Local estrogen treatment in patients with urogenital symptoms.

OBJECTIVES: Determination of the efficacy and safety of vaginally administered low dose (25 microg) micronized 17beta-estradiol in the management of patients with urogenital symptoms. METHODS: A total of 1612 patients with urogenital complaints were randomized to receive 25 microg of micronized 17beta-estradiol (n=828) or placebo (n=784) in a multicenter double-blind placebo-controlled study running for 12 months. Female patients were treated once a day over a period of 2 weeks, and then twice a week for the remaining of the 12 months with an active or placebo tablet. The assessment included full history-questionnaire, micturition diary, gynecologic and cystometric examination, transvaginal ultrasound, and serum 17beta-estradiol level determination. It was carried out at the beginning, and after 4 and 12 months of treatment. RESULTS: The overall success rate of micronized 17beta-estradiol and placebo on subjective and objective symptoms of postmenopausal women with vaginal atrophy was 85.5%, and 41.4%, respectively. A significant improvement of urinary atrophy symptoms was determined in vaginal ERT group as compared with the beginning of the study (51.9% vs. 15.5%, P=0.001). The maximal cystometric capacity (290 ml vs. 200 ml, P=0.023), the volume of the urinary bladder at which patients first felt urgency (180 vs. 140, P=0.048), and strong desire to void (170 ml vs. 130 ml, P=0.045) were significantly increased subsequent to the micronized 17beta-estradiol treatment. The number of patients with uninhibited bladder contractions significantly decreased following micronized 17beta-estradiol as compared with pretreatment values (17/30, P=0.013). Side effects were observed in 61 (7.8%) patients treated with low dose micronized 17beta-estradiol. Therapy with 25 microg of micronized 17beta-estradiol did not raise serum estrogen level nor stimulated endometrial growth. CONCLUSIONS: Local administration of 25 microg of micronized 17beta-estradiol is an effective and a safe treatment option in the management of women with urogenital complaints.

Ultrasonic assessment of the peri- and postmenopausal ovary.

OBJECTIVE: To analyze the role of B-mode ultrasound, color and power Doppler examination and three-dimensional ultrasound with and without power Doppler facilities in evaluation of the peri- and postmenopausal ovaries. METHODS: Review of the literature on ultrasonic assessment of the ovaries in postmenopausal patients. RESULTS: Ultrasound has assumed an increasing role in the evaluation of the peri- and postmenopausal ovaries. Ovarian lesions are a cause of great concern because of their malignant potential and the limited ability to distinguish accurately between benign and malignant lesions using morphological sonographic criteria alone. Superimposed color Doppler imaging enables the visualization of vascularization and blood flow characteristics of the ovaries and/or ovarian lesions. It becomes possible to detect normal, suspicious and pathologic blood flow characteristics even in small blood vessels. Owing to superb quality of the images, three-dimensional sonography enables precise assessment of the ovarian lesions. Three-dimensional power Doppler ultrasound permits effective targeting and rendering of ovarian tumor vasculature, thus providing new diagnostic possibilities for early detection of ovarian malignancy and differentiation of benign from malignant ovarian pathology. CONCLUSIONS: Further technological development will contribute to more objective evaluation of ovarian tumor morphology and vasculature which will give us tremendous support in clinical decision making and planning the management.

Extraovarian steroid cell tumor 'not otherwise specified' as a rare cause of virilization in twelve-year-old girl.

BACKGROUND: We present a 12-year-old girl with a 5-year history of progressive virilization. RESULTS: Regarding elevated plasma levels of 17-hydroxyprogesterone (17-OHP) and androgens, normal ultrasound and CT scan of ovaries and adrenal glands, the nonclassic form of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency was presumed the cause of virilization. As the glucocorticoid therapy did not normalize high levels of 17-OHP and androgens, and the DNA analysis did not demonstrate a mutation causing CAH, a laparotomy was performed. Near the right ovary a tumor was found and extirpated. Pathohistological studies determined it to be a rare steroid cell tumor, 'not otherwise specified'. Within the next months the signs of virilization resolved and menarche occurred. CONCLUSIONS: Steroid cell tumor should be considered in differential diagnosis of virilization in childhood. Regarding the age of our patient and pathohistological findings of the tumor, her prognosis is favorable.

