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Intracytoplasmic sperm injection (ICSI), developed 30 years ago in our Center, allowed men with low to extremely low sperm quality to become the genetic father of their child. Overall, ICSI is now applied in up to 70% of all ART cycles and has resulted in the birth of millions of children worldwide. ICSI circumvents natural sperm selection mechanisms and concerns related to transgenerational inheritance of male infertility following ICSI could not be fully answered till today, given the young age of the offspring born after ICSI. Since genetic abnormalities account for one third of all cases of male factor infertility, it is essential to identify these genetic abnormalities because of the potential risk of their transmission to the offspring through the use of ART. As ICSI is nowadays performed even in couples with normal semen parameters, the disentanglement of procedure-related and parental background-related factors on the health of the offspring is important. Due to the safety concerns regarding the health of children born after ART, a longitudinal follow-up program was set up in our Center in the 80’s, resulting in data of more than 50 000 pregnancies. To date, more than 25 000 children are conceived after ICSI in our Center. We have not only the worldwide eldest cohort of ICSI offspring but also a uniform cohort of children and young adults conceived by ICSI because of (severe) male factor infertility. We present the results of these prospective studies with focus on the reproductive health in ICSI children between 8 and 22 years and compare these with available data from others. First of all, we will show data on the gonadal function assessed both clinically and by means of biomarkers at pre-pubertal and pubertal ages in ICSI boys. Secondly, we will present findings on the reproductive status of young adult men and women conceived after ICSI between 1992 and 1996: results of hormonal work-up, physical examination and semen parameters for ICSI men and results of hormone levels and antral follicle counts for ICSI women. Finally, we will discuss the findings in these cohorts of children and young adults conceived because of male infertility in view of the available literature regarding reproductive function in offspring conceived following ICSI.

On behalf of Neelke De Munck, Herman Tournaye and all colleagues BrusselsIVF (UZBrussel) and Vrije Universiteit Brussel (1980-2022) In the early years of ART, IVF was successful to alleviate female-factor or idiopathic infertility, yet limited results were obtained in male-factor infertility. In the 80s, assisted fertilization procedures using micromanipulation were introduced, i.e. zona drilling and partial zona dissection. As these procedures did not perform well, at BrusselsIVF we investigated subzonal insemination (SUZI) in mice and eventually we introduced SUZI in our clinic for couples with impaired sperm parameters. IRB approval was obtained in 1991 but required a prospective follow-up protocol of all pregnancies and children born. During SUZI, it occurred that sperm was injected in the oolemma, after which normal fertilization and embryo development was observed. The first pregnancy leading to a live birth in January 1992 was established when 11 oocytes had no fertilization after SUZI but one oocyte was fertilized after “failed SUZI” and was eventually transferred. In the following period, we were able to demonstrate that ICSI was more efficient than SUZI and from July 1993 we only applied ICSI in patients in need of assisted fertilization. We deliberately choose to share our experience widely with our peers and thereafter ICSI was performed in many centres worldwide and ICSI has been mushrooming. The 2017 ICMART World Report mentions an annual increase of ART cycles with 2 million cycles in 2017 resulting in 330.000 deliveries. The percentage of cIVF versus ICSI is very different depending on the region but on average 70% of cycles are ICSI. This indicates that ICSI is also widely used for non-male infertility. The occurrence of about 15% failed fertilization cycles after cIVF is often the reason for using ICSI. This is for sure the case in countries where only a limited number of cycles is reimbursed by social security or where no reimbursement is in place. ICSI proved to relieve infertility in couples with severely impaired characteristics of ejaculated sperm including cryptozoospermia. For these patients ICSI results were similar as cIVF for other indications. ICSI with epididymal or testicular sperm could also solve the problem in patients with obstructive azoospermia. Patients with non-obstructive azoospermia can also be helped if sperm cells can be found by testicular sperm extraction using conventional TESE or microTESE. However, the chance to have a child in NOA couples embarking for both TESE and eventually ICSI is much lower if expressed in terms of Intention to Treat: 13,4% in NOA patients and 10,1% in Klinefelter patients. Most of these couples will embark on treatment even after full counselling before considering the use of donor sperm. At BrusselsIVF, ICSI is also used on in-vitro matured oocytes following culture of immature cumulus-oocyte complexes collected from antral follicles in PCOS patients. For couples undergoing Preimplantation Genetic Testing with biopsy of day-3 cleaving embryos or day-5 or day-6 blastocysts, ICSI is still the preferred procedure in order to avoid contamination at the time of biopsy by remnants of sperm or cumulus cells when cIVF is used. From the start in 1980 we have invested in clinical, translational and fundamental research. As of now this research is carried out by more than 250 individuals from BrusselsIVF, the centre for reproductive medicine of the UZBrussel and different research units at the VUB. Research focuses on different aspects of infertility, medical genetics, human reproduction, early human development and human pluripotent cells.

Elke gezondheidswerker vandaag is enthousiast over en respecteert theoretische en praktische educatie en hoe die samen met wetenschappelijk onderzoek voortdurend bijdragen tot vooruitgang in preventie, behandeling en eventueel genezing van ziekten. De Commissie Internationalisering van de Koninklijke Academie voor Geneeskunde van België (KAGB) en haar zusterorganisatie Académie Royale de Médecine de Belgique (ARMB) hebben een reeks praktische aanbevelingen geformuleerd ter bevordering van opleidingen van zorgverleners door middel van internationale uitwisselingen. Dat programma, ter beschikking gesteld in beide landstalen, is internationaal toegankelijk en ook in het Engels raadpleegbaar via het internet. Het is bruikbaar en toepasbaar voor betrokkenen in alle disciplines in de biomedische sector en functioneert onder de naam “Hippocrates Medicine Program” of “Hippocrates Geneeskunde Programma”. De aanbevelingen hebben als doel om internationale opleidingen en trainingen te stimuleren en om de status van onderzoekers in opleiding en zelfs specialisten alsook biomedisch academische stafleden aan universiteiten en opleidingsziekenhuizen te verbeteren tijdens stages in het buitenland.

