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Research question Ovarian stimulation during IVF cycles involves close monitoring of oestradiol, progesterone and ultrasound measurements of follicle growth. In contrast to blood draws, sampling saliva is less invasive. Here, a blind validation is presented of a novel saliva-based oestradiol and progesterone assay carried out in samples collected in independent IVF clinics. Design Concurrent serum and saliva samples were collected from 324 patients at six large independent IVF laboratories. Saliva samples were frozen and run blinded. A further 18 patients had samples collected more frequently around the time of HCG trigger. Saliva samples were analysed using an immunoassay developed with Salimetrics LLC. Results In total, 652 pairs of saliva and serum oestradiol were evaluated, with correlation coefficients ranging from 0.68 to 0.91. In the European clinics, a further 237 of saliva and serum progesterone samples were evaluated; however, the correlations were generally poorer, ranging from –0.02 to 0.22. In the patients collected more frequently, five out of 18 patients (27.8%) showed an immediate decrease in oestradiol after trigger. When progesterone samples were assessed after trigger, eight out of 18 (44.4%) showed a continued rise. Conclusions Salivary oestradiol hormone testing correlates well to serum-based assessment, whereas progesterone values, around the time of trigger, are not consistent from patient to patient.

Study question Is this incidence of early stage abnormal cleavage events different between embryos created following ICSI compared with IVF? Summary answer Embryos derived from ICSI are more likely to exhibit abnormal cleavage compared with those from IVF. This difference is most marked in women ≥35 years. What is known already Time lapse imaging (TLI) has been instrumental in allowing detailed annotation of early embryo development to provide an objective aid for embryo selection in ART cycles. Amongst several abnormal cleavage events reported, rapid cleavage and multichotomous mitosis/direct cleavage, during the first days after fertilisation have been demonstrated to be associated with lower blastulation rates, reduced implantation potential, increased aneuploidy and poor pregnancy outcomes. With ICSI being utilised commonly, and being the insemination method of choice in some clinics, the incidence of abnormal cleavage was investigated in association with insemination method, ICSI or IVF. Study design, size, duration The incidence of abnormal cleavage events was evaluated in a large multicentre retrospective analysis of 36,671 embryos from 6689 patients treated in 8 IVF clinics enabled with time lapse imaging, between 2011 - 2019. This constituted 10931 IVF embryos and 25740 ICSI embryos. Participants/materials, setting, methods Following ICSI or after IVF fertilisation check, embryos were time-lapse imaged every 10 minutes and annotated using the EmbryoScope. Second cell cycle durations were calculated as follows: time to reach 3-cell (t3) from 2-cell (t2) (t3-t2 = cc2). These were analysed using a welch t-test as three groups of abnormal cleavage: direct cleavage/trichotomous mitosis (DC) - where cc2=0 hours(h), rapid cleavage within 2h (R2) - where 0 Main results and the role of chance The incidence of DC, R2 and R5 in the whole cohort of embryos was 5%, 8% and 9% respectively. In the subpopulation of IVF embryos the incidence of DC, R2 and R5 was 4%, 8% and 9% respectively. In the subpopulation of ICSI embryos the incidence of DC, R2 and R5 was 6%, 8% and 9% respectively. The incidence of DC was significantly higher in ICSI embryos compared with IVF (p Male-factor diagnoses showed no significant differences in abnormal cleavage values between ICSI or IVF in all three abnormal cleavage categories. Limitations, reasons for caution This analysis could not control for all potential confounders therefore it is possible that the increased abnormal cleavages observed in this investigation are a result of another, or combination of factors. Despite quality assurance programs being in place across all clinics, there is a risk of annotation bias. Wider implications of the findings: There is a higher incidence of early abnormal cleavage in embryos derived from ICSI, particularly in those from women of increased age and this research may help elucidate the reasons for this and add to the debate regarding the appropriateness of the increasing use of ICSI. Trial registration number Not applicable

Study question Are safety and effectiveness of Ovaleap® (follitropin alfa), and Gonal-f®, comparable in one treatment cycle of ART in routine clinical practice? Summary answer Safety in terms of incidence proportions of OHSS and OHSS severity, as well as pregnancy and live birth rates, were similar between Ovaleap® and Gonal-f®. What is known already Ovaleap® (Theramex), a r-hFSH, is a biosimilar medicinal product to Gonal-f® (Merck). As a biosimilar, it went through a rigorous series of physio-chemical, in vitro, in vivo tests and confirmatory Phase I and III studies, to demonstrate similarity/equivalence in quality, safety and efficacy to the reference medicinal product, per the European Medicines Agency (EMA) guidelines. Ovaleap® was approved by the EMA in 2013 for use at the same dose and for the same therapeutic indications as Gonal-f®. Further outcome data from a broader patient population on safety and live birth outcomes provides clinically important insights on newly introduced FSH medicines. Study design, size, duration SOFIA (Safety of Ovaleap® Follitropin alfa in Infertile women undergoing superovulation for Assisted reproductive technologies) was a multi-national, comparative, non-interventional, prospective cohort study. The study was performed at 56 centers specializing in ART from six European countries, (Belgium, France, Germany, Italy, Spain, and the United Kingdom) from January 2017 to September 2019 and comprised of 817 infertile women undergoing controlled ovarian hyperstimulation in one treatment cycle for ART Participants/materials, setting, methods The study population comprised of infertile women undergoing controlled ovarian hyperstimulation for ART, who were administered Ovaleap® or Gonal-f® and were naïve to any FSH containing products. Eligible patients were enrolled at a ratio of approximately 1:1, both within and between countries. They were followed up to 30 days after the last FSH dose administration. Women who had a confirmed clinical pregnancy were followed until the end of the pregnancy or until delivery. Main results and the role of chance A total of 408 and 409 women who were administered Ovaleap® or Gonal-f®, respectively, were eligible for analysis. A total of 382 patients (94%) in the Ovaleap® and 390 patients (95%) in the Gonal-f® cohort completed FSH treatment (up to oocyte maturation triggering), respectively. The two cohorts were generally similar with regard to demographic and baseline characteristics. The incidence proportion of OHSS was 5.1% (95% CI: 3.4, 7.7) in the Ovaleap® and 3.2% (95% CI: 1.9, 5.4) in the Gonal-f® cohort. This difference in OHSS incidence proportion between the two cohorts was not statistically significant neither before (p = 0.159) nor after univariate adjustment for each potential confounder (p > 0.05). The incidence proportion of OHSS severity grades was similar in the two treatment groups (3.4% versus 2.0% for Grade I, 1.2% versus 1.0% Grade II, and 0.5% versus 0.2% Grade III, in the Ovaleap® and Gonal-f® cohorts, respectively) and without a significant statistical difference (p = 0.865, for each grade). Among patients who had embryo transfer, clinical pregnancy rates were 33% and 31%, live birth rates 27% and 26% in the Ovaleap® and Gonal-f® cohorts, respectively. Limitations, reasons for caution Since treatment was non-randomised, the study may have been susceptible to selection bias. This was addressed at both the design stage, by balancing recruitment to a 1:1 ratio for Ovaleap® and Gonal-f® treatments, and also at the analysis stage in which, a univariate analysis was performed. Wider implications of the findings: Findings from this first large European prospective comparative real-world SOFIA study demonstrated that effectiveness (pregnancy and delivery rates) and safety (risk and severity of OHSS), were similar between Ovaleap®and Gonal-f® treatments. Ovaleap, a biosimilar r-hFSH is therefore a suitable option for follicular stimulation in routine clinical practice. Trial registration number EUPAS17328

