Effect of prolactin on follicle-stimulating hormone receptor binding and progesterone production in cultured porcine granulosa cells.

OBJECTIVE: To determine the effect of prolactin (PRL) on follicle-stimulating hormone receptor (FSH-R) binding and progesterone (P) production in cultured porcine granulosa cells. DESIGN: Controlled experiment. SETTING: Academic research laboratory. INTERVENTION(S): Immature granulosa cells were cultured in a serum-free medium. All cell populations were supplemented with porcine (p) FSH and cultured in the absence or presence of ovine (o) PRL. MAIN OUTCOME MEASURE(S): Specific pFSH-R binding and P in medium. RESULT(S): In the control cells, FSH-R binding increased 31-fold and P production increased 700-fold by day 4. Physiologic levels of oPRL potentiated the action of pFSH and resulted in a further 50% increase in pFSH-R binding and P production by day 4 over that in controls. In contrast, higher concentrations of oPRL blocked the rise in both pFSH-R binding and P production. The alteration in pFSH-R binding was associated with a change in FSH-R number. CONCLUSION(S): Physiologic levels of PRL amplify the stimulatory effects of FSH on the acquisition of the FSH-R and P production in cultured granulosa cells. Higher concentrations of PRL cause a decrease in FSH-R binding and P production. Prolactin may act as a "co-gonadotropin" and fine-tune the process of folliculogenesis by altering the acquisition of granulosa FSH receptors.

Angiotensinogen genotype and plasma volume in nulligravid women.

OBJECTIVE: To determine if nonpregnant plasma volume is altered in women who are homozygous for the T 235 coding angiotensinogen allele, which predisposes women to an increased risk of preeclampsia. METHODS: We measured plasma volume by Evans blue dilution and analyzed it as a function of angiotensinogen genotype in 15 nulligravid women during midfollicular phase of 26 menstrual cycles. Eleven women were evaluated during two cycles, and four women were evaluated in one cycle. Fourteen women were white, and one was Asian. No subjects had illnesses or were taking medication. The range of body mass index (BMI [kg/m2]) was 20.2-31.0. Plasma volume (mL) was reported as plasma volume divided by BMI to control for variations in body sizes. Statistical analysis was performed by analysis of variance with post hoc testing using Fisher least significant difference test for multiple comparisons (P < .05 accepted for significance). RESULTS: Angiotensinogen genotype analysis showed five women homozygous for M 235, three women homozygous for T 235, and seven women who were heterozygous (MT 235). T 235 homozygotes had significantly lower plasma volume divided by BMI compared with women who were homozygous for M 235 and women who were heterozygous for MT 235 (mean + standard deviation [SD] [71.2 + 8.8, 86.6 + 5.2, 95.8 + 15.6, respectively, P < .05]). There was a tendency toward higher plasma volume in heterozygote MT 235 compared with homozygote M 235 carriers, but it was not statistically significant. CONCLUSION: We conclude that the homozygous T 235 coding angiotensinogen genotype is associated with reduced plasma volume in nulligravid women during the follicular phase of the menstrual cycle compared with M 235 homozygotes and heterozygotes. This association of the T 235 coding genotype might contribute to fetal growth restriction in preeclampsia.

In vivo estrogen regulation of epidermal growth factor receptor in human endometrium.

The effects of estrogen and progesterone on the expression of epidermal growth factor receptor (EGFR) in human endometrium were studied in hypogonadal women under conditions that simulated a normal menstrual cycle. All women received the same regimen of estrogen and progesterone and underwent serial biopsies. In one group of women (group I), a biopsy was obtained before receiving estrogen (CD0) and after 11 days (CD11) of estrogen replacement. A second group of women was biopsied on CD11 and CD21 to assess the combined effects of progesterone and estrogen (group II). Immunohistochemistry was used to test for the presence of EGFR, and a ribonuclease protection assay was used to assess the amounts of EGFR messenger ribonucleic acid (RNA) relative to ribosomal RNA in the tissue. In group I, a significant increase in EGFR messenger RNA from CD0 to CD11 was observed. A similar increase was observed to occur between CD11 and CD21 in group II. Immunostaining for EGFR was absent in all CD0 biopsies, but was present in all estrogen-exposed endometrium. No difference in immunostaining was noted between CD11 and CD21. We conclude that estrogen stimulates the synthesis of EGFR in human endometrium and that progesterone does not appear to modulate this effect. The examination of other parameters in hormone-replaced hypogonadal subjects will be valuable in understanding the complex physiological regulation of the human endometrium.