Let G be a connected complex reductive group. A well known theorem of I. Losev's says that a smooth affine spherical G-variety X is uniquely determined by its weight monoid, which is the set of irreducible representations of G that occur in the coordinate ring of X. In this paper, we use the combinatorial theory of spherical varieties and a smoothness criterion of R. Camus to characterize the weight monoids of smooth affine spherical varieties., v1: 33 pages. v2: 36 pages, improved exposition, updated references and citations

Background: It can be difficult to motivate children to participate in physical exercises, especially when they have oncological disorders. Purpose: examine the effect of a humanoid robot and music on motivation to participate in physical activity in typically developing children (TDC) and children with oncological disorders (COD). Methods: Two cohort studies were conducted; in an elementary school in Belgium (N=75TDC, 4-13years) and pediatric cancer ward of Ghent University Hospital (N=13COD, 3-15years). Participants completed four gross motor exercise conditions; with a human or humanoid robot instructor and with or without music. Motivation was assessed using the Smileyometer (amount of fun), Again score (amount of chosen repetitions) and Fun Sorter (which condition was most fun). Results: TDC indicated higher Smileyometer-scores in the two conditions with music. The Again score revealed no preference in either group. In TDC and COD, the results of the Fun Sorter indicated that conditions with music or the robot were more fun than without music or with the human instructor. The combination of the robot and music was most preferred. Conclusions: Music or a humanoid robot seemed to increase children's initial motivation to participate in physical activity, also in hospitalized COD. Further research should evaluate the long-term effects. This project was supported by the agency for Innovation by Science and Technology ('Agentschap voor Innovatie door Wetenschap en Technologie', Flanders). PM was supported by a grant of the Research Foundation of Flanders ('Fonds Wetenschappelijk Onderzoek -Vlaanderen'; 'Krediet aan Navorsers' [grant number 1503915N]). PM was supported by the European Commission as a Marie Sklodowska-Curie fellow (part of the EU Framework Programme for Research and Innovation, Horizon 2020; [proposal 660458]). The funding sponsors had no role in any part of the research conducted, including study design, collection, analysis, or interpretation of data, writing the manuscript, and the decision to submit the manuscript for publication.

Let G be a complex connected reductive group. I. Losev has shown that a smooth affine spherical G-variety X is uniquely determined by its weight monoid, which is the set of irreducible representations of G that occur in the coordinate ring of X. In this paper we use a combinatorial characterization of the weight monoids of smooth affine spherical varieties to classify: (a) all such varieties when G is $\mathrm{SL}(2) \times \mathbb{C}^{\times}$ and (b) all such varieties for G simple which have a G-saturated weight monoid of full rank. We also use the characterization and F. Knop's classification theorem for multiplicity free Hamiltonian manifolds to give a new proof of C. Woodward's result that every reflective Delzant polytope is the moment polytope of such a manifold., Comment: v1: 31 pages. v2: 33 pages, implemented changes requested by referees, changed title, to appear in Acta Mathematica Sinica

Male births exceed female births by 5-6% (for a sex ratio at birth of 1.05-1.06) while a women's life expectancy, on a global scale, is about 6 years longer. Thus within various age groups the male:female ratio changes over time. Until age 50 years men outnumber women; thereafter their numbers show a sharp decline. Consequently at age 80 years, there are many more women than men. An estimated 25% of this male excess mortality is due to biological causes, the rest being explained by behavioural, cultural and environmental factors. For both women and men, the main health risks related to lifestyle are smoking, alcohol, unhealthy diet and physical inactivity. In the year 2010, overweight (BMI: 25-29 kg/m(2)) and obesity (BMI:30 kg/m(2)) were responsible for over 3 million deaths, with similar relative risks in men and women for overweight and obesity. Smoking and alcohol are the major causes of the global gender gap in mortality. For women in some parts of the world however pregnancy is also hazardous. On a global scale, in 2013 about 300 000 deaths were related to pregnancy, with sub-Saharan Africa registering the highest maternal mortality: over 500 maternal deaths per 100 000 births. Additional woman's health risks arise from gender discrimination, including sex-selective abortion, violence against women and early child marriage. Providers should be aware of the effect that these risks can have on both reproductive and general health.

Objective: To present an overview of the numbers and types of human embryos used in research projects in Belgium from 2007 to 2015. Design: Analysis of all research proposals approved by the Federal Commission for Medical and Scientific Research on Embryos In Vitro. Setting: Not applicable. Patient(s): Not applicable. Main Outcome Measure(s): Number of embryos used for research, number of embryos created for research, and areas of embryo research. Result(s): Since 2007, 15,811 embryos were used for 36 research projects. In total, 10,492 (66%) fresh supernumerary embryos (unfit for transfer or freezing) were used, 4,083 (26%) frozen supernumerary embryos (donated by parents whose child wish was completed or abandoned), and 1,236 (8%) embryos created for research. Most projects focused on research into embryo development. Fresh supernumerary embryos were mainly used for human embryonic stem cell (hESC) research. Frozen supernumerary embryos were almost exclusively used for research into embryo development and for hESC research. Embryos created for research were used for research into embryo development, oocyte research, research into cryopreservation of oocytes, and hESC research. Conclusion(s): Having concrete data on embryo research is crucial for an informed debate. Moreover, these data are necessary to find out trends in research such as the numbers of embryos needed and the areas of research. Data collection requires a sufficiently clear definition of "research'' and "embryo.'' These conceptual questions frequently reveal lack of clarity in legislation. (C)2017 by American Society for Reproductive Medicine.

Study question What is the semen quality of young adult men who were conceived 18-22 years ago by ICSI for male infertility? Summary answer In this cohort of 54 young adult ICSI men, median sperm concentration, total sperm count and total motile sperm count were significantly lower than in spontaneously conceived peers. What is known already The oldest ICSI offspring cohort worldwide has recently reached adulthood. Hence, their reproductive health can now be investigated. Since these children were conceived by ICSI because of severe male-factor infertility, there is reasonable concern that male offspring have inherited the deficient spermatogenesis from their fathers. Previously normal pubertal development and adequate Sertoli and Leydig cell function have been described in pubertal ICSI boys; however, no information on their sperm quality is currently available. Study design, size, duration This study was conducted at UZ Brussel between March 2013 and April 2016 and is part of a large follow-up project focussing on reproductive and metabolic health of young adults, between 18 and 22 years and conceived after ICSI with ejaculated sperm. Results of both a physical examination and semen analysis were compared between young ICSI men being part of a longitudinally followed cohort and spontaneously conceived controls who were recruited cross-sectionally. Participants/materials, setting, method Results of a single semen sample in 54 young adult ICSI men and 57 spontaneously conceived men are reported. All young adults were individually assessed, and the results of their physical examination were completed by questionnaires. Data were analysed by multiple linear and logistic regression, adjusted for covariates. In addition, semen parameters of the ICSI fathers dating back from their ICSI treatment application were analysed for correlations. Main results and the role of chance Young ICSI adults had a lower median sperm concentration (17.7 million/ml), lower median total sperm count (31.9 million) and lower median total motile sperm count (12.7 million) in comparison to spontaneously conceived peers (37.0 million/ml; 86.8 million; 38.6 million, respectively). The median percentage progressive and total motility, median percentage normal morphology and median semen volume were not significantly different between these groups. After adjustment for confounders (age, BMI, genital malformations, time from ejaculation to analysis, abstinence period), the statistically significant differences between ICSI men and spontaneously conceived peers remained: an almost doubled sperm concentration in spontaneously conceived peers in comparison to ICSI men (ratio 1.9, 95% CI 1.1-3.2) and a two-fold lower total sperm count (ratio 2.3, 95% CI 1.3-4.1) and total motile count (ratio 2.1, 95% CI 1.2-3.6) in ICSI men compared to controls were found. Furthermore, compared to men born after spontaneous conception, ICSI men were nearly three times more likely to have sperm concentrations below the WHO reference value of 15 million/ml (adjusted odds ratio (AOR) 2.7; 95% CI 1.1-6.7) and four times more likely to have total sperm counts below 39 million (AOR 4.3; 95% CI 1.7-11.3). In this small group of 54 father-son pairs, a weak negative correlation between total sperm count in fathers and their sons was found. Limitations, reasons for caution The main limitation is the small study population. Also, the results of this study where ICSI was performed with ejaculated sperm and for male-factor infertility cannot be generalized to all ICSI offspring because the indications for ICSI have nowadays been extended and ICSI is also being performed with non-ejaculated sperm and reported differences may thus either decrease or increase. Wider implications of the findings These first results in a small group of ICSI men indicate a lower semen quantity and quality in young adults born after ICSI for male infertility in their fathers. Study funding/competing interests This study was supported by Methusalem grants and by grants from Wetenschappelijk Fonds Willy Gepts, all issued by the Vrije Universiteit Brussel (VUB). All co-authors except M.B. and H.T. declared no conflict of interest. M.B. has received consultancy fees from MSD, Serono Symposia and Merck. The Universitair Ziekenhuis Brussel (UZ Brussel) and the Centre for Medical Genetics have received several educational grants from IBSA, Ferring, Organon, Shering-Plough and Merck for establishing the database for follow-up research and organizing the data collection. The institution of H.T. has received research grants from the Research Fund of Flanders (FWO), an unconditional grant from Ferring for research on testicular stem cells and research grants from Ferring, Merck, MSD, Roche, Besins, Goodlife and Cook for several research projects in female infertility. H.T. has received consultancy fees from Finox, Abbott and ObsEva for research projects in female infertility.