Comparison of protective media and freezing techniques for cryopreservation of human semen.

OBJECTIVE: To evaluate the influence of cryopreservation medium and freezing-thawing techniques on human sperm motility and morphology. STUDY DESIGN: 63 semen samples were obtained from 39 donors to the artificial insemination programme. Possible effects of the sperm dilution with cryomedium on the motility were examined 10 min after exposure of 24 high initial quality semen samples to TEST-yolk ¿zwitterion-citrate-egg yolk extender containing TES [N-Tris (hydroxymethyl) methylaminoethane sulfonic acid] and Tris [(hydroxymethyl) aminomethane]¿ and human sperm preservation medium (HSPM). Post-thaw sperm motility from 24 frozen semen samples was examined comparing the cryoprotective efficacy of TEST-yolk and HSPM following different freezing techniques (vapour freezing, fast programmable freezing and slow programmable freezing). The relationship of sperm morphology to the effects of freezing was investigated on 39 semen samples following different freezing techniques. Post-thaw sperm motility from 39 frozen semen samples was compared among three groups divided according to the percentage of morphologically normal cells (<40, 40-50 and >50%) in fresh semen. RESULTS: Exposure of spermatozoa to cryomedia for 10 min at room temperature significantly reduced motility in TEST-yolk treatment group for 9% and in HSPM treatment group for 18% (P<0.01). The recovery of motile sperms (mean+/-standard deviation) was 49+/-15.7, 43+/-15.2 and 52+/-16.8% when TEST-yolk was used and 34+/-17.8, 32+/-18.2 and 50+/-13.6% when HSPM was used as a cryopreservative following vapour freezing, and fast and slow programmable freezing, respectively. Following vapour freezing and also following fast programmable freezing, the recovery of motile sperm was significantly higher (P<0.05) after addition of TEST-yolk medium than after addition of HSPM. Post-thaw motility of the sperm cryopreserved in HSPM showed significant differences (P<0.05) after three different freezing techniques. The recovery of motile sperms was 57+/-26.4, 38+/-8.6 and 38+/-17.3% in groups with >50, 40-50 and <40% morphologically normal cells, respectively. The percentage of morphologically normal spermatozoa was reduced 8% after vapour freezing and 6 and 3% after fast and slow programmable freezing, respectively. The results were statistically analysed using SAS/STAT software. CONCLUSIONS: Slow programmable freezing was superior to vapour freezing and fast programmable freezing as a method for sperm cryopreservation. However, none of these methods of freezing had discernible effects on sperm morphology. Motility of spermatozoa decreased due to the exposure of semen to cryomedium. TEST-yolk was a superior cryomedium to HSPM. Fresh semen with more than 50% of morphologically normal cells showed the best recovery of motile cells after freezing and thawing.

Combined interstitial and intrauterine pregnancies after in-vitro fertilization and embryo transfer.

A case of combined interstitial and intrauterine pregnancies after bilateral salpingectomy and in-vitro fertilization with embryo transfer is reported. The case was incorrectly diagnosed ultrasonographically as intrauterine triplets at 7 weeks gestation. The patient suffered from intra-abdominal bleeding at 14 weeks gestation. At laparotomy, a ruptured left interstitial pregnancy with a non-viable fetus was found in the left abdominal quadrant. The uterine defect was successfully repaired and gestational contents and blood were evacuated from the abdominal cavity. The intrauterine twin pregnancy progressed without incident, and a Caesarean section was performed at 36 weeks gestation, resulting in healthy male and female infants.

Simultaneous bilateral tubal pregnancy following in-vitro fertilization and embryo transfer.

A case of simultaneous bilateral tubal pregnancy resulting from in-vitro fertilization and embryo transfer is presented. Repeated transvaginal ultrasound examinations confirmed an intrauterine sac but no fetus. A diagnosis of early missed abortion was incorrectly made and a curettage was performed. The pathological examination showed the presence of decidua and Arias-Stella phenomenon but no chorionic villi. Diagnostic laparoscopy and laparotomy performed 40 days after embryo transfer (eighth week of gestation), revealed bilateral tubal pregnancy. Bilateral salpingectomy was performed.

[National consensus on hyperandrogenemia]. / Nacionalni konsenzus o hiperandrogenemiji.