Transvaginal ultrasonographic assessment of Hyskon or lactated Ringer's solution instillation after laparoscopy: randomized, controlled study.

We sought to evaluate two common fluids placed in the pelvis after pelvic surgery for their ability to remain in the pelvis for a time thought adequate for prevention of adhesions. Thirteen patients undergoing operative laparoscopy were randomized to receive 250 ml 32% dextran 70 (Hyskon), 250 ml lactated Ringer's solution, or no fluid (control) at the end of surgery. Serial transvaginal ultrasonograms were obtained at 1 hr, 3 hr, 6 hr, 24 hr, 96 hr (4 days), and 168 hr (7 days) after surgery. Patients were asked about side effects of fluid instillation. The volume of lactated Ringer's solution declined rapidly after instillation, with no significant difference from control at 24 hr (12 ml versus 7 ml). The volume of Hyskon did not decline rapidly by 24 hr and remained higher than the volume in controls or those receiving lactated Ringer's solution (188 ml, P = 0.003). Although the volume of Hyskon remained higher than that of lactated Ringer's solution or fluid volume in control patients by days 4 and 7, this difference did not reach statistical significance (45 ml versus 7 ml and 14 ml respectively, P = 0.39, on day 4). Patients in all groups noted abdominal pain. One patient who received Hyskon developed severe vulvar edema and another developed dyspnea. We conclude that the volume of Hyskon in the peritoneal cavity after laparoscopy does not decline as rapidly as does that of lactated Ringer's solution; however, significant side effects may limit its usefulness. Transvaginal ultrasonography is useful in monitoring fluids placed in the pelvis for prevention of adhesions.

The association of intrauterine filling defects on hysterosalpingogram with endometriosis.

OBJECTIVE: To determine whether there is a significant association between the presence of polyps or polypoid endometrium on hysterosalpingogram (HSG) with the presence of pelvic or peritoneal endometriosis. DESIGN: Retrospective chart review. SETTING: Department of Reproductive Endocrinology and Infertility at the University of Vermont, College of Medicine. PATIENTS: One hundred twenty patients, all of whom had a HSG, hysteroscopy, and laparoscopy by a member of our department between 1989 and 1993. MAIN OUTCOME MEASURE: Hysterosalpingograms were assessed for the presence of hypertrophic or polypoid endometrium. The presence of endometriosis was documented laparoscopically and scored according to The American Fertility Society classification and HSG findings were documented by hysteroscopy. RESULTS: Endometriosis was found in 27 of 32 women with polyps or polypoid endometrium but in only 19 of 88 women without. chi 2 analysis revealed a significant association between the demonstration of polyps or polypoid endometrium on HSG and the presence of endometriosis, chi 2(1) = 33.97. The predictive value of a positive test was 84% and the negative predictive value was 75%. CONCLUSION: The presence of polyps or polypoid endometrium on HSG is significantly associated with the presence of pelvic or peritoneal endometriosis.

Development of a sensitive porcine granulosa cell bioassay for follicle-stimulating hormone (FSH).

