarian steroids were measured in daily serum samples from nonconceptive and conceptive natural cycles. Profiles of relaxin and glycodelin were closely associated, with the onset of relaxin preceding glycodelin secretion by 1-2 days in nonconceptive cycles, and the pregnancy-associated increases in each hormone differing by about 2 days. The second study tested the hypothesis that relaxin stimulates glycodelin secretion. Samples were obtained from patients injected with human relaxin for 28 days. In subjects demonstrating ovarian cyclicity, glycodelin secretion was elevated, but it was not detected in subjects without ovarian cyclicity or in placebo-treated control subjects. This study reveals a close temporal and quantitative relationship between relaxin and glycodelin profiles in the late luteal phase and early pregnancy. It also demonstrates that relaxin administration can stimulate glycodelin production from a developed endometrium. This is the first report of a nonsteroidal ovarian factor that controls glycodelin secretion, and these results suggest a function for relaxin during early pregnancy. Glycodelin is a potent inhibitor of sperm zona pellucida binding by virtue of its extensive carbohydrate structure, but it is normally at a nadir in the periovulatory period. The data demonstrate that relaxin can stimulate glycodelin secretion throughout the menstrual cycle, including the periovulatory period, when relaxin-induced glycodelin secretion could have a contraceptive effect.

Characterization of human chorionic gonadotropin in normal and abnormal pregnancies.

In this study circulating human chorionic gonadotropin (hCG) levels in the peri-implantation period of natural cycles of normal women were characterized. The hypothesis that the bioactivity of hCG in abnormal pregnancies is different from that in normal pregnancies was tested daily through serum hCG measurements in two immunoenzymometric assays, a radioreceptor assay and a human luteinizing hormone/human chorionic gonadotropin--cyclic adenosine monophosphate--luciferase bioassay. In normal pregnancies, the levels of immunoreactive and bioactive hCG were higher than in abnormal pregnancies (p < 0.05). In addition, the slopes of the rise in concentration of hCG as measured in all four assays were significantly greater for normal pregnancies than for abnormal pregnancies (p < 0.01). The rate of rise of hCG bioactivity was significantly greater than the rise of immunoreactivity or receptor-binding activity in normal pregnancies, but this difference was not observed in abnormal pregnancies. These results indicate that: (1) A steep rise of bioactive hCG is a consistent feature of the peri-implantation period of normal pregnancies. (2) Abnormal pregnancies had a deficiency in the production of hCG, which was reflected in both the hCG levels and the rate of increase, even in the first week of implantation. (3) Some abnormal pregnancies produced hCG with lower biological activity than the hCG of normal pregnancies.

The relationship of corpus luteum volume to relaxin, estradiol, progesterone, 17-hydroxyprogesterone and human chorionic gonadotropin levels in early normal pregnancy.

Our purpose was to characterize the growth pattern of the corpus luteum of early normal human pregnancy and correlate this growth with the corpus luteum hormone products: relaxin, progesterone, estradiol and 17-hydroxyprogesterone. A prospective study of seven patients was initiated at a mean gestational age of 4 weeks and 2 days. Corpus luteum volume and hormone concentrations were determined for each study patient every 48 h for a period of 2 weeks. Transvaginal imaging of the corpus luteum was performed by a single observer. Corpus luteum volume was calculated using the formula for an ellipsoid (4/3 pi abc/8). Correlation between corpus luteum volume and hormone concentrations was tested using Pearson's r. There was a mean three-fold increase in corpus luteum volume between 4 and 6 weeks' gestational age. Concomitantly, relaxin and estradiol concentrations increased, 17-hydroxyprogesterone declined slightly, progesterone remained stable and human chorionic gonadotropin (hCG) increased exponentially. Mean positive correlations were shown between corpus luteum volume and relaxin (r = 0.72), corpus luteum volume and hCG (r = 0.68), and hCG and relaxin (r = 0.82). However, there was a lack of correlation between corpus luteum volume and estradiol, progesterone and 17-hydroxyprogesterone. We have shown that a rapid increase in the corpus luteum volume occurs in early normal human pregnancy without a parallel rise in the classic corpus luteum steroid products. We interpret these findings to suggest that growth of the corpus luteum in early human pregnancy is largely derived from the proliferation of non-steroid secreting cells. The precise role of these cells in controlling steroidogenesis in this gland has yet to be defined.