Controlled ovarian stimulation (COS) with gonadotropins to produce multiple follicular development and high-quality oocytes is the cornerstone of assisted reproductive technology. Today, recombinant human follicle-stimulating hormone (r-hFSH) is widely used for COS. A long-acting r-hFSH and a combination of r-hFSH and recombinant human luteinizing hormone have recently become available. Formulations of purified urinary FSH with or without luteinizing hormone activity (provided by human chorionic gonadotropin) are also available. COS protocols can now be individualized to optimize efficacy and safety - defined as singleton pregnancies with a low incidence of ovarian hyperstimulation syndrome. This is facilitated by an estimation of ovarian response using the antral follicle count and/or serum anti-Müllerian hormone levels; anti-Müllerian hormone is viewed as the most reliable single marker. However, an efficient management strategy for poor responders to COS is still required. Developments in biomarkers and other techniques for accurate identification of viable oocytes and embryos and optimal uterine receptivity are expected.

The main objective of this user experience testing study was to evaluate the impact of human factors on the use of a disposable pen containing follitropin alfa by patients and nurses with special focus on the convenience, safety and ease of use, in different types of stimulation protocols.Infertile women trying to conceive, and specialist nurses were recruited across 6 European countries. In total 18 patients and 19 nurses took part in the testing, which included both nurse-patient pairings and in-depth interviews. A standardized list of expected and pre-defined critical steps according to the Instructions for Use (IFU), was used to assess the correct handling of the pen.During the user experience testing, no critical errors, related to the use of the pen, which could affect the success of the injection process were identified. In general, both nurses and patients found the pen very easy to learn, use and would be confident using the pen for self-injection. Nurses also found the pen very easy to train the patients.The study provides valuable information on the pen from both patient and nurse perspectives in different simulated scenarios reflecting standard practice.

In this open-label study, women aged 36–40 years undergoing ovarian stimulation were randomized to recombinant human FSH (rhFSH) plus recombinant human luteinizing hormone (rhLH) from stimulation day 1 (group A; n = 103), or rhFSH alone (days 1–5) followed by rhFSH plus rhLH from day 6 (group B; n = 99). The primary objective was equivalence in number of oocytes retrieved per patient. The mean (±SD) number of oocytes retrieved was 9.7 (±6.9) in group A and 10.9 (±6.5) in group B; the estimated difference between groups (−1.28 oocytes [95% confidence interval: −3.15 to 0.59]) did not reach the predefined limit of equivalence (±3 oocytes). The study's primary objective was therefore not met. In both groups, a mean (±SD) of 1.9 (±0.6) embryos were transferred per patient. Implantation rates were 24.7% in group A and 13.3% in group B. Clinical pregnancy rates per started cycle and per embryo transfer were 31.6% and 34.4% in Group A, 17.2% and 18.9% in Group B. Ovarian hyperstimulation syndrome was reported in four (group A) and five (group B) patients. The potential benefit of initiating LH supplementation earlier during ovarian stimulation in older women is of interest, warranting further exploration.

The prediction of extremes of ovarian response to stimulation and the irreversibility of reduced ovarian reserve remain important clinical and basic science research issues of IVF treatment. Recommending commencement of ovarian stimulation using any of the available exogenous compounds without knowledge of individual ovarian potentials is simplistic and dangerous because of the possible adverse consequences for the woman. The identification of groups of patients likely to benefit from one protocol than another is central to the workup process of IVF. Determining the agents for ovarian stimulation as well as the combination of them, the daily dose and duration according to some background information should be seen as the way to enhance safety and cost-effectiveness. This discussion paper aims to introduce the concept of individualized ovarian stimulation in routine clinical practice and to generate interest for tailored stimulation protocols.

