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STUDY QUESTIONCan risks associated with novelties in assisted reproduction technologies (ARTs) be assessed in a systematic and structured way?SUMMARY ANSWERAn ART-specific risk assessment tool has been developed to assess the risks associated with the development of novelties in ART (EuroGTP II-ART).WHAT IS KNOWN ALREADYHow to implement new technologies in ART is well-described in the literature. The successive steps should include testing in animal models, executing pre-clinical studies using supernumerary gametes or embryos, prospective clinical trials and finally, short- and long-term follow-up studies on the health of the offspring. A framework categorizing treatments from experimental through innovative to established according to the extent of the studies conducted has been devised. However, a systematic and standardized methodology to facilitate risk evaluation before innovations are performed in a clinical setting is lacking.STUDY DESIGN, SIZE, DURATIONThe EuroGTP II-ART risk assessment tool was developed on the basis of a generic risk assessment algorithm developed for tissue and cell therapies and products (TCTPs) in the context of the project ‘Good Practices for demonstrating safety and quality through recipient follow-up European Good Tissue and cells Practices II (EuroGTP II)’. For this purpose, a series of four meetings was held in which eight ART experts participated. In addition, several tests and simulations were undertaken to fine-tune the final tool.PARTICIPANTS/MATERIALS, SETTING, METHODSThe three steps comprising the EuroGTP II methodology were evaluated against its usefulness and applicability in ART. Ways to improve and adapt the methodology into ART risk assessment were agreed and implemented.MAIN RESULTS AND THE ROLE OF CHANCEAssessment of the novelty (Step 1), consisting of seven questions, is the same as for other TCTPs. Practical examples were included for better understanding. Identification of potential risks and consequences (Step 2), consisting of a series of risks and risk consequences to consider during risk assessment, was adapted from the generic methodology, adding more potential risks for processes involving gonadic tissues. The algorithm to score risks was also adapted, giving a specific range of highest possible risk scores. A list of strategies for risk reduction and definition of extended studies required to ensure effectiveness and safety (Step 3) was also produced by the ART experts, based on generic EuroGTP II methodology. Several explanations and examples were provided for each of the steps for better understanding within this field.LIMITATIONS, REASONS FOR CAUTIONA multidisciplinary team is needed to perform risk assessment, to interpret results and to determine risk mitigation strategies and/or next steps required to ensure the safety in the clinical use of novelties.WIDER IMPLICATIONS OF THE FINDINGSThis is a dynamic tool whose value goes beyond assessment of risk before implementing a novel ART in clinical practice, to re-evaluate risks based on information collected during the process.STUDY FUNDING / COMPETING INTEREST(S)This study was called EUROGTP II and was funded by the European Commission (Grant agreement number 709567). The authors declare no competing interests concerning the results of this study.

Pierre Zitoun,1 Jaya Parikh,2 Martine Nijs,3 Wenjie Zhang,4 Rachel Levy-Toledano,5 Boxiong Tang6 1American Hospital of Paris, Neuilly-sur-Seine, France; 2Lister Fertility Clinic, London, UK; 3Nij Geertgen, Elsendorp, the Netherlands; 4WG Group, New York, NY, USA; 5Theramex HQ UK Limited, London, UK; 6Teva Pharmaceuticals, Frazer, PA, USA Purpose: The purpose of this study was to assess product-specific features for a variety of self-administered injection devices and identify key factors that patients and nurses in select European markets find most important when selecting injection devices for self-administration of recombinant human follicle-stimulating hormone and urinary human follicle-stimulating hormone for fertility/reproductive therapy.Patients and methods: Patients (N=402) in France, Italy, Spain, Germany, the UK, the Netherlands, and Belgium, as well as reproductive/fertility nurses (N=40) in Germany, Italy, France, Spain, and the Czech Republic were surveyed. All patients were previously prescribed a follicle-stimulating hormone (FSH) treatment for either in vitro fertilization or ovulation induction. Patient and nurse preferences for attributes across all injection devices in the market were obtained via an online questionnaire and evaluated using the maximum differential scaling (MaxDiff) and conjoint analyses, which captured the relative importance of the selected FSH injection device attributes to determine specific qualities in overall product preference.Results: Both the MaxDiff and conjoint analyses indicated that, for patients and nurses, the ideal FSH injection device would be a highly accurate, multi-use reusable pen injector with a dial-back function that would be easy for both use and education/instruction. Patients and nurses each selected attributes pertinent to their own experiences with the FSH injection device. Categorically, patients valued factors that resulted in minimal impact on daily life, including reduced injection volume to minimize injection-site pain, as well as a reusable device that would be easy to use; nurses placed greater value on a device that would be easy to teach in order to instruct the greatest number of patients while minimizing risk.Conclusion: Patient and nurse preferences were aligned on certain selected attributes of the FSH products. Although this study was an unbranded examination of attributes across all injection devices currently in the market, results demonstrated that the preferred product attributes were all characteristics of the Ovaleap® Pen. Keywords: infertility, prefilled pen device, recombinant human follicle-stimulating hormone, urinary follicle-stimulating hormone, preference