We determine, under a certain assumption, the Alexeev-Brion moduli scheme M_S of affine spherical G-varieties with a prescribed weight monoid S. In [ arXiv:1008.0911 ] we showed that if G is a connected complex reductive group of type A and S is the weight monoid of a spherical G-module, then M_S is an affine space. Here we prove that this remains true without any restriction on the type of G., v1: 37 pages. v2: 43 pages, implemented changes requested by referee, a version without the appendix will appear in Algebras and Representation Theory

Within joint venture De Groene Boog, BESIX will design and construct a 2.2 km long tunnel in the North of Rotterdam for a new highway connection. Its design process entails the structural design of many similar tunnel sections and thousands of foundation and sheet piles, as well as the production of a detailed 3D Building Information Model and numerous technical drawings. To effectively perform these tasks and efficiently cope with design changes, an automation strategy has been developed that benefits from the tunnel’s repetitive geometry. The digital models are set up parametrically, while software innovations allow to dynamically transfer data between and combine the strengths of multiple software packages. Through standardization we aim to automate structural calculations and drawing production.This paper will present the proposed automation strategy for the ongoing design process of the tunnel and will evaluate its benefit during the detailed design phase compared to traditional methods. The present project forms the first application of parametric design and automation of such magnitude being developed in-house at the BESIX Engineering Department. It reflects our ambition to use these techniques for tackling the ever increasing challenges within the construction industry and simultaneously boosting productivity and the quality of the final product.

We study Alexeev and Brion's moduli scheme $M_\Gamma$ of affine spherical varieties with weight monoid $\Gamma$ under the assumption that $\Gamma$ is free. We describe the tangent space to $M_\Gamma$ at its `most degenerate point' in terms of the combinatorial invariants of spherical varieties and deduce that the irreducible components of $M_\Gamma$, equipped with their reduced induced scheme structure, are affine spaces.

Infertility care has improved remarkably over the last few decades and has received growing attention from health care providers. Several treatments, including expensive options such as Assisted Reproductive Techniques, are now widely available for routine clinical use. In most cases, adoption of these treatments has occurred without robust cost-effective analyses. IVF for unexplained infertility and ICSI in the absence of semen abnormalities are two examples of this gradual technology creep. More in-depth economic analyses in the field of infertility are undoubtedly warranted. However, performing these analyses is challenging because infertility care poses a number of unique challenges. Studies of cost-effectiveness are open to criticism because there is a lack of consensus about the outcomes of choice and the appropriate perspective. The use of quality adjusted life years (QALYs) to allow comparisons with other clinical conditions is also controversial because the value associated with infertility care cannot be easily captured in QALYs. Moreover, their use triggers the crucial question of whose QALYs merit consideration—an individual's, a couple's or a child's. In conclusion, economic analysis in infertility represents a peculiar but crucial challenge. If management of infertility is to become an integral part of publicly or privately funded health care systems worldwide, better quality data and a shared vision about the costs and benefits of infertility treatments are needed.

This monograph, written by the pioneers of IVF and reproductive medicine, celebrates the history, achievements, and medical advancements made over the last 40 years in this rapidly growing field. (C) 2018 by American Society for Reproductive Medicine.

We apply the combinatorial theory of spherical varieties to characterize the momentum polytopes of polarized projective spherical varieties. This enables us to derive a classification of these varieties, without specifying the open orbit, as well as a classification of all Fano spherical varieties. In the setting of multiplicity free compact and connected Hamiltonian manifolds, we obtain a necessary and sufficient condition involving momentum polytopes for such manifolds to be K��hler and classify the invariant compatible complex structures of a given K��hler multiplicity free compact and connected Hamiltonian manifold., v1: 32 pages. v2: 47 pages, fixed errors, improved exposition, expanded Section 7. v3: 47 pages, implemented changes and corrections requested by referee

Background: Networks have become a widely-used approach to address the complexity of issues around international health and health equity, and many types of networks have been described. The added value of networking is not straightforward and it should be documented. Be-Cause Health (BCH) is a unique, multistakeholder network linking the Belgian actors of the health and development sector, creating a partnership to support their commitment to equitable health. Consequently, BCH has very broad objectives with creative and dynamic outcomes. The aim of this study was to explore if the structure and process of the BCH network could be adapted to improve its effectiveness. Methods: The overall framework of the project included a narrative literature review combined with qualitative semi-structured interviews. The literature review looked at the structure and process of networks and the means of evaluating network effectiveness. The results of this review were used to inform the interviews of key Be-Cause Health members. Results: The reviewed articles illustrate the diversity of networking activities and the complexity of their evaluation, suggesting the use of participative evaluation methods to capture the intangible outcomes as experienced by the members themselves. The interviewees reported that the major benefits of their BCH membership were information sharing and connecting with peers. They felt the growth of the BCH network was mostly related to its culture of trust among members, to the inspiring leadership, to the concrete activities it could organise, to its flexible structure tailored to their needs,and to the support of the secretariat and to the embeddedness of the network in a wider institution with secured public funding. They identified a need for improved internal and external communication and for more concrete objectives of the working groups. Conclusion: The BCH network is a unique response to the complex challenges of health equity and development. The outcomes of such complex networks cannot be measured with a simple quantitative method, and require participative methods of evaluation. There is no formula to predict the change of effectiveness that would result from a change of structure.

