Abstract: OBJECTIVE: To assess the impact of oral contraceptive pill (OCP) pretreatment in ovarian stimulation for in vitro fertilization (IVF) using gonadotropin releasing hormone (GnRH) antagonists. DESIGN: Systematic review and meta-analysis of randomized controlled trials (RTC). SETTING: University IVF center. PATIENT(S): Infertile patients (n = 847), treated in four trials. INTERVENTION(S): Systematic literature search (MEDLINE, EMBASE, CENTRAL COCHRANE, conference proceedings, reference lists) for randomized trials on OCP treatment before stimulation with gonadotropins and GnRH antagonist for IVF. Meta-analysis of data yielding pooled odds ratios (OR) or weighted differences of the means (WMD) and 95% confidence intervals (CI). MAIN OUTCOME MEASURE(S): Ongoing pregnancy rate per randomized woman. RESULT(S): Ongoing pregnancy rate per randomized woman was not found to be significantly different between patients with and those without OCP pretreatment (OR: 0.74, 95% CI: 0.53 to 1.03). Duration of gonadotropin stimulation (WMD: +1.41 days, 95% CI: +1.13 to +1.68) and gonadotropin consumption (WMD: +542 IU, 95% CI: +127 to +956) were significantly increased after OCP pretreatment. No significant differences were observed regarding the number of cumulus-oocyte complexes (COCs) and the fertilization rates. CONCLUSION(S): A significant difference in ongoing pregnancy rates between patients who received OCP pretreatment and those who did not is currently not present, although further studies are necessary for more solid conclusions on pregnancy likelihood to be drawn.

Abstract: The aim of this study was the comparison of liquid-based cytology (ThinPrep, TS) to conventional smears (CS) in the investigation of subfertile men with testicular fine-needle aspiration (FNA). Between January and December 2004, testicular FNA biopsies were performed in 30 subfertile men. Both TS and CS were diagnosed according to Meng classification. Features specifically recorded in each smear included sample adequacy, tissue cells preservation, contamination with red blood cells, quality of smear background, ease of cell recognition, and the cytological diagnosis.There was agreement in the cytological diagnosis between TS and CS (P = 0.88) and sample adequacy (P = 0.73). TS was superior to CS regarding cell preservation, presence of red blood cells or tissue artifacts, quality of the smear background, and cell recognition (P < 0.0001). In testicular FNA cytology, TS appear to be superior to CS in respect to cell preservation, absence of red blood cells, background quality, and cell recognition. These advantages, however, are not translated in improved cytological diagnosis.

Abstract: In recent years there is increasing evidence of an interaction between metabolic syndrome and testicular function. Metabolic syndrome, in particular obesity, affects testicular function by reducing total testosterone and sex hormone-binding globulin levels, as well as having a detrimental effect on spermatogenesis. On the other hand, hypogonadism further increases insulin resistance, which is the main pathophysiological feature of metabolic syndrome. There are implications that testosterone replacement can improve not only testicular function, but also parameters of the metabolic syndrome. Although the exact pathophysiological mechanisms remain unclear, leptin, resistin and ghrelin appear to play crucial roles in the interaction between metabolic syndrome and testicular function. All of this evidence supports the notion that the metabolic syndrome is a complex clinical entity characterized by pathophysiological mechanisms that affect the endocrine system as a whole; for these reasons it has been proposed to rename it 'metabolic-neuroendocrine syndrome'.

Abstract: OBJECTIVE: To examine the literature systematically in order to identify prospective comparative trials answering the following question: Is vitrification of human embryos associated with a higher postthawing survival rate as compared with slow freezing? DESIGN: Systematic review and meta-analysis. SETTING: University-based hospital. PATIENT(S): Not applicable. INTERVENTION(S): Vitrification versus slow freezing for cryopreservation of human embryos. MAIN OUTCOME MEASURE(S): Postthawing survival rate. RESULT(S): Four eligible studies were identified, three of which were randomized controlled trials. Overall, the current review summarizes information from 8,824 cryopreserved human cleavage stage embryos/blastocysts (vitrification: n = 7,482; slow freezing: n = 1,342). Survival rate of cleavage stage embryos was significantly higher after vitrification as compared with slow freezing (odds ratio 15.57, 95% confidence interval 3.68-65.82; random effects model). Postthawing survival rate of vitrified blastocysts was significantly higher compared with that observed with slow freezing (odds ratio 2.20, 95% confidence interval 1.53-3.16; fixed effects model). CONCLUSION(S): Vitrification appears to be associated with a significantly higher postthawing survival rate than slow freezing. Further prospective trials are necessary to confirm the above results and, in addition, allow the evaluation of the two cryopreservation methods in terms of pregnancy achievement.

Abstract: AIM: To determine stimulated serum anti-Mllerian hormone (AMH) levels in men with different causes of subfertility. SUBJECTS AND METHODS: We prospectively studied 82 subfertile men and 31 controls. The subfertile men underwent a diagnostic procedure to identify the causes of subfertility. Study parameters included testicular volume, levels of follicle-stimulating hormone, luteinizing hormone, total testosterone, prolactin, inhibin B and AMH, and sperm parameters. RESULTS: Clinical diagnoses in subfertile men were idiopathic non-obstructive azoospermia (n = 26, 32%), idiopathic non-obstructive dyspermia (n = 17, 21%), varicocele (n = 16, 20%), cryptorchidism (n = 10, 12%) and other diagnoses (n = 13, 16%). Serum AMH levels in subfertile men were 60% lower than in controls [median (interquartile range) 4.6 (3.6) vs. 11.6 (7.7) ng/ml, p 0.001], with no significant differences among the different groups of subfertile men. CONCLUSIONS: Serum AMH levels differentiate control from subfertile men but not men with different causes of subfertility.

Abstract: OBJECTIVE: To evaluate the risk of gestational diabetes mellitus (GDM) in women with polycystic ovary syndrome (PCOS). DESIGN: Systematic review and meta-analysis of observational studies. SETTING: Tertiary Department of Reproductive Endocrinology. PATIENT(S): Five thousand two hundred ninety-three pregnant women (721 with PCOS and 4,572 controls without PCOS). INTERVENTION(S): Literature search in the electronic databases MEDLINE, EMBASE, and CENTRAL, study of the references of all relevant trials or reviews, and manual search of the abstracts from the major meetings in the field of human reproduction. MAIN OUTCOME MEASURE(S): Gestational diabetes mellitus odds ratio. RESULT(S): Women with PCOS demonstrated a significantly higher risk for the development of GDM as compared with women without PCOS (odds ratio 2.89, 95% confidence interval [CI] 1.68-4.98), yet with significant statistical heterogeneity (I(2) = 59.3%), durable to sensitivity analysis. In the subgroup of cohort studies, this finding remained robust (7.11, 95% CI 2.95-17.12), whereas in the subgroup of case-control studies, it did not (0.89, 95% CI 0.38-2.06). Metaregression modeling revealed a linear dependence of the outcome on study type and baseline risk (post hoc). CONCLUSION(S): Significant heterogeneity among studies and dependence of the outcome on study type make the higher risk of GDM in women with PCOS a questionable finding. The conduction of properly designed studies should precede any recommendation to pregnant women with PCOS in regard to the risk of GDM.

Abstract: OBJECTIVE: To present a series of six consecutive women with adenomyomas who were successfully managed with a standard treatment strategy to elucidate the feasibility and the effectiveness of laparoscopic treatment of adenomyomas. DESIGN: Cross-sectional case series. SETTING(S): Obstetrics and gynecology department of a tertiary academic hospital and endoscopic unit of a private hospital. PATIENT(S): The six cases described in this report were nonpregnant women of reproductive age (mean age, 34.8 years old; range, 29-38 years) who presented in the outpatient gynecological clinic for yearly routine visit (one patient), dysmenorrhea and menorrhagia (three patients), and history of pregnancy loss (two patients). INTERVENTION(S): Laparoscopic excision of uterine adenomyomas. MAIN OUTCOME MEASURE(S): Feasibility and effectiveness of laparoscopic management of adenomyomas. RESULT(S): The average operating time was 100.5 minutes, and the average estimated blood loss was 163 mL. No event complicated the intraoperative and the postoperative course of these cases, and no case was converted to laparotomy. The mean follow-up was 13.7 months, with complete regression of the symptoms. CONCLUSION(S): Excision of adenomyomas presents intraoperative peculiarities involving difficulties in their dissection and manipulation. Laparoscopic management of these lesions appears to be safe and feasible with good follow-up results and limited recurrence rates.

Abstract: The aim of the present study was to investigate whether endometriosis and cancer share common molecular characteristics. Tissue samples were collected prospectively during diagnostic laparoscopy of patients with primary infertility. Using high-density oligonucleotide microarrays, (Affymetrix Gene Chip HG-U133 Set) the genome-wide gene expression profile of advanced ovarian endometriosis was analyzed compared with matched normal endometrium. Expression of TERT, the gene encoding the telomerase reverse transcriptase subunit, and telomerase activity were analyzed in eutopic and ectopic endometrium. Genome-wide, high-resolution array-CGH was used to screen for genomic aberrations in endometriosis. Expression microarray data were validated quantitatively with RT-PCR. The genes RARRES1 and RARRES2 (retinoic acid receptor responder 1 and 2) were found to be up-regulated in endometriosis, suggesting a high degree of differentiation. Consistently, down-regulated genes included those involved in the cell cycle, cell metabolism and homeostasis. Expression of TERT and telomerase activity were present in eutopic but absent in ectopic endometrium. Array-CGH revealed a normal genomic pattern without gross amplifications and deletions. In conclusion, these data suggest that advanced ovarian endometriosis represents a highly differentiated tissue with minimal or no malignant potential.

Abstract: BACKGROUND: Both cleavage-stage and blastocyst-stage embryo transfer policies have advantages and drawbacks. The number of embryos transferred, however, is a crucial parameter that needs to be considered before attempting any comparison. METHODS: An extensive literature search yielded initially 282 studies from which 8 randomized controlled trials met the inclusion criteria: (i) truly randomized design (ii) policy to transfer equal number of embryos in both the cleavage-stage and the blastocyst-stage groups and (iii) published as full text in a peer-review journal. Primary outcome was the live birth rate and secondary outcomes were clinical pregnancy rate, multiple pregnancy rate, cancellation rate and cryopreservation rate. RESULTS: A total of 1654 patients were reviewed. Live birth rate per randomized patient was significantly higher (n = 6 studies) in patients who had a blastocyst-stage transfer as compared to patients with cleavage-stage embryo transfer [odds ratio (OR): 1.39, 95% confidence interval (CI): 1.10-1.76; P = 0.005]. Clinical pregnancy rate (OR: 1.27, 95% CI: 1.03-1.55; P = 0.02) and cancellation rate per patient randomized (OR: 2.21, 95% CI: 1.47-3.32; P = 0.0001) were significantly higher in patients with a blastocyst-stage embryo transfer as compared to patients in whom a cleavage-stage embryo transfer was performed. The cryopreservation rate was significantly higher in the cleavage-stage group (OR: 0.28, 95% CI: 0.14-0.55; P = 0.0002). CONCLUSIONS: The best available evidence suggests that the probability of live birth after fresh IVF is significantly higher after blastocyst-stage embryo transfer as compared to cleavage-stage embryo transfer when equal number of embryos are transferred in the two groups compared.

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Abstract: OBJECTIVE: To compare FSH, inhibin B (INHB), and antimüllerian hormone (AMH) as predictors of the recovery of sperm in testicular fine-needle aspiration biopsy (FNA) performed in men with azoospermia. DESIGN: Cross-sectional, clinical study. SETTING: Academic Unit of Reproductive Endocrinology. PATIENT(S): Fifty-one men with azoospermia and 31 controls. INTERVENTION(S): Testicular FNA. MAIN OUTCOME MEASURE(S): Serum FSH, INHB, and AMH levels. RESULT(S): Clinical diagnoses in men with azoospermia were idiopathic nonobstructive azoospermia (n = 34, 67%), cryptorchidism (n = 4, 8%), varicocele (n = 3, 6%), and other diagnoses (n = 10, 16%). In pairwise comparison of receiver operating characteristic curves, none of FSH (area under curve 0.716), INHB (0.610), AMH (0.565), or volume of the larger testis (0.693) was proved to be superior to the others as predictor of sperm retrieval during an FNA procedure. Similarly, in a logistic regression analysis, none of FSH, AMH, INHB, or volume of the larger testis could predict presence of sperm in FNA. CONCLUSION(S): Serum INHB and AMH, as well as their combination, are not superior to FSH as predictors of the presence of sperm in testicular FNA in men with azoospermia and should not be used for this purpose.

Abstract: OBJECTIVE: To systematically review the literature to identify randomized controlled trials, which evaluate interventions aiming to improve the probability of pregnancy in poor responders undergoing in vitro fertilization (IVF). DESIGN: Systematic review and meta-analysis. SETTING: University-based hospital. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Pregnancy rate. RESULT(S): Twenty-two eligible randomized controlled trials were identified that evaluated in total 15 interventions to increase pregnancy rates in poor responders. Based on limited evidence, the only interventions that appear to increase the probability of pregnancy were the addition of GH to ovarian stimulation (odds ratio for live birth: 5.22, confidence interval: 95% 1.09-24.99) and the performance of embryo transfer on day 2 compared with day 3 (ongoing pregnancy rate: 27.7% vs. 16.3%, respectively; difference: +11.4, 95% confidence interval: +1.6 to +21.0). CONCLUSION(S): Insufficient evidence exists to recommend most of the treatments proposed to improve pregnancy rates in poor responders. Currently, there is some evidence to suggest that addition of GH, as well as performing embryo transfer on day 2 versus day 3, appear to improve the probability of pregnancy.

Abstract: This paper analyses the ethical aspects of cross-border reproductive care. Ethical questions are raised by some of the main reasons of cross-border travelling, i.e. law evasion and unequal access to treatment. The phenomenon also generates possible conflicts linked to the responsibility of the professionals. Three points are discussed: the moral obligation of the physician to refer the patient, his/her duty to provide information and counselling and the acceptability of fee-splitting. The recommendations focus on measures to reduce or limit the number of patients that have to travel abroad and on steps to guarantee the safety and quality of the treatment wherever it is provided.

Abstract: OBJECTIVE: Inhibin-B (Inh-B) is produced by Sertoli cells and controls Follicle Stimulating Hormone (FSH) secretion through a negative feedback mechanism. The primary aim of this study was to compare Iotanh-B with FSH as predictors of the recovery of sperm in testicular fine needle aspirate in men with azoospermia. DESIGN: In 51 men with azoospermia basal values of Luteinizing Hormone (LH), FSH, prolactin and testosterone as well as Inh-B values before and 24 h and 48 h after the administration of 300 IU recombinant human FSH were determined. Testicular Fine Needle Aspiration (FNA) was also carried out. Thirty-one young healthy men were also enrolled in the study as controls. RESULTS: There was significant difference between men with azoospermia and controls with regard to the basal Inh-B levels [median (interquartile range) 37.2 (36) vs. 103.0 (90) pg/mL, respectively, p=0.003] but not to the stimulated Inh-B levels [40.5 (41) vs. 73.0 (44) pg/mL, p=0.113 at 24 h and 34.3 (34) vs. 82.0 (50) pg/mL, p=0.098 at 48 h)]. The Area Under Curve in Receiver Operating Characteristic curves were similar for Inh-B and FSH (0.610 vs. 0.716, respectively, p=0.151) as far as prediction of sperm retrieval is concerned. CONCLUSIONS: Basal serum Inh-B values are significantly lower in men with azoospermia compared to controls. However, Inh-B is not superior to FSH in predicting the presence of sperm in testicular fine needle aspirate.

Abstract: Justice and access are among the most urgent questions for medically assisted reproduction. This paper analyses this question not only for people suffering from infertility, but also for people who need assistance to prevent the birth of a child with a specific genetic disorder. Based on the impact of not being able to have a child on the quality of life of a person, the position is defended that infertility treatment should be at least partially reimbursed. Simultaneously, the medical professionals have an obligation towards their patients and the health care system to bring down the costs as far as reasonably possible.

Abstract: BACKGROUND: The purpose of the present systematic review and meta-analysis was to examine whether the probability of pregnancy is increased by adding estrogen to progesterone for luteal phase support in patients treated by in vitro fertilization (IVF). METHODS: A literature search covering MEDLINE, EMBASE, CENTRAL, meeting proceedings and reference lists of published articles was performed to identify relevant RCTs. Data were extracted for meta-analysis yielding pooled relative risks (RR) and 95% confidence intervals (CI). Sensitivity analyses by including studies with pseudo-randomization or unclear method of randomization were also performed (n=1141 patients in total). RESULTS: Four RCTs (n=587 patients) were eligible for inclusion. No statistically significant differences were present between patients who received a combination of progesterone and estrogen for luteal support when compared with those who received only progesterone, in terms of positive hCG rate (RR: 1.02, 95% CI: 0.87-1.19), clinical pregnancy rate (RR: 0.94, 95% CI: 0.78-1.13) and live birth rate (RR: 0.96, 95% CI: 0.77-1.21) per woman randomized. These results did not materially differ in the sensitivity analyses performed. CONCLUSIONS: The currently available evidence suggests that the addition of estrogen to progesterone for luteal phase support does not increase the probability of pregnancy in IVF. However, there is an obvious need for further RCTs that will assess, with more confidence, the effect of estrogen addition to progesterone during the luteal phase on the probability of pregnancy.

Abstract: PURPOSE OF REVIEW: Taking into consideration the increasing interest on hyaluronan and its biological as well as physiological properties, this review will focus on the role of this molecule in human embryo implantation. RECENT FINDINGS: Several studies have been performed up to date in order to assess whether the addition of hyaluronan in the human embryo culture system can improve pregnancy and implantation rates, including one retrospective and six randomized controlled trials. On the one hand, four of those studies showed significant increase in clinical pregnancy and/or implantation rates after using embryo transfer medium containing high concentration of hyaluronan. On the other hand, three studies did not demonstrate any significant improvement in clinical pregnancy and implantation rates. However, regardless of statistical significance, almost all studies demonstrate higher pregnancy and implantation rates after using embryo transfer medium containing high concentration of hyaluronan. SUMMARY: Up to date, the results regarding the role of hyaluronan in human embryo implantation are still conflicting and, thus, further prospective randomized clinical trials are necessary to draw solid conclusions.