With the successful use of Assisted Reproduction, in particular intracytoplasmic sperm injection (ICSI), to treat infertile couples we have become less discriminating with the quality of spermatozoa we use to treat our patients. Numerous studies have shown the presence of nuclear DNA strand breaks in human ejaculated spermatozoa. The reason why human spermatozoa, in particular from men with abnormal semen parameters, possess these abnormalities in their nuclear DNA is still not clear. Two processes that have been linked to the presence of nuclear DNA strand breaks in spermatozoa are anomalies in apoptosis during spermatogenesis or problems in the packaging of the chromatin during spermiogenesis. Understanding the mechanisms responsible for producing abnormal spermatozoa in the human will improve our knowledge about certain causes of male infertility. More importantly, the impact of such sperm, if selected to perform ICSI, needs to be better understood so that any detrimental paternal effects can be avoided.

In this preliminary prospective randomized study of 420 patients undergoing ovarian stimulation for IVF/intracytoplasmic sperm injection (ICSI), 17 patients (4%) developed moderate to severe ovarian hyperstimulation syndrome (OHSS). Re-evaluation for OHSS on day 4 and 6 after oocyte retrieval identified one patient with continuous risk for severe OHSS, resulting in cancellation of the transfer (1/17, 5.8%). Prospectively, two of three patients had the zona pellucida of the blastocyst removed by pronase exposure prior to transfer. Significantly more patients became pregnant when a zona-free blastocyst was transferred in comparison to transfer of a single zona-intact embryo (9/11 or 82% versus 1/5 or 20%; P < 0.01). Higher ongoing singleton pregnancy rates were obtained when the zona pellucida was removed prior to the transfer (6/11 and 1/5 respectively). This preliminary prospective randomized study indicates that by prolonging the evaluation time for patients at risk of developing OHSS for up to 6 days after the oocyte retrieval, those patients at risk for developing severe OHSS can be identified. Transferring a single zona-free day 5 embryo (blastocyst) and freezing of the supernumerary embryos offers the patient with moderate OHSS an optimal chance for a singleton pregnancy, while avoiding the serious maternal complications of ovarian hyperstimulation syndrome.

Since the first reports of successful pregnancies in humans after treatment with intracytoplasmic sperm injection (ICSI), intensive investigations have focused on several important aspects of this form of assisted reproductive technology. In addition to the technical development of ICSI and increasing understanding of the biochemical and biophysical processes involved during fertilization after injection of an immobilized sperm, studies have aimed to define the indications for patients for a first-line ICSI treatment. One of the major concerns is of course the safety of the technique in terms of the health and reproductive life of the babies born after ICSI. The rhesus monkey is an excellent model to investigate all aspects of this micromanipulation technique. This article provides an evaluation of ICSI.

The sperm-hyaluronan binding assay (HBA) is a diagnostic kit for assessing sperm maturity, function and fertility. The aim of this prospective cohort pilot study was to evaluate the relationship between HBA and WHO sperm parameters (motility, concentration and detailed morphology) and possible influence of sperm processing on hyaluronic acid binding. A cohort of 68 patients undergoing a first combo in vitro fertilisation/intracytoplasmic sperm injection treatment after failure of three or more intrauterine insemination cycles were included in the study. Outcome measures studied were fertilisation rate, embryo quality, ongoing pregnancy rate and cumulative pregnancy rate. HBA outcome improved after sperm preparation and culture, but was not correlated to detailed sperm morphology, concentration or motility. HBA did not provide additional information for identifying patients with poor or absent fertilisation, although the latter had more immature sperm cells and cells with cytoplasmic retention present in their semen. HBA outcome in the neat sample was significantly correlated with embryo quality, with miscarriage rates and ongoing pregnancy rates in the fresh cycles, but not with the cumulative ongoing pregnancy rate. No threshold value for HBA and outcome in combo IVF/ICSI treatment could be established. The clinical value for HBA in addition to routine semen analysis for this patient population seems limited.

A microfluorimetric assay was designed to monitor beta-glucuronidase (beta-gluc) activity during the early development of mouse embryos and was also applied to measure beta-gluc activity in human sperm cells and single human oocytes. Mouse oocytes and preimplantation embryos at different stages of development, failed fertilized human oocytes, and human sperm cells were collected, and beta-gluc activity was analysed. In the mouse, beta-gluc activity could be measured starting from the morula stage on in a pooled sample of 15 embryos and at blastocyst formation in one single embryo. beta-Gluc activity was low in human sperm cells. The enzyme could be readily demonstrated in 87 per cent of non-fertilized human oocytes. Considerable variation in beta-gluc activity was noticed between individual oocytes.