Croatian Endocrine Society and Croatian Academy of Medical Sciences organized Symposium on Hyperandrogenaemia on March 22nd, 1996. Different aspects of this syndrome were discussed: epidemiology, classification and clinical features, steroid biosynthesis in the adrenal gland and ovarium, the genetics of polycystic ovarian syndrome (PCOS), clinical significance of testosterone and dihydrotestosterone metabolism, androgen excess and metabolic syndrome (syndrome X), insulin disturbances in PCOS, increased risk for development of non insulin dependent diabetes mellitus, androgen effects on serum lipoproteins, insulin like growth factors and function of ovarium, Doppler parameters in PCOS, treatment of hyperandrogenaemia, skin changes in PCOS, tests for adrenal and ovarial function, arterial hypertension and hyperinsulinism. National Board of Hyperandrogenaemia has been elected.

Number of follicles, oocytes and embryos in human in vitro fertilization is relative to serum estradiol and progesterone patterns during different types of ovarian hyperstimulation.

Preovulatory serum estradiol and progesterone levels as well as their ratio were compared in different types of ovulation induction in order to determine whether these findings could be used to predict the number of preovulatory follicles, number of oocytes aspirated and embryos obtained. Significantly more oocytes were retrieved by follicular aspiration and significantly more embryos developed in patients receiving gonadotropin-releasing hormone agonist and human menopausal gonadotropins than in those given other ovulation inductors. On days -2 and -1 of the cycle, serum estradiol levels were significantly lower in pure follicle-stimulating hormone induction. Serum progesterone was significantly higher in pure follicle-stimulating hormone cycles on days -4 and -3. In clomiphene citrate and human menopausal gonadotropin induction, progesterone levels were significantly lower on days -2 and -1, and on the day of follicular aspiration. Ratios of estradiol/progesterone were lower in pure follicle-stimulating hormone group from day -3 to day -1 of the cycle. A significant correlation was found between estradiol and progesterone serum levels and the numbers of preovulatory follicles, oocytes and embryos. The study revealed the usefulness of serum estradiol and progesterone determinations in assisted reproduction.

Follicular fluid contents of hyaluronic acid, follicle-stimulating hormone and steroids relative to the success of in vitro fertilization of human oocytes.

OBJECTIVES: To determine the concentrations of hyaluronic acid, FSH, P, and E2 in the follicular fluid (FF) obtained from IVF-ET patients and to assess the value of these measurements in predicting the outcome of fertilization. DESIGN: One hundred eleven samples were retrospectively analyzed for the hyaluronic acid and hormone contents. SETTING: University-based tertiary care center. PATIENTS: Preovulatory FF samples were collected from 67 women undergoing IVF-ET treatment because of tubal absence or obstruction. MAIN OUTCOME MEASURES: The FF hyaluronic acid and hormone concentrations were compared according to the type of ovulation induction, follicular development, and IVF outcome. RESULTS: According to the type of ovulation induction, a significantly lower hyaluronic acid concentration was found in FF harvested from the patients treated with GnRH agonist-hMG. No significant correlation was found between FF hyaluronic acid and either morphological maturity of the oocyte-cumulus complex or fertilizability of oocytes. The level of FSH was significantly higher in FF, yielding a mature oocyte-cumulus complex and from which the oocyte obtained successfully fertilized and cleaved. A significant increase in the E2 concentration was found in FF in which mature cumuli oophori were present. The levels of hyaluronic acid significantly correlated with FSH in FF. CONCLUSIONS: Expansion of the human oocyte-cumulus cell complex is an FSH-dependent phenomenon. The data are in agreement with the hypothesis that intrafollicular FSH plays an important role in the secretion of hyaluronic acid by granulosa cells and may act synergistically with E2 to enhance cytoplasmic maturation, resulting in successful fertilization.

Histochemical demonstration of a delta 5,3 beta-hydroxysteroid dehydrogenase activity of cumulus cells related to the maturity and developmental potential of recovered oocytes.