Objective: To investigate the effectiveness of treatment with transdermal testosterone gel (TTG) before controlled ovarian stimulation (COS) using GnRH antagonist multiple-dose protocol (MDP) in low responders undergoing IVF/intracytoplasmic sperm injection (ICSI). Design: Prospective randomized controlled trial. Setting: University-affiliated infertility clinic. Patient(s): A total of 110 low responders, who were defined as patients who failed to produce %3 follicles with a mean diameter of R16 mm with the result that %3 oocytes were retrieved despite the use of a high gonadotropin dose (>2,500 IU) in a previous failed IVF/ICSI cycle. Intervention(s): Patients were randomized into TTG pretreatment group or control group. For TTG pretreatment group, 12.5 mg TTG were applied daily for 21 days in the cycle preceding COS for IVF. Main Outcome Measure(s): COS results and IVF outcome. Result(s): There were no differences in patients’ characteristics between the two groups. Total dose and days of rhFSH used were significantly fewer in the TTG pretreatment group than in the control group. The numbers of oocytes retrieved, mature oocytes, fertilized oocytes, and good-quality embryos were significantly higher in the TTG pretreatment group. Embryo implantation rate and clinical pregnancy rate per cycle initiated also were significantly higher in the women pretreated with TTG. No patient reported adverse effects attributed to TTG use. Conclusion(s): TTG pretreatment might be beneficial in improving both response to COS and IVF outcome in low responders undergoing IVF/ICSI. (Fertil Steril� 2011;95:679‐83. � 2011 by American Society for Reproductive Medicine.)

This pilot study compared the efficacy and safety of two simple dosing algorithms, one based on anti-Müllerian Hormone (AMH) and the other on the antral follicle count (AFC), to determine the starting dose of recombinant FSH (rFSH) for ovarian stimulation in 348 women. Patients were randomized to a predefined AMH- or AFC-based algorithm. The proportion of cycles with the desired response was similar when rFSH dose was determined using AMH or AFC (35.2% versus 28.4%). There was a significant difference between the groups in the proportion of cycles with a hyperresponse (8.6% and 17.4%, but the incidence of ovarian hyperstimulation syndrome was similar (1.1% and 4.6%). There were no significant differences between two groups in outcomes, including implantation (19.3% versus 19.0%), clinical pregnancy (38.0% versus 46.9%), multiple pregnancy (16.5% versus 15.2%) and miscarriage (7.0% versus 8.3%). However, statistically significant differences in ovarian response were evident among the AMH and AFC subgroups: for AMH, Desired and Hypo; for AFC, Hypo and Hyper. This pilot study provides information for developing protocols to further validate the use of either AMH or AFC to guide the starting dose of rFSH in ovarian stimulation. The ideal outcome for couples undergoing IVF treatment is the birth of a healthy baby. One factor that might influence this is retrieving an adequate number of eggs, which are obtained using various treatment protocols. A group of drugs called gonadotrophins have been used for more than 20years to stimulate the ovaries to produce eggs. However, the dose to start treatment has not been clearly defined. A few studies have looked at ways to use the best gonadotrophin dose for each woman, but to be useful in the clinic any approach needs to be simple and easy to use. This study compared the effectiveness and safety of two simple approaches to determining the starting dose of recombinant FSH (rFSH) for ovarian stimulation in women undergoing IVF. One was based on the concentration of a hormone secreted by developing eggs (anti-Müllerian hormone; AMH) and the other on the number of developing follicles (antral follicle count; AFC). The number of cycles achieving the desired response in terms of number of eggs was similar when rFSH dose was guided using AMH or AFC, and the incidence of ovarian hyperstimulation syndrome was also similar. In addition, rates of clinical pregnancy, multiple pregnancy and miscarriage did not differ between the two groups. However, patients with low AMH concentrations or low AFC had a poor response to ovarian stimulation. This pilot study provides useful information from which new studies can further assess these approaches to personalizing treatment during IVF.

In their quest for a child, infertile couples embark on a journey that is full of expectations and hopes. Over recent years, treatment procedures for assisted conception have become safer and more efficient. However, couples undergoing treatment can still experience some degree of emotional stress due to disappointment if pregnancy is not achieved, or if treatment cycles may have to be cancelled due to a low- or hyper-response. Strategies aimed at minimizing the variability of ovarian response or overall treatment outcome can be expected to significantly reduce this emotional stress. New developments have led to the production of follitropin alfa filled by mass. This is a highly consistent FSH preparation improving the consistency of ovarian response and reducing the risk of cycle cancellation. The impact of this new FSH preparation for assisted reproduction treatments is discussed in this review.

In this randomized, controlled, open-label, phase IV study, ovarian response after a follitropin alfa starting dose determined by the CONSORT calculator was compared with a standard dose (150 IU). Normo-ovulatory women (aged 18-34 years) eligible for assisted reproductive techniques were recruited (23 centres: nine European countries and Chile); 200 women were randomized (CONSORT [n = 96]; standard dosing [n = 104]). Significantly lower mean daily (121.5 versus 167.4 IU; P < 0.001) and total (1288.5 versus 1810.0 IU; P < 0.001) doses of follitropin alfa were administered in the CONSORT group. Clinical pregnancy rates were CONSORT (36.0%) and standard dosing (35.5%); estimated difference (confidence interval 0.6%; -13.5 to 14.6). Ovarian hyperstimulation syndrome occurred in 6.3% and 12.5% of patients in the CONSORT and standard-dosing groups, respectively. The primary efficacy analysis found a significantly lower mean [SD] number of oocytes retrieved in the CONSORT (10.0 [5.6]; P = 0.037) versus standard-dosing group (11.8 [5.3]). Although the CONSORT calculator was statistically inferior to standard dosing in the number of oocytes retrieved, clinical pregnancy rates (fresh embryo transfers) were similar in both groups, and incidence of ovarian hyperstimulation syndrome was lower in the CONSORT group.