This study presents a case of intracytoplasmic sperm injection (ICSI) with ejaculated immotile spermatozoa from a patient with immotile cilia syndrome. Semen analysis of the patient suffering from immotile cilia syndrome revealed an extreme oligoasthenoteratozoospermia (OAT: count

Objective To determine the influence of ejaculatory abstinence on within-subject semen parameters and DNA fragmentation. Design Prospective study. Setting Private infertility institute and university-based research laboratory. Patient(s) Sixteen consenting male volunteers undergoing infertility investigation. Intervention(s) None. Main outcome measure(s) Within-subject analysis of World Health Organization semen parameters and sperm DNA fragmentation and chromatin packaging after 1, 3, 5, and 8 days' abstinence. Result(s) Of 16 men recruited, data for 11 men were included for statistical analysis because 5 men did not strictly comply with abstinence criteria. Duration of abstinence had a statistically significant positive influence on sperm concentration and semen volume. Abstinence had no statistically significant influence on pH, viability, total and grade A motility, or morphology. The percentage of DNA fragmentation remained unchanged relative to abstinence. The percentage of sperm with immature chromatin was statistically significantly increased with 1 day of abstinence. Conclusion(s) This is the first study to report on within-subject semen parameter, DNA fragmentation, and chromatin packaging variations after specified target days of abstinence. Sperm numbers and semen volume increased with duration of abstinence. Abstinence did not influence pH, viability, morphology, total or grade A motility, or sperm DNA fragmentation. A short (24-hour) abstinence period negatively influenced chromatin quality.

There is good evidence in literature that intrauterine insemination (IUI) is the best first line treatment and most cost-effective procedure for moderate male factor subfertility. It seems very difficult to identify individual semen parameters predicting the likelihood of pregnancy after IUI. This can be explained by a lack of standardization of semen analysis, but many other methodological variables may also influence IUI success rates such as the patient selection, type of ovarian stimulation and number of inseminations per cycle. A review of the literature confirmed that sperm morphology using strict criteria and the inseminating motile sperm count (IMC) after sperm preparation are the two most important sperm parameters to assess the real impact of semen quality on IUI outcome. A universal threshold level above which IUI can be performed with acceptable pregnancy rates has not been determined yet, although IUI success seems to be impaired with

This is the first report of an intrauterine pregnancy following timed coitus, resulting from transperitoneal sperm and/or oocyte migration as the oocyte originated from an ectopic (undescended) ovary. The patient was treated in the infertility clinic after a history of primary infertility for 2 years. Diagnosis of a moderate teratozoospermia and the presence of a unicornuate uterus with one right-sided normal ovary was made. During the first intrauterine insemination (IUI) treatment cycle with clomiphene citrate stimulation, a discrepancy between oestradiol concentrations and follicular growth was observed. Magnetic resonance imaging (MRI) was carried out because the presence of an ectopic ovary was suspected. The diagnosis of a left-sided undescended ovary was made, containing several follicles, the largest measuring 16 mm in diameter. Because of a spontaneous LH surge 2 days later without substantial follicular growth in the normal right-sided ovary, IUI was cancelled and timed intercourse was planned. Surprisingly, and although the largest follicle in the normally located ovary reached a maximum diameter of only 12 mm on repetitive ultrasound monitoring, this patient became pregnant and gave birth to a healthy baby boy.

Freezing of human sperm is considered a routine procedure in assisted reproductive technology (ART) laboratories. This article considers various aspects of cryopreservation of human sperm. Human sperm show a specific cryophysical behaviour and different sperm freezing protocols have been developed to avoid damage to the sperm cells. The damage can range from impaired motility and reduced viability to damage to the cellular organelles and effects at the molecular level, resulting in an impaired fertilizing potential. As testicular sperm are immature and only a small number can be retrieved, special techniques are required for successful freezing and thawing of these samples. Banking of human sperm has to be performed in a safe and controlled way and different guidelines are necessary to ensure that this is achieved.

beta-N-Acetylhexosaminidase is a lysosomal enzyme; mutation in this protein leads to Tay - Sachs or Sandhoff disease. We have developed an assay for the beta-N-acetylhexosaminidase in mouse oocytes and preimplantation embryos. Activity was low in oocytes and zygotes and started to rise from the late 2-cell stage. By using -amanitin, an inhibitor of DNA-dependent RNA polymerase, we were able to show that the first embryonic mRNA transcriptions of the fl-N-acetylhexosaminidase genes take place between 38 and 46 h post HCG (early 2-cell stage) and between 46 and 54 h post HCG (late 2-cell stage).

The health of a preimplantation embryo can be diagnosed in one or more cells biopsied from the conceptus. Here, we tried to evaluate the impact of biopsy of some trophectoderm cells from hatching mouse blastocysts on their further in vitro implantation and early egg cylinder formation. Of 374 blastocysts evaluated 112 hours after hCG, 34% initiated hatching with a small number of mural, polar, or intermediate trophectoderm cells. Half of 59 embryos that underwent induction of hatching by zona puncturing herniated some cells through this opening. After removal of cells with a glass microneedle from spontaneously hatching blastocysts, viability assessed by vital FDA staining was impaired, as well as the in vitro zona pellucida shedding and implantation. When polar trophectoderm cells were biopsied, a significantly lower number of embryos reached the egg cylinder stage.