Abstract: The present paper focuses on oocyte donation for non-reproductive purposes, i.e. research and future therapy. The general principles of research ethics apply to these interventions. The proportionality principle demands that any possible harms to the oocyte donors should be proportionate to the possible benefits for society. The non-maleficence principle states that every reasonable effort should be made to minimize risks for donors. The position is adopted that, mutatis mutandis, women who donate oocytes for research should be treated similarly to research participants in clinical trials. This implies, among other things, that oocyte donors for research should receive reimbursement for all costs of the procedure and should get compensation for the time lost and inconvenience suffered during the treatment. In order to avoid malpractice and exploitation of poor women, a number of measures are proposed such as a ban on the import of oocytes.

Abstract: Gonadotropin-releasing hormone (GnRH) agonists were introduced in ovarian stimulation for in-vitro fertilization to suppress the premature surge of luteinizing hormone (LH). Although agonist use is accompanied by a series of disadvantages, including hypoestrogenaemia, cyst formation, a requirement for a prolonged period of downregulation, and an increase in follicle-stimulating hormone (FSH) and LH as soon as the agonist is administered, agonists became well accepted in clinical practice as their use was also associated with increased rates of pregnancy. However, the recent development of side-effect-free GnRH antagonists, characterized by an immediate mode of action and a shorter period of administration. Provides clinicians with flexibility in terms of administration, and offers patients a friendlier method of ovarian stimulation. Comparative studies between the two analogues have suggested that the use of antagonists is associated with a shorter duration of FSH stimulation and a decreased incidence of hospital admission due to the occurrence of ovarian hyperstimulation syndrome, while the probability of a live birth does not depend on the type of analogue used.

Abstract: The role of progesterone elevation on in vitro fertilization (IVF) outcome has remained a debatable issue for several years. The aim of this systematic review and meta-analysis was to evaluate whether progesterone elevation on the day of human chorionic gonadotrophin (hCG) administration is associated with the probability of pregnancy. Eligible studies were considered those in which patients did not participate more than once. A literature search in MEDLINE, EMBASE and CENTRAL identified 12 eligible studies, 10 of which were retrospective. The majority (n = 10) of these studies did not detect a statistically significant association between progesterone elevation and the probability of pregnancy. Meta-analysis was performed only for the studies (n = 5) that provided data on clinical pregnancy per patient reaching hCG administration for final oocyte maturation. No statistically significant association between progesterone elevation and the probability of clinical pregnancy was detected (Odds ratio: 0.75, 95% confidence interval 0.53-1.06; P = 0.10). This finding persisted in the sensitivity analyses performed, which excluded the studies that did not report clearly that measurement of progesterone did not affect patients' management and those that did not report definition of clinical pregnancy. In addition, subgroup analyses were conducted on the basis of type of gonadotrophin-releasing hormone GnRH analogue used and on the value of serum threshold used to classify patients in those with or without progesterone elevation. These analyses, however, did not materially change the results obtained. In conclusion, the best available evidence does not support an association between progesterone elevation on the day of hCG administration and the probability of clinical pregnancy in women undergoing ovarian stimulation with GnRH analogues and gonadotrophins for IVF.

Abstract: OBJECTIVE: To examine whether a high concentration of recombinant hyaluronan in the embryo transfer (ET) medium can influence pregnancy rates in humans. DESIGN: Retrospective, matched group study. SETTING: Private and university hospital in vitro fertilization (IVF) units. PATIENT(S): Couples undergoing IVF or intracytoplasmic sperm injection (ICSI) were divided in two groups. In group 1 (402 patients), ET was performed with an ET medium containing a high concentration of recombinant hyaluronan in the presence of recombinant human serum albumin (HSA). In group 2 (425 couples), ET was performed with a medium containing a lower concentration of hyaluronan and HSA. INTERVENTION(S): IVF and ICSI. MAIN OUTCOME MEASURE(S): The two groups were compared for pregnancy rates according to the woman's age, the technique used (IVF or ICSI), the number of embryos transferred, and the day of ET. RESULT(S): Between the two groups, no statistically significant differences were observed in the total pregnancy rates after IVF or ICSI. Similarly, the pregnancy rates did not differ between the two groups when subdivided according to the woman's age, the number of embryos transferred (one to four), or the day of ET (2, 3, or 5 after oocyte pick up). CONCLUSION(S): The use of an ET medium with a high concentration of hyaluronan does not appear to have any statistically significant effect on pregnancy rates.

Abstract: The aim of this systematic review and meta-analysis was to assess whether the addition of recombinant luteinizing hormone (LH) increases live birth rate, among patients treated with follicle stimulating hormone (FSH) and gonadotrophin-releasing hormone (GnRH) analogues for in vitro fertilization (IVF). Eligible studies were randomized controlled trials (RCTs) answering the research question that contained sufficient information to allow ascertainment of whether randomization was true and whether equality was present between the groups compared, regarding baseline demographic characteristics, gonadotrophin stimulation protocol, number of embryos transferred and luteal phase support administered. A literature search identified seven RCTs (701 patients) that provided the information of interest, among which five reported agonist and two antagonist cycles. The reported outcome measure, clinical pregnancy, was converted to live birth using published data in one study. No significant difference in the probability of live birth was present with or without rLH addition to FSH (odds ratio [OR]: 0.92, 95% confidence interval (CI): 0.65-1.31; P = 0.65). This finding remained stable in subgroup analyses that ordered the studies by dose of rLH added, the type of analogue used to inhibit premature LH surge, the time rLH was added during the follicular phase, the age of patients analysed, the presence of allocation concealment and by the way the information on live birth was retrieved. In conclusion, the available evidence does not support the hypothesis that the addition of recombinant LH increases the live birth rate in patients treated with FSH and GnRH analogues for IVF.

Abstract: PURPOSE OF REVIEW: To summarize the available data regarding the value of oral contraceptive pill addition in ovarian stimulation schemes used for in-vitro fertilization. RECENT FINDINGS: In agonists cycles, a decreased incidence of ovarian cyst formation is expected in patients pretreated with the oral contraceptive pill after gonadotropin-releasing hormone agonist administration compared to those treated according to a long follicular protocol. In antagonist cycles, oral contraceptive pill pretreatment appears to be feasible and has been used for programming cycle initiation. Solid evidence regarding its effect on the probability of pregnancy is, however, currently lacking. SUMMARY: The optimal use of oral contraceptive pretreatment as well as its effect on in-vitro fertilization outcome have not yet been fully explored. The effect of oral contraceptive pill pretreatment is worth further investigation in properly designed trials.

Abstract: The analysis of the welfare of the child in the context of medically assisted reproduction is divided in two parts: the first part addresses the risks associated with the would-be parent(s), the second part focuses on possible risks inherent in the technologies and treatments themselves. The risk factors connected with the intended parents may be medical and psychosocial. Two important considerations are raised here: the necessity to avoid discrimination and respect for the privacy of the patients. The second part concerns the ethical questions involved in the rapid dissemination of new technologies in assisted conception. Technology and research must always be subordinate to the welfare of the future offspring. The different steps involved in the responsible application of medical technologies to treat infertility are analysed.

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Abstract: PURPOSE: To explore the possible relationship between sperm quality and embryo development, pregnancy and implantation rates, in patients undergoing intracytoplasmic sperm injection (ICSI). METHODS: Fertilization and cleavage rates, quality of embryos, blastocyst development, pregnancy and implantation rates were analyzed in 1020 embryos from 219 couples undergoing first ICSI treatment cycle. The couples were allocated in five groups, according to semen parameters: Group 1: patients with normal semen parameters, Group 2: patients with mild oligo-astheno-teratozoospermia, Group 3: patients with severe oligo-astheno-teratozoospermia, Group 4: patients with obstructive azoospermia, Group 5: patients with non-obstructive azoospermia. RESULTS: Fertilization and cleavage rates, quality of embryos as well as blastocyst development rates were significantly reduced, as semen quality decreased. However, no significant differences were observed in clinical pregnancy and implantation rates. CONCLUSION: Overall, a negative relationship was observed between semen quality and embryo development, even before activation of the embryonic genome, suggesting that sperm can affect embryogenesis from a very early stage.

Abstract: BACKGROUND: A possible impact of paternal sperm quality on the outcome in children born after assisted reproductive technologies, especially ICSI, has been discussed. The objective of this study was to assess whether sperm concentration has any influence on growth and cognitive development in children born with a gestational age more than 32 weeks after ICSI or IVF. METHODS: Singleton children born after ICSI (n = 492) or IVF (n = 265) from five European countries were examined at age 5 years. The ICSI group was divided into five subgroups according to paternal sperm origin and sperm concentration: (1) epididymal and testicular sperm group, (2) ejaculated sperm < 1 x 10(6)/ml, (3) ejaculated sperm 1-4.99 x 10(6)/ml, (4) ejaculated sperm 5-19.99 x 10(6)/ml and (5) ejaculated sperm > or = 20 x 10(6)/ml. The IVF group was divided into two subgroups: (1) < 20 x 10(6)/ml and (2) > or = 20 x 10(6)/ml. Growth parameters at birth and age 5 were evaluated. Cognitive development was assessed with the Wechsler Preschool and Primary Scale of Intelligence-Revised. RESULTS: No significant difference was found for gestational age, birth weight and birth weight standard deviation scores (SDS) between the ICSI and IVF sperm groups. No significant difference in height and weight at age 5 or SDS weight or height or BMIs at age 5 was found. There was no significant difference in total intelligence quotient (IQ)--performance or verbal IQ--between the groups. CONCLUSION: We found no indication that growth and cognitive development in ICSI and IVF children differed depending on paternal sperm concentration.

Abstract: OBJECTIVE: To report a translocation between an autosome and the Y chromosome. DESIGN: Amniocentesis of a fetus because of mother's advanced age followed by karyotype and PCR analysis. SETTING: Tertiary health center. PATIENT(S): A phenotypically normal twin male infant. INTERVENTION(S): Karyotype with G and Q banding and amplification of testis-specific protein 1-Y and of azoospermia factor (AZF) a, AZFb, AZFc, and distal AZFc regions of Y chromosome. MAIN OUTCOME MEASURE(S): Karyotype, PCR. RESULT(S): We report a phenotypically normal twin male infant with de novo 46,ChiY,t(1;Y)(p22;p11) that was found in amniocentesis. In genetic counseling, it was recommended that the fetus be monitored through a detailed prenatal ultrasonographic examination, which did not indicate any pathological findings. A phenotypically normal male baby was born who is now a 12-month-old healthy infant. The karyotype was confirmed in the peripheral blood with G and Q banding. Amplification of testis-specific protein 1-Y, AZFa, AZFb, AZFc, and distal AZFc regions of the Y chromosome did not reveal any deletions. CONCLUSION(S): We cannot predict whether this male infant will have oligospermia or azoospermia as an adult and, furthermore, whether in case of fertility there is a risk for unbalanced autosome;Y translocations in the offspring, with congenital malformations and dysmorphic features. This case illustrates the complexities in counseling for prenatally diagnosed de novo autosome;Y translocations and the need for additional cases to be reported.

Abstract: The aim of this systematic review was to evaluate, among women with normal ovulation or World Health Organization (WHO) II oligoanovulation who undergo ovarian stimulation for IVF using GnRH analogues, whether endogenous LH levels predict the likelihood of ongoing pregnancy beyond 12 weeks. A literature search identified six studies that answered the research question, among which two were prospective studies (one in GnRH agonist and one in GnRH antagonist cycles). None of the retrospective studies suggest that low endogenous LH levels are associated with a significantly decreased probability of ongoing pregnancy beyond 12 weeks in such patients. In the two prospective studies high endogenous LH levels during down-regulation were associated with a decreased probability of ongoing pregnancy beyond 12 weeks. Until further prospective studies modify the existing evidence summarized here, an adverse effect of low endogenous LH levels on the probability of ongoing pregnancy beyond 12 weeks is not a sensible rationale for LH supplementation during ovarian stimulation for IVF using GnRH analogues.

Abstract: BACKGROUND: The effect of recombinant human LH (r-hLH; lutropin alfa) in women undergoing controlled ovarian stimulation with recombinant human FSH (r-hFSH) prior to IVF was investigated. METHODS: After down-regulation with the GnRH agonist, buserelin, 114 normo-ovulatory women (aged 18-37 years) received r-hFSH alone until the lead follicle reached a diameter of 14 mm. Patients were then randomized in a double-blind fashion to receive r-hFSH in addition to r-hLH, 75 IU s.c., or placebo daily for a maximum of 10 days prior to oocyte retrieval and IVF. The primary end-point was the number of metaphase II oocytes. RESULTS: There were no significant differences between treatment groups for the primary end-point. Serum estradiol concentrations on the day of HCG administration were significantly higher in the group receiving r-hLH plus r-hFSH than in the group receiving r-hFSH alone (P = 0.0001), but there were no significant differences between the groups in dose and duration of r-hFSH treatment required, oocyte maturation, fertilization rate, pregnancy rate and live birth rate. CONCLUSION: In this patient population, the addition of r-hLH during the late follicular phase of a long GnRH agonist and r-hFSH stimulation cycle provides no further benefit in terms of oocyte maturation or other end-points.

Abstract: The present review describes, on the basis of the currently available evidence, the consensus reached by a group of experts on the use of gonadotropin-releasing hormone (GnRH) antagonists in ovarian stimulation for IVF. The single or multiple low-dose administration of GnRH antagonist during the late-follicular phase effectively prevents a premature rise in serum luteinizing hormone (LH) levels in most women. Although controversy remains, most comparative studies suggest a slight, not significant reduction in the probability of pregnancy after IVF using GnRH antagonist versus GnRH agonist co-treatment. Published meta-analyses suggest that this slight difference in pregnancy rates is not attributed to chance. Further studies applying varying treatment regimens and outcome measures are required. Data are not in favour of a need to modify the starting dose of gonadotropins. Data are not in favour of increasing gonadotropin dose at GnRH antagonist initiation. The addition of LH from the initiation of ovarian stimulation or from GnRH antagonist administration does not appear to be necessary. Replacement of human chorionic gonadotropin (HCG) by GnRH agonist for triggering final oocyte maturation is associated with a lower probability of pregnancy. The optimal timing for HCG administration needs to be explored further. GnRH antagonist initiation on day 6 of stimulation appears to be superior to flexible initiation by a follicle of 14-16 mm, although earlier GnRH antagonist administration is worth further evaluation. Luteal phase supplementation in GnRH antagonist protocols remains mandatory in IVF. Effects of GnRH antagonist co-treatment on the incidence of ovarian hyperstimulation syndrome remains uncertain, although a trend is present in favour of the GnRH antagonists. The role of GnRH antagonists in ovarian stimulation for IVF appears to be promising, although many questions regarding preferred dose regimens and effects on clinical outcomes remain.

Abstract: OBJECTIVE: To examine the effects of coping and of infertility-specific and nonspecific stress on pregnancy outcomes after IVF treatment. DESIGN: Cross-sectional design controlling for potential confounding variables. Psychosocial measures were assessed on the day of embryo transfer. SETTING: Fertility clinic in Greece. PATIENT(S): Three hundred forty-two women. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): A clinically recognized pregnancy or a biochemical pregnancy. RESULT(S): Of 342 women participating in the study, 79 became pregnant, representing a success rate of 23.3%. There were no statistically significant differences between women who became pregnant and those who did not in terms of duration of infertility, causes of infertility, previous IVF cycles, and infertility-specific or nonspecific stress. The only factor that was significantly associated with pregnancy outcome was emotionally expressive coping (adjusted odds ratio, 1.272; 95% confidence interval, 1.06-1.52). Women who coped by expressing their emotions were less likely to get pregnant than were women who did not. CONCLUSION(S): The link between psychological processes and reproductive outcomes of IVF treatment is less likely to be identified through the assessment of stress but rather is likely to be identified through the assessment of the strategies that people use to deal with stress. Emotionally expressive coping may be a risk factor for reduced pregnancy rates in women undergoing IVF.

Abstract: This systematic review and meta-analysis aimed to answer the following clinical question: among patients treated for IVF with gonadotrophins and GnRH analogues, is the probability of live birth per randomized patient dependent on the type of analogue used? Eligible studies were randomized controlled trials (RCTs), published as a full manuscript in a peer-reviewed journal, that contained sufficient information to allow ascertainment of whether randomization was true and whether equality was present between the groups compared. A literature search identified 22 RCTs comparing GnRH antagonists and GnRH agonists that involved 3176 subjects. Where live birth was not reported in a study that fulfilled the inclusion criteria, an effort was made to contact the corresponding authors to retrieve the missing information. If this was not possible, the reported outcome measure, clinical pregnancy or ongoing pregnancy was converted to live birth in 12 studies using published data (Arce et al., 2005). No significant difference was present in the probability of live birth between the two GnRH analogues [odds ratio (OR), 0.86; 95% confidence intervals (CI), 0.72 to 1.02]. This result remains stable in subgroup analysis that ordered the studies by type of population studied, gonadotrophin type used for stimulation, type of agonist protocol used, type of agonist used, type of antagonist protocol used, type of antagonist used, presence of allocation concealment, presence of co-intervention and the way the information on live birth was retrieved. In conclusion, the probability of live birth after ovarian stimulation for IVF does not depend on the type of analogue used for pituitary suppression.

Abstract: This article analyses the different ethical aspects of posthumous assisted reproduction. Two situations are distinguished: cases in which the gametes or embryos are used by the surviving partner and cases in which the gametes or embryos are made available for third persons. The moral evaluation of the procedure depends on whether the act is restricted to the existing parental project. A major difficulty for the moral evaluation is the inconclusiveness of the empirical data on the psychosocial development of children born after this procedure. The Task Force concluded that posthumous reproduction by a partner is acceptable if the following conditions are met: written consent has been given by the deceased person, the partner received extensive counselling and a minimum waiting period of 1 year is imposed before a treatment can be started. For use by third parties, the usual conditions for gamete and embryo donation apply.