A simple and rapid histochemical technique is described for demonstration of delta 5,3 beta-hydroxysteroid dehydrogenase activity in cumulus cells from preovulatory follicles aspirated for in vitro fertilization (IVF) of corresponding oocytes. Histochemical activity of delta 5,3 beta-hydroxysteroid dehydrogenase was demonstrated in samples of cumulus obtained from 62 oocytes recovered from 24 women. Patients were treated with clomiphene citrate in combination with human menopausal gonadotropins and human chorionic gonadotropin injections. The cumulus was found to contain small and large cell types. Small cells possessed more delta 5,3 beta-hydroxysteroid dehydrogenase activity predominantly in the area near the oocyte. Cytoplasmic vacuolation has been noted in large, pale cells with moderate or low enzyme activity. The most active cells were predominant in cumulus from which oocytes were fertilized. Significant differences have been found between high and low delta 5,3 beta-hydroxysteroid dehydrogenase activity of cumulus cells from mature oocyte-corona-cumulus complexes leading to a successful fertilization and cleavage of oocytes and between groups with different histochemical activity when aspirated complexes were scored immature and the IVF of oocytes has failed.

Protein concentration in pre-ovulatory follicular fluid related to ovarian stimulation.

Sixty follicular fluids obtained from 26 women with either clomiphene citrate and human menopausal gonadotropins (hMG) or hMG-induced ovulation were analyzed for the contents of total proteins, fibrinogen, plasminogen, antithrombin III, ceruloplasmin, alpha-2 macroglobulin, alpha-1 antitrypsin and immunoglobulins (IgG, IgA, IgM). Concentrations of these proteins was correlated to the type of ovarian follicle growth induction. Follicular fluids from patients stimulated with clomiphene citrate-hMG contained significantly higher concentrations of ceruloplasmin than those treated with hMG alone. No significant differences in the concentrations of other proteins were noted between the two types of ovarian induction. A multivariate data analysis resulted in three Varimax factors (VRX I) suggesting that proteins with antiprotease activity in the follicular fluid may play a role in human follicle maturation. Follicular fluid Ig may reflect the degree of follicular wall permeability under hMG treatment. Accordingly, it may be assumed that a combination of different proteins described by VRX factors could be used for evaluation of ovarian stimulation.

[Validity of the direct immunofluorescent monoclonal antibody test in the diagnosis of chlamydial infection]. / Valjanost direktne imunofluorescencije monoklonskim protutijelima u dijagnostici klamidijske infekcije.

The sensitivity and specificity of direct fluorescent monoclonal antibody staining were compared with the isolation of the organisms in McCoy cells. For both diagnostic methods cervical smears were taken in 120 women. Of 120 women 25 (20.8%) were infertile, 60 (50.0%) healthy fertile women and 35 (29.8%) adolescents who came for control gynecological examination. C. trachomatis was isolated in 5/25 (20.0%) of infertile women, in 5/60 (8.3%) of healthy fertile women and in 4/35 (11.4%) adolescents. With the cell culture as the reference method, the antigen detection test had a sensitivity of 92.8%, a specificity of 98.1%, a positive predictive value of 86.7% and a negative predictive value of 99.1%.

Chromosomal preparations of human triploid zygotes and embryos fertilized in vitro.

Forty-eight zygotes with more than two pronuclei were identified after in vitro fertilization, representing 6.1% of all fertilized oocytes. The chromosome preparations from pronuclear stage to the cleaved human embryos were examined. Prophase was found in eight out of ten zygotes. The spreading of chromosomes allowed an adequate counting in only two cases. Six of the eight preparations displayed a late prophase. In this stage each haploid group of chromosomes can be analysed separately. Kariogamy usually occurred 4 to 5 h after the pronuclei had disappeared, and polyploid number of chromosomes were found in well-spread metaphases. The chromosomal preparations were made for eleven human embryos arising from zygotes with three pronuclei. Out of ten preparations, where the chromosomes could be counted, seven embryos (70%) contained hypodiploidic groups of chromosomes. In two of the cases, however, triploid metaphases were found, and in the last one a triploid/diploid mosaicism.

Lipid and lipoprotein contents of human follicular fluid.