Gonadotrophin-releasing hormone (GnRH) antagonists have recently been introduced into clinical practice. They appear to offer a promising alternative to the long-established GnRH agonist regimens for prevention of a premature LH surge during ovarian stimulation for assisted reproductive techniques. Clinical outcomes achieved with antagonists are comparable with those of a long GnRH agonist protocol, while treatment times and gonadotrophin requirements are reduced and safety is improved. In particular, the antagonists appear to be associated with a lower risk of ovarian hyperstimulation syndrome (OHSS) than do agonists. Patient surveys suggest a preference for antagonist over agonist treatment cycles. These benefits suggest that GnRH antagonists have the potential to replace agonists as the treatment of choice in ovarian stimulation for assisted reproductive techniques. Two agents, cetrorelix and ganirelix, are currently in clinical use. Cetrorelix is available in single- and multiple-dose formulations, offering increased flexibility compared with ganirelix.

BACKGROUND A prospective randomized study was carried out in two centres to compare the number of oocytes retrieved after two different starting doses of recombinant human FSH (rhFSH) (Gonal-F) in women undergoing ovarian stimulation for IVF/intracytoplasmic sperm injection (ICSI) cycles using the multiple dose regimen of the gonadotrophin-releasing hormone (GnRH) antagonist cetrorelix (Cetrotide) to prevent induction of the premature LH surge. METHODS Sixty women were randomized to receive rhFSH 150 IU ('low'), and 60 women to receive rhFSH 225 IU ('high') as the starting dose for the first 5 days of stimulation. From stimulation day 6 and onwards, including the day of human chorionic gonadotrophin (HCG) administration, the women received 0.25 mg of cetrorelix as a daily dose. The primary endpoint was the number of oocytes retrieved. RESULTS The mean number (+/- SD) of oocytes was 9.1 +/- 4.4 and 11.0 +/- 4.6 in the 'low' and 'high' groups respectively (P = 0.024). The mean number of 75 IU ampoules of rhFSH was significantly lower in the 'low' group (23.0 +/- 6.3 versus 30.5 +/- 5.6, P < 0.0001). The ongoing pregnancy rate per started cycle and per embryo transfer were 25.9 and 28.8% versus 25.4 and 26.8% respectively in the 'low' and 'high' rhFSH groups (P = NS). CONCLUSIONS When using a starting dose of 225 IU rhFSH combined with the multiple dose of 0.25 mg cetrorelix from stimulation day 6, significantly more oocytes were obtained than with a starting dose of 150 IU rhFSH.

This prospective, double-blind, randomized, multicentre study compared the efficacy and safety of recombinant human follicle stimulating hormone (r-hFSH; Gonal-F((R))) versus highly purified urinary FSH (u-hFSH HP; Metrodin((R)) HP) in women undergoing ovarian stimulation for in-vitro fertilization, including intracytoplasmic sperm injection. A total of 278 patients began a long gonadotrophin-releasing hormone agonist protocol, then 139 received r-hFSH and 139 u-hFSH HP, 150 IU/day administered s.c., for the first 6 days of treatment. On day 7, the dose was adjusted, if necessary, according to ovarian response. Human chorionic gonadotrophin (HCG, 10 000 IU, s.c.) was administered once there was more than one follicle 18 mm in diameter and two others >/=16 mm. Oocyte retrieval was performed 36-38 h after HCG injection: 128 patients (92%) receiving r-hFSH and 113 (81%) receiving u-hFSH HP had at least one oocyte retrieved. Among patients receiving r-hFSH, there was a significantly higher mean (+/- SD) number of oocytes retrieved (11.0 +/- 5.9 versus 8.8 +/- 4.8 with u-hFSH HP; P = 0. 002) and mean number of embryos obtained (5.1 +/- 3.7 versus 3.5 +/- 2.9 with u-hFSH HP; P = 0.0001). With r-hFSH, significantly fewer FSH treatment days (11.7 +/- 1.9 versus 14.5 +/- 3.3) and 75 IU ampoules (27.6 +/- 10.2 versus 40.7 +/- 13.6) were required than with u-hFSH HP (P = 0.0001). Embryo replacement on day 2-3 after oocyte retrieval resulted in 36 liveborn children in the Gonal-F((R)) group and 33 in the Metrodin HP((R)) group (not significant). There were seven cases (5.0%) of ovarian hyperstimulation syndrome in the r-hFSH group and three (2.2%), in the u-hFSH HP group (not significant). It is concluded that r-hFSH is more effective than u-hFSH in inducing multiple follicular development.

Objective: To compare the efficacy and the safety of recombinant human FSH (hFSH) with urinary hFSH for stimulating follicular development in women undergoing IVF-ET. Design: Multicenter, prospective, randomized, open, parallel group, clinical study. Setting: Eight European academic IVF units and one private IVF unit. Patient(s): Infertile female patients aged 18 to 38 years suffering from tubal disease, mild endometriosis, or unexplained infertility. Intervention(s): Pretreatment with buserelin acetate was followed by recombinant or urinary hFSH treatment started at an initial dose of 225 IU FSH/d. Dose adjustment was allowed after 5 days of FSH. After administration of hCG, a standard IVF-ET procedure was performed. Main Outcome Measure(s): Follicular development, oocyte retrieval, fertilized oocytes, duration and dose of FSH, and pregnancy. The hypothesis formulated before the study was that no difference was expected between the two FSH preparations. Result(s): Sixty patients were treated with recombinant hFSH and 63 with urinary hFSH. The mean number (±SD) of growing follicles (mean diameter >10 mm) was 10.3 ± 4.9 and 11.2 ± 5.2, of follicles (mean diameter >14 mm) was 7.8 ± 3.6 and 9.2 ± 4.5, of retrieved oocytes was 9.3 ± 5.0 and 10.7 ± 5.3, and of fertilized oocytes was 5.6 ± 3.8 and 6.5 ± 4.3, for recombinant and urinary hFSH, respectively. The duration of FSH treatment was 9.9 ± 2.3 and 9.4 ± 1.8 days and the average total dose was 2270 ± 714 and 2095 ± 591 IU of FSH, for recombinant and urinary hFSH, respectively. Thirteen pregnancies were recorded in the recombinant hFSH group and 11 in the urinary hFSH group. Nine patients delivered 13 live infants in the recombinant hFSH group and eight delivered 13 live infants in the urinary hFSH group. In terms of safety, no difference was recorded between the groups and no anti-FSH antibodies were found in any of the patients. Conclusion(s): This clinical study shows that recombinant hFSH is as safe and effective as urinary hFSH in stimulating ovarian follicular development.