Intrauterine insemination (IUI) with or without ovarian stimulation is a common treatment for infertility. Despite its popularity, the effectiveness of IUI treatment is not consistent, and the role of IUI and in vitro fertilization (IVF) treatment in practice protocols has not been clarified.Medline searches were done by individual topics (utilization, procedures, effectiveness of partner but not donor IUI and related endocrine issues). Effectiveness of IUI was evaluated in relevant randomized controlled trials, using meta-analysis and meta-regression where necessary.Stimulated IUI is ineffective in male infertility and the effect on other diagnoses is small. With clomiphene citrate and IUI, the most common IUI protocol, pregnancy rates average 7% per cycle. FSH ovarian stimulation and IUI treatment is only modestly better than observation only with pregnancy rate 12% per cycle but multiple birth rates averaging 13%. Mildly stimulated (1-2 follicles) cycles might reduce the cost and multiple birth rates but may require more cycles of treatment. Prevention of premature luteinizing hormone surges and luteal phase support do not appear to be major requirements in IUI cycles.IUI treatment requires ovarian stimulation to achieve modest results, but the high multiple pregnancy rates mean that it is no more than a poor substitute for IVF treatment. More trials are needed on IUI treatment with mild stimulation and on the order of IUI and other treatments.

In vitro fertilization and embryo transfer or gamete (or zygote) intra-Fallopian transfer after ovum donation were performed in 16 patients with primary or secondary amenorrhea, associated with chromosome abnormalities. The patients showed the wide range of (mostly X) chromosome abnormalities characteristic for women with primary or premature ovarian failure. Four of these patients became pregnant and three of them have delivered healthy infants with a normal karyotype. This pregnancy rate is far superior to the accepted fertility figure in these patients. When these results were compared with the fertility treatment results of three other groups of women with absent ovarian function (1. ovarian dysgenesis; 2. surgical castration; 3. premature menopause) but with a normal 46,XX karyotype, no difference in treatment efficiency could be detected. These results offer a promising approach for the treatment of infertility in agonadal patients with chromosome aberrations.

BACKGROUND: Preimplantation genetic diagnosis or screening (PGD, PGS) involves embryo biopsy on Day 3. Opting for one- or two-cell biopsy is a balance between the lowest risk for misdiagnosis on the one hand and the highest chance for a pregnancy on the other hand. METHODS: A prospective controlled trial was designed and 592 ICSI cycles were randomly assigned to the one-cell (group I) or the two-cell group (group II). Primary outcomes were diagnostic efficiency and embryonic development to delivery with live birth (analysed by cycle). The false-positive rate for the PCR cycles is presented as a secondary outcome (analysed by embryo). RESULTS: A strong significant correlation was observed between embryonic developmental stage on Day 3 and post-biopsy in vitro development on Day 5 (P < 0.0001). The influence of the intervention on Day 3 was less significant (P = 0.007): the biopsy of one cell is less invasive than the biopsy of two cells. PCR diagnostic efficiency was 88.6% in group I and 96.4 % in group II (P = 0.008). For the fluorescence in situ hybridization (FISH) PGD cycles no significant difference in efficiency was obtained (98.2 and 97.5% in group I and II, respectively). Similar delivery rates with live birth per started cycle were obtained [58/287 or 20.2% in group I versus 52/303 or 17.2% in group II, P = 0.358; the absolute risk reduction = 3.05%; 95% confidence interval (CI): -3.24,9.34]. Post-PGD PCR reanalysis showed six false positives in 97 embryos (6.2%) in group II and none in group I (91 embryos reanalysed). No false negatives were found. CONCLUSIONS: While removal of two blastomeres decreases the likelihood of blastocyst formation, compared with removal of one blastomere, Day 3 in vitro developmental stage is a stronger predictor for Day 5 developmental potential than the removal of one or two cells. The biopsy of only one cell significantly lowers the efficiency of a PCR-based diagnosis, whereas the efficiency of the FISH PGD procedure remains similar whether one or two cells are removed. Delivery rates with live birth per started cycle were not significantly different.

Background Oocyte cryopreservation remains a realistic objective, provided that more systematic approaches are applied, such as thorough analysis of the oocyte oolemma permeability to water and diverse cryoprotectants. Methods We prospectively investigated volume changes over time at different temperatures (30 degrees C, 22 degrees C and 8 degrees C) of human metaphase II (MII) oocytes (obtained in stimulated ICSI cycles and matured in vitro from the germinal vesicle stage) when exposed to changes in external osmolality. We also investigated human in vitro matured (IVM) oocytes membrane permeability characteristics at 22 degrees C to 1,2-propanediol (PG) and dimethylsulphoxide (DMSO) and at 30 degrees C, 22 degrees C and 8 degrees C to ethylene glycol (EG), and calculated corresponding oocyte oolemma permeability coefficients (Lp and Pcpa). Furthermore, we investigated the osmotic tolerance limits of IVM oocytes exposed to changes in external osmolality as assessed by their developmental competence during the course of 72 h after ICSI. Results The results of our studies describe human oocyte membrane permeability coefficients for EG at 30 degrees C (2.85+/-0.15x10(-3) cm/min), 22 degrees C (1.17+/-0.60x10(-3) cm/min) and 8 degrees C (0.37+/-0.15x10(-3) cm/min). Furthermore, at 22 degrees C the EG oolemma permeability coefficient was lower than that of PG and DMSO (1.17+/-0.60x10(-3) cm/min versus 2.15+/-0.70x10(-3) and 1.56+/-0.38x10(-3) cm/min, respectively). Our results also indicate, that human IVM MII oocytes tolerated exposure to solutions in the range of 39-2264 mOsmol/kg H2O as assessed by the oocytes' developmental competence after exposure. Conclusions The results of the present study may contribute to a better understanding of the biology and cryobiology of human oocytes, and to the design of better and more robust cryopreservation (freezing or vitrification) protocols.