The concentrations of total cholesterol, phospholipids, triacylglycerols and lipoproteins were measured in 87 follicular fluids obtained from 35 women undergoing in vitro fertilization and embryo transfer. The results were correlated with the levels of progesterone in follicular fluid. Two different types of ovarian stimulation were used. High density lipoproteins were the dominant lipoproteins found in the preovulatory follicular fluid. Low density lipoproteins were absent or appeared in trace amounts. Significantly higher triacylglycerol and high density lipoprotein levels were found when stimulation with human menopausal gonadotropins and chorionic gonadotropin was applied, as compared to the clomiphene citrate-menopausal gonadotropin-chorionic gonadotropin menstrual cycle. In both groups, extracorporal fertilization resulted in cleavage of oocytes and embryo transfer. No significant correlation between any follicular fluid lipid and progesterone concentration was found. The lipids estimated in the follicular fluid appeared to have no influence on the oocyte fertilizability. The presence of triacylglycerols and high density lipoproteins in the follicular fluid may indicate follicular wall permeability under the treatment with menopausal gonadotropins.

Prostaglandin F2 alpha, progesterone and estradiol concentrations in human follicular fluid and their relation to success of in vitro fertilization.

Prostaglandin F2 alpha (PGF2 alpha), progesterone (P4) and estradiol-17 beta (E2) in follicular fluid were measured by radioimmunoassay in patients undergoing in vitro fertilization and embryo transfer. Follicular growth was induced using clomiphene citrate-hMG-hCG (15 patients) and FSH-hMG-hCG (4 patients). There was no significant difference in follicular fluid PGF2 alpha and P4 concentrations relative to oocyte maturity as assessed morphologically. The highest PGF2 alpha concentration was found in fluid from FSH-hMG-hCG cycles where fertilization occurred. The value is significantly higher (p less than 0.002) than in fluid from clomiphene-hMG-hCG cycles whether fertilization took place or not. There was no significant difference in P4 and E2 levels in relation to the type of ovarian induction or success in fertilization. Positive correlation between P4 and E2 in follicular fluid was found (r = 0.404). The positive correlation between total dose of hMG given to the patients and PGF2 alpha concentration in their preovulatory follicular fluid (r = 0.434) suggests that PGF2 alpha is secreted locally as the result of hMG and hCG stimulation. It is proposed that PGF2 alpha could be a biochemical marker for assessing the success of in vitro fertilization.

[Fertilization in vitro in the treatment of male infertility]. / Fertilizacija in vitro u lijecenju muske neplodnosti.

Thirty marital couples were included in an in vitro fertilization study of male infertility. Second-degree oligospermia (less than or equal to 5 X 10(6)/ml mobile sperms) was found in 3 couples, first-degree oligospermia (less than or equal to 12 X 10(6)/ml mobile sperms) in 21 couples, and asthenospermia (less than 20% of mobile sperms) in 6 couples. The semen for insemination was prepared by the standard procedure and the overlay technique. After the separation (removal) of seminal fluid and capacitation the sperms were found to be significantly more mobile. Ovarian cells obtained by laparoscopy were inseminated with 50,000 to 250,000 mobile sperms. The results were compared with those in a group of patients with normospermia (greater than or equal to 12 X 10(6)/ml mobile sperms). The number of fertilized ovarian cells proved to be significantly lower in patients with second-degree oligospermia (P less than 0.001). There was no difference in the rate of fertilization and ovarian cell division either in patients with first-degree oligospermia or asthenospermia or normospermia. Seven clinical pregnancies were recorded: 2 in the second-degree oligospermia group, 4 in the first-degree oligospermia group, and 1 in the asthenospermia group.

[Pregnancy after fertilization in vitro and embryo transfer]. / Trudnoce nakon izvantjelesne oplodnje i prijenosa zametka u materiste.

The first 13 pregnancies established by in vitro fertilization and embryo transfer. A tubal factor was the main indication for in vitro fertilization (IVF) in 10 patients, whereas in three couples infertility was caused by the infertile sperm. The mean age and duration of infertility in female patients were 33.5 and 11.4 years, respectively. Multiple follicular growth was stimulated by either clomiphene citrate (100-150 mg during 5 days), clomiphene citrate + HMG + HCG, or HMG + HCG. Timing of ovulation was determined by assaying serum estradiol, progesterone and LH levels. The ultrasound monitoring of follicular growth was determined on a daily basis. Endogenous LH surge was observed in one patient, whereas in others 5000-10,000 IU HCG were needed to induce ovulation. All oocytes were recovered by laparoscopy. There were 3.8 oocytes per patient on an average. Fifty-six percent of oocytes were preovulatory. All oocytes were allowed to mature in vitro for 3 to 7 hours (mean 5.7 hours). The fertilization and cleavage rates were 76% and 72% respectively. Only one polyspermic fertilization was observed. Forty-five hours after the insemination the embryos between the 2-cell and 6-cell stages were reimplanted into the uterus. Using various ET cannulas, one to seven embryos per patient (2.7 embryos on the average) were replaced into the uterine cavity, 5 mm below the fundus. Only four embryo replacements were completely atraumatic. No medications were used for the luteal phase support. The corpus luteum function and early pregnancy were controlled by serial measurements of serum progesterone, estradiol and beta HCG. Three clinical miscarriages and full-term deliveries were recorded.(ABSTRACT TRUNCATED AT 250 WORDS)