The Sixth Evian Annual Reproduction (EVAR) Workshop Group Meeting was held to evaluate the impact of IVF/intracytoplasmic sperm injection on the health of assisted-conception children. Epidemiologists, reproductive endocrinologists, embryologists and geneticists presented data from published literature and ongoing research on the incidence of genetic and epigenetic abnormalities and congenital malformations in assisted-conception versus naturally conceived children to reach a consensus on the reasons for potential differences in outcomes between these two groups. IVF-conceived children have lower birthweights and higher peripheral fat, blood pressure and fasting glucose concentrations than controls. Growth, development and cognitive function in assisted-conception children are similar to controls. The absolute risk of imprinting disorders after assisted reproduction is less than 1%. A direct link between assisted reproduction and health-related outcomes in assisted-conception children could not be established. Women undergoing assisted reproduction are often older, increasing the chances of obtaining abnormal gametes that may cause deviations in outcomes between assisted-conception and naturally conceived children. However, after taking into account these factors, it is not clear to what extent poorer outcomes are due to the assisted reproduction procedures themselves. Large-scale, multicentre, prospective epidemiological studies are needed to investigate this further and to confirm long-term health consequences in assisted-conception children. Assisted reproduction treatment is a general term used to describe methods of achieving pregnancy by artificial means and includes IVF and sperm implantation. The effect of assisted reproduction treatment on the health of children born using these artificial methods is not fully understood. In April 2011, fertility research experts met to give presentations based on research in this area and to look carefully at the evidence for the effects of assisted reproduction treatment on children's health. The purpose of this review was to reach an agreement on whether there are differences in the health of assisted-conception children with naturally conceived children. The researchers discovered no increased risk in birth defects in assisted-conception children compared with naturally conceived children. They found that IVF-conceived children have lower birth weights and higher fat under the skin, higher blood pressure and higher fasting glucose concentrations than naturally conceived children; however, growth, development and cognitive function are similar between groups. A very low risk of disorders of genetic control was observed in assisted-conception children. Overall, there did not appear to be a direct link between assisted reproduction treatment and children's health. The researchers concluded that the cause of some differences in the health of children conceived using assisted reproduction treatment may be due to the age of the woman receiving treatment. Large-scale, research studies are needed to study the long-term health of children conceived using assisted reproduction treatment. © 2013, Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

A prospective, randomized, comparative, assessor-blind study was carried out in two centres to compare the efficacy and safety of recombinant human follicle stimulating hormone (r-hFSH; Gonal-F) versus highly purified urinary FSH (u-hFSH HP; Metrodin HP), both administered s.c. in women undergoing ovarian stimulation for in-vitro fertilization including intracytoplasmic sperm injection (ICSI). A total of 235 patients started a long gonadotrophin-releasing hormone agonist protocol: 119 received r-hFSH and 114 received u-hFSH HP (150 IU/day) for the first 6 days. Two patients were excluded from the study because they mistakenly received the incorrect treatment combination. Human chorionic gonadotrophin (HCG; 10000 IU, s.c.) was administered once there was at least one follicle 18 mm in diameter and two others > or = 16 mm. In all, 119 (100%) and 102 (89%) of the patients respectively in the r-hFSH and u-hFSH HP groups achieved the criteria for HCG. The mean numbers (+/- SD) of oocytes recovered (the primary endpoint) were 12.2 +/- 5.5 and 7.6 +/- 4.4 in the r-hFSH and u-hFSH HP groups respectively (P < 0.0001). However, the number of FSH treatment days (11.0 +/- 1.6 versus 13.5 +/- 3.7) and the number of 75 IU ampoules (21.9 +/- 5.1 versus 31.9 +/- 13.4) used were significantly less (P < 0.0001) in the r-hFSH group than in the u-hFSH HP group. In patients treated using ICSI (63 patients in each group), no difference in oocyte maturation was observed. The mean numbers of embryos obtained were 8.1 +/- 4.2 and 4.7 +/- 3.5 (P < 0.0001), in favour of the r-hFSH group. In the majority of patients (96 and 99% respectively) only one or two embryos were replaced (mean 2.0 +/- 0.2 and 1.9 +/- 0.1 respectively) in the r-hFSH and u-hFSH HP groups. The clinical pregnancy rates per started cycle and per embryo transfer were 45 and 36%, and 48 and 47%, respectively in the r-hFSH and u-hFSH HP groups (not significant). There were six (5.1%) and two (1.7%) cases of ovarian hyperstimulation syndrome respectively. In conclusion, it was found that r-hFSH was more effective than u-hFSH at inducing multiple follicular development. However, the high rate of low ovarian response in the u-hFSH group compared with our general experience was unexpected. The availability of a gonadotrophin with less inter-batch variation would be beneficial for clinicians. r-hFSH seems to fulfil such a requirement.