Clinical evaluation of luteal function.

OBJECTIVE: To determine the ability of luteal phase length determined by basal body temperature (BBT) pattern and a midluteal serum progesterone level to predict the result of an endometrial biopsy in a subsequent cycle. METHODS: We performed a retrospective analysis of 141 women with a history of infertility who were being evaluated for luteal function. The luteal phase length determined from a BBT chart of one menstrual cycle was compared to a single midluteal serum progesterone level from a second menstrual cycle. These findings were compared to a luteal phase endometrial biopsy performed in a third menstrual cycle. Subjects were divided into four groups depending upon luteal phase length (normal 11 or more days) and serum progesterone level (normal at least 10 ng/mL). The four groups were designated "normal," "short luteal phase," "low progesterone," and "abnormal," depending upon the results of the two tests. The frequency of in- and out-of-phase endometrial biopsy results in the four groups was compared. RESULTS: There was no difference in the occurrence of an in- or out-of-phase endometrial biopsy when the four groups were compared. CONCLUSION: Neither luteal phase length nor a single midluteal serum progesterone level was predictive of subsequent in-phase or out-of-phase endometrial biopsy.

An unusual late presentation of imperforate hymen.

BACKGROUND: The imperforate hymen is a rare gynecologic abnormality that may not be detected until the onset of menses, when hematocolpos causes symptoms due to an expanding pelvic mass. CASE: A 19-year-old woman had primary amenorrhea, originally misdiagnosed as müllerian agenesis. Because of a combination of late onset of menses, vaginal dilation, and sexual intercourse, her hematocolpos remained relatively asymptomatic. CONCLUSION: Proper diagnosis of the imperforate hymen could have been made earlier by contrasting the ultrasound findings with the absence of associated genitourinary and skeletal anomalies. Magnetic resonance imaging can detect a uterus not previously seen on ultrasound. An early imaging study may further prevent the misdiagnosis of congenital absence of the uterus.

Semen preparation with the Sperm Select System versus a washing technique.

OBJECTIVE: To compare sperm migration through sodium hyaluronate with simple washing as methods for preparing sperm for IUI. DESIGN: Ten normal semen specimens were prospectively collected and samples were prepared by simple washing and by migration into sodium hyaluronate using the Sperm Select System (Select Medical Systems, Williston, VT). The semen and each treatment group were evaluated for sperm concentration, percent motile, viability, acrosomal status, longevity, and computer-aided semen analysis (CASA) parameters. SETTING: University reproductive endocrinology facility. RESULTS: The recovery of motile sperm was significantly higher for the washing method (mean +/- SEM 75% +/- 7%) than for the hyaluronate method (10% +/- 1%). The number of motile sperm recovered by migration into hyaluronate was independent of the percentage of motile sperm in the semen specimen and positively correlated with sperm concentration. The hyaluronate method produced greater percentages of motile, viable, and morphologically normal sperm, with lower proportions of premature acrosome reactions, higher sperm velocity, and greater linearity. CONCLUSIONS: The Sperm Select System method of sperm separation provides a highly uniform specimen with improved sperm quality. However, the recovery of motile sperm is considerably lower than for simple washing methods.

Classification of adverse reproductive effects can be improved by measurements of multiple biomarkers for ovarian toxicity and early fetal loss.

Hormone assays have been developed and applied for monitoring reproductive function using self-collected urine samples in non-clinical populations of women. Early pregnancy loss, menstrual dysfunction, reduced fertility as well as the site of toxicity can now be assessed using daily early morning urine samples. The understanding of the specific limitations of individual assay systems is important, however, to make the best use of these systems. The use of multiple end-points and computer algorithms is suggested to avoid misclassification of adverse reproductive events.