[The effect of delayed insemination on fertilization in vitro and the development of human embryos]. / Ucinak odogodene inseminacije na oplodnju in vitro i razvoj humanih zametaka.

Extracorporeal fertilization of 363 oocytes was applied in 148 patients with tubal infertility. The aspiration of the follicles was performed 36 to 38 hrs after the onset of the endogenous LH rise or 36 hrs after injection of hCG. Oocytes maturity was evaluated on the basis of the morphological appearance of the cumulus oophorus. The eggs were incubated in culture medium before insemination. The duration of delayed insemination had a marked effect on the proportion of fertilized oocytes and the development of embryos (X2 = 10.34, P = 0.05). After maturation of 1.5 to 3.5 hrs, minimal preovulatory oocytes were fertilized and developed to embryos. Maturation of 3.5 to 5.5 hrs increased the number of fertilized oocytes from 56% to 71.7% and the number of embryos from 50% to 55.1%. After 5.5 to 7.5 hrs of maturation significantly more fertilized oocytes (t = 2.4, P less than 0.02) and developed embryos (t = 2.41, P less than 0.02) were obtained. Preincubation of intermediary preovulatory or immature oocytes had no beneficial effect on the fertilization rate and the development of embryos.

[Persisting suppression of hyperprolactinemia after bromcriptine withdrawal]. / Trajna supresija hiperprolaktinemije nakon lijecenja bromokriptinom.

The 22-years old patient with secondary amenorrhea, galactorrhea and macroprolactinoma of hypophysis was observed. After the therapy with Parlodel, which lasted 34 months, her menstrual cycle became regular and the level of prolactin was normal. The quantity of Parlodel used daily for the normalization of prolactin was 20 mg at the beginning, and only 2,5 mg later on. This quantity was taken intermittently, only until the ovulation. She conceived 5 months after therapy withdrawal. During the pregnancy period, prolactin in serum was not higher then physiological value. The pregnancy did not cause any growth of tumor. After the delivery and the break of lactation, hyperprolactinemia persisted, but only 29,1-34,4% of the value before the beginning of the therapy.

[Human beta chorionic gonadotropin in the early diagnosis of extrauterine pregnancy]. / Humani beta korionski gonadotropin u ranoj dijagnozi izvanmaternicne trudnoce.

In 1979 and 1980, at the University Hospital Department of Gynecology and Obstetrics, 115 patients with extrauterine pregnancy were surgically treated. In 26 patients the value of the radioimmunologic determination of beta chorionic gonadotropin (HCG) in the serum was analysed. Along with it, also other clinicodiagnostic procedures (immunochemical test for pregnancy, culdocentesis, curettage) wire carried out. Not in a single case of extrauterine pregnancy in which the serum beta HCG level was determined, was the result falsely negative. The immunochemical test for pregnancy was falsely negative in 32.6% and the result of culdocentesis was falsely negative in 13.6% of the patients examined. In patients with negative culdocentesis laparoscopy was performed (10.4%), showing ectopic pregnancy. In the course of extrauterine pregnancy the elimination of beta HCG was significatly reduced in relation to its level in normal pregnancy. The concentration of beta HCG in the observed patients with extrauterine pregnancy ranged from 27 to 3924 IU/L. On the basis of a low beta HCG concentration in the serum it is possible to determine with much certainty that pregnancy is not quite normal. Of great help is the observation of changes in the elimination of this hormone over a short period of time. A timely diagnosis of extrauterine pregnancy improves prognosis of future fertility.