Objective: To analyze the role of serum LH and of E 2 secretion on IVF-ET outcome in patients stimulated with FSH. Design: Using data from three studies, we analyzed ovarian response to FSH and IVF-ET outcome from two standpoints: [1] serum LH and [2] serum E 2 on the day of hCG administration divided by the number of retrieved oocytes. Setting: Twenty-six academic and private clinical centers. Patient(s): Three hundred twenty-three ovulatory patients eligible for IVF-ET. Intervention(s): Patients were treated with a GnRH agonist and FSH and underwent IVF-ET. Main Outcome Measure(s): Follicular development, embryos, and pregnancy rate (PR). Result(s): Serum LH levels did not correlate significantly with number of oocytes retrieved, E 2 -oocyte ratio, fertilization rate, or PR. Five patient subpopulations were identified on the basis of E 2 -oocyte ratios: 0 to 70 (A), 70 to 140 (B), 140 to 210 (C), 210 to 280 (D), and >280 (E) pg/mL per oocyte (conversion factor to SI unit, 3.671). Pregnancy rates were significantly different, i.e., 5.3%, 31.3%, 18.1%, 23.9%, and 20.4% for groups A, B, C, D, and E, respectively. Groups were not different in terms of baseline characteristics, number of follicles, fertilization rates, and number of embryos transferred. Conclusion(s): Patients with very low levels of LH respond to FSH alone as well as those with higher LH. The E 2 -oocyte ratio is a strong index of success rate.

A multicentre, multinational study was carried out between November 1990 and February 1992 to assess the safety and efficacy of a new highly purified urinary human follicle stimulating hormone (FSH; Metrodin HP) which is practically devoid of luteinizing hormone (LH) activity. Metrodin HP was administered s.c. to stimulate multiple follicular development in women undergoing in-vitro fertilization (IVF) and embryo transfer. A total of 139 women were recruited from 10 participating centres. Of these, 135 underwent pituitary desensitization with a long gonadotrophin-releasing hormone (GnRH) agonist protocol and following determination of ovarian inactivity (mean +/- SD of 12.9 +/- 3.2 days), Metrodin HP s.c. stimulation was started; 122 patients were fully eligible for efficacy analysis and 118 of these (97%) received up to 10,000 IU human chorionic gonadotrophin (HCG) to induce final follicular maturation and timed oocyte recovery. Mean plasma LH concentrations at the beginning of Metrodin HP treatment were 1.6 +/- 0.8 mIU/ml and by the day of HCG administration were significantly (P < 0.001) reduced (1.2 +/- 0.8 mIU/ml). The mean plasma oestradiol and inhibin concentrations on the day of HCG were 6173 +/- 3567 pmol/l and 8.2 +/- 4.4 IU/ml respectively. There was a positive correlation (r = 0.49, P < 0.001) between individual oestradiol and inhibin concentrations on the day of HCG. In the 118 patients who received HCG, the mean number of oocytes recovered was 8.4 +/- 4.7 following stimulation with 36 +/- 10, 75 IU ampoules of Metrodin HP over 12.2 +/- 2.1 days.(ABSTRACT TRUNCATED AT 250 WORDS)

This study was designed to examine ovarian performance, i.e. follicular growth, normal steroidogenesis and luteal phase function, following the administration of multiple increasing doses of human follicle stimulating hormone (FSH) with a constant low dose of luteinizing hormone (LH) in women with isolated hypogonadotrophic hypogonadism. Human menopausal gonadotrophin (HMG) was used in the first treatment cycle, starting with 150 IU of LH and 150 IU of FSH per day, for 7 days. The dose was increased daily with 75 IU of LH and 75 IU of FSH for another 7 days if no response was detected by serial ultrasound measurements and serum oestradiol determinations. In the second treatment cycle, a constant dose of 75 IU of LH (using HMG) was administered per day and up to 150 IU of FSH (using urofollitrophin) was supplemented. If no response was detected after 7 days of treatment, the dose of FSH was increased. For the final stage of ovulation induction, human chorionic gonadotrophin (HCG) was administered in the presence of at least one follicle > 17 mm in diameter but with no more than three follicles > 16 mm in diameter. To verify the adequacy of the luteal phase, a pharmacokinetic/pharmacodynamic study of beta-HCG, oestradiol and progesterone was performed following the second treatment cycle only. Ovarian stimulation using a constant dose of 75 IU of LH and increasing doses of FSH up to 225 IU, resulted in normal follicular growth and hormonal milieu. Both women showed normal luteal phase oestradiol and progesterone production and both women conceived following the second treatment cycle.

Background The Fifth Evian Annual Reproduction (EVAR) Workshop Meeting discussed knowledge regarding contemporary genetics in female reproduction. Methods Specialist reproductive medicine clinicians and geneticists delivered presentations based on published literature and current research. The content of this report is based on the expert presentations and subsequent group discussions that took place during this Workshop. Results Numerous ovarian genes with a role in infertility have been identified. Future challenges for genetic screening of patients, such as those with polycystic ovary syndrome, primary ovarian insufficiency or endometriosis, include the identification of high-throughput strategies and how to apply these findings to infertile patients. The identification of high-quality embryos in IVF using objective technologies remains a high priority in order to facilitate single-embryo transfer. Gene expression profiling of cumulus cells surrounding the oocyte, and proteomic and metabolomic approaches in embryo culture media may significantly improve non-invasive embryo quality assessment. Conclusions The way forward in advancing the knowledge of genes involved in reproduction was considered to be through genome-wide association studies involving large numbers of patients. Establishing international collaboration is required to enable the application of such technologies in sufficient numbers of patients.