Abstract: This article is a systematic review of the literature on utilization of gonadotrophin-releasing hormone antagonists (GnRH-ant) for ovarian stimulation for IVF in special patient groups. Summarized by meta-analysis are the data from randomized controlled trials (RCT) in which GnRH-agonist (GnRH-a) and GnRH-ant were compared (eight RCT for poor response, four RCT for PCOS). Also reviewed are the data from two RCT and 13 retrospective or observational trials in which patients at risk of ovarian hyperstimulation syndrome (OHSS) were triggered with GnRH-agonist instead of HCG. For poor responders, no differences in clinical outcomes were found, except a significantly higher number of cumulus-oocyte complexes in GnRH-antagonist multiple dose protocol as compared to GnRH-agonist long protocol (P=0.05). For PCOS patients, no differences in outcomes were found, except a significantly shorter duration of stimulation, when GnRH-antagonist multiple dose protocol and GnRH-agonist long protocol are compared (P<0.01). However, sample sizes are still small and power to detect subtle differences is therefore limited. For OHSS risk patients triggered with GnRH-agonist, reports on the efficacy of this measure vary in the literature. GnRH-agonist triggering appears to be associated with a reduction in the incidence of mild and moderate OHSS. For prevention of severe OHSS, as yet, only very limited evidence is available.

Abstract: OBJECTIVE: To date, very few studies have been conducted on the neurodevelopmental well-being of children conceived through intracytoplasmic sperm injection (ICSI). The limitations of these studies often include a lack of comparison with a demographically matched, naturally conceived (NC) group and the investigation of only very young children, with relatively small samples sizes. One study showed that there were no differences in IQ scores among ICSI-conceived, in vitro fertilization (IVF)-conceived, and NC children at 5 years of age. Unfortunately, psychomotor development was not assessed in that study. Because findings regarding these children's cognitive and motor development are inconclusive, the aim of this study was to shed more light on the cognitive and motor development of 5-year-old ICSI-conceived children. METHODS: A total of 511 ICSI-conceived children were compared with 424 IVF-conceived children and 488 NC controls. Children were recruited in 5 European countries, ie, Belgium, Denmark, Greece, Sweden, and the United Kingdom. Participation rates ranged from 45% to 96% in the ICSI and IVF groups and from 34% to 78% in the NC group. Cognitive and motor development was assessed with the Wechsler Preschool and Primary Scale of Intelligence-Revised (WPPSI-R) and McCarthy Scales of Children's Abilities (MSCA) Motor Scale, respectively. The WPPSI-R consists of 2 major scales, ie, Verbal and Performance, each including 6 subtests. The 6 Performance Scale subtests are object assembly, geometric design, block design, mazes, picture completion, and animal pegs. The 6 Verbal Scale subtests are information, comprehension, arithmetic, vocabulary, similarities, and sentences. Scores on the Performance and Verbal Scale subtests are summed to yield the performance IQ (PIQ) and verbal IQ (VIQ), respectively. Scores on both the Performance Scale and the Verbal Scale yield the full-scale IQ (FSIQ). IQ scales have a mean score of 100 and a SD of 15. Each subtest has a mean score of 10 and a SD of 3. The MSCA consists of 6 scales, ie, Verbal, Perceptual-Performance, Quantitative, General Cognitive, Memory, and Motor Scale. In this study, only the Motor Scale was administered. This scale assesses the child's coordination during performance of a variety of gross- and fine-motor tasks. Leg coordination, arm coordination, and imitative action tests provide measures of gross-motor ability. Draw-a-design and draw-a-child assess fine-motor coordination, as revealed by the levels of hand coordination and finger dexterity. The mean score for this test is 50, with a SD of . RESULTS: No differences were identified among ICSI, IVF, and NC children with respect to VIQ, PIQ, or FSIQ scores of the WPPSI-R. Furthermore, there were no differences between groups regarding the discrepancy between VIQ and PIQ scores. These results were not influenced by gender, country, or maternal educational level. However, in the subgroup of firstborn children with mothers who gave birth at an older age (33-45 years), NC children obtained significantly better VIQ and FSIQ scores than did children conceived through assisted reproductive technologies. These differences in VIQ and FSIQ scores between ICSI/IVF and NC children were relative, because NC children scored <1 IQ point higher than ICSI/IVF children. Therefore, these scores show no clinical relevance. For Verbal Scale subtests, variables such as age of the mother at the time of the birth, educational level of the mother, and gender and nationality of the child interacted with mode of conception, resulting in clinically irrelevant differences between scores for the ICSI/IVF and NC groups on the arithmetic, vocabulary, and comprehension subtests. For Performance Scale subtests, these same demographic factors interacted with mode of conception for the block design, object assembly, and animal pegs subtests, again resulting in clinically irrelevant differences among groups. In the 3 groups (ICSI, IVF, and NC), we observed equal numbers of children scoring below 1 SD from the mean on the WPPSI-R and the MSCA. CONCLUSIONS: This study includes a substantial number of children from several European countries. Apart from a few interaction effects between mode of conception and demographic variables, no differences were found when ICSI, IVF, and NC scores on the WPPSI-R and MSCA Motor Scale were compared. Nevertheless, the aforementioned interaction effects could indicate that demographic variables such as maternal age at the time of the birth and maternal educational level play different roles in the cognitive development of IVF and ICSI children, compared with NC children. Additional research is needed to explore and verify this finding. Previous studies revealed that ICSI children, in comparison with NC children, more frequently obtained scores below 1 SD from the mean on 3 subtests of the Performance Scale (object assembly, block design, and mazes) or showed a trend of 5.2% of ICSI children, compared with 2.5% of IVF children and 0.9% of NC children, obtaining a score below 1 SD from the mean, but those findings were not confirmed in this study. Here no differences were found among the 3 groups in the numbers of children scoring below 1 SD from the mean on the VIQ, PIQ, and FSIQ tests and the Verbal and Performance Scale subtests. Motor development results were somewhat more conclusive. There were no differences between the scores of ICSI, IVF, and NC children on the MCSA Motor Scale. No interaction effects were found between mode of conception and demographic variables, indicating that these results are not influenced by gender, nationality, maternal educational level, or maternal age at the time of the birth. Furthermore, equal proportions of children in all 3 groups scored below 1 SD from the mean. The results of this study are reassuring for parents who conceived through ICSI (or IVF). The findings indicate that the motor and cognitive development of their offspring is very similar to that of NC children. However, demographic factors such as maternal educational level and maternal age at the time of the birth might play different roles in the cognitive development of ICSI and IVF children, compared with NC children.

Abstract: The late 20th century trend to delay birth of the first child until the age at which female fecundity or reproductive capacity is lower has increased the incidence of age-related infertility. The trend and its consequences have also stimulated interest in the possible factors in the female and the male that may contribute to the decline in fecundity with age; in the means that exist to predict fecundity; and in the consequences for pregnancy and childbirth. In the female, the number of oocytes decreases with age until the menopause. Oocyte quality also diminishes, due in part to increased aneuploidy because of factors such as changes in spindle integrity. Although older male age affects the likelihood of conception, abnormalities in sperm chromosomes and in some components of the semen analysis are less important than the frequency of intercourse. Age is as accurate as any other predictor of conception with assisted reproductive technology. The decline in fecundity becomes clinically relevant when women reach their mid-30s, when even assisted reproduction treatment cannot compensate for the decline in fecundity associated with delaying attempts at conceiving. Pregnancies among women aged >40 years are associated with more non-severe complications, more premature births, more congenital malformations and more interventions at birth.

Abstract: This 9th statement of the ESHRE Taskforce on Ethics and Law considers ethical questions and specific dilemmas concerning preimplantation genetic diagnosis for human leukocyte antigen typing of embryos. This application is particularly complex because the interests of the sick child needing a transplantation should be balanced against the interests of the future donor child who may result from the technique. It is concluded that, if parents intend to love the child, the creation and use as a donor is not inherently disrespectful.

Abstract: We conducted a retrospective, cross-sectional study to evaluate the efficacy, safety, and effect of first-trimester multifetal pregnancy reduction on procedure-related complications and obstetrical outcome in multiple pregnancies with a monochorionic component. Although procedure-related complications were relatively common, the obstetrical outcome was favorable in most cases when the monochorionic twin component was reduced.

Abstract: PURPOSE: The understanding of the public's knowledge on human cloning (HC) and its acceptability are considered important for the development of evidence-based policy making. The aim of this research study was to investigate the demographic and socioeconomic variables that affect the public's knowledge and intention to use HC in urban areas of Greece. Additionally, the possible association of religiousness with the knowledge and the intention to use HC were also investigated. METHODS: Individual interviews were conducted with 1020 men and women of urban areas in Greece. Stratified random sampling was performed to select the respondents. Several scientists, experts in HC, evaluated the content of the instrument initially developed. The final questionnaire was consequently the result of a pilot study. RESULTS: Almost half of the respondents (51.5%) believed that "HC is a sort of in vitro fertilization" and 42.9% that "it has already been applied to human being." They were not aware that "the cloned fetus grows in the woman's uterus" (41.5%) and that "HC could regenerate human organs" (41.7%). The acceptability of human cloning for the cure of terminal diseases and transplantation need is very high (70.7% and 58.6%, respectively). The public's intention to have recourse to cloning on the grounds of "bringing" back to life a loved person or because of reproductive disorders was reported as desire by 35% and 32.5%, respectively. The occupational category (scientists, self-employed, and artists), the Intention to use HC, and the number of children are highly significant predictors of valid knowledge about HC. Low rates of church attendance appeared to relate with high reported Intention to use HC, and increasing scores of valid knowledge about HC increased the public's Intention to use HC. CONCLUSIONS: A number of specific demographic and socioeconomic characteristics and high scores of knowledge provide a persuasive justification in demonstrating intention toward HC. The current study suggests that these findings should receive further attention by policymakers and scientists within the Greek context.

Abstract: BACKGROUND: Over a million children have been born from assisted conception worldwide. Newer techniques being introduced appear less and less 'natural', such as intracytoplasmic sperm injection (ICSI), but there is little information on these children beyond the neonatal period. METHODS: 540 ICSI conceived 5-year-old children from five European countries were comprehensively assessed, along with 538 matched naturally conceived children and 437 children conceived with standard IVF. RESULTS: Of the 540 ICSI children examined, 63 (4.2%) had experienced a major congenital malformation. Compared with naturally conceived children, the odds of a major malformation were 2.77 (95% CI 1.41-5.46) for ICSI children and 1.80 (95% CI 0.85-3.81) for IVF children; these estimates were little affected by adjustment for socio-demographic factors. The higher rate observed in the ICSI group was due partially to an excess of malformations in the (boys') urogenital system. In addition, ICSI and IVF children were more likely than naturally conceived children to have had a significant childhood illness, to have had a surgical operation, to require medical therapy and to be admitted to hospital. A detailed physical examination revealed no further substantial differences between the groups, however. CONCLUSIONS: Singleton ICSI and IVF 5-year-olds are more likely to need health care resources than naturally conceived children. Assessment of singleton ICSI and IVF children at 5 years of age was generally reassuring, however, we found that ICSI children presented with more major congenital malformations and both ICSI and IVF children were more likely to need health care resources than naturally conceived children. Ongoing monitoring of these children is therefore required.

Abstract: The present review summarizes existing knowledge on the use of gonadotropin releasing hormone (GnRH) antagonists based on experience gathered after the completion of phase III comparative trials with GnRH agonists. Available data suggest that prolongation of the follicular phase significantly decreases the probability of pregnancy. Moreover, patients with elevated progesterone at initiation of stimulation have significantly fewer chances of achieving an ongoing pregnancy. Luteal support remains mandatory, while the replacement of human chorionic gonadotrophin by GnRH agonist does not appear to be feasible. Although not conclusive, existing data are not in favour of increasing the starting dose of gonadotrophins, of LH supplementation or of using a flexible antagonist protocol.

Abstract: OBJECTIVE: To assess laterality in a group of European children conceived with the help of assisted reproductive therapy, or naturally conceived. DESIGN: Prospective cohort study. SETTING: Five European countries. POPULATION: A total of 1525 five-year-old children divided into three groups according to conception status: naturally conceived; intracytoplasmic sperm injection (ICSI); in vitro fertilisation (IVF) group. METHODS: All children were assessed using the McCarthy Scale of Children's Abilities (Motor Scale) with additional items using a comb, a spoon and an indirect measure was made of the parental handedness scores, using a questionnaire. MAIN OUTCOME MEASURES: Child handedness for drawing and writing. RESULTS: Observed handedness did not differ significantly between natural conception, IVF and ICSI in girls or boys, except for ICSI-conceived girls, fewer of whom were left-handed than the naturally conceived controls, 7.0%versus 12.4% (P < 0.05), however, this was close to the population norm of 8%. The degree of parental handedness did not significantly vary in the three groups. CONCLUSIONS: Laterality, as assessed by handedness, may be marginally less common in girls conceived with assisted reproductive therapies than with children conceived naturally.

Abstract: A case of a nontuplet pregnancy that was successfully reduced to twins resulting in a good neonatal outcome is described. A 37-year-old woman achieved a nontuplet pregnancy after ovarian stimulation with GnRH-analogues and gonadotrophins in the short protocol and artificial insemination. Reduction of the nontuplet pregnancy to twins was successfully performed in three attempts and delivery by caesarian section was followed on the 32nd week of pregnancy. Two alive and healthy neonates with birth weights 1235 g and 1515 g were born. Multifetal pregnancy reduction is medically justified in the very high order pregnancies. The first approach, though, to the problem of multiple gestation should be prevention through carefully monitored infertility treatment, as the elective embryo reduction procedure carries with it medical, ethical and psychological issues for both the parents and the physicians involved.

Abstract: This 10th statement of the Task Force on Ethics and Law considers ethical questions specific to varied surrogacy arrangements. Surrogacy is especially complex as the interests of the intended parents, the surrogate, and the future child may differ. It is concluded that surrogacy is an acceptable method of assisted reproductive technology of the last resort for specific medical indications, for which only reimbursement of reasonable expenses is allowed.

Abstract: BACKGROUND: In the era of human genome research, there is a large theoretical debate among scientists and authorities on the ethical dimension based on the moral liberty of the individuals and the scientific and economic dimension based on the freedom and the independence of the scientific and technological activities. Meanwhile, the understanding of beliefs on human cloning (HC) and its acceptability are important for the development of evidence-based policy making. However, previous research in the field of public beliefs towards human genetics is limited. Therefore, in the present study, we investigated the role of public beliefs as predictors of the acceptability of the respondents to use HC. METHODS: Personal interviews were conducted with 1020 men and women of urban areas in Greece. Stratified random sampling was performed to select participants. Several scientists, experts in HC, evaluated the content of the instrument initially developed. The final questionnaire was the result of a pilot study. RESULTS: The acceptability of HC for the cure of incurable diseases and transplantation need is very high (70.7 and 58.6%, respectively). Public's intention to have recourse to HC because of "bringing" back to life a loved person or because of reproductive disorders was reported by 35 and 32.5%, respectively. With respect to the role of beliefs: increasing scores of reasons of social benefits, moral/religious reasons and legislative reasons increased the public's intention to have recourse to HC; inversely, decreasing scores of reasons of human commodification/exploitation increased public's intention to have recourse to HC. Additionally, low rates of church attendance appeared to be correlated with high reported acceptability of HC. CONCLUSION: There is great public concern regarding the application of HC, which probably reflects the existing ambivalence over the relationship between technology and society. Scientists and policymakers should take into account these indicators of public disquiet and should manage the public involvement in policy decisions, from which they have so far been excluded.

Abstract: Cervical agenesis or dysgenesis (fragmentation, fibrous cord and obstruction) is an extremely rare congenital anomaly. Conservative surgical approach to these patients involves uterovaginal anastomosis, cervical canalization, and cervical reconstruction. In failed conservative surgery, total hysterectomy is the treatment of choice. We report what we believe to be the first successful end-to-end cervico-cervical anastomosis of an unusual case of congenital cervical fragmentation. A 15 year old Caucasian female presented complaining of primary amenorrhoea and cyclic, monthly abdominal pain. At laparotomy, a symmetrical transverse disruption of the cervix in a central and distal part was found and an end-to-end cervico-cervical anastomosis of the two cervical fragments was performed with the use of a 16F Foley catheter as a stent into the endocervical canal. One month after surgery the patient had normal menses. We conclude that a thorough investigation of the patient with suspected cervical anomaly is necessary and conservative surgical treatment should be applied as a first-line treatment option. In the presence of functional and intact cervical segments, the reconstruction of the cervical canal with an end-to-end cervico-cervical anastomosis is feasible and effective.

Abstract: The widespread application of protocols using gonadotropin-releasing hormone (GnRH) agonists or antagonists in assisted reproduction treatment has led to an increasing number of pregnancies exposed to these drugs. This issue has raised scepticism as to the safety of these medications, concerning both pregnant women and their offspring. The main parameters that can be studied to ensure the safety of GnRH analogues include: a) systemic and local reactions to the medication; b) incidence of ovarian hyperstimulation syndrome (OHSS); c) direct effect on oocytes and embryos; and d) the health of those children exposed. So far, no systemic side effects and no major local reactions have been reported following the use of GnRH agonists or third-generation antagonists. On the other hand, the incidence of OHSS seems to be higher with GnRH agonist protocols compared to conventional or GnRH antagonist protocols. The recent cloning of the GnRH receptor has led to the demonstration of GnRH receptor gene expression in the human ovary, although the existence of GnRH receptors per se remains controversial. Similarly, the potential direct effect of GnRH analogues on the follicles and oocytes remains a matter of debate. The incidence of miscarriage and the health of children born as a result of in vitro fertilisation (IVF) treatment do not appear to be influenced by the GnRH agonist treatment. This also seems to be the case for the GnRH antagonists, although the available information on this issue is still limited. Therefore, most of the accumulated data concerning the safety of the GnRH analogues are encouraging, and no serious side effects have been reported. On the other hand, as no definite conclusions about the safety of these drugs can be drawn until now, continued assessment of the aforementioned parameters in long-term follow-up studies is recommended.