Feeder-free human embryonic stem cell (hESC) culture is associated with the presence of mesenchymal-like cells appearing at the periphery of the colonies. The aim of this study was to identify this early differentiation process. Long-term feeder-free hESC cultures using matrigel and conditioned medium from mouse and from human origin revealed that the appearance of mesenchymal-like cells was similar regardless of the conditioned medium used. Standard characterization confirmed the preservation of hESC properties, but the feeder-free cultures could not be maintained longer than 37 passages. The early differentiation process was characterized in the short term after switching hESCs cultured on feeders to feeder-free conditions. Transmission electron microscopy showed an epithelium-like structure inside the hESC colonies, whereas the peripheral cells revealed the acquisition of a rather mesenchymal-like phenotype. Immunochemistry analysis showed that cells at the periphery of the colonies had a negative E-cadherin expression and a positive Vimentin expression, suggesting an epithelial-mesenchymal transition (EMT). Nuclear staining of beta-catenin, positive N-cadherin and negative Connexin 43 expression were also found in the mesenchymal-like cell population. After RT-PCR analysis, Slug and Snail, both EMT-related transcription factors, were detected as up-regulated in the mesenchymal-like cell population. Taken together, our data suggest that culturing hESCs in feeder-free conditions enhances an early differentiation process identified as an EMT.

Although few ethical concerns exist regarding the use of adult stem cells, the field of embryonic stem cell research is fraught with moral qualms. Several alternative sources of pluripotent stem cells have recently been presented that try to sidestep the destruction of human embryos. The goal of these new proposals is to avoid embryo destruction, the main objection to embryonic stem cell research and thus introduce a type of stem cell research that would gain widespread approval and support. This article suggests that most embryo-saving alternatives fail to reach this goal given the concessions they require with regard to the speed of progress, technical complexity, safety and security of applications, degree of dependence on limited resources and extent of the field of application. The second part of the article identifies and analyses the two main strategies that alternative sources of pluripotent stem cells are based on and points out their shortcomings.

Let G be a complex reductive group. A normal G-variety X is called spherical if a Borel subgroup of G has a dense orbit in X. Of particular interest are spherical varieties which are smooth and affine since they form local models for multiplicity free Hamiltonian K-manifolds, K a maximal compact subgroup of G. In this paper, we classify all smooth affine spherical varieties up to coverings, central tori, and C*-fibrations., v1: 23 pages, uses texdraw; v2: 25 pages, introduction updated, Lemma 7.2 fixed, references added, typos corrected

BACKGROUND: Apart from research applications, testicular stem cell transplantation (TSCT) may one day also have valuable clinical applications. Therefore, it is important to investigate whether this technique is a safe method to have progeny. This controlled study aims at evaluating the fetuses and the live born offspring obtained after TSCT in male mice. METHODS: Male mice were mated with wild-type (WT) females after TSCT to produce offspring. First, fetuses were evaluated on the 17th gestational day. The length, weight and morphological age were compared to those of control mouse fetuses. The live born offspring were then investigated for their reproductive potential over three generations. RESULTS: The litter sizes after TSCT were decreased compared to controls. Fetuses showed developmental retardation of a quarter of a day, but no major external abnormalities were observed. The live born pups were able to produce normal litter sizes, at least until the third generation. CONCLUSIONS: Transplanted animals are able to reproduce naturally. Although litter sizes are lower and development is retarded, no major morphological or procreative abnormalities were observed.

BACKGROUND: Since its introduction in 1994, testicular stem cell transplantation (TSCT) has been widely used for research. This technique may also become important for preserving fertility in pre-pubertal cancer patients. Therefore, it is necessary to investigate the safety aspects of reproduction using spermatozoa obtained after TSCT. In this study, preimplantation development of mouse embryos, using spermatozoa obtained after TSCT, was examined. METHODS: TSCT-derived spermatozoa were used for IVF and ICSI. Embryos were cultured for five days until they reached blastocyst stage and were evaluated by differential staining. RESULTS: IVF revealed significantly lower fertilization and development rates after TSCT-IVF compared to control-IVF. Blastocysts derived from TSCTIVF had significantly lower inner cell mass numbers (ICMs) and lower ICM/trophectoderm (TE) ratios compared to control-IVF blastocysts. No differences in fertilization and development rates were observed between TSCT-ICSI and control-ICSI, and blastocyst quality in the transplanted group was similar to that of the control blastocysts. CONCLUSION: Our study showed that after TSCT-IVF, fertilization and preimplantation development were disturbed and blastocysts showed reduced ICM and ICM/TE ratio. However, after TSCT-ICSI, both fertilization and preimplantation development were normal and blastocyst formation was comparable to control-ICSI.

Patients with recurrent IVF failure are defined as patients who are younger than 37 years and who had at least three consecutive unsuccessful IVF/intracytoplasmic sperm injection (ICSI) cycles with good quality embryos. These patients might be predisposed to chromosome errors in their embryos and therefore might benefit from preimplantation genetic diagnosis for aneuploidy screening (PGD-AS). This technique is, however, expensive and some normal embryos might be lost due to the error rate. The aim of this retrospective study was to define those patients who would benefit most from it. One hundred and twenty-one first PGD-AS cycles for recurrent IVF failure were analysed. The aneuploidy rate, 'no embryo transfer' rate, live birth rate per embryo transfer and implantation rate were respectively 48.3, 22.3, 29.7 and 19.5%. A multivariate logistic regression analysis gave us a predictive model demonstrating that to have a 90% probability of having an embryo transfer after PGD-AS, the patient should have at least 10 mature oocytes, eight normally fertilized oocytes and six embryos for biopsy. This study suggests that most patients with recurrent IVF failure may benefit from PGD-AS. Future studies, however, should more strictly define this heterogeneous group of patients, so that comparison is easier.

BACKGROUND: The xenografting of pre-pubertal human testicular tissue to an immunodeficient mouse is a theoretical strategy for restoring fertility in childhood cancer patients, while circumventing the risk of malignant recurrence. This study aimed at comparing the grafting of pre-pubertal and adult murine testicular tissue, as well as that of human adult testicular tissue, to two immunodeficient recipients, i.e. Swiss Nude mice and SCID-NOD mice. MATERIALS AND METHODS: In this study, we evaluated the survival of pre-pubertal and adult murine testicular tissues, and that of adult human testicular tissue after subcutaneous grafting to immunodeficient mice. RESULTS: After allografting pre-pubertal testicular tissue pieces, meiotic cells were observed in 69.1% of the grafts, while complete spermatogenesis was observed in 30.9%. All grafts of adult murine testicular tissue and 59.5% of the adult human testicular grafts showed sclerosis. However, in 21.6% of the adult human testicular grafts, spermatogonia were still observed, with increasing sclerosis in time. No significant differences were observed between the two mouse models under evaluation. CONCLUSION: After xenografting human adult testicular tissue to a recipient mouse, spermatogonia were maintained over a period of >195 days. However, in order to prove xenografting as a method for external germ line storage, the transplants should have a more immature developmental stage. Moreover, not only the developmental status of the tissue at the time-point of grafting, but also the structural organisation of the seminiferous epithelium, might influence the development of the testicular tissue.