To obtain direct evidence for FSH-stimulated paracrine signaling in the ovary, 21-day-old intact or hypophysectomized female Wistar rats received four sc injections of recombinant human FSH (rhFSH; total dose, 16-72 IU) at 12-h intervals. Ovaries were removed 48 h after the first injection to extract total RNA for Northern analysis of 17-hydroxylase/C17-20-lyase (cytochrome P450c17 alpha) mRNA or to isolate thecal/interstitial cells for assessment of basal and hLH-responsive androgen synthesis in vitro. In situ hybridization with a 35S-labeled cytochrome P450c17 alpha cRNA probe confirmed that expression of the cytochrome P450c17 alpha gene was specific to thecal/interstitial cells. The approximately 2.0-kilobase P450c17 alpha mRNA signal in ovarian total RNA from intact animals was increased approximately 5-fold by treatment with rhFSH (total dose, 72 IU) or PMSG (15 IU). This effect was shown to be dose dependent, with a approximately 2-fold increase in response to 16 IU (total dose) rhFSH. P450c17 alpha mRNA levels in isolated granulosa and thecal/interstitial cell total RNA from intact animals were compared to establish which was the principal cellular site of P450c17 alpha mRNA expression. The P450c17 alpha mRNA signal was undetectable in control granulosa cells and only barely discernible after treatment with 72 IU (total dose) rhFSH. In contrast, P450c17 alpha mRNA was abundant in control thecal/interstitial mRNA, and its level was increased 4- to 6-fold by treatment with rhFSH. Treatment of hypophysectomized animals with rhFSH did not consistently alter ovarian P450c17 alpha mRNA levels. During culture for 48 h in serum-free medium, basal androgen (androstenedione plus androsterone) production by thecal/interstitial cells from intact animals was unaffected by treatment with rhFSH in vivo, but hLH-stimulated androgen production by these cells was enhanced approximately 2-fold. Neither basal nor hLH-responsive androgen production by thecal/interstitial cells from hypophysectomized animals was altered by previous treatment with rhFSH in vivo. Treatment of thecal/interstitial cell cultures from both intact and hypophysectomized animals with inhibin (0.1-30 ng/ml), a putative granulosa-derived paracrine factor, did not measurably affect basal androgen synthesis, but potently enhanced LH-responsive androgen synthesis in vitro. Similarly, treatment of thecal/interstitial cell cultures with conditioned medium from FSH-treated granulosa cell cultures significantly enhanced LH-responsive, but not basal, androgen production. We conclude that treatment of pituitary-intact rats with "pure" FSH modulates thecal/interstitial cell androgen synthesis. Granulosa cells, but not thecal cells, possess FSH receptors, and thecal/interstitial cells are the principal ovarian sites of P450c17 alpha expression.(ABSTRACT TRUNCATED AT 400 WORDS)

PURPOSE OF REVIEW: Use of preimplantation genetic diagnosis to improve in-vitro fertilization outcomes is reviewed. RECENT FINDINGS: Many embryos produced in vitro contain chromosomal abnormalities and have little potential for forming a viable pregnancy. The most commonly used method for preimplantation genetic diagnosis involves embryo biopsy on day 3 of development, followed by fluorescence in-situ hybridization analysis of 5-12 chromosomes. However, positive results have been more common with single-cell biopsy and the analysis of nine or more chromosomes, including 15, 16, 21, and 22. Comparative genomic hybridization, array-comparative genomic hybridization, and single-nucleotide polymorphism arrays analyze all chromosomes and, although technically demanding and requiring experience for successful use, improve the selection potential of preimplantation genetic diagnosis and minimize error rates. Recent data suggest that biopsy at the blastocyst stage may allow sampling of representative genetic material without compromising embryo viability. The optimal strategy for aneuploidy screening using preimplantation genetic diagnosis seems to be blastocyst biopsy at 5 days and comprehensive chromosome analysis (comparative genomic hybridization, array-comparative genomic hybridization, single-nucleotide polymorphism array). SUMMARY: The use of preimplantation genetic diagnosis to assist the identification and preferential transfer of healthy euploid embryos should improve implantation rates, reduce miscarriages and trisomic offspring, and ultimately lead to an increase in live birth rates.

To identify baseline characteristics related to successful ovulation induction, data were analysed from oligo- or anovula- tory patients undergoing their first cycle of human recombinant FSH (r-hFSH; follitropin alfa) in a chronic low-dose (75 IU starting dose), step-up protocol in two clinical trials (n = 446). Patients were grouped according to response: group A, ovulated within 14 days (75 IU follitropin alfa); group B, ovulated after 14 days (>75 IU follitropin alfa); group C, not administered human chorionic gonadotrophin (HCG) because of poor response; group D, cycle cancelled due to over-response (HCG not administered); group E, spontaneous ovulation prior to obtaining criteria for administration of HCG. Mean body mass index (BMI) of group A (25.0 kg/m 2 ) was significantly lower than groups B (27.1 kg/m 2 , P < 0.001) or C (28.2 kg/m 2 , P < 0.0001), but similar to group D (24.3 kg/m 2 ). Mean antral follicle count (AFC) of group A was also significantly lower than group C (18.3 versus 22.7; P = 0.018), but not signifi- cantly different from groups B (21.5) or D (19.5); group E had the highest mean AFC (35.7). Comparatively low BMI, low AFC and higher (although still within the normal range) FSH concentration at baseline were associated with successful ovulation induction in infertile women undergoing a chronic low-dose, step-up stimulation protocol. RBMOnline