PURPOSE: Our objective was to develop a sensitive in vitro bioassay for follicle-stimulating hormone (FSH) that does not require the housing of animals in a research facility. MATERIALS AND METHODS: Porcine granulosa cells from 1- to 3-mm follicles were cultured on laminin for 48 hr in serum-free medium in the absence or presence of FSH or with other purified pituitary hormones, supplemented with 19-OH androstenedione. Estradiol accumulation in medium per microgram of DNA of cells was determined as a reflection of FSH-induced aromatase activity. RESULTS: FSH (0.01-10 ng/ml) caused a dose-dependent increase in estradiol production per microgram of DNA, with 1, 10, and 100 ng/ml significantly higher than control. Porcine FSH was approximately two fold more biopotent than rat FSH in this system. Higher doses of FSH (100 ng/ml) caused less estradiol accumulation, presumably reflecting FSH receptor down regulation. No other pituitary hormone produced significant estradiol accumulation. Unextracted serum from a patient with premature ovarian failure (10-50 microliters) was tested in parallel to purified rat FSH (0-50 ng/ml) in this system, resulting in similar estradiol accumulation per microgram of DNA. CONCLUSIONS: We have developed a porcine granulosa cell bioassay for FSH which is sensitive, is specific for FSH, and does not require the housing of animals on site. It can be completed by a technician within 4 working days and can detect FSH in a sample of human serum.

Reproductive outcome after tubal reversal in women 40 years of age or older.

OBJECTIVE: To determine the reproductive outcome of women who received a microsurgical tubal anastomosis operation at age 40 years or older. DESIGN: Multicenter retrospective cohort study. SETTING: Four university teaching hospitals. PATIENTS: Fifty-two women having undergone tubal sterilization reversal at age > or = 40 years. MAIN OUTCOME MEASURES: Pregnancy and live birth rate. RESULTS: Of the 52 women, 10 were lost to follow-up. Of those traced, 18 of 42 (42.8 percent) conceived. Of those 18, 6 patients had a live birth, 10 patients had a first trimester spontaneous abortion, 1 patient had an ectopic pregnancy, and 1 patient had an elective termination. Overall, the live birth rate was 14.3 percent, spontaneous abortion rate was 23.8 percent, and ectopic pregnancy rate was 2.4 percent. CONCLUSIONS: Microsurgical tubal anastomosis is a justifiable alternative to IVF-ET in women age 40 years or older.

Hysteroscopy versus hysterectomy for the treatment of abnormal uterine bleeding: a comparison of cost.

OBJECTIVE: To compare the costs of hysteroscopic treatment of abnormal uterine bleeding with the costs associated with abdominal and vaginal hysterectomy. DESIGN: Retrospective review. SETTING: Academic medical center and outpatient gynecology practice. PATIENTS: A list of all women undergoing an endometrial ablation (n = 60), hysteroscopic myomectomy (n = 14), abdominal (n = 192), or vaginal (n = 37) hysterectomy between June 1, 1987 and June 1, 1992 for the treatment of abnormal uterine bleeding was generated by a computer search of billing records using the appropriate ICD-9-CM codes. MAIN OUTCOME MEASURE: The direct cost per case for each patient were defined as the sum of the surgeon's fee, all related hospital and operating room charges, the anesthesiologist's fee, and the cost of preoperative medications. When the initial procedure failed, the cost associated with any additional surgery was accounted for in the determination of the direct cost per case. Indirect costs per case were calculated based on known demographic data, recovery time, and the cost in lost productivity during recuperation. Mean direct and indirect costs per case were determined for each procedure and compared. RESULTS: The mean direct cost per case for endometrial ablation was $5,159 and for myomectomy and ablation was $5,525. The direct cost per case was not different between the hysteroscopic procedures but both were significantly less expensive than either vaginal ($8,132) or abdominal ($8,833) hysterectomy. Total hospital cost also was significantly less for the hysteroscopic procedures than for hysterectomy by either technique. The indirect costs associated with the hysteroscopic procedures were significantly less than those incurred by abdominal and vaginal hysterectomy. CONCLUSIONS: The total direct and indirect cost per case of an hysteroscopic approach to the treatment of abnormal uterine bleeding is significantly less than that associated with vaginal or abdominal hysterectomy. This difference persists when the cost of failure of an hysteroscopic procedure is accounted for.

Color flow pulsed Doppler ultrasound in diagnosing luteal phase defect.