Abstract: BACKGROUND: ICSI is used with increasing frequency, but there is less information about the children born following this method of assisted reproduction than other forms of IVF. Some authors have suggested that it may contribute to more family stress than IVF. METHODS: ICSI conceived children were compared with IVF conceived children and naturally conceived (NC) controls. They were selected in five European countries: Belgium, Denmark, Greece, Sweden and the UK, and seen for psychological testing and a paediatric examination when they were 5 years old. In all countries, except Greece, mothers and fathers were asked to complete questionnaires about parental well-being, family relationships, parenting and child behaviour. RESULTS: Very few differences were found between the ICSI and NC group or the ICSI and IVF group. The only significant differences were that mothers in the ICSI conceived group reported fewer hostile or aggressive feelings towards the child and higher levels of commitment to parenting than the mothers of NC children. CONCLUSIONS: The study confirms the results of previous work with IVF families. This should be encouraging for families using these techniques in the future.

Abstract: In this 8th statement of the ESHRE Taskforce on Ethics and Law, the focus is on the ethical issues raised by (assisted) reproduction for HIV positive men and women. Due to treatment improvements, their life expectancy has increased substantially. This, in combination with the reduction of perinatal transmission to the child, makes the creation of a family more acceptable. Recommendations are made concerning methods of avoiding HIV transmission in the couple and to their offspring. It is concluded that, if certain precautions are taken, medical assistance to reproduction of HIV positive people is ethically acceptable. For the time being, only cases of serodiscordant couples should be considered.

Abstract: Idiopathic non-obstructive azoospermia or severe oligozoospermia (INOA) consists a special group of men characterized by eugonadism, primary infertility, low testicular volume, azoospermia or severe oligozoospermia and high follicle stimulating hormone values. Aims of this study were to describe the clinical, hormonal, sperm and histological characteristics of men with INOA and to define if and to what extend men with the INOA phenotype carry Y chromosome long arm (Yq) microdeletions. Sixty-three men with INOA were studied through clinical examination, spermiograms, hormonal profile, polymerase chain reaction for Yq microdeletions, karyotype and testicular fine-needle aspiration biopsy. Sixty-seven men with infertility of known causes and sixty fertile men served as controls. Men with INOA had significantly lower total testosterone levels than fertile men as well as higher prevalence of loss of libido, higher luteinizing hormone levels and lower sperm volume whereas men with infertility of known causes had intermediate values. The prevalence of Yq microdeletions was 3% in men with INOA, 3% in men with infertility of known causes (all of them with abnormal karyotype) and 0% in fertile men. In conclusion, men with INOA have more severe testiculopathy than these with infertility of known causes. These men may be at increased risk of developing partial androgen deficiency of the aging male.

Abstract: Interstitial pregnancy is a rare but life-threatening condition. A case of a 28-year-old woman with a partially ruptured interstitial pregnancy treated with operative laparoscopy is presented. A laparoscopic cornual resection and a left salpingectomy were performed uneventfully. Serum beta-human chorionic gonadotrophin concentrations were measured serially at weekly intervals until resolved on day 20 postoperatively. It seems, therefore, that laparoscopic treatment is still an effective option for management even in ruptured interstitial pregnancy, preserving the anatomical integrity of the uterus and future fertility, and that rupture of interstitial ectopic pregnancy is not a contra-indication for laparoscopy.

Abstract: The European Society of Human Reproduction and Embryology (ESHRE) Ethics Task Force sets out a recommended multidisciplinary approach to the application of preimplantation genetic diagnosis (PGD). The statement includes consideration of fundamental ethical principles, specific problems in cases of high genetic risk, and PGD for aneuploidy screening, HLA typing and sex selection for non-medical reasons.

Abstract: Poor response is not a rare occurrence in ovarian stimulation. Although not fully accepted, the most dominant criteria for poor ovarian response are small numbers of follicles developed or oocytes retrieved, and low estradiol (E2) levels after the use of a standard stimulation protocol. There is no ideal predictive test as the poor responder is revealed only during ovulation induction; however, increased levels of day 3 FSH and E2 as well as decreased levels of inhibin B can be used to assess ovarian reserve. Several protocols have been proposed for clinical management of low ovarian response in IVF. Although high doses of gonadotrophins have been used by the vast majority of authors, results have been controversial and prospective randomized studies have shown little or no benefit. The few available relevant studies do not indicate that recombinant FSH improves outcome. Flare-up GnRH agonist protocols (including all dosage varieties) produce better results than standard long luteal protocols. Luteal initiation GnRH agonist 'stop' protocols were shown to improve ovarian response according to prospective studies with historical controls, but this was not confirmed by well-designed prospective, randomized, controlled studies. The few available data obtained with GnRH antagonists have not shown any benefits. Adjuvant therapy with growth hormone (GH) or GH-releasing factors results in no significant improvement. The use of corticosteroids reduces the incidence of poor ovarian response in women undergoing IVF treatment. The limited data obtained with nitric oxide donors are encouraging. Pretreatment with combined oral contraceptives prior to stimulation may help ovarian response. No benefit was observed with standard use of ICSI or assisted hatching of zona pellucida. Finally, natural cycle IVF has produced results which are comparable with those obtained with stimulated cycles in true poor responders. Well-designed, large-scale, randomized, controlled trials are needed to assess the efficacy of these different management strategies.

Abstract: Fertility, defined as the ability to achieve a pregnancy, declines gradually over the woman's lifespan. Although this decline seems to begin from the age of 30 years, it is more obvious between 35 and 40 and increases dramatically thereafter. The age of 41 is considered to be the point when fertility stops and sterility starts. The actual menopause occurs approximately 10 years after the substantial loss of conception potential. Thus, the biological rather than the chronological age of the woman can predict more accurately her fertility potential. This decline in female reproductive potential correlates with ovarian factors, although a slight contribution from the uterus itself and from the neuroendocrine axis cannot be excluded. The ovarian reserve decreases with advancing age, while a parallel decrease in the quality of the oocytes is present, as indicated by the increased incidence of oocyte aneuploidy. The endocrine function of the ovary also declines with age, the later becoming unable to sustain its normal function in the neuroendocrine axis. Additionally, the role of the various endometrial factors remains controversial. On the other hand, exposure to toxic factors and the increased prevalence of infertility-related diseases like endometriosis and PID, may also contribute. Spontaneous conception rates are minimal in perimenopausal women, mainly due to a qualitative and quantitative loss of female gametes. In the rare case of spontaneous conception achievement, complications are more likely. The application of classic ovulation induction and IVF may serve some selected cases, where the woman's ovarian biological age does not correspond to her chronological one. However, the implantation, clinical pregnancy, and live birth rates in women of advanced age undergoing IVF treatment, show very poor results. On the other hand, preimplantation genetic diagnosis is an accurate diagnostic tool for exclusion of genetically deficient embryos prior to embryo transfer. Oocyte donation seems to be the most reliable option of the perimenopausal woman, since the cumulative birth rates after four treatment cycles is approximately 80%. Cryopreservation of ovarian tissue may be an alternative in nulliparas women <40 years of age who want to have children in the future or women with the same desire who, unfortunately, have had pelvic radiotherapy, chemotherapy, oophorectomy, or premature menopause. This technique has given encouraging results in animals, but has not achieved pregnancies in humans. In the future, the use of drugs to block oocyte depletion as well as recent techniques, such as cytoplasmic or germinal vesicle transfer, will be more widely tested and may offer an option to the perimenopausal woman who wishes to conceive.

Abstract: PURPOSE OF REVIEW: To provide the clinician with updated knowledge of the most recent findings on the clinical use of gonadotropin-releasing hormone antagonists. RECENT FINDINGS: Gonadotropin-releasing hormone antagonists, which have recently been introduced in clinical practice, cause an immediate suppression of gonadotropin secretion by competitive blocking of pituitary gonadotropin-releasing hormone receptors. Thus, they are effective in preventing the premature luteinizing hormone surges during ovarian stimulation for in-vitro fertilization and may improve the patient's response to lower doses of gonadotropins. Better patient acceptance, shorter treatment cycles and fewer follicles and oocytes are also reported. Data existing so far concerning the necessity of luteal phase support after the use of gonadotropin-releasing hormone antagonists show that it might not be mandatory when used in clomiphene citrate costimulated cycles or in intrauterine insemination cycles. The use of gonadotropin-releasing hormone antagonists seems to be safe for pregnant women and their offspring. All sex-hormone-dependent disorders, currently treated with gonadotropin-releasing hormone agonists, may in future be indications for a gonadotropin-releasing hormone antagonist, including endometriosis, leiomyoma, and breast cancer in women, benign prostatic hypertrophy and prostatic carcinoma in men, and central precocious puberty in children. The vast majority of the available clinical data up till now, however, are in assisted reproduction and prostate cancer. SUMMARY: It is expected that the availability of gonadotropin-releasing hormone antagonist will lead to the use of 'softer' ovarian stimulation protocols, which will be shorter, cheaper and safer compared with the conventional protocols.

Abstract:

Abstract: This study was performed to examine the contribution of genetic polymorphism of oestrogen and androgen receptor (AR) genes in male infertility. We have studied in total 173 Greek men, 109 infertile patients and 64 controls (group A). Patients were divided in to three subgroups: group B (n=29) with idiopathic moderate oligospermia, group C (n=42) with azoospermia or idiopathic severe oligospermia and group D (n=38) with azoospermia or oligospermia of various known aetiologies. All patients and controls were genotyped for two polymorphisms of the oestrogen receptor alpha (ERalpha) gene and also for the (CAG)n repeat length polymorphism of the X-linked androgen receptor (AR)gene. The control group had statistically significant difference from group C regarding the XbaI polymorphism of ERalpha gene. Despite the fact that we did not observe any statistically significant differences in the mean and range of the CAG repeat number, the frequency of the higher repeats of the nucleotide repeat sequence (CAG)n of the AR gene was 2-4 times higher in groups B and C compared with the control group A. Our results indicate that both ERalpha and AR gene play significant role in male fertility. It is possible that a synergy may exist between unfavourable genotypes of these two genes in male infertility.

Abstract: The wide application of gonadotrophin-releasing hormone (GnRH) analogues in assisted reproduction has raised concerns about their potential extra-pituitary actions. Although data from animal studies support the presence of ovarian GnRH receptors and of direct ovarian effects of GnRH analogues, this has not been unequivocally demonstrated in humans. Available evidence suggests, however, the existence of GnRH receptors in human granulosa-luteal cells, while the majority of the studies performed showed that progesterone is inhibited by GnRH agonists in vitro. On the other hand, no difference seems to exist between agonists and antagonists in terms of granulosa-luteal cell steroidogenesis under basal conditions. So far, no studies have suggested an adverse effect of GnRH analogues on the developing human follicle. Moreover, the physiological significance of a direct in-vitro ovarian effect of GnRH analogues remains to be established.

Abstract: beta-thalassemia (beta-thal) is characterized by disturbances of the reproductive system. The aim of the present study was: 1) to assess the hypothalamic- pituitary-gonadal axis in patients with beta-thal in relation to their phenotype and 2) to determine prognostic features of current gonadal status. We studied 135 patients (67 males and 68 females) with beta-thal through history, physical examination, spermiograms and GnRH test. These patients were divided into beta-thal major (51 males and 62 females) and beta-thal intermedia phenotypes (16 males and 6 females). Male patients with beta-thal major were subdivided into three groups a) eugonadal (35%, Tanner's stage V, normal testicular volume, normal spermiograms, normal basal and stimulated hormone values), b) patients with hypogonadotrophic hypogonadism (HH) of late onset (24%, Tanner's stage II-V, low-normal testicular volume, abnormal spermiograms, normal basal gonadotrophin values and abnormal response to GnRH test) and c) patients with HH of early onset (41%, Tanner's stage I, small testicular volume, abnormal spermiograms, abnormal basal and stimulated hormone values). Female patients with beta-thal major were subdivided into: a) eugonadal (32%, Tanner's stage V, regular menstruation, normal basal and stimulated hormone values), b) patients with hypogonadotrophic hypogonadism (HH) of late onset (34%, Tanner's stage II-V, secondary amenorrhea, subnormal basal and stimulated gonadotrophin values) and c) patients with HH of early onset (34%, Tanner's stage I, primary amenorrhea, subnormal basal and stimulated hormone values). Patients with beta-thal intermedia were subdivided into eugonadal (75% of males, 33% of females) and hypogonadal (25% of males, 67% of females). Current gonadal status could not be predicted by means of transfusion or chelation parameters. In conclusion, beta-thal patients could be eugonadal or develop early or late onset HH. trade mark-thal intermedia patients have a more favorable profile than beta-thal major individuals. Current gonadal status of beta-thal patients cannot be predicted by means of history, clinical or laboratory parameters.

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Abstract: Uterine malformations consist of a group of miscellaneous congenital anomalies of the female genital system. Their mean prevalence in the general population and in the population of fertile women is approximately 4.3%, in infertile patients approximately 3.5% and in patients with recurrent pregnancy losses approximately 13%. Septate uterus is the commonest uterine anomaly with a mean incidence of approximately 35% followed by bicornuate uterus (approximately 25%) and arcuate uterus (approximately 20%). It seems that malformed uterus and especially septate uterus is not an infertility factor in itself. However, it may have a part in the delayed natural conception of women with mainly secondary infertility. On the other hand, patients with uterine malformations seem to have an impaired pregnancy outcome even as early as their first pregnancy. Overall term delivery rates in patients with untreated uterine malformations are only approximately 50% and obstetric complications are more frequent. Unicornuate and didelphys uterus have term delivery rates of approximately 45%, and the pregnancy outcome of patients with untreated bicornuate and septate uterus is also poor with term delivery rates of only approximately 40%. Arcuate uterus is associated with a slightly better but still impaired pregnancy outcome with term delivery rates of approximately 65%. Women who have undergone hysteroscopic septum resection and have been reported in the different series comprise a highly selected group of symptomatic patients with term delivery and live birth rates of only approximately 5%. Hysteroscopic treatment seems to restore an almost normal prognosis for the outcome of their pregnancies with term delivery rates of approximately 75% and live birth rates of approximately 85%. It seems, therefore, that hysteroscopic septum resection can be applied as a therapeutic procedure in cases of symptomatic patients but also as a prophylactic procedure in asymptomatic patients in order to improve their chances for a successful delivery.

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Abstract: OBJECTIVE: To present a case of Marfan syndrome and concurrent congenital obstructive azoospermia. DESIGN: Case report. SETTING: Tertiary-care academic hospital. PATIENT(S): A man with Marfan syndrome and obstructive azoospermia. INTERVENTION(S): The patient was evaluated for azoospermia that was proved to be due to congenital absence of large segments of vas deferens and epididymis bilaterally. MAIN OUTCOME MEASURE(S): Evaluation for testicular sperm extraction and ICSI procedure. RESULT(S): The patient is a candidate for testicular sperm extraction and ICSI. CONCLUSION(S): The Marfan syndrome and azoospermia may be associated in sporadic cases.

Abstract: OBJECTIVE:To describe clinical and laboratory features of a patient with 45,X/46,XY mosaic karyotype and Y chromosome microdeletions and to discuss the diagnostic problems in his management. DESIGN:Case report. SETTING:University department. PATIENT(S):A 17-year-old man with ambiguous genitalia, 45,X/46,XY mosaic karyotype, and Y chromosome microdeletions. INTERVENTION(S):Testicular ultrasonography, karyotype, open testicular biopsy, polymerase chain reaction (PCR) screening for cystic fibrosis, PCR screening for Y chromosome microdeletions in peripheral blood and testicular tissue, and reverse transcriptase PCR in testicular tissue for Y chromosome microdeletions. MAIN OUTCOME MEASURE(S):Avoidance of dissemination of testicular cancer. RESULT(S):The patient was referred for bilateral orchiectomy. CONCLUSION(S):45,X/46,XY mosaic karyotype is associated with a broad spectrum of phenotypes that includes female with Turner syndrome, male with mixed gonadal dysgenesis, male pseudohermaphroditism, and apparently normal male. Microdeletions of the long arm of the Y chromosome may be associated with Y chromosomal instability, leading to formation of 45,X cell lines. 45,X/46,XY males carry an increased risk for gonadal tumors and must be followed closely.

Abstract: Oocyte donation is an effective treatment modality for women lacking functioning ovaries, but also for women in whom repetitive in-vitro fertilization (IVF) cycles did not result in the development of adequate number of oocytes as well as for those at risk of transmitting genetic diseases. In women with ovarian failure, artificial menstrual cycles are required in order to produce endometrial growth and differentiation similar to that in women with normal ovarian function. Synchronization of donor's and recipient's cycles is mandatory, since the window of implantation is rather limited. The excellent results of oocyte donation treatment confirm that this assisted reproduction technique can provide a novel approach for the treatment of infertility in these groups of patients. Nevertheless, pregnancies in women of advanced reproductive age are associated with significantly more obstetrical complications and higher perinatal morbidity and mortality rates. Furthermore, aging parents have considerably higher chances to develop serious or life-threatening diseases. Thus, careful medical screening and extensive counselling is mandatory, taking into account the psychosocial ramifications of the procedure and, especially, the best interest of the child-to-be.

Abstract: Lymphocyte cultures from patients who had previously undergone at least three unsuccessful procedures of assisted reproduction were analysed for cytogenetic abnormalities. A total of 12,657 metaphases from 33 individuals (15 patients and 18 healthy controls with two normal offsprings) were studied. A significantly higher incidence (P<0.001) of chromosome aberrations was found in the patients (6.79+/-0.68%; x+/-SD) as compared to the controls (1.72+/-0.3%; P<0.001). Chromosomal breakages, particularly at the centromere region, were also observed with significantly increased frequency in the patients than in the controls (6.18+/-0.65 vs. 1.42+/-0.27%, respectively; P<0.001). It is possible that the high rate of centromere breakages in the ART patients (3.18+/-0.47 vs. 0.26+/-0.12%, P<0.001) may predispose to meiotic chromosomal abnormalities. A single cell aberration was demonstrated in 0.61+/-0.21% of ART patients' lymphocytes versus 0.3+/-0.12% in the controls (P<0.01). Structural rearrangements and chromosomal breaks predominantly affected the bands containing genes for the immune response and the cell cycle. Mosaic karyotypes were found in six patients. One of them had a karyotype 46,XX/46,XX,r(14) and the others had sex chromosomal mosaicisms. The prenatal diagnosis could be essential in these cases. It is concluded that chromosomal aberrations could play a role in the repeated failure of ART procedures.