The columns considered in this paper consist of a central mast; a split-up cross arm hinged or fixed to the central mast and optimally stressed stays. The buckling load of these columns is calculated and the influence of the geometrical parameters is examined. The indicator of volume, W, is proposed as a measure of the efficiency of different structures that are designed to transmit the same load P.This paper demonstrates that the stayed columns with a split-up cross arm have a number of advantages with respect to the stayed columns examined previously. It further reveals that both the sections of the stays and the cross arm length influence the buckling load and the efficiency decisively. Compared to other columns with one cross arm, an efficiency increase of more than 20% can be achieved. The efficiency is defined as the minimal volume of material needed to sustain a given load P.The theory of morphological indicators is used to propose a design procedure for stayed columns. This procedure takes constraints (such as local buckling, yielding, choice of the morphology...) into consideration and proposes the most efficient column. Furthermore, it takes imperfection into account as prescribed by the construction code (Eurocode 3) [Eurocode Steel/3. Belgian Standard NBN ENV 1993-1-1: 1992/AX (1992)].

Objective To provide background information about the average aneuploidy and implantation rates of older patients after IVF with preimplantation genetic diagnosis for aneuploidy screening (PGD-AS) when the patients are subdivided into age categories; and to compare pregnancy outcome data after PGD-AS in this group of patients with a similar control group. Design Retrospective clinical study. Setting Patients in an academic reproductive medicine unit. Patient(s) All patients 37 years or older who had PGD-AS between October 1999 and December 2003 and all pregnant patients 37 years or older who had IVF/intracytoplasmic sperm injection without PGD-AS during the same period of time. Intervention(s) IVF with PGD-AS. Main Outcome Measure(s) Aneuploidy rate, miscarriage rate, live birth rate, implantation rate, multiple pregnancy rate, and prenatal testing. Result(s) Three hundred ninety-four PGD-AS cycles of patients between 37 and 46 years of age were analyzed. The aneuploidy rate gradually increased with age. The implantation rate remained similar over all age groups. There was a trend to a lower miscarriage and multiple pregnancy rate in the PGD-AS group and a higher delivery/live birth rate. There were five elective terminations of pregnancy after prenatal testing and three late miscarriages due to prenatal testing in the control group. Conclusion(s) Preimplantation genetic diagnosis for aneuploidy screening can give valuable information to older patients concerning the reason why their IVF cycles are unsuccessful and whether it is worthwhile to continue IVF treatment, and it can help patients to avoid the emotional trauma that can occur after prenatal testing during the second trimester of pregnancy.

Background Eliciting an endogenous LH surge by GnRH-agonist for the induction of final oocyte maturation may be more physiological compared with the administration of HCG. However, the efficacy of this intervention in patients treated for IVF with GnRH antagonists remains to be assessed. Methods 106 patients were randomized to receive either 10 000 IU urinary HCG or 0.2 mg Triptorelin for triggering final oocyte maturation. Ovarian stimulation for IVF was performed with a fixed dose of 200 IU recombinant FSH and GnRH antagonist was started on stimulation day 6. Luteal phase was supported with micronized vaginal progesterone and oral estradiol. The study was monitored continuously for safety and stopping rules were established. Results No significant differences were present in the number of cumulus-oocyte complexes retrieved, in the proportion of metaphase II oocytes, in fertilization rates or in the number and quality of the embryos transferred between the two groups. However, a significantly lower probability of ongoing pregnancy in the GnRH agonist arm prompted discontinuation of the trial, according to the stopping rules established (odds ratio 0.11; 95% confidence interval 0.02-0.52). Conclusions Lower probability of ongoing pregnancy can be expected when GnRH agonist is used for triggering final oocyte maturation instead of HCG in patients undergoing ovarian stimulation for IVF with GnRH antagonists.

Background: Prolongation of follicular phase by delaying hCG administration has been reported to result in a significantly lower ongoing pregnancy rate that did not seem to be due to an embryonic factor. The aim of this prospective randomized study was to assess the effect of delaying hCG administration on endometrial histology. Methods: Ten oocyte donors underwent endometrial biopsy on the day of oocyte retrieval and endometrial histology was assessed by Noyes' criteria. Ovarian stimulation was performed with recombinant (r)FSH and daily GnRH antagonist starting on day 6 of stimulation. Patients were randomized by a computer-generated list to receive 10 000 IU of hCG either as soon as ≥3 follicles ≥17 mm were present on ultrasound (early-hCG group, n=5) or 2 days after this criterion was met (late-hCG group, n=5). Results: When hCG was delayed, endometrial advancement was present in all samples examined (median advancement 3 days, range 2-3 days). On the contrary, no secretory changes were observed when the follicular phase was not prolonged (difference in the proportion of patients with advancement between the early-hCG and the late-hCG group: 100%, 95% CI: 38-100). Conclusions: Prolongation of follicular phase by delaying hCG administration results in a higher incidence of endometrial advancement on the day of oocyte retrieval in GnRH antagonist cycles. © The Author 2005. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved.

In humans, the Deleted in Azoospermia Like (DAZL) gene is believed to function in the development of primordial germ cells and in germ cell differentiation and maturation because the expression of DAZL is only found in the germ and non-germ lineage of the reproductive system and in embryonic stem (ES) cells. The present study examined the presence of DAZL transcripts in the last stages of oocyte maturation, in ES cells, and throughout the preimplantation development; the link between gametes and ES cells. The finding of DAZL transcripts in the last stages of oogenesis and during the first two cell cycles of the preimplantation development was expected, because DAZL is a germ cell marker and the transcripts present at that time are generally encoded by the maternal genome. During the third cell cycle, DAZL showed a variable expression pattern, which may point to the maternal to embryonic transition. After the third cell cycle, transcripts were again consistently detected, suggesting embryonic DAZL transcription. In blastocysts, DAZL transcripts were only detected in those of good quality and this as well in the inner cell mass (ICM) as in the trophectoderm (TE). The presence of DAZL transcripts in the ICM and in ES cells was not surprising since both can lead to the formation of germ cells, but TE cells cannot. The quality-related expression of DAZL in blastocysts, and especially its trophectodermal expression, might imply other functions for DAZL beyond germ cell development.

PGD is an alternative to prenatal diagnosis that circumvents therapeutic abortion. Diagnosis is carried out on single cells obtained from three-day-old embryos, and only those that are free of the disease under consideration are transferred to the mother. Neurofibromatosis type 1 (NF1) is a common neurocutaneous disorder, inherited as an autosomal dominant trait and caused by mutations in the NF1 gene. For some patients, PGD may be the only acceptable manner to ensure the birth of unaffected children. Because of the large number of known NF1 mutations, the development of mutation-specific single-cell protocols is impractical, labour-intensive and expensive. This paper discusses the development of five PGD protocols, three of which are based on multiplex PCR for microsatellite-markers linked to the NF1 gene. After a linkage study, the diagnosis can be established through the markers, thereby obviating the need to detect the mutation itself. This not only ensures the accurate diagnosis of the embryos, but also a prompt acceptance of PGD referrals since one protocol can be useful for several couples. In addition, two mutation-specific PCRs were developed for two couples where a marker-based protocol was not applicable. In total, 16 PGD cycles were carried out for six couples, which resulted in one ongoing pregnancy and the delivery of a healthy unaffected boy.