OBJECTIVE: To determine whether color flow pulsed Doppler analysis of corpus luteum blood flow in normal cycles differs from cycles with a luteal phase defect. DESIGN: A prospective study of natural ovarian cycles. SETTING: The University of Vermont Reproductive Endocrinology and Infertility Service. PATIENTS: Ten women with regular menstrual cycles and at risk for luteal phase defect (LPD) four with unexplained infertility, two with recurrent abortion, and four with age > 35 years. INTERVENTIONS: All women were examined by transvaginal color flow pulsed Doppler during the early follicular, late follicular, early luteal, midluteal, and late luteal phase of the menstrual cycle. Venous blood for P concentration was drawn on each day of Doppler exam. Urine testing for LH surge and endometrial biopsy during the late luteal phase were performed on each patient. MAIN OUTCOME MEASURES: Lowest resistance index associated with the highest amplitude signal from intraovarian vessels of each ovary, dated endometrial biopsies, serum P. RESULTS: Mean resistance indexes in LPD patients (n = 3) were significantly higher compared with normal women (n = 6) throughout the follicular and luteal phases. One patient remained anovulatory and was excluded from statistical analysis. Although systolic and diastolic velocities generally were observed to be lower in LPD patients compared with normal women, these differences were not statistically significant. High correlations were observed between P and resistance index within each luteal time point, achieving its highest value during the midluteal phase. CONCLUSIONS: This initial study provides evidence that color flow pulsed Doppler analysis of blood flow impedance to the corpus luteum may aid in assessing luteal phase adequacy.

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.

Prognostic significance of morphologic changes of the corpus luteum by transvaginal ultrasound in early pregnancy monitoring.

OBJECTIVE: To determine whether the ultrasound appearance, size, or change in size of the corpus luteum of early pregnancy correlated with serum progesterone, estradiol (E2), or 17-hydroxyprogesterone or were predictive of pregnancy outcome. METHODS: Transvaginal ultrasound assessment of the corpus luteum was performed prospectively on 55 women between 4-8 weeks' gestation. Forty-five (82%) subjects conceived in spontaneous cycles and ten (18%) conceived in cycles stimulated with clomiphene citrate. Fifty-three of 55 (96.4%) women had a second ultrasound assessment 5-8 days later (mean 6.7). Blood was drawn from each patient on the day of the ultrasound examination to measure hormone concentration. RESULTS: The appearance of the corpus luteum (macrocystic [more than 50% cystic], microcystic [less than 50% cystic], or noncystic) was not predictive of hormone concentration or pregnancy outcome. A nonviable pregnancy occurred in five of six (83%) women in whom a corpus luteum was undetectable by ultrasound and in 15 of 49 (31%) women in whom a corpus luteum was present (P = .01). There was no specific corpus luteum volume which could predict pregnancy failure. However, when a decreasing volume from first to second ultrasound examination was observed, 11 of 20 (55%) pregnancies resulted in nonviable outcomes compared to five of 27 (19%) when an increasing volume was observed (P < .01). There was no significant positive correlation between corpus luteum volume and plasma progesterone or 17-hydroxyprogesterone. A weak correlation was observed between corpus luteum volume and E2 (r = 0.38, P = .04). CONCLUSIONS: Our data reveal a lack of correlation between the size of the corpus luteum on ultrasound examination and known steroid products in pregnancies conceived during spontaneous cycles. Corpus luteum volume and steroid products were higher in those patients whose ovulation was induced with clomiphene citrate. Also, different morphologic appearances of the corpus luteum in early human pregnancy, based on the amount of cystic component, have no functional significance. However, a decreasing corpus luteum volume before 8 weeks' gestation is associated with a higher probability of early pregnancy loss.

Efficacy and safety of single-dose systemic methotrexate in the treatment of ectopic pregnancy.