Abstract: The widespread application of intracytoplasmic sperm injection (ICSI) has raised concern about the efficacy and safety of this novel technique. The European Society of Human Reproduction and Embryology (ESHRE) has established an ICSI Task Force to collect annually the clinical results, the outcome of pregnancy, and the follow-up of children after ICSI using ejaculated, epididymal, and testicular sperm in order to address these important issues in a relatively short time. Over a 3-year span (1993-1995), the number of centers for ICSI increased from 35 to 101, and the total number of ICSI cycles per year rose from 3,157 to 23,932. The incidence of oocytes damaged by the procedure remained low (< 10%), whereas the fertilization rates obtained with ejaculated, epididymal, and testicular spermatozoa for 1995 were 64%, 62%, and 52%, respectively. Thus, 86-90% of the couples had embryo transfer, and the viable pregnancy rate was 21% for ejaculated, 22% for epididymal, and 19% for testicular sperm, while the incidence of multiple gestations was 29%, 30%, and 38%, respectively. It is noteworthy that no difference was found in ICSI results concerning the etiology of azoospermia, for example, obstructive (congenital or acquired) or nonobstructive. Furthermore, 3,149 transfers of frozen-thawed embryos after ICSI with ejaculated, epididymal, or testicular sperm were performed, and in 11%, 9%, and 7% of them, respectively, a viable pregnancy was achieved. The ICSI results were similar during this 3-year period, irrespective of the origin of the sperm. The perinatal outcome of children born after ICSI was not different from that after in vitro fertilization or natural conception and was only affected by multiplicity. Moreover, the incidence of major or minor malformations was not increased, but the chromosomal, especially the sex chromosomal, aberration rate was slightly elevated (approximately 2%). Therefore, ICSI has opened new horizons in the treatment of male infertility. The achievement of pregnancy after ICSI using ejaculated, epididymal, or testicular sperm is very satisfactory. The procedure seems to be safe, but further follow-up of the children is necessary to more accurately assess its safety.

Abstract: The combination of aspermia and obstructive azoospermia in the same infertile man is a rather rare entity. In the case reported here, all diagnostic criteria as well as subsequent recovery following two operations are compatible with an inflammatory origin. In such cases assisted reproduction should be recommended. However, in this case, an early spontaneous pregnancy rendered this unnecessary.

Abstract: The clinical application of gonadotrophin-releasing hormone (GnRH) antagonists instead of GnRH agonists, to prevent spontaneous premature luteinizing hormone surge during ovarian stimulation for assisted reproduction treatment has been advocated. A recent, double-blind, dose-finding study, including six dosages of the GnRH antagonist ganirelix, in women undergoing ovarian stimulation with recombinant follicle stimulating hormone (FSH), has indicated that high doses of GnRH antagonist (1 or 2 mg once daily) are associated with a low implantation rate. This follow-up study reports on the pregnancy rate after replacement of cryopreserved embryos obtained in stimulation cycles of the above-mentioned trial. Ovarian stimulation was initiated on day 2 of the cycle, with daily injections of 150 IU recombinant FSH. Ganirelix (0.0625, 0.125, 0.25, 0.5, 1.0 or 2.0 mg) was administered once daily from stimulation day 6 onwards, up to and including the day of human chorionic gonadotrophin. Retrieved oocytes were fertilized by in-vitro fertilization (IVF) or intracytoplasmic sperm injection and a maximum of three fresh embryos was transferred. Excess embryos were frozen, and subsequently used in either natural or programmed cycles. Until June 1998, 11 ongoing pregnancies (12-16 weeks after embryo transfer) were achieved from 46 cycles in which embryos had been first frozen (23.9% per transfer). Six of these 11 patients had been treated with a high dose of ganirelix (1.0 or 2.0 mg) during the IVF cycles in which the embryos were obtained. In conclusion, our data suggest that high dosages of ganirelix do not adversely affect the potential of embryos to establish clinical pregnancy in freeze-thaw cycles.

Abstract: The introduction and widespread application of assisted reproduction techniques have raised major concern about the outcome of resulting pregnancies, as well as about the offspring's health. It seems that pregnancies achieved after standard in-vitro fertilization (IVF) bear an increased risk for prematurity and low birth weight. On the other hand, pregnancy outcome of singletons achieved after intracytoplasmic sperm injection (ICSI) and after the transfer of frozen-thawed embryos is similar to that of spontaneously conceived singletons, and pregnancy complications are related only to gestation multiplicity. The incidence of congenital and chromosomal anomalies after standard IVF seems to be similar to that expected in the general population. The prevalence of congenital malformations does not seem to be higher after ICSI. On the other hand, there is a slightly increased risk for transmission of chromosomal aberrations of paternal origin and a higher risk of de novo, mainly sex-chromosomal aberrations. Postnatal growth and development of children born after standard IVF and cryopreservation seem to be within the normal ranges. Growth of ICSI children is also normal while their mental development needs further investigation. Family functioning in assisted reproduction families is better.

Abstract: PURPOSE: This study was undertaken to correlate the follicular levels of interleukin (IL)-1 alpha, IL-2, tumor necrosis factor-alpha (TNF-alpha), and leukotriene (LT) B4 with oocyte maturity, fertilization, and achievement of pregnancy. METHODS: The material was obtained from 22 women undergoing IVF, 8 of whom became pregnant and 14 of whom did not. RESULTS: All of the studied cytokines and LT B4 were found in follicular fluids, but there were no significant differences according to oocyte maturity, fertilization, embryo quality, and achievement of pregnancy. On the other hand, a significant positive correlation was found between IL-1 alpha and TNF-alpha, IL-1 alpha, and LT B4 as well as between TNF-alpha and LT B4 in follicular fluids with subsequently fertilized oocytes. CONCLUSIONS: It seems that IL-1 alpha, TNF-alpha and LT B4 may take part in the process of follicle wall degradation, and their follicular correlations may suggest more optimal follicular and oocyte development and maturation.

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Abstract: Intracytoplasmic sperm injection (ICSI) has revolutionized the treatment of male infertility, since normal fertilization and ongoing pregnancies can be achieved with severely affected spermatozoa. Hence, the application of ICSI is rapidly expanding around the world, necessitating an accurate assessment of the efficacy and safety of this novel technique. The European Society of Human Reproduction and Embryology (ESHRE) Task Force is gathering data annually on the clinical results, the pregnancy outcome and the follow-up of children born after ICSI using ejaculated, epididymal and testicular spermatozoa, in order to be able to provide reliable information on these important issues. During the 3 years 1993-1995, the number of centres performing ICSI increased from 35 to 101, and the total number of ICSI cycles performed per year rose from 3157 to 23932. The incidence of oocytes damaged by the procedure remained low (<10%) and the fertilization rates obtained with ejaculated, epididymal and testicular spermatozoa in 1995 were 64, 62.5 and 52% respectively. Thus, approximately 90% of the couples had an embryo transfer and the viable pregnancy rate was 21% for ejaculated, 22% for epididymal and 19% for testicular spermatozoa (with 25-30% multiple pregnancies). Furthermore, 3149 transfers of frozen-thawed embryos were performed and 7-11% of them resulted in a viable pregnancy. The ICSI results were similar during this 3 year period, irrespective of the origin of the spermatozoa. The perinatal outcome of children born after ICSI was not different from those born after in-vitro fertilization (IVF) or natural conception, and was only affected by multiplicity. Moreover, the incidence of major or minor malformations was not increased, but the chromosomal, especially the sex-chromosomal, aberration rate was slightly elevated. To summarize, a very high success rate is obtained by ICSI independently of the source of the spermatozoa, verifying the superiority of ICSI over conventional IVF. The procedure seems to be safe, but further follow-up of the children is necessary in order to be able to assess its safety more accurately.

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Abstract: Since 1992, intracytoplasmic sperm injection has been established as the treatment of choice for severe male subfertility. Worldwide results with regard to fertilization, pregnancy and pregnancy outcome, collected by the EHSRE Task Force on ICSI, are reported in this short survey. Aspects of genetic inheritance that are of major concern are addressed briefly.

Abstract: Ovarian pregnancy is a rare variant of ectopic implantation. A case of an intrafollicular ovarian pregnancy after ovulation induction/intrauterine insemination is presented. The woman had primary infertility of 4 years. Diagnostic laparoscopy revealed endometriosis and adhesions. After adhesiolysis and laser vaporization of endometriotic implants, the patient underwent ovulation induction with artificial insemination by husband/intrauterine insemination; she conceived at her second attempt. The pregnancy proved to be an ovarian intrafollicular one. She was treated by right partial ovariectomy. Three months later she conceived spontaneously with an intrauterine pregnancy which is still ongoing. The diagnostic problems resulting from the coexistence of ovarian hyperstimulation and the intrafollicular development of pregnancy are discussed. A re-evaluation of the criteria for the diagnosis of ovarian pregnancy based on the currently available diagnostic methods is proposed. Moreover, the pathophysiology of ovarian and especially intrafollicular implantation is reviewed.

Abstract: OBJECTIVE: To present a case of immotile cilia syndrome, a very rare cause of male infertility and to evaluate the role of the recently suggested treatment by intracytoplasmic sperm injection (ICSI). DESIGN: Case report. SETTING: Tertiary-care academic hospital. PATIENT(S): One man with immotile cilia syndrome, showing no motile spermatozoa despite normal morphology and viability. INTERVENTION(S): The patient's partner underwent two cycles with IVF of the oocytes achieved by ICSI. MAIN OUTCOME MEASURE(S): Evaluation of ICSI procedure in cases of immotile cilia syndrome. RESULT(S): Thirty-three percent of the oocytes were fertilized and subsequently divided enabling ET in both cycles. CONCLUSION(S): Intracytoplasmic sperm injection seems to represent a promising approach to the problem of infertility in men with immotile cilia syndrome.

Abstract: The aim of this study was to evaluate the clinical, hormonal and biochemical characteristics of infertile men with azoospermia. A total of 187 azoospermic out of 2610 infertile men (7.2%) were studied. Mean testicular volume and basal plasma levels of FSH were the most useful parameters concerning the evaluation of azoospermia. Basal plasma levels of LH and T were useful only in azoospermic men with hypogonadism, whereas plasma PRL levels, semen volume, and seminal plasma fructose levels were not found to be of common use except in selected cases.

Abstract: Delayed puberty and hypogonadism are frequently observed in patients with homozygous beta-thalassaemia. We evaluated the pituitary-testicular axis in 30 thalassaemic men, aged from 17 to 35 years who were regularly transfused and underwent chelation therapy, while emphasis was given to pituitary reserves of gonadotrophins and the correlation of hormones with serum ferritin (SF). The investigation included endocrinological examination, evaluation of serum basal levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), free testosterone and gonadotrophin-releasing hormone (GnRH) test and also spermiograms. According to the results, patients were divided into three groups: group A, which included 18 eugonadal patients with moderately elevated SF, group B which included six patients who had hypogonadotrophic hypogonadism and excessive elevation of SF, and group C, which included six patients characterized as intermediate, with regard to sexual maturation and SF levels. In conclusion, beta-thalassaemia major leads to variable pituitary iron overload and thus hypophyseal damage. This endocrine disturbance is becoming less frequent nowadays with early and intensive chelation therapy.

Abstract: In an in-vitro fertilization (IVF)/embryo transfer programme granulosa cells obtained from 59 individual preovulatory follicles were analysed using multiparameter image analysis cytometry, in an attempt to determine whether their morphometric and DNA-cytometric parameters could prove useful in assessing follicle and oocyte maturity and in predicting fertilizability and outcome of these IVF cycles. Almost all morphometric and DNA-cytometric parameters were not correlated with either the patient's age or oocyte maturity, and did not predict oocyte fertilization or occurrence of a clinical pregnancy. The only possible relevant parameter which, despite its inverse correlation to total luteinizing hormone administration, also proved to be inversely correlated to pregnancy outcome (in the seven cases in which a pregnancy occurred), was the percentage of granulosa cell nuclei with increased DNA content (> 5c). Finally, if granulosa cells do not reveal euploid polyploidization in spontaneous or induced ovulatory cycles, the detected cells with increased DNA content should be interpreted as aneuploid, i.e. with chromosomal aberrations, and so their presence could also be discussed in connection with the hypothetical risk of prospective neoplastic transformation of the tissue.

Abstract: The application of intracytoplasmic sperm injection (ICSI) is rapidly becoming more popular around the world. The European Society of Human Reproduction and Embryology (ESHRE) Task Force is aiming to collect annually the clinical results and the pregnancy outcomes of ICSI using ejaculated, epididymal and testicular spermatozoa to enable the provision of reliable information on the efficacy and safety of this novel technique. This review summarizes the activities of the ESHRE Task Force on ICSI during the last 2 years. The number of centres performing ICSI as well as the number of ICSI cycles increased significantly from 1993 to 1994. The incidence of oocytes damaged by the procedure was low (7.2-10.6%), whereas the fertilization rate achieved with ejaculated, epididymal and testicular spermatozoa was high (51.1-60.8%), even with extremely impaired semen quality. Thus, 89-93% of patients had an embryo transfer and 21-31% of them achieved a viable pregnancy, irrespective of the origin of the spermatozoon. ICSI results were similar in 1993 and 1994. The follow-up of children born after ICSI revealed no increase in the incidence of major congenital malformations or chromosomal aberrations. These findings are quite reassuring, although the numbers are still too few. Therefore, efforts need to be continued to enhance the database and thus provide a reliable assessment of this new treatment modality.

Abstract: Concerns have been raised recently about the possible association between superovulation and ovarian cancer. In order to contribute to the limited literature on this important issue, two cases of ovarian tumours in women who had undergone multiple ovulation inductions are presented. In the first case, the patient had secondary anovulatory infertility. She was treated with human menopausal gonadotrophin (HMG) alone and in combination with clomiphene citrate or buserelin for six cycles. She then underwent ovarian stimulation with buserelin/HMG in the long protocol for in-vitro fertilization (IVF) and embryo transfer. In preparation for a new IVF/embryo transfer attempt, 8 months later, the screening ultrasound revealed a cystic formation of the left ovary and an enlargement of the right. During laparotomy, both ovaries were found to bear large tumours (approximately 6 x 5 x 4 cm) which were removed. Histological examination showed that they were epithelial tumours (serous-papillary cystadenomas) of borderline malignancy. The patient conceived spontaneously 1.5 years after the operation. In the second case, the patient presented with secondary anovulatory infertility. She underwent ovulation induction with clomiphene/HMG and with buserelin/HMG in the long protocol, and intra-uterine insemination with husband's spermatozoa and conceived (singleton pregnancy). She was delivered by Caesarean section, during which a cystic tumour of the left ovary was removed. Histological examination revealed a benign mucous cystadenoma of the ovary.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: The effect of pentoxifylline (PF) treatment on human sperm motility and fertilizing capacity was assessed in 43 couples undergoing IVF/ET, as well as in 84 couples treated with AIH/IUI. Sperm motility increased following PF treatment in asthenospermic samples (n = 12) from 39.2 +/- 1.5 to 45.5% +/- 2% (p = .05), in oligoasthenospermic (n = 40) from 38.1 +/- 1.6 to 43.6 +/- 1.7% (p = .001), and in total (n = 127) from 50 +/- 1 to 52.5 +/- 1% (p < .001). In addition, PF incubation of sperm samples resulted in higher sperm motility values compared to swim-up in all categories of sperm samples. Furthermore, progressive motility increased in all sperm groups following PF treatment (astheno-spermic: +16.7%; oligoasthenospermic: +14%, p < .001; oligozoospermic: +23.5%, p < .001; normozoospermic: +15.3%, p < .05; total +19.2%, p < .001). Sperm preparation with PF resulted in higher fertilization rates in all categories of sperm samples compared to swim-up (total: 46.6 vs. 29.1%, respectively; p < .05). Moreover, PF treatment of human sperm resulted in 9 viable pregnancies (IVF/ET: 5, AIH/IUI: 4) and one biochemical (IVF/ET). PF improving sperm motility and fertilizing capacity appears to be a promising motility stimulant.

Abstract: Increased interest has arisen recently about the possible association between ovarian stimulation and ovarian tumours. In this article, the current knowledge on the epidemiology, pathogenesis and aetiology of ovarian tumours is extensively reviewed in relation to the existing literature on the relationship between ovulation induction and ovarian neoplasia. The available data from epidemiological studies and case reports do not support a direct causal relationship between ovarian stimulation and ovarian cancer. However, it is possible that ovarian stimulants may have an augmenting role for special categories of tumours, e.g. sex-cord stromal tumours. A definite answer to this important issue may be reached through large prospective epidemiological studies or large retrospective well-designed case-control studies.