BACKGROUND: Ovarian hyperstimulation syndrome (OHSS) in IVF/ICSI cycles may occur either as an early (early onset) or a late pattern (late onset). This observational study was designed to identify whether the onset pattern of OHSS is associated with the occurrence of pregnancy and the early pregnancy outcome. METHODS: Among 4376 consecutive IVF/ICSI cycles, 113 patients were hospitalized for OHSS after IVF/ICSI treatment and were included in the study. The setting was the Dutch-speaking Brussels Free University Hospital, between June 2000 and September 2002. RESULTS: Early OHSS occurred in 53 patients, and late OHSS complicated 60 patients. A total of 96.7% of the late OHSS cases occurred in a pregnancy cycle and were more likely to be severe than the early cases ( P< 0.05). Although in the early group there initially was a 41.5% positive HCG rate per cycle, the clinical pregnancy rate fell to 28,3% as a result of a significantly ( P< 0.05) increased preclinical pregnancy loss rate compared with the non-OHSS patients (31.8 versus 88.3%, respectively). The ongoing pregnancy rate per cycle was 14.4% in the early and 26.4% in the late group. Multiple pregnancy rates were high in both groups (40 and 45.5%, respectively), but only in the late group did the incidence reach significance compared with the non-OHSS population (45.5 versus 29.1%, P 5 0.02). Estradiol levels and number of follicles on the day of HCG were significantly higher in the early OHSS group. However, there was no difference in estradiol values on the day of hospital admittance between the two groups. In addition, the number of follicles on the day of HCG administration appears to be a better prognostic indicator for the occurrence of severe OHSS than the estradiol values (87% of the severe cases had $14 or follicles of a diameter $11 mm, whereas only 50% of them had an estradiol value $3000 ng/l). CONCLUSIONS: The early OHSS pattern is associated with exogenously administered HCG and a higher risk of preclinical miscarriage, whereas late OHSS may be closely associated with the conception cycles, especially multiple pregnancies, and is more likely to be severe. Further clarification of these two different clinical entities could have implications for research protocols as well as for preventive and management strategies for OHSS.

BACKGROUND: This was a retrospective study to determine if diagnostic testicular biopsy followed by cryopreservation should be the procedure of choice for all patients with testicular failure. METHODS: The first part of the study analysed 97 ICSI cycles scheduled with frozen‐thawed testicular sperm for 69 non-obstructive azoospermia (NOA) patients. The second part focused on a subgroup of 32 patients who underwent 42 ICSI cycles with frozen and 44 cycles with fresh testicular sperm. Sperm characteristics, fertilization, embryo quality, pregnancy and implantation rates were evaluated. RESULTS: Part I: The average time needed to find sperm was 113min per cycle and 17min per individual sperm. Fertilization rate, embryo transfer rate, ongoing pregnancy and implantation rates were 58.4%, 83%, 20.8% and 11.3%, respectively. Part II: The search time per sperm was higher (P 5 0.016) in frozen (18min) than in fresh suspensions (13min). A higher embryo transfer rate was observed in fresh cycles than in frozen cycles (93.2% vs 76.2%, P 5 0.028). Fertilization, ongoing pregnancy and implantation rates were comparable for the two groups. CONCLUSIONS: Even in a programme with low-restrictive criteria for patient allocation and for sperm cryopreservation, diagnostic testicular biopsy followed by cryopreservation can be the procedure of choice for patients with testicular failure.

BACKGROUND: It is generally accepted that the age-related increased aneuploidy rate is correlated with reduced implantation and a higher abortion rate. Therefore, advanced maternal age (AMA) couples are a good target group to assess the possible benefit of preimplantation genetic diagnosis for aneuploidy screening (PGD-AS) on the outcome after assisted reproductive technology (ART). METHODS: A prospective randomized controlled clinical trial (RCT) was carried out comparing the outcome after blastocyst transfer combined with PGD-AS using fluorescence in situ hybridization (FISH) for the chromosomes X, Y, 13, 16, 18, 21 and 22 in AMA couples (aged ≥37 years) with a control group without PGD-AS. From the 400 (200 for PGD-AS and 200 controls) couples that were allocated to the trial, an oocyte pick-up was performed effectively in 289 cycles (148 PGD-AS cycles and 141 control cycles). RESULTS: Positive serum HCG rates per transfer and per cycle were the same for PGD-AS and controls: 35.8% (19.6%) [%/per embryo transfer (per cycle)] and 32.2% (27.7%), respectively (NS). Significantly fewer embryos were transferred in the PGD-AS group than in the control group (P < 0.001). The implantation rate (with fetal heart beat) was 17.1% in the PGD-AS group versus 11.5% in the control group (not significant; P = 0.09). We observed a normal diploid status in 36.8% of the embryos. CONCLUSIONS: This RCT provides no arguments in favour of PGD-AS for improving clinical outcome per initiated cycle in patients with AMA when there are no restrictions in the number of embryos to be transferred.

The CTG repeat at the 3' untranslated region of the dystrophia myotonica protein kinase (DMPK) gene shows marked intergenerational and somatic instability in patients with myotonic dystrophy (DM1), when the repeat is expanded to more than approximately 55 repeats. Intensive research has yielded some insights into the timing and mechanism of these intergenerational changes: (1) increases in expansion sizes occur during gametogenesis but probably not during meiosis, (2) the marked somatic mosaicism becomes apparent from the 2nd trimester of development onward and increases during adult life, and (3) DNA repair mechanisms are involved. We have performed preimplantation genetic diagnosis for DM1 since 1995, which has given us the unique opportunity to study the expanded CTG repeat in affected embryos and in gametes from affected patients. We were able to demonstrate significant increases in the number of repeats in embryos from female patients with DM1 and in their immature and mature oocytes, whereas, in spermatozoa and embryos from male patients with DM1, smaller increases were detected. These data are in concordance with data on other tissues from adults and fetuses and fill a gap in our knowledge of the behavior of CTG triplet expansions in DM1.