Background Women have been able to delay childbearing since effective contraception became available in the 1960s. However, fertility decreases with increasing maternal age. A slow but steady decrease in fertility is observed in women aged between 30 and 35 years, which is followed by an accelerated decline among women aged over 35 years. A combination of delayed childbearing and reduced fecundity with increasing age has resulted in an increased number and proportion of women of greater than or equal to 35 years of age seeking assisted reproductive technology (ART) treatment. Methods Literature searches supplemented with the authors' knowledge. Results Despite major advances in medical technology, there is currently no ART treatment strategy that can fully compensate for the natural decline in fertility with increasing female age. Although chronological age is the most important predictor of ovarian response to follicle-stimulating hormone, the rate of reproductive ageing and ovarian sensitivity to gonadotrophins varies considerably among individuals. Both environmental and genetic factors contribute to depletion of the ovarian oocyte pool and reduction in oocyte quality. Thus, biological and chronological ovarian age are not always equivalent. Furthermore, biological age is more important than chronological age in predicting the outcome of ART. As older patients present increasingly for ART treatment, it will become more important to critically assess prognosis, counsel appropriately and optimize treatment strategies. Several genetic markers and biomarkers (such as anti-Müllerian hormone and the antral follicle count) are emerging that can identify women with accelerated biological ovarian ageing. Potential strategies for improving ovarian response include the use of luteinizing hormone (LH) and growth hormone (GH). When endogenous LH levels are heavily suppressed by gonadotrophin-releasing hormone analogues, LH supplementation may help to optimize treatment outcomes for women with biologically older ovaries. Exogenous GH may improve oocyte development and counteract the age-related decline of oocyte quality. The effects of GH may be mediated by insulin-like growth factor-I, which works synergistically with follicle-stimulating hormone on granulosa and theca cells. Conclusion Patients with biologically older ovaries may benefit from a tailored approach based on individual patient characteristics. Among the most promising adjuvant therapies for improving ART outcomes in women of advanced reproductive age are the administration of exogenous LH or GH.

Circulating levels of luteinizing hormone (LH) are essential for the production of steroid hormones that regulate the timing of ovulation and target tissue responses, as well as maintenance of the corpus luteum and therefore early pregnancy. Clinical and basic science observations show that elevated levels of serum LH during the follicular phase of the menstrual cycle are not only unnecessary for follicular maturation but are deleterious to normal reproductive processes. These elevations may occur as a result of administration of exogenous LH or through an endogenous pathological process (i.e. polycystic ovarian disease, PCOD). Resting levels of LH, synergizing with locally produced IGFs, inhibin and perhaps other growth factors, are adequate for normal follicular growth and steroidogenesis. Elevations in serum LH above these resting levels may result in increased androgen production that diminishes follicular function and reduces early embryo viability. Elevated LH levels during the preovulatory period may also negatively influence post-ovulatory events such as conception and implantation. With these facts in mind, the best results for ovulation induction would be expected with purified follicle-stimulating hormone (FSH) administration to women following gonadotrophin releasing hormone (GnRH) down-regulation. It is hoped that this review provides the reader with an analysis of the complex series of events that regulate normal follicular maturation. The reevaluation of the two cell-two gonadotrophin theory suggests that during the preovulatory period, resting levels of LH are adequate for normal follicular maturation. Indeed, overstimulation of the ovary with excessive amounts of LH may diminish the ability of that target organ to produce fertile ova.(ABSTRACT TRUNCATED AT 250 WORDS)

Objective To provide recommendations for the standardized use of the Antral follicle count (AFC) which is used to predict ovarian response to gonadotrophin stimulation during assisted reproductive technology treatment. However, the nature of the follicles that are visualized by ultrasound and the competence of the oocytes held within are largely unknown. In addition, there is considerable variability in the clinical definitions and technical methods used to count and measure antral follicles in both published studies and clinical practice. Design and Setting In December 2007, specialist reproductive medicine clinicians and scientists attended a workshop in an effort to address these issues. Literature concerning the physiology and measurement of ovarian antral follicles was reviewed, clinical and technical considerations regarding antral follicle measurement were discussed, and an operational definition of AFC was developed. Patient(s) None. Intervention(s) None. Outcome Measures Simple recommendations were established for the standardization of AFC assessment in routine clinical practice. The basic clinical and technical requirements required for AFC evaluation were agreed upon, and a systematic method of measuring and counting antral follicles in routine practice was proposed. Conclusion(s) The use of a standardized approach according to the practical recommendations for antral follicle counting as presented is encouraged in future clinical trials and routine practice. The authors also advocate a systematic evaluation of these recommendations as standardized study data become available.

Laboratory Procedures: 1: Establishing and Maintaining an IVF Laboratory 2: Gamete Collection, Preparation and Selection 3: Micromanipulation 4: Culture, Selection and Transfer of the Human Embryo 5: Cryopreservation 6: Diagnosis of Genetic Disease in Preimplantation Embryos 7: Implantation Clinical Techniques: 9: Quality Management Systems 10: Patient Investigation and the Use of Drugs 11: Stimulation Protocols 12: Technical Procedures and Outcomes 13: Special Medical Conditions 14: Complications of Treatment 15: Egg Donation and Surrogate Motherhood 16: Future Directions and Clinical Applications 17: The Support Team 18: Ethics and Legislation.

Sequential exogenous oestradiol and progesterone are often used to prepare the endometrium in frozen embryo transfer (FET) cycles. This open-label, randomized study compared the efficacy and acceptability of self-administered once daily vaginal progesterone gel and vaginal micronized progesterone tablets (three times daily) for luteal support in FET cycles. An Asian population of women (agedor = 45 years) were assigned randomly to receive progesterone gel (90 mg, once daily, n= 100) or vaginal micronized progesterone tablets (200 mg, three times daily, n= 100). All received oestradiol from day 2 of the menstrual cycle, for at least 10 days (or until endometrial thickness wasor =8 mm), before self-administering progesterone for 2 days before FET and up to 14 weeks afterwards if pregnancy occurred. Clinical pregnancy rates (31% for gel and 28% for tablets) and implantation rates (9.8% and 8.8%, respectively) were not significantly different between the treatment groups. Asian women using once daily progesterone gel found the gel easy to use and comfortable, and preferred it to their previous experience of vaginally administered tablets. In summary, once-daily vaginal progesterone gel has similar efficacy to vaginal tablets and is associated with high patient satisfaction.