Abstract: One of the main endocrinological disturbances in patients with polycystic ovarian syndrome (PCOS) is the increased baseline concentrations of luteinizing hormone (LH) and consequently a high LH:follicle-stimulating hormone (FSH) ratio. The aim of this study was to assess the relationship between the baseline LH:FSH ratio with the stimulation response and the miscarriage risk in PCOS women stimulated for assisted reproduction techniques (ART) with and without gonadotrophin-releasing hormone analogue (GnRHa). Two groups of PCOS patients were analysed retrospectively. Group A (n = 20, 20 cycles) consisted of women stimulated with human menopausal gonadotrophin (HMG), and group B (n = 128, 162 cycles) comprised women stimulated with buserelin-long/HMG. LH and FSH concentrations were measured during the early follicular phase (days 4-6) in a preceding spontaneous or progestin-induced cycle. The following parameters were assessed: number of follicles developed, number of oocytes obtained and percentage of mature oocytes, as well as number of abortions and live births. In group A, the baseline LH:FSH ratio was correlated inversely with the number of follicles developed (P < 0.05), the number of oocytes obtained (P < 0.05) and the percentage of mature oocytes (P < 0.05). In group B, no correlation was found between the LH:FSH ratio and the number of follicles and oocytes, because their numbers were relatively constant irrespective of the baseline LH:FSH ratio, but a significant inverse correlation was noted with the percentage of mature oocytes (P < 0.001). However, a comparison of the slopes of the curve indicated a better correlation between the LH:FSH ratio and the percentage of mature oocytes in group A than in group B (P < 0.05). These findings were also confirmed when patients were subdivided according to the LH:FSH ratio (< 3 or > or = 3). Furthermore, in women who miscarried, the mean LH:FSH ratio was significantly higher than in women having a live birth. In conclusion, in PCOS patients stimulated with HMG, a high basal LH:FSH ratio appears to have an adverse effect on the number of follicles and oocytes, as well as on oocyte maturity. On the other hand, the administration of GnRHa in the long protocol seems to reverse this detrimental effect on follicle and oocyte development. Furthermore, a higher LH:FSH ratio seems to predict a greater possibility for miscarriage, despite the use of GnRHa.

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Abstract: In this study we evaluated retrospectively the efficacy of five different ovarian stimulation protocols in an in vitro fertilization program, in which 512 women were involved. Ovulation was induced by the following protocols: group I (271 cycles): buserelin short protocol (1 mg/day, intranasally) with human menopausal gonadotropin/human chorionic gonadotropin (hMG/hCG); group II (45 cycles): buserelin (short protocol) with pure follicle stimulating hormone (p-FSH)/hMG/hCG; group III (24 cycles): clomiphene citrate (100 mg/day) with hMG/hCG; group IV (122 cycles): hMG (3 ampules/day) and hCG; group V (113 cycles): hMG/hCG and prednisolone (7.5 mg/day) after cycle programming with oral contraceptives. The lowest cancellation rate (3.3%) was noted in group I, followed by group V (9.7%). The highest number of follicles was observed in groups I (8.3 +/- 0.3; mean +/- SEM) and V (7.8 +/- 0.5). Also, more oocytes were retrieved in group I (7.2 +/- 0.3, p < 0.001), which were of good quality based on oocyte maturity as well as on the fertilization rate, and more embryos (4.5 +/- 0.3, p < 0.05) were developed. The correlation between estradiol and the total follicular volume on the day of hCG administration was also examined in the five groups. The best correlation (r = 0.6502) was found in group I, followed by group V (r = 0.5810). Significant differences were observed in the five groups with regard to the number of hMG ampules administered (p < 0.0001, F = 15.393) and the stimulation days (p < 0.0001, F = 35.32). Sixty-six clinical pregnancies were achieved: 37 (17.5%) in group I, seven (25.9%) in group II, one (10%) in group III, ten (15.6%) in group IV and 11 (15.5%) in group V (differences were not statistically significant). In conclusion, all five protocols were satisfactory in ovarian stimulation for in vitro fertilization, and gonadotropin releasing hormone (GnRH) analogs seemed to be more advantageous by reducing the cancellation rate, enhancing the number of oocytes retrieved and embryos developed and by improving the pregnancy rates.

Abstract: PURPOSE: The administration of two GnRH analogues (buserelin and leuprolide acetate) in long and short protocols was evaluated prospectively. In the long protocol, the analogues were given from day 1 of the cycle for at least 14 days and then hMG administration was started, while in the short protocol the analogues were initiated from cycle day 1, adding hMG from day 3. The patients were divided into four groups according to the protocol used: Group I, buserelin-long; Group II, buserelin-short; Group III, leuprolide-long; and Group IV, leuprolide-short. Serum E2 levels on the day of hCG injection and the number of follicles observed, oocytes retrieved, and embryos obtained, as well as implantation rates, were significantly higher (P < 0.001) in the long protocols than in the short ones. RESULTS: Pregnancy rates were similar in all groups, although a trend for better results was observed in buserelin-long compared to buserelin-short. There were no differences in the results achieved with buserelin or leuprolide. CONCLUSION: The administration of GnRH analogues (buserelin and leuprolide acetate) in long protocols induced a more intense ovarian response and was associated with significantly higher implantation rates and also a trend for higher pregnancy rates, although this difference was not statistically significant.

Abstract: The aim of this study was to examine the occurrence of ovarian cysts during the administration of three different gonadotrophin-releasing hormone analogues (GnRHa) in the long protocol as well as their characteristics, management and outcome compared with patients with no cyst formation. A total of 172 in-vitro fertilization (IVF) cycles in which GnRHa was administered at menstruation were analysed. Group B consisted of 72 cycles in which buserelin was used. Of these, 10 (13.9%) were with cysts (group B1) and 62 (86.1%) without cysts (group B2). Group T included 49 cycles in which triptorelin was injected. Of these, seven (14.2%) were with cysts (group T1) and 42 (85.7%) without cysts (group T2). Group L comprised 51 cycles in which leuprolide was administered. Of these, eight (15.7%) were with cysts (group L1) and 43 (84.3%) without cysts (group L2). All women with ovarian cysts had higher serum oestradiol concentrations and all except five underwent cyst aspiration with no complication. No differences were observed in the number of follicles and oocytes between groups B, T and L or between the groups with cysts and those without cysts. The pregnancy rate was similar in all groups. In conclusion, follicle cyst formation does not seem to be related to the use of a specific GnRHa, its short- or long-acting form or to the mode of administration. In addition, follicle cyst aspiration is a safe and successful solution to the problem of functionally active ovarian cysts.

Abstract:

Abstract: Psychosocial impacts of infertility were investigated in couples undergoing different treatment procedures in our clinic. Couples were interviewed in a semi-structured way by a psychologist or a psychiatrist and responded to three specially structured questionnaires: the Life Events Scale, the Marlowe-Crowne/Taylor Scale and the Side Effect Checklist. The data were analysed in terms of demographic characteristics as well as treatment procedure. The psychosocial, psychosexual and emotional outcomes of their infertility problem and Greek traditional culture laws are discussed. Stress has been identified in both sexes, depression mostly in women, while men showed a tendency towards repressed anxiety and thus a greater risk of psychosomatic illness, a finding supported by their response to the Side Effect Checklist. Women showed a high defensive anxiety and also reported numerous psychosomatic symptoms. These couples seem to have special needs and fears, both general and treatment specific. Very few of our couples would be considered as severely emotionally disturbed. Women seem to have more difficulties in social adjustment. Sexual dysfunction was reported by almost half of our subjects, although this was associated with a degree of deterioration in their marriage. Guilt feelings, particularly connected with previous abortions, seem to be torturing most women. Finally, both partners seem to have psychological problems irrespective of the one in whom the aetiological problem was found. Moreover, traditional rules seem to impose a special burden on people coming from rural areas. Our results strongly support the belief that infertile couples undergoing different treatments need psychological counselling and supportive psychotherapy.

Abstract: The combined administration of the gonadotrophin-releasing hormone (GnRH) agonist buserelin and human menopausal gonadotrophin (HMG) was evaluated in 527 cycles (428 patients) of an assisted reproduction programme. All women were randomly allocated according to the ovulation induction protocol into two groups: group I (short protocol; 318 cycles) was given buserelin (1 mg/day) intranasally from cycle day 1 and HMG (2 ampoules/day) from day 3 until human chorionic gonadotrophin (HCG) administration: group II (long protocol; 209 cycles) was given buserelin (1 mg/day) intranasally from cycle day 1 for at least 14 days and then 2 ampoules HMG/day were added, increasing progressively according to the ovarian response. The number (mean +/- SEM) of follicles developed was higher in group II than in group I (9.1 +/- 0.4 versus 7.7 +/- 0.3, respectively; P < 0.05). More oocytes were retrieved in group II (8.4 +/- 0.5) than in group I (6.5 +/- 0.3) (P < 0.001), as well as more embryos (6.3 +/- 0.5 and 4.0 +/- 0.3, respectively; P < 0.001). Moreover, in group II there was a better correlation between oestradiol and the total follicular volume (r = 0.5391) on cycle day 0 compared with group I (r = 0.458), while oestradiol values were similar between the two groups. No differences were observed in the cancellation rate, fertilization rate and maturity of the oocytes between the two groups. The pregnancy rate per transfer was slightly better in group II (25.8%) than in group I (19.4%), but this difference was not significant.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Follicular fluid samples and oocytes were obtained from 75 women (87 cycles), who participated in an assisted conception programme. Determinations of the concentration of oestradiol, progesterone, testosterone and growth hormone were performed in all follicular fluid samples. Patients were stimulated with the following regimes: group A (24 cycles, 94 samples), human menopausal gonadotrophin (HMG) (three ampoules/day) and human chorionic gonadotrophin (HCG); group B (23 cycles, 53 samples), HMG/HCG with prednisolone (7.5 mg/day) after cycle programming with oral contraceptives; group C (40 cycles, 60 samples), buserelin with HMG/HCG. Oestradiol concentrations (mean +/- SEM) were significantly higher (P < 0.05) in group A (320.1 +/- 27.3 ng/ml) and those of growth hormone in both groups A and C (3.8 +/- 0.2 and 3.2 +/- 0.15 ng/ml, respectively), as compared to the other groups, whereas progesterone and testosterone concentrations were similar in all groups. The mean concentrations of oestradiol, progesterone, testosterone and growth hormone were significantly higher (P < 0.01) in follicular fluid with oocytes of intermediate maturity than with mature oocytes (382.5 ng/ml, 7847.5 ng/ml, 1704.5 ng/dl and 3.7 ng/ml versus 217.8 ng/ml, 5488.4 ng/ml, 1313.6 ng/dl and 2.7 ng/ml, respectively). On the other hand, only oestradiol concentrations were significantly higher in follicular fluid of fertilized compared to non-fertilized oocytes. Concentrations of the other hormones analysed, except growth hormone, were similar in follicular fluid from pregnant and non-pregnant women after assisted reproduction. Growth hormone, on the other hand, was significantly lower (P < 0.05) in follicular fluid from pregnant compared to non-pregnant women (2.8 versus 3.5 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract:

Abstract: The purpose of the present study was to evaluate the kinetics of human sperm acrosome reaction in vitro using the triple stain technique. Acrosome reaction was studied in sperm samples from 16 fertile men 2, 6, and 9 h after ejaculation, following incubation in culture medium (CM; Ham's F-10), with a mixture of CM and follicular fluid (FF), or with FF only. Incubation of sperm samples without the influence of any medium served as control. The highest proportion of living acrosome-reacted sperm after a 2-h incubation period occurred in samples incubated with FF (18%), followed by samples incubated with the mixture (15.2%), and then with CM (11.8%). The proportion of living sperm that had undergone the acrosome reaction in the control group was significantly lower (5.7%, p < 0.05). After 6 h of incubation, live acrosome-reacted sperm in FF had increased to 39%, in the mixture to 35.5%, and in CM to 30.5%, whereas in the control group the increase was only 6.3% (p < 0.05). After 9 h of incubation, the percentage of living reacted sperm showed a decline compared with the percentage at 6 h. This decline was greater in samples incubated with FF (from 39 to 19.8%) than in samples incubated with the mixture (from 35.5 to 23.6%). Samples incubated in CM only showed a small decrease from 30.5 to 28.4%, while in the control group this percentage decreased from 6.3 to 2.3%. In conclusion, incubation of sperm in suitable media favorably influences the acrosome reaction, inducing an increase in the percentage of live acrosome-reacted sperm.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Sperm concentration and percentage motility values generated by the HT M-2030 system (CASA) were compared with those obtained by subjective semen analysis (SSA). Three calibrations arbitrarily designated as A, M, and D for gates and discriminators and three calibrations arbitrarily designated LI, MI, and HI for default pixel count (DPC) and default intensity (DI) were evaluated. The best correlation between CASA and SSA was observed utilizing M calibration (n = 114) with which sperm concentration was +0.3 x 10(6)/mL (r = .96) and motility was -6.3% (r = .89) compared to the values obtained by SSA. It was found that 35.9% of sperm concentration values and 34.2% of sperm motility values were within 10% of the values obtained by SSA. When sperm concentration was between 50 and 100 x 10(6)/mL the difference in motility was reduced (-3.2%) while the difference in sperm concentration was reversed (-2.6 x 10(6)/mL). LI calibration (DPC = 4, DI = 86) gave acceptable results with M calibration for sperm concentration (+2.1 x 10(6)/mL) and motility (-6.9%) compared to the values obtained by SSA. In the presence of sperm clumping, the difference between CASA and SSA was reversed for sperm concentration (+0.56 x 10(6)/mL for normal samples vs. -2.2 x 10(6)/mL for samples with clumping) and was reduced for sperm motility (-7.14% vs. -4.55%, respectively). HT M-2030 under proper calibration can be used as a rapid, objective, and reliable alternative to conventional semen analysis in routine and for research purposes.

Abstract: Gonadotrophin releasing hormone (GnRH) agonists have had a profound impact on infertility treatment by in vitro fertilization and related procreation techniques. Its combination with human menopausal gonadotrophin (HMG) for superovulation has increased the success rate in patients with premature luteinizing hormone (LH) surges, a poor response, endometriosis or polycystic ovarian disease. Although most in vitro fertilization (IVF) centres routinely use GnRH agonists for reasons of a better planning of activities, its superiority as a first-line treatment over the more conventional regimes of clomiphene citrate (CC)/HMG or HMG alone is still debated. An increase of the number and maturity of recovered oocytes was reported by most groups, but an inverse relationship between the number of oocytes and embryo quality and pregnancy rate after cryopreservation has been frequently mentioned. The possible complications of the combined use of a GnRH agonist with HMG include the formation of ovarian 'follicle' cysts and ovarian hyperstimulation syndrome; luteal phase defects are also common. Routine luteal phase support with progesterone and oestradiol valerate or human chorionic gonadotrophin (HCG) is required. GnRH agonists have proved to be safe drugs, side-effects are minor and sporadic and the congenital malformation rate in GnRHa/HMG treatment is comparable to the spontaneous rates reported in national IVF registers.

Abstract: Seminal plasma transferrin levels were estimated in 51 men with various infertility problems and in 15 fertile subjects. The estimation of transferrin was carried out by using the radial immunodiffusion technique on LC-partigen transferrin plates. The mean value of transferrin decreased (p less than .05) in cases of azoospermia (91 +/- 51 micrograms/ejaculate). No significant difference was found in other sperm quality disorders (asthenoteratospermia = 227 +/- 141, mild OTA = 186 +/- 96, severe OTA = 247 +/- 137 micrograms/ejaculate). The mean value of transferrin decreased (p less than .05) in obstructive azoospermia cases (73 +/- 70 micrograms/ejaculate), whereas in the other causes of infertility no statistical differences were found. Transferrin was correlated with sperm count, sperm volume, and serum LH.

Abstract: Although healthy babies have been born after preimplantation diagnosis for sex determination, this technique is still in an experimental phase. To date, removal by micromanipulation of one or two blastomeres from an eight-cell embryo seems an acceptable procedure. Since a reasonable number of biopsied embryos will implant and develop in utero, the methodology for the analysis of the blastomeres at the chromosomal, gene product or DNA level, needs to be improved before further clinical application. In the meanwhile, more information about early embryo development will become available.

Abstract: beta-N-acetylhexosaminidase is a lysosomal enzyme, which has two isoenzymes: beta-Hex A, a trimer consisting of one alpha-chain and two beta-chains (alpha beta 2) and beta-Hex B, a tetramer formed of four beta-chains (beta 2 beta 2). Genetic defects in the alpha-chain lead to Tay-Sachs disease, whereas mutations in the beta-chain gene lead to Sandhoff disease. In a previous study we developed a microassay for total beta-N-acetylhexosaminidase and used this for measuring activities in mouse oocytes and preimplantation embryos. In this study, to assess the feasibility of transferring this technique to the human for the purposes of preimplantation diagnosis for Tay-Sachs and Sandhoff disease, beta-Hex activity was assayed in human oocytes and embryos and in the medium in which they had been cultured. We showed that although the activity of beta-N-acetylhexosaminidase in human oocytes and embryos was > 500 times higher than in the mouse, it was not detectable in the culture medium and the activity in oocytes and embryos remained virtually constant throughout human preimplantation development, making it difficult to distinguish embryonic from maternal enzyme activity. In the absence of this distinction it would be inappropriate to use beta-N-acetylhexosaminidase activity for the purposes of preimplantation diagnosis of Sandhoff or Tay-Sachs disease. These experiments demonstrate that measuring the beta-N-acetylhexosaminidase activity in human embryos cannot be used at present for preimplantation diagnosis.

Abstract: In this study, we examined the efficacy of intrauterine insemination with washed spermatozoa from the husband (AIH/IUI) in the treatment of infertility. A total of 127 treatment cycles were completed (1.95 cycles per patient). The indications for AIH/IUI were male subfertility (group I: 53 couples), cervical factor (group II: four couples), male and cervical factor (group III: six couples) and unexplained infertility (group IV: two couples). Sperm washing caused a reduction of the sperm concentration from 52 x 10(6)/ml to 44 x 10(6)/ml (P less than 0.08) and motility from 49% to 45% (P less than 0.03). Twelve pregnancies were achieved (18.5% per couple and 9.5% per cycle): eight pregnancies in group I (15.1%), one in group II (25%) and three in group III (50%). Of the 12 pregnancies, eight occurred in the first insemination cycle, three in the second and one in the third (1.4 cycles/pregnancy). Sperm motility was significantly higher in the pregnant than in the non-pregnant group (65.5% versus 46.4%, respectively; P less than 0.004), whereas no significant difference was observed in the sperm concentration between these two groups (39 x 10(6)/ml and 54 x 10(6)/ml, respectively). In conclusion, AIH/IUI is a technically easy and non-invasive procedure which offers a satisfactory pregnancy rate in selected couples with male subfertility and/or cervical factor infertility.