Objective To assess the effect of altering the timing of hCG administration on ongoing pregnancy rates in patients stimulated with recombinant FSH (rec-FSH) and GnRH antagonists for IVF. Design Prospective, randomized, controlled trial. Setting Tertiary referral center. Patient(s) Four hundred thirteen patients undergoing IVF. Intervention(s) Rec-FSH stimulation starting on day 2 of the cycle combined with daily GnRH antagonist starting on day 6 of stimulation. Patients were randomized to receive 10,000 IU of hCG either as soon as at least three follicles were ≥17 mm on ultrasound (early-hCG group, 208 patients) or 2 days later after this criterion was met (late-hCG group, 205 patients). Main outcome measure(s) Ongoing pregnancy rate. Result(s) Fertilization rates and number and quality of embryos transferred did not differ between the two groups. However, a significantly lower ongoing pregnancy rate was present in the late-hCG as compared with the early-hCG group (25.0% vs. 35.6%, respectively). Conclusion(s) Prolongation of the follicular phase in patients stimulated with rec-FSH and GnRH antagonists for IVF does not affect oocyte or embryo quality but is associated with a significantly lower ongoing pregnancy rate.

Missing information about reproductive processes is an important barrier to conception and live birth among infertile couples. In vitro and genetic techniques have been informative, but not enough is known about potential defects in fertilization, implantation and early embryo development to define explicit infertility diagnoses and direct specific treatment. Study of gamete quality is needed because endocrine and ultrasound measurements in the female cannot determine oocyte quality and semen analysis in the male is a limited predictor of fertilizing ability. Also, more study would help in understanding the conditions in the Fallopian tube and elsewhere that foster fertilization and embryo development. Research in related subjects is also needed: after many years of enquiry, there exists little or no evidence to determine how and if endometriosis causes subfertility. In contrast, the recently described impact of smoking in males is large, clinically important and possibly reversible. Missing information limits the choice of specific treatment, puts a ceiling on the overall prognosis, increases the likelihood of persistent infertility and forms a barrier to understanding the broader meanings of success in the management of infertility.

Primary infertility is a key issue in the developed world, while the developing world has high rates of secondary infertility. The impact of HIV/AIDS on fertility is insufficiently explored. One of the most important barriers to access to infertility treatment is cost; at the same time the role of social and cultural factors in restricting access should not be underestimated. IVF has become the standard therapy for female infertility, and ICSI for infertility of the male partner. However, the use of these therapies should not be initiated without a thorough investigation and, whenever possible, individual diagnosis of the underlying causes of infertility. Multiple gestation remains one of the most challenging and controversial issues in the treatment of infertility. Current IVF practices are often blamed for this; in this respect, attention should also be focused on the role of ovarian stimulation in ovulation induction. National guidelines and national registries for assisted reproductive technology (ART) are becoming more widespread and are expected to play an important role in promoting best practice in ART in the future.

Background Beta-Thalassaemia results from co-inheritance of two mutant beta-globin alleles. Allogeneic cord blood cell transplantation (CBT) from an HLA-identical sibling donor is an excellent treatment option for beta-thalassaemia. In families with an affected child and willing to have another child, IVF followed by preimplantation genetic diagnosis (PGD) can be applied to exclude affected embryos. Furthermore, healthy embryos could be HLA matched with the affected child so that cord blood from the future newborn can be used to transplant the affected sibling. Methods We developed an indirect single-cell HLA typing technique based on the use of a bank of seven microsatellite markers within the HLA locus from which four informative and evenly distributed markers were selected. Results The methodology was validated in three beta-thalassaemia families having six ovarian stimulation cycles in view of IVF and PGD. Six PGD cycles were performed in two families. On 58 embryos tested, the combined PCR was successful in 54 (93%). Two transfers were done and one clinical pregnancy was obtained. Using confirmatory analysis on 50 embryos, the accuracy for HLA typing was 100%. Conclusion This strategy offers a new therapeutic option for patients with beta-thalassaemia and other monogenic diseases that can be cured with CBT.

A cohort follow-up study was designed to assess the efficacy of an intracytoplasmic sperm injection cryopreservation programme through analysis of cumulative live birth rates in successive frozen–thawed cycles in a tertiary referral centre. There were 2013 patients and they underwent 2680 frozen–thawed embryo transfer cycles. The follow-up period was between 1992 and 2001. Only frozen–thawed embryo transfer cycles up to the fourth trial were included. Crude cumulative live birth rates were calculated in five age subgroups, i.e.

Imprinting is an epigenetic mechanism leading to mono-allelic expression of imprinted genes. In order to inherit the differential epigenetic imprints from one generation to the next, these imprints have to be erased in the primordial germ cells and re-established in a sex-specific manner during gametogenesis. The exact timing of the imprint resetting is not yet known and the use of immature gametes in assisted reproductive technologies may therefore lead to abnormal imprinting and related diseases. Imprinting is associated with differential allelic methylation in a CpG-context. We studied the methylation patterns of the imprint control (IC) region of the human SNRPN-gene in human spermatozoa, oocytes in different developmental stages [germinal vesicle (GV), metaphase I and metaphase II oocytes] and in preimplantation embryos using the bisulphite sequencing technique. In the spermatozoa, almost all potential methylation sites were unmethylated whereas mainly methylated patterns were found in the oocytes at different developmental stages. In the embryos, an average methylation pattern of 53% was found indicating that the imprints, which have been set during gametogenesis, are stably maintained in the preimplantation embryo. Our results indicate that the maternal imprints for the IC-region of the human SNRPN-gene are already re-established at the GV stage and that they are not re-established in a late oocyte stage or after fertilization as previously reported. Recent advances in assisted reproductive technologies raise questions concerning safety and the epigenetic risks involved. Our study was the first to check the methylation imprints in human pre-implantation embryos and oocytes at different developmental stages.

BACKGROUND: The purpose of this study was to assess cumulative delivery rates in patients with non-obstructive or obstructive azoospermia following treatment by testicular sperm extraction (TESE)‐ICSI. METHODS: A cohort follow-up study was conducted. Between January 1994 and December 2000, 364 couples with obstructive azoospermia underwent a total of 609 fresh TESE‐ICSI treatment cycles. In addition, 303 fresh TESE‐ICSI treatment cycles were performed in 235 couples for non-obstructive azoospermia. This study included only patients in whom sperm was recovered. In the non-obstructive group, only patients with maturation arrest, atrophic sclerosis and germ cell aplasia were included. The main outcome measure was a delivery beyond 25 weeks gestation. RESULTS: In patients with obstructive azoospermia, the crude delivery rate after three cycles was 35% while the expected cumulative delivery rate was 48% [95% confidence interval (CI), 41‐55]. On the other hand, in patients with non-obstructive azoospermia, the crude cumulative delivery rate after three treatment cycles was 17% while the expected delivery rate was 31% (95% CI, 15‐46). A high dropout rate in couples with both non-obstructive and obstructive azoospermia was observed (75 and 50% respectively, after the first cycle). CONCLUSION: This study shows that there is a value in performing several TESE‐ICSI attempts in patients with obstructive and non-obstructive azoospermia. The estimates of the non-obstructive group beginning from the third cycle are less reliable due to fewer patients. However, overall, the obstructive group performed better than the non-obstructive group.