More than an add-on? The Europeanization of the Dutch civil service

[From the introduction]. European integration does not stop to fascinate political scientists. Many of us are excited about this institution that transcends national interests, overcomes collective action problems, and presents member states with such a durable and authoritative framework that they slowly but unrecognizably loose authority to model their own policies as desired. But does it? Despite our excitement, many of us have troubles escaping the reflexes caused by the years of international relations hegemony in studying the EU. Does the EU really have the clout to force member states to adopt unwanted policies? Then how about the never-ending stories about non-compliance, the European Commission’s hesitance in adopting a tough stance on reluctant member states, the difficulties of monitoring actual application and enforcement on the ground? The tension between member state dominance and supranational control continues to offer a well of fascinating research topics. In order to demonstrate the success of the EU in transcending member states’ institutions and policies, or even the domestic interests underlying them, we are advised to answer at least three questions. First, we should answer the question of the extent to which Europe matters for the member states. Because even if we can identify compliance by initially reluctant member states, this may not be very meaningful if the EU’s share in national matters is only minimal. Even though interesting from a theoretical viewpoint, the societal relevance of massive research attempts to explain the fate of EU intervention in member states is slight when it affects only a minimal terrain of national policy making. Second, we should try to answer the question to what extent any processes of Europeanization we observe are truly affecting the core of what member states are doing or are just added on to existing structures and policies. That is, if we believe that the EU really is capable of overriding member state concerns, the adaptations made by member states should be far from ‘easy’. The adoption of coordination structures, for instance, is an interesting phenomenon, but it does not constitute evidence of the EU’s transformative effect as coordination structures may simply be added on to existing organizational arrangements and can perfectly well co-exist with domestic institutions that were already out there. Finally, we should answer the question of how the European Union impacts on member states. Under what conditions does the EU succeed in bringing about domestic change, and when do member states carry on their business as usual

Female subfertility

The WHO defines female subfertility as failure to achieve a clinical pregnancy after 12 months of regular intercourse or due to impairment of a woman’s capacity to reproduce. This PrimeView highlights some of the mechanisms that may contribute to this condition

Comparison of DNA methylation patterns of parentally imprinted genes in placenta derived from IVF conceptions in two different culture media

Study question: Is there a difference in DNA methylation status of imprinted genes in placentas derived from IVF conceptions where embryo culture was performed in human tubal fluid (HTF) versus G5 culture medium? Summary answer: We found no statistically significant differences in the mean DNA methylation status of differentially methylated regions (DMRs) associated with parentally imprinted genes in placentas derived from IVF conceptions cultured in HTF versus G5 culture medium. What is known already: Animal studies indicate that the embryo culture environment affects the DNA methylation status of the embryo. In humans, birthweight is known to be affected by the type of embryo culture medium used. The effect of embryo culture media on pregnancy, birth and child development may thus be mediated by differential methylation of parentally imprinted genes in the placenta. Study design, size, duration: To identify differential DNA methylation of imprinted genes in human placenta derived from IVF conceptions exposed to HTF or G5 embryo culture medium, placenta samples (n = 43 for HTF, n = 54 for G5) were collected between 2010 and 2012 s as part of a multi-center randomized controlled trial in the Netherlands comparing these embryo culture media. Placenta samples from 69 naturally conceived (NC) live births were collected during 2008-2013 in the Netherlands as reference material. Participants/materials, setting, methods: To identify differential DNA methylation of imprinted genes, we opted for an amplicon-based sequencing strategy on an Illumina MiSeq sequencing platform. DNA was isolated and 34 DMRs associated with well-defined parentally imprinted genes were amplified in a two-step PCR before sequencing using MiSeq technology. Sequencing data were analyzed in a multivariate fashion to eliminate possible confounding effects. Main results and the role of chance: We found no statistically significant differences in the mean DNA methylation status of any of the imprinted DMRs in placentas derived from IVF conceptions cultured in HTF or G5 culture medium. We also did not observe any differences in the mean methylation status per amplicon nor in the variance in methylation per amplicon between the two culture medium groups. A separate surrogate variable analysis also demonstrated that the IVF culture medium was not associated with the DNA methylation status of these DMRs. The mean methylation level and variance per CpG was equal between HTF and G5 placenta. Additional comparison of DNA methylation status of NC placenta samples revealed no statistically significant differences in mean amplicon and CpG methylation between G5, HTF and NC placenta; however, the number of placenta samples exhibiting outlier methylation levels was higher in IVF placenta compared to NC (P < 0.00001). Also, we were able to identify 37 CpG sites that uniquely displayed outlier methylation in G5 placentas and 32 CpG sites that uniquely displayed outlier methylation in HTF. In 8/37 (G5) and 4/32 (HTF) unique outliers CpGs, a medium-specific unique outlier could be directly correlated to outlier methylation of the entire amplicon. Limitations, reasons for caution: Due to practical reasons, not all placentas were collected during the trial, and we collected the placentas from natural conceptions from a different cohort, potentially creating bias. We limited ourselves to the DNA methylation status of 34 imprinted DMRs, and we studied only the placenta and no other embryo-derived tissues. Wider implications of the findings: It has often been postulated, but has yet to be rigorously tested, that imprinting mediates the effects of embryo culture conditions on pregnancy, birth and child development in humans. Since we did not detect any statistically significant effects of embryo culture conditions on methylation status of imprinted genes in the placenta, this suggests that other unexplored mechanisms may underlie these effects. The biological and clinical relevance of detected outliers with respect to methylation levels of CpGs and DMR require additional analysis in a larger sample size as well. Given the importance and the growing number of children born through IVF, research into these molecular mechanisms is urgently needed. Study funding/competing interest(s): This study was funded by the March of Dimes grant number #6-FY13-153. The authors have no conflicts of interest. Trial registration number: Placental biopsies were obtained under Netherlands Trial Registry number 1979 and 1298

Comparing the cumulative live birth rate of cleavage-stage versus blastocyst-stage embryo transfers between IVF cycles:a study protocol for a multicentre randomised controlled superiority trial (the ToF trial)

Introduction In vitro fertilisation (IVF) has evolved as an intervention of choice to help couples with infertility to conceive. In the last decade, a strategy change in the day of embryo transfer has been developed. Many IVF centres choose nowadays to transfer at later stages of embryo development, for example, transferring embryos at blastocyst stage instead of cleavage stage. However, it still is not known which embryo transfer policy in IVF is more efficient in terms of cumulative live birth rate (cLBR), following a fresh and the subsequent frozen-thawed transfers after one oocyte retrieval. Furthermore, studies reporting on obstetric and neonatal outcomes from both transfer policies are limited. Methods and analysis We have set up a multicentre randomised superiority trial in the Netherlands, named the Three or Fivetrial. We plan to include 1200 women with an indication for IVF with at least four embryos available on day 2 after the oocyte retrieval. Women are randomly allocated to either (1) control group: embryo transfer on day 3 and cryopreservation of supernumerary good-quality embryos on day 3 