Abstract: Renin-like activity and angiotensin II/III immunoreactivity in follicular fluids from 34 women stimulated with human menopausal gonadotropin and human chorionic gonadotropin (56.8 +/- 6.5 ng angiotensin I per milliliter per hour and 187 +/- 21 pg/ml [mean +/- SEM], respectively) were much higher (p less than 0.001) than in follicular fluids from 12 unstimulated preovulatory women (1.41 +/- 0.37 ng angiotensin I per milliliter per hour and 58.5 +/- 13.7 pg/ml) and in simultaneously drawn plasma (4.47 +/- 0.73 ng angiotensin I per milliliter per hour and 31.8 +/- 11.6 pg/ml, respectively; p less than 0.001). Plasma renin-like activity and angiotensin II/III immunoreactivity in stimulated cycles did not differ from unstimulated cycles. Follicular fluid angiotensin II/III immunoreactivity correlated significantly with follicular fluid renin-like activity in stimulated (r = 0.72; p less than 0.01) and in unstimulated samples (r = 0.86; p less than 0.01). Significant correlation was found also between follicular fluid renin-like activity and estradiol. A sharp preovulatory rise of renin-like activity and angiotensin II/III immunoreactivity was noted in unstimulated follicular fluid samples collected on cycle days 13 and 14 compared to days 9 through 12 (p less than 0.01). The findings that follicular fluid renin-like activity and angiotensin II/III immunoreactivity are correlated, and that gonadotropins have a stimulatory effect on follicular fluid concentrations support our concept of a physiologic intrinsic ovarian renin-angiotensin system.

Abstract: Stimulation of multiple follicular development was induced concurrent with a viable early intrauterine pregnancy; the infant was subsequently delivered at term. Despite the pregnancy, the ovaries responded successfully to gonadotropin stimulation. In addition, three recovered oocytes were successfully fertilized, and two subsequently cleaved. Thus pregnancy does not create a state of ovarian refractoriness or a hormonal milieu totally deleterious to the initial stages of embryo development.

Abstract: Fertilization and cleavage rates were compared in 1024 heparin-exposed and nonexposed human oocytes recovered from 183 consecutive in vitro fertilization (IVF) cycles. Heparinized Ham's F-10 medium, (Gibco, Grand Island, NY) 1.0 ml (2.0 mg heparin/ml) was added to bloody follicular fluid; clear follicular aspirates did not receive heparin. Fertilization and cleavage rates for heparin-exposed (n = 714) and nonexposed (n = 310) oocytes were not significantly different: 63.9 versus 61.6% fertilized (chi 2 = 0.472); 89.3 versus 87.4% of fertilized eggs cleaved (chi 2 = 0.445). A subset of 100 patients, each contributing both heparin-exposed and nonexposed oocytes, also was evaluated. Fertilization and cleavage rates were not significantly different: 59.1 versus 60.8% fertilized (chi 2 = 0.192); 87.6 versus 87.2% of fertilized oocytes cleaved (chi 2 = 0.014). A modified triple stain was used to evaluate the acrosome reaction rate of sperm that had been coincubated with 76 oocytes from ten patients. There was no significant difference in the proportion of viable acrosome-reacted sperm following incubation with heparin-exposed (1.9 +/- 1.0%) versus nonexposed (2.3 +/- 1.2%) (mean +/- standard deviation [SD]) oocytes. The addition of heparin to follicular fluid at the time of oocyte recovery for IVF has no apparent effect on fertilization or cleavage in vitro, nor any influence on the acrosome reaction.

Abstract: The glycosaminoglycans (GAGs) chondroitin sulfate (CS) and heparan sulfate (HS) were assayed in fluid from 178 individual follicles obtained from women after human menopausal gonadotropin ovulation induction for subsequent in vitro fertilization. CS and HS concentrations ranged from 0 to 3.9 and 0 to 18.6 mg/ml, respectively. The percentage of oocytes fertilized was significantly higher when CS concentrations ranged from 0.3 to 0.8 mg/ml (P less than 0.03). As HS levels increased, maturation scores by visual assessment of the egg-corona-cumulus cell complexes increased (P less than 0.05), but the percentage of oocytes fertilized was not affected. Cleavage rates of developing embryos were not related to the concentrations of GAGs.

Abstract: Semen preparation is an important step of in vitro fertilization (IVF) and can affect the success of this procedure. Prior to oocyte insemination, spermatozoa are washed to remove seminal plasma which is believed to contain decapacitation factors. This study was undertaken to evaluate the effect of preparation on semen quality and subsequent successful IVF. Oocytes were recovered from 12 hMG/hCG-stimulated women by laparoscopy, and 6 h later semen specimens were obtained from the male partners. After liquefaction, 1 ml of semen was centrifuged twice in Ham's F10 medium supplemented with 10% of homologous serum, and the final suspension was used to inseminate the preovulatory eggs. In the initial and washed-sperm suspensions, motility was evaluated by the MEP method, and the occurrence of acrosome reaction and sperm viability were evaluated by the triple-stain technique. Fertilization was documented by the formation of two pronuclei. Washing caused a significant decrease in the percentage of motile sperm from 68% to 59% but significantly increased mean sperm velocity from 26 to 29 micron/sec (p less than 0.01). The mean fertilization rate was 65%, and no correlation was found with any of the parameters of semen quality before or after washing. Semen preparation for IVF is associated with a decrease in the percentage of motile sperm that does not seem to affect the fertilizing ability of normal spermatozoa but may be of importance in patients with abnormal semen.

Abstract: Human ovarian follicular fluid (hFF) has angiogenic activity, although the causative factors are unknown. We recently found that hFF contains renin activity which converts renin substrate to angiotensin I (AI). Since the enzymatic cleavage product of AI, angiotensin II (AII), is a potent stimulator of new vessel formation, we have examined Sephadex G-25 column fractions of hFF and extracted hFF and plasma from individual patients for AII-like immunoreactivity (AII-IR). Eluent fractions from Sephadex G-25 column chromatography of hFF had significant AII-IR which eluted in the same fractions as synthetic AII. Individual, extracted FF samples contained approximately 10 times higher levels of AII-IR than extracts of plasma from the same patients. Serial dilutions of the Sephadex column fractions and extracted FF and plasma inhibited binding of 125I-AII to rabbit anti-AII serum in a manner parallel to the inhibition caused by synthetic AII, indicating that the detected immunoreactivity was not due to non-specific assay interference. In summary, the results indicate the presence of significant AII-like immunoreactivity in hFF. AII may now be considered as a potential mediator of the angiogenic activity present in hFF and may play an important paracrine and/or autocrine role in physiologic events in the ovary.

Abstract: One hundred forty-four women suffering from obstructive tubal disease underwent laparoscopy for in vitro fertilization and embryo transfer (IVF-ET). Ovulation was induced by a human menopausal gonadotropin/human chorionic gonadotropin (hMG/hCG) protocol. The patients were divided according to their estradiol (E2) response to hMG/hCG administration: group A, an increase of E2 levels on the day after hCG administration (day +1); group B, a decrease of E2 levels on the day after hCG administration. Each of these groups was further subdivided into high responders (E2 greater than 500 pg/ml on day 0), designated A-1 and B-1, respectively, and low responders (E2 less than 500 pg/ml on day 0), designated A-2 and B-2, respectively. Patients responding in an A-1 pattern had significantly higher E2 levels from day 0 to day +8. Fertilization and cleavage rates did not differ among the groups, nor did luteal-phase progesterone. Twenty-one pregnancies were achieved from 126 transfers (17%), of which 16 (75%) occurred in women responding in an A pattern. We recommend that laparoscopy be deferred in women responding with a B pattern, because this preselection may increase pregnancy rates per IVF-ET cycle.

Abstract: Baseline and gonadotropin-stimulated estradiol production were examined in long term cultures of human granulosa-luteal cells isolated from women undergoing in vitro fertilization. Estradiol production declined by 70% during the first 48 h in culture and was minimally stimulated by the addition of hCG to the culture medium. During subsequent culture from 48-120 h estradiol production was significantly increased over control levels by hCG concentrations greater than 0.1 IU/ml. Incubation with testosterone stimulated estradiol production 100-fold in the presence and absence of gonadotropin. hCG (0.01-10 IU/ml) stimulated a 3- to 13-fold increase in progesterone production. However, at hCG concentrations greater than 1 IU/ml, coincubation with testosterone (10(-7) M) significantly inhibited progesterone production. Dihydrotestosterone also inhibited progesterone production, but to a lesser extent than testosterone. Freshly isolated granulosa-luteal cells specifically bound small amounts of [125I]hCG (less than 1,000 cpm/10(5) cells). Glycine buffer wash was shown to reversibly remove more than 88% of bound hCG and, in freshly isolated cells, increased [125I]hCG binding by 100%. In 5-day cultures, specific [125I] hCG binding nearly doubled from 52,000 cpm/10(5) cells in control cultures to 87,000 cpm/10(5) cells in cultures treated with hCG (0-5 IU/ml). At the highest concentration of hCG (5 IU/ml), testosterone (10(-7) M) significantly inhibited the amount of [125I]hCG specifically bound. In summary, estradiol production in long term cultures of granulosa-luteal cells appears to be gonadotropin dependent. In addition, the presence of testosterone (10(-7) M) antagonizes hCG-stimulated progesterone and LH receptor production by these cells.

Abstract:

Abstract: The present study was designed to determine the accuracy of the cervical score for the timing of human chorionic gonadotropin (hCG) administration in in vitro fertilization patients compared to the timing of hCG administration based upon estradiol and ultrasound measurements. Forty-two patients undergoing ovulation induction with human menopausal gonadotropin (hMG)/hCG for in vitro fertilization were studied. A total of 192 cervical scores was obtained together with plasma estradiol levels and pelvic sonography. hCG was given based on estradiol and ultrasound results without knowing the cervical score. An independent decision to give hCG was made based on the cervical score only following a mean of 2.5 days of a cervical score of 9-12 without the examiner's knowledge of estradiol and ultrasound results. This decision was theoretic and was not considered for the actual timing of hCG. In 16 (38%) of the cases this decision coincided with that based on estradiol and ultrasound. In 24 (57%) of the cases there was a 1-day gap. A direct correlation was found among the cervical score, the plasma estradiol concentration, and the mean diameter of the two leading follicles (P less than 0.001). The overall distribution of the cervical score grouped according to estradiol levels (less than 200, 201-400, and greater than 401 pg/ml) was significantly different (P less than 0.001). The proportion of "mature" cervical scores (score of 11-12) in the three different estradiol groups was 15.6, 34.5, and 68.0%, respectively. In conclusion, independent decisions to give hCG based on cervical score or estradiol and ultrasound were coincident +/- 1 day in 95% of the cases.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: The attainment of synchronous follicular development in human menopausal gonadotropin/human chorionic gonadotropin-stimulated cycles for in vitro fertilization (IVF) continues to be a perplexing problem. Two regimens of follicle stimulation for IVF cycles were, therefore, compared. Twenty-nine patients commenced human menopausal gonadotropin (hMG) therapy on day 1 of the menstrual cycle (Group I), while 30 women received hMG from the third day of the cycle (Group II). The hMG therapy was tailored to the individual patients's response, based on ultrasonographic measurements of follicular size and serum estradiol (E2) levels. Both groups of patients received a mean of 19.6 +/- 1.4 ampules of hMG over a mean of 6.1 +/- 0.2 days. The pattern of serum E2 and progesterone levels in the periovulatory and luteal phase was not affected by the day of initiation of hMG therapy, although Group I patients demonstrated lower (P less than 0.05) E2 levels on the 2 days prior to human chorionic gonadotropin (hCG) administration. In terms of follicle growth, Group II follicles consistently demonstrated a significantly (P less than 0.01, chi 2 test) larger proportion of medium- and large-sized follicles compared to Group I follicles on almost all of the days when ultrasonographic measurements were taken. In addition, Group II follicles demonstrated an earlier shift (day-1) to the larger follicles than Group I follicles (day 0). Significantly (P less than 0.001) more oocytes were recovered per aspirated follicle in Group II patients, but the fertilization rate per oocyte was greater (P less than 0.003) for Group I oocytes. Nevertheless, pregnancy rates did not differ between the two groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Consideration of reproductive potential following multiple ectopic pregnancies is important in counseling patients and when choosing a conservative surgical approach (salpingotomy, salpingostomy), partial salpingectomy, or in vitro fertilization. The cases of 336 patients at Yale-New Haven Hospital who had ectopic pregnancies between 1976 and 1981 were evaluated. Thirty-two patients (9.5%) had two ectopic pregnancies. Twenty-three patients (71.8%) were followed. Six of these (26.1%) were not able to conceive because of surgical sterilization, and four (17.4%) were not trying to conceive. Of those 13 patients (56.5%) actively trying to conceive and having at least one tube remaining, 4 (30.8%) had term intrauterine gestations. One had a third ectopic gestation, which represented 20% of all conceptions, or 7.7% of those individuals trying to conceive. Our results indicate that although the reproductive potential after two ectopic pregnancies is poor, viable pregnancies do occur, and the repeat ectopic pregnancy rate is not high enough to preclude a repeat conservative surgical approach.

Abstract: We suggest that regression of the corpus luteum is an active process induced by PGF2 alpha, GnRH, and a peptide of ovarian origin whose action GnRH mimics (20). The initial events involved in luteolysis occur within minutes, and they are intimately linked to inhibition of LH action. Membrane receptor binding of luteolytic hormones activates production of a second messenger (such as a product of PI turnover) that stimulates release of sequestered, intracellular Ca2+ by a mechanism linked to inhibition of microsomal Ca2+-ATPase activity. The increase in cytosolic Ca2+ inhibits adenylate cyclase activity by blocking GTP-dependent activation of adenylate cyclase. As a result, the cell response to LH is abolished and function is lost.

Abstract:

Abstract: Because laparoscopic ovum harvest for IVF is becoming more common, efforts must be made at the time of prior infertility surgery to render the ovaries accessible for ovum recovery. In this report, a simple technique of ovarian suspension in preparation for eventual IVF is described. Seventeen infertile patients undergoing lysis of adhesions underwent ovarian suspension by plication of the suspensory ligament of the ovary with a running suture of 3-0 Prolene, thus apposing the proximal pole of the ovary to the anterior-lateral surface of the uterus. Because the fallopian tube is independent of the sutured structures, the tubal anatomy was undisturbed. Fourteen of these 17 patients have since undergone laparoscopy during IVF. In 12 cases, despite the recurrence of adhesions, the ovaries were accessible for ovum recovery from all aspects. Two patients conceived normally after the lysis of adhesions. This simplified ovarian suspension represents an effective technique for facilitating ovum recovery for IVF which may not disturb fallopian tube ovum pickup or transport mechanisms.

Abstract: cAMP, estradiol (E2), and progesterone levels were determined in 24 follicular fluid samples obtained from 8 women who conceived after in vitro fertilization and in 47 samples from 26 women who did not. Follicular development was induced by human menopausal gonadotropin, and maturation of retrieved oocytes was assessed by the degree of cumulus mucification and corona dispersal. The mean follicular fluid cAMP concentration was significantly (P less than 0.001) lower in women who became pregnant than in those who did not (106 vs. 241 pmol/ml), while the mean E2 level was significantly (P less than 0.01) higher (727 vs. 497 ng/ml), and the progesterone to E2 ratio was significantly (P less than 0.05) lower (9.5 vs. 18.0). Overall, follicles of immature, intermediate, and mature oocytes did not differ in cAMP content. However, intermediate and mature oocytes from women who became pregnant were derived from follicles containing significantly (P less than 0.01) lower cAMP levels than those of women who did not become pregnant (66 and 122 vs. 233 and 288 pmol/ml, respectively). Furthermore, fertilized oocytes leading to conception originated from follicles with significantly (P less than 0.001) lower cAMP concentrations than the follicles that yielded nonfertilized oocytes or fertilized oocytes not leading to conception (92 vs. 270 and 240 pmol/ml, respectively). Similarly, significantly (P less than 0.05) lower cAMP levels were found in the follicular fluid of cleaved oocytes resulting in a pregnancy compared to those that did not (86 vs. 236 pmol/ml). It is concluded that low levels of cAMP are associated with successful fertilization and cleavage of human oocytes in vitro resulting in viable pregnancies and may, therefore, be used as a marker of optimal follicular development in in vitro fertilization cycles.

Abstract: In vitro fertilization has offered new insights into our understanding of ovulation induction, folliculogenesis, and luteal phase events. This new information is provided by the ability to precisely study these cycles in a frequent and sequential fashion through the use of peripheral blood markers, ultrasound evaluation, and follicular fluid constituents and cell culture techniques, as well as direct observation of the oocyte, fertilization, and cleavage. In these stimulated cycles the follicular phase serum estradiol levels in conjunction with ultrasound were evaluated; a poor correlation was shown between follicle size and number and estrogen production. This distinct dyssynchrony suggests the recruitment of a number of cohorts of follicles in each stimulated cycle. From the biochemical markers in follicular fluid, cyclic adenosine monophosphate has a distinct predictive value in regard to pregnancy in in vitro fertilization-embryo transfer cycles. In the luteal phase, the mass effect of aspiration of great numbers of granulosa cells, the effect of supplemental progesterone, and the influence of high follicular phase estradiol levels remain controversial and, therefore, a less clear cut pattern emerges. Variations in the protocol have not greatly improved the major problems of folliculogenesis associated with ovulation induction and an in vitro fertilization-embryo transfer program, that is, follicular asynchrony and luteal phase deficiency.