OBJECTIVE: To evaluate the safety and efficacy of single-dose systemic methotrexate (MTX) in the treatment of ectopic pregnancy (EP). DESIGN: A database was started and continued prospectively for 35 patients meeting criteria for MTX therapy from June 1991 to October 1993. Follow-up was performed retrospectively on all patients with EPs (n = 82) by evaluating hospital and clinic records and by contacting affiliated physicians and individual patients. SETTING: The University of Vermont Reproductive Endocrinology Service. INTERVENTIONS: Methotrexate 50 mg/m2 was administered IM; blood samples were collected on days 0, 4, and 7 of MTX therapy and weekly thereafter until hCG titers became < 4 mIU/mL. RESULTS: Thirty-five of 82 (42.7%) patients diagnosed with EP were treated with MTX. The mean hCG concentration on day of treatment was 1388.1 +/- 463.5 (+/- SE) mIU/mL, and mean time to complete resolution of hCG was 23.1 +/- 2.9 days. Thirty of 35 (85.7%) were successfully treated with a single dose of MTX. Five of 35 (14.3%) failed therapy and required laparoscopic surgery. Twelve of 35 (34.3%) experienced mild side effects that resolved spontaneously. Ten of 13 (76.9%) demonstrated tubal patency at follow-up hysterosalpingogram. Of the 15 patients seeking pregnancy, 3 of 15 (20.0%) conceived, resulting in 3 term deliveries and 2 spontaneous abortions. CONCLUSIONS: Our results support the use of single-dose systemic MTX for the treatment of unruptured EP in carefully selected patients.

Septate uterus with cervical duplication: a rare malformation.

This previously unreported case of complete uterine septum, cervical duplication, and a longitudinal vaginal septum is best explained by the theory of Muller et al. (5), not by classically held views of unidirectional müllerian development. According to their theory, this anomaly could develop from failure of fusion of the most caudad müllerian ducts, resulting in a normal uterine fundus, with a complete septum, cervical duplication, and a longitudinal vaginal septum. This anomaly was accurately characterized using an endoscopic approach.

Pregnancy after laparoscopic neosalpingostomy in a patient with atresia of the distal fallopian tubes.

Structural congenital anomalies of the fallopian tube are interesting clinical problems that are infrequently reported in the literature. We report a patient with congenital absence of the distal fallopian tubes. After surgical correction by laparoscopic neosalpingostomies using a new intussusception technique, a normal IUP occurred and was delivered at term.

Reproductive performance following a second ectopic gestation.

OBJECTIVES: Our purpose was to determine the obstetric outcome following a second ectopic gestation in women actively trying to conceive, with a review of the literature. METHODS: Charts of 37 patients coded for having at least two ectopic pregnancies between 1986 and 1989 were reviewed. Duration of follow-up ranged from 7 months to 7 years with a mean follow-up time of 25 months. RESULTS: We report a 45.4% intrauterine pregnancy rate, 27.3% live birth rate, and 36.4% recurrent ectopic pregnancy rate. Review of the literature shows an intrauterine pregnancy rate of 26-50%, live birth rate of 25-31.2%, and recurrent ectopic pregnancy rate of 7.7-40%. CONCLUSIONS: Four of five studies report the risk of a third ectopic gestation is less than an intrauterine gestation. This may prove helpful in counseling patients with a history of recurrent ectopic gestation in choosing IVF or attempting conception naturally.

Ovulation after intravenous and intramuscular human chorionic gonadotropin.