Abstract: Seventy-one oocyte--corona--cumulus complexes (OCCCs) were obtained from eight women who conceived and delivered following in vitro fertilization (IVF) and eight women who were matched for the number of embryos transferred into the uterus but failed to conceive following the procedure. The steroid secretion of these OCCCs was assessed during a 24-hr culture period. Intermediate (N = 35) and mature (N = 36) complexes did not differ in steroid secretion, and progesterone was the major steroid secreted. OCCCs associated with a fertilized oocyte (N = 59) produced significantly (P less than 0.02) higher levels of progesterone than those associated with nonfertilized oocytes (N = 12) (515 +/- 53 vs 231 +/- 70 ng/OCCC/24 hr) but comparable levels of estradiol (9.17 +/- 1.14 vs 6.55 +/- 2.0 ng/OCCC/24 hr) and testosterone (0.34 +/- 0.09 vs 0.22 +/- 0.05 ng/OCCC/24 hr). Steroid production by OCCCs of oocytes which continued to cleave was not different from that of oocytes which did not cleave following fertilization, but the estradiol-to-testosterone ratio was significantly higher (31.4 +/- 3.5 vs 19.1 +/- 3.3; P less than 0.02) in the former group. The degree of cleavage was not related to the levels of steroid secretion. Cleaved oocytes associated with viable pregnancies (N = 28) were derived from OCCCs which secreted significantly less testosterone than those which did not result in a pregnancy following transfer (N = 26) (0.27 +/- 0.04 vs 0.41 +/- 0.04 ng/OCCC/24 hr).(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Angiogenesis is a feature of ovarian follicle and corpus luteum development. Ovarian homogenates and follicular fluid (FF) contain factors that stimulate new vessel formation. Because the renin-angiotensin system has been shown to facilitate angiogenesis, renin activity was measured in FF and plasma from 20 normal, ovulatory women who were undergoing in vitro fertilization and were therefore stimulated with human menopausal gonadotropin/human chorionic gonadotropin. Serum estradiol (E2) and FF E2 and progesterone (P) were also determined and correlated with FF and plasma renin activity. FF renin activity was significantly higher than plasma renin activity (55.8 +/- 7.9 versus 3.8 +/- 0.7 ng of angiotensin I/ml/hour, P less than 0.001). Positive correlations were found between FF E2 and renin activity (r = 0.53, P less than 0.05). There was also a correlation between plasma renin activity and serum E2 (r = 0.69, P less than 0.05). No correlation was present between FF P and renin activity. The high renin-like activity present in FF after stimulation with gonadotropins could be involved in the mechanism of angiogenesis and may play an important role in events related to reproductive processes.

Abstract: The pattern of periovulatory and luteal phase levels of serum estradiol (E2) and progesterone (P) were compared between 8 conception and 28 nonconception cycles of patients undergoing in vitro fertilization (IVF). Ten additional women served as control subjects and did not undergo follicular aspiration. Follicle growth was induced with an individualized Pergonal (human menopausal gonadotropin) regimen, and laparoscopy was performed 36 hours after human chorionic gonadotropin administration. The length of the luteal phase did not differ significantly among the three groups and was between 14 and 15 days in duration. When IVF conception cycles were compared with nonconception cycles, although no difference in the number of large follicles was observed (4.25 +/- 0.45 versus 3.6 +/- 0.25), the patterns of E2 and P differed significantly. Daily serum E2 levels tended to be higher in the periovulatory phase in conception cycles when compared with nonconception cycles, and were significantly (P less than 0.05) higher in the early, mid, and late luteal phases. Serum P levels were significantly higher (P less than 0.05) in conception cycles from the midluteal phase onward. A decline in both serum E2 and P in the midluteal phase in conception cycles suggested some degree of corpus luteum deficiency. It is suggested that high E2 levels in the periovulatory phase may be an indicator of better follicular development under human menopausal gonadotropin stimulation and that the deficiency observed in the late luteal phase is overcome with the establishment of pregnancy.

Abstract: One hundred six women suffering from obstructive tubal disease not corrected by previous surgery were treated in an in vitro fertilization (IVF) program. Ovulation was induced by 3 amps of human menopausal gonadotropin (HMG)/day starting on the third day of the cycle for 5 days. In most of the patients the regimen was continued for another 1-3 days, depending on the individual's ovarian response (mean, 20 +/- 5 amps/cycle). Monitoring consisted of daily follicular ultrasonography and serum estradiol measurements. Human chorionic gonadotropin (HCG), 10,000 IU, was administered when more than two large (1.5 to 1.8 cm in diameter) follicles were visualized. Using this regimen, a mean of five follicles per woman was aspirated, from which a mean of 3.9 ova was recovered. The oocytes were pre-incubated for 8 or 24 hr, according to the morphological degree of mucification and dispersal of the oocyte-corona-cumulus complex. Following exposure to washed spermatozoa for 16 hr, a 68% fertilization rate was obtained. Oocytes were transferred into the uterus 48 hr after laparoscopy. Ninety-nine transfers (93% of the women) of 1-8 embryos (mean, 2.9/woman) were performed and resulted in 16 clinical pregnancies. No pregnancies occurred in 14 women transferred with one to three oocytes in the pronuclear stage and only one pregnancy (7.1%) was obtained in 14 women transferred with one cleaved oocyte. Over 70% of the women were transferred with two or more cleaved oocytes: in this group the pregnancy/transfer rate was 21%. Of the pregnant women 5 of 16 (31%) aborted between 6 and 10 weeks of gestation and 1 (6%) had an ectopic pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: A simple apparatus to collect moving sperm by non-traumatic means which can be used for artificial insemination is described. The technique is based on enhancing the process of migration from the seminal fluid into a top-layered artificial medium in an ordinary test tube. This has been achieved by controlling 3 main variables: 1) The dilution of migrated sperm was minimize by using only 0.5 ml of the medium layered onto 1 ml semen; 2) increasing the surface area between these media by turning the test tube from a vertical to almost a horizontal position; 3) stimulating sperm activity by incubation at 37 degrees C under air: 5% CO2 for 30 min. When restored to a vertical position approximately 0.3 ml medium, sufficient for most AIH or IVF procedures, was gently aspirated. The effects of these variables on the rate of sperm migration was tested one at a time, and increments that ranged from 20% to as much as 10-fold were detected. When these 3 variables were optimized and 58 semen specimens analyzed, it was found that motility increased from 42 to 87%, velocity from 24.5 to 27.3 micron per sec, whilst abnormal forms dropped from 37 to 15%. The final concentration of motile sperm was 23 X 10(6)/ml compared to an original mean concentration of 34 X 10(6)/ml, indicating a relative recovery of 68%. Oligoasthenospermic specimens revealed similar changes in sperm motility, velocity, morphology and recovery. However, due to the low initial content of moving sperm (4.8 X 10(6)/ml), their final concentration was also low (2.7 X 10(6)/ml). Such specimens required additional preliminary preparation to increase the sperm concentration prior to the migratory procedure.

Abstract: Correct timing of human chorionic gonadotropin (hCG) administration in induced cycles for in vitro fertilization is of crucial importance to oocyte maturation and normal luteal function. The purpose of this work was to compare the effect of hCG timing on follicular development, oocyte maturation, and fertilization in vitro, as well as on the pattern of luteal phase hormone secretion. Ovulation was induced in 32 normally cycling women by human menopausal gonadotropin (hMG)/hCG administration. In the first group (17 women) 10,000 IU hCG was administered 24 hours after the last injection of hMG and in the second group (15 women) 48 to 72 hours after the last hMG injection. Serum estradiol levels prior to oocyte aspiration were similar in both groups, as were the numbers of large follicles on the day of hCG administration (4.5 +/- 2.3 versus 4.1 +/- 1.9 follicles/woman, respectively). The distribution of oocyte-corona-cumulus complexes was similar in both groups and was comprised of 11% immature, 43% intermediate, and 45% mature complexes. The fertilization rate, however, was significantly (P less than 0.001) reduced in the group treated by delayed hCG injection (57% versus 84%), and the percentage of degenerated oocytes was increased (9% versus 1%). Luteal phase length as well as progesterone and estradiol levels were comparable in both groups. It is concluded that an interval longer than 24 hours between the last injection of hMG and the administration of an ovulatory dose of hCG does not affect follicular and luteal phase serum steroid patterns but may result in a decreased oocyte fertilization rate, possibly due to atretic changes in the follicles.

Abstract: A simple and atraumatic method for concentrating washed, motile spermatozoa from normal and subnormal semen specimens is described. It incorporates a modified technique of centrifugation in which sperm are spun onto a soft, fluid cushion, thereby minimizing mechanical damage. Following initial semen dilution to 5-10 ml in an artificial medium, the mixture is transferred to a test tube and layered onto 1 ml of oily contrast medium (Lipiodol). After centrifugation at 300 g for 10 min all but 0.5 ml of the supernatant is discarded, and the unshaken test tube is incubated at 37 degrees C for 15-20 min, during which time the motile sperm migrate into the upper 0.5 ml. After this incubation, 0.3 ml of the upper layer is removed which is sufficient for most IVF and AIH purposes. It contains concentrated, washed, motile spermatozoa that are free of debris and most abnormal forms. No change in pH or osmolarity and no diffusion of any iodine from the oil base into the top layer were detected.

Abstract: One hundred seventeen oocyte-cumulus-corona cell complexes (CCCs) were obtained from 15 women undergoing in vitro fertilization after human menopausal gonadotropin/human chorionic gonadotropin follicular stimulation. In each woman, five oocyte-CCCs were left intact, and the remaining one to five oocytes were freed of their CCCs by hyaluronidase (300 IU/ml) dispersal treatment. Dispersal time for individual CCCs correlated well with their degree of mucification and was significantly shorter in 20 mature CCCs as compared with 24 intermediate CCCs (1.20 +/- 0.05 versus 2.35 +/- 0.13 minutes, respectively; P less than 0.001). Oocyte maturation at collection and after 8 hours of in vitro incubation was not related to the CCC type present at harvest. Maturation of oocytes progressed regardless of CCC type; so that after 8 hours of preincubation, 73% of the oocytes attained a polar body while 20% were still at the germinal vesicle breakdown stage and 7% at the germinal vesicle stage. Overall, the 44 denuded oocytes and 72 intact oocyte-CCCs were comparable in rate of fertilization (64% versus 68%) and subsequent cleavage (75% versus 82% of all fertilized oocytes). It is concluded that in human menopausal gonadotropin-stimulated cycles, asynchrony between individual CCC mucification and oocyte maturation may occur, and that the absence of CCC does not seem to affect in vitro fertilization and cleavage rates.

Abstract: A procedure for continuous culture of rat conceptuses during organogenesis with a number of advantages over existing methods has been established. In this method, rat conceptuses of pregnancy Day 10 (embryonic age 9.5 days; Witschi Stage 13) with embryos at pre- or early somite neurula stage were cultured for 96 h in roller bottles fitted with New Brunswick swivel caps. These caps have 5 inlets which permit continuous gassing of culture bottles and withdrawal of samples or supply of growth medium. The culture medium used in this study was immediately centrifuged, heat-inactivated fresh male rat serum. Continuous gassing of roller bottles with humidified gas mixtures of 5% CO2 and increasing O2 concentrations (5, 20, 40 and 95%), and balanced N2 provided optimal progressive conceptus development and differentiation. The average pO2 of the medium rose from 73.4 to 427.3 mm Hg, while the pCO2 and pH remained relatively stable. During the 96-h culture period, growth and differentiation of conceptuses were considerable, reaching Witschi Stage 27/28. Cultured embryos developed 48-52 somites with extensive differentiation of various organs: brain and sensory organs, heart and circulatory system, limb bud and hepatic prominence, and numerous internal visceral organs. Embryonic DNA and protein contents increased 100- to 200-fold from the initial values. Therefore, this improved procedure with periodic progressive increases in pO2 and stable low pCO2 and physiologic pH in the medium permits growth and differentiation of rat conceptuses in vitro over a prolonged period of time.

Abstract: The production of 17 beta-estradiol and progesterone (Prog) by human granulosa-luteal cells obtained from 24 aspirated follicles of 11 women undergoing laparoscopy in an in vitro fertilization program was studied. Follicular growth was stimulated with an individualized human menopausal gonadotropin regimen begun on either day 1 (group I; n = 5) or day 3 (group II; n = 6) of the menstrual cycle, and laparoscopy was performed 36 h after hCG administration. Granulosa-luteal cells were cultured for 2 h in culture medium alone or in the presence of either pregnenolone (10(-7) M) or testosterone (10(-7) M). Aromatase activity was present in the granulosa-luteal cells, as evidenced by a significant (P less than 0.001) increase in E2 production in the presence of testosterone. The addition of pregnenolone did not augment Prog production. Granulosa-luteal cells derived from Group II patients produced significantly (P less than 0.001) more Prog than those derived from group I patients. In addition, group II granulosa-luteal cells associated with mature oocyte-coronacumulus complexes produced significantly (P less than 0.001) more Prog than those in group I. Fertilization and pregnancy correlated with Prog production, in that granulosa-luteal cells associated with oocytes that were fertilized produced significantly (P less than 0.001) less Prog than those associated with nonfertilized oocytes. Granulosa-luteal cells from the 2 patients in this series who conceived demonstrated a further significant (P less than 0.02) reduction in Prog production. It appears that administration of human menopausal gonadotropin early (day 1) in the follicular phase results in incomplete maturation of the granulosa cells. Furthermore, the optimal oocyte, in terms of successful fertilization, may be one derived from a follicle that has undergone appropriate stimulation resulting in adequate maturation, but has not surpassed that point.

Abstract: Ovarian ultrasonography is a new diagnostic technique which has become almost essential in monitoring ovulation. Recent improvements in ultrasound technology have allowed for accurate assessment of the number and size of the developing follicles and their rate of growth, as well as ovulation and postovulatory events. In the nonstimulated cycle follicular size correlates well with optimal maturation and there is a linear correlation between follicular diameter and plasma estradiol (E2 levels). In stimulated cycles, because of asynchrony of various recruited cohorts of follicles, these rules are not as steadfast. These observations indicate that when there is endogenous gonadotropic activity, follicular growth stimulated by human menopausal gonadotropins (hMG) does not develop synchronously. The aim should be, therefore, not only to improve the monitoring system but mainly to synchronize the cohorts of follicles recruited for development in any one cycle by a better regimen for ovulation induction. In view of the high success rate of hMG treatment in patients without endogenous gonadotropin secretion, it is tempting to speculate that inducing similar conditions in women with endogenous gonadotropin production may have a significant change in the pattern of follicular development in their conception rate.

Abstract: These studies were undertaken to explore the roles of both hCG and PRL in the modulation of early luteal function in the human. Human granulosa-luteal cells isolated during cycles stimulated by human menopausal gonadotropin hCG were obtained at the time of follicle aspiration and cultured to determine the effects of hCG and PRL on both progesterone and hCG receptor binding. Progesterone production by hCG-stimulated granulosa-luteal cells was increased 3.5-fold over unstimulated levels after 120 h, with maximal stimulation at hCG concentrations greater than 1 IU/ml. [125I]hCG binding to granulosa luteal cells was increased 3-fold in cells cultured with hCG (10 IU/ml) after both 48 h (P less than 0.03) and 96 h (P less than 0.02) in culture. hCG (1 IU/ml) stimulated a significant increase in progesterone production above basal levels after 72 h of culture, which continued to increase until 96 h of culture; 20 alpha-dihydroprogesterone (20 alpha-OH progesterone) production also was increased by hCG (1 IU/ml) at 72 h of culture, but unlike progesterone production, showed no further increase. In both the presence and absence of hCG, granulosa-luteal cells cultured with PRL (100 ng/ml) produced significantly more 20 alpha-OH progesterone (P less than 0.04 and P less than 0.02, respectively) after several days than cells cultured without PRL. In addition, progesterone production in the presence of hCG (10 IU/ml) decreased significantly (P less than 0.04) as 20 alpha-OH progesterone levels increased. Equivalent amounts of [125I]hCG were bound by human granulosa-luteal cells cultured with and without PRL (100 ng/ml). These results show that cultured human granulosa-luteal cells are responsive to hCG, with parallel increases in both progesterone production and [125I]hCG receptor binding. The presence of PRL (100 ng/ml) had no effect on [125I]hCG binding. In both the presence and absence of hCG, PRL resulted in an increase in 20 alpha-OH progesterone production and, in the presence of hCG (10 IU/ml), a decrease in progesterone production after several days in culture.

Abstract: In the present study the interaction of aminoglycoside antibiotics with oxytocin and prostaglandin E2 on the contractility of the rat uterus was investigated. It was found that oxytocin and prostaglandin E2 were able to completely reverse the uterine relaxing effect of aminoglycoside antibiotics and when administered concomitantly they exert a protective action. These findings support the view that aminoglycoside antibiotics affect superficially bound extracellular calcium only, since oxytocin and prostaglandins primarily act on intracellular calcium.

Abstract: In the present study the inhibitory effect of five aminoglycoside antibiotics on the contractility of rat uterus was investigated. We found that all the antibiotics tested inhibited the contractility of the isolated rat uterus and only quantitative differences existed among them. The inhibitory potency of the aminoglycoside antibiotics appears to be as follows: sisomicin greater than gentamicin greater than tobramycin greater than dibekacin greater than amikacin. Calcium not only has the ability to restore the normal contractility of the uterus but also to exert protective action against the inhibitory effect of aminoglycoside antibiotics on the uterus contractility.

Abstract: The effect of serum from patients with severe chronic renal failure on the motility of ejaculated spermatozoa was objectively evaluated by the multiple exposure photography method. Uremic serum was not found to have a deleterious effect on the motility of ejaculated spermatozoa. On the contrary it caused a significant increase of spermatozoal velocity and a slighter increase in the percentage motility. These effects cannot be only attributed to the lowering of seminal viscosity and to the reduction of sperm concentration, but to various substances contained in blood serum.

Abstract: In order to investigate the effect of fresh blood serum and normal saline on the motility of ejaculated spermatozoa, semen samples were collected from 7 fertile males, aged 25-39 yrs. Each sample was divided into 5 aliquots: one aliquot remained undiluted, and fresh blood serum from fertile men and normal saline were added to the others to produce 1:1 and 3:1 dilutions. Spermatozoal motility was evaluated objectively, every hour and for 4 h after ejaculation, by the multiple exposure photography method. It was found that serum from fertile men as well as normal saline caused a significant increase of spermatozoal velocity. However, spermatozoal velocity under the influence of serum from fertile men changed with time in a different way than in the undiluted aliquot and those diluted with normal saline. Thus, the beneficial effect of fresh blood serum on spermatozoal velocity, cannot only be attributed to the lowering of seminal viscosity and the reduction of sperm concentration caused by dilution, but to various substances present in blood serum.