OBJECTIVE: To define the time interval from intravenous and intramuscular hCG administration to follicular wall rupture and the endocrinologic events associated with ovulation. DESIGN: Subjects were studied in two cycles and received hCG either 10,000 IU IM or 500 IU IV in a random sequence with an intervening spontaneous menstrual cycle. PATIENTS: Thirty women from the University of Vermont Reproductive Endocrinology Service with unexplained, male, or cervical factor infertility. INTERVENTIONS: Subjects underwent superovulation with clomiphene citrate followed by hCG administration when the lead follicle reached a mean diameter of 18 mm. Follicular rupture was determined by ultrasound monitoring every 2 hours starting 31 and 30 hours after intravenous and intramuscular hCG, respectively. Serum samples were obtained hourly for hormone measurements. The study was completed 2 hours after follicular rupture or 48 hours after hCG administration. RESULTS: Twenty-five subjects received both intramuscular and intravenous hCG. The mean time to ovulation was 40.4 hours after intramuscular hCG (range, < or = 36 to > or = 48 hours) and 38.3 hours after intravenous hCG (range, 33 to > or = 48 hours). No differences were noted in the time interval to ovulation or rate of change in circulating E2 and P levels after IM versus IV hCG administration. CONCLUSIONS: These findings suggest (1) ovulation occurs over a broad range of time after hCG administration; (2) ovulation does not occur in a more specific time interval after intravenous than intramuscular hCG; and (3) the rate of change in circulating E2 and P levels are not different after intravenous than intramuscular hCG.

In vitro CA-125 secretion by endometrium from women with advanced endometriosis.

OBJECTIVE: To determine if the endometrium of women with endometriosis is a potential source of the elevated serum concentrations of CA-125 associated with endometriosis. DESIGN: Secretion of CA-125 by the endometrium in tissue culture was documented and measured in response to treatment with progesterone (P), 17 beta-estradiol (E2), and progesterone and estradiol (E2+P). Sequential biopsies of the endometrium from the early and late luteal phase were obtained from women with laparoscopically documented normal anatomy or moderate to severe endometriosis. SETTING: Division of Reproductive Endocrinology, Department of Obstetrics and Gynecology at the University of Vermont, College of Medicine, Burlington, Vermont. MAIN OUTCOME MEASURES: Secretion of CA-125 by endometrium in vitro was determined by radioimmunoassay in spent media using a commercially available kit (Centicor CA-125 Kit; Centicor, Malvern, PA). RESULTS: Secretion of CA-125 by endometrium in vitro is inhibited by the presence of P and E2+P in both the early and the late luteal phase. The amount of CA-125 secreted increases significantly from the early to the late luteal phase in all treatment groups. When compared with controls, the endometrium of women of with endometriosis secreted two to four times more CA-125 in all treatment groups, in both the early and late luteal phase. CONCLUSIONS: When compared with control endometrium, the endometrium of women with advanced endometriosis secretes significantly more CA-125 and represents a potential source of the elevated serum levels of CA-125 seen in these women.

CA-125 levels in human uterine fluid.

OBJECTIVE: To measure uterine fluid CA-125 concentration and to determine if any menstrual cycle phase dependent changes exist in its level. Serum levels are measured for comparison. DESIGN: CA-125 levels in uterine fluid were measured during the follicular and luteal phases of the menstrual cycle. In a sequential study, paired uterine fluid and serum samples were obtained once in both midfollicular and midluteal phases of the same menstrual cycle. RESULTS: CA-125 in uterine fluid during the follicular phase (n = 14) ranged from 16.4 x 10(3) to 616.5 x 10(3) U/mL, and from 6.2 x 10(3) to 567.3 x 10(3) U/mL in the luteal phase (n = 11). In the paired sequential uterine fluid and serum samples, (1) the means (+/- SEM) CA-125 in uterine fluid were 81.5 x 10(3) +/- 37.9 x 10(3) U/mL and 91.4 x 10(3) +/- 56.8 x 10(3) U/mL in the midfollicular and midluteal phases, respectively (P = 0.75); (2) the CA-125 levels in serum increased in the midluteal phase (P less than 0.05); and (3) compared with serum, uterine fluid levels were greater with a wider range. CONCLUSIONS: When compared with serum CA-125, uterine fluid contains high concentrations varying over a wide range without fluctuation between the follicular and luteal phases of the menstrual cycle.

Recurrent spontaneous abortion.

Recurrent pregnancy loss is a condition with many different etiologies. The proper evaluation and care of couples suffering from this form of reproductive failure requires that the clinician be well versed in many aspects of the physiology of normal and abnormal early pregnancy. Topics relevant to the subject of recurrent spontaneous abortion, with special attention to recent advances, are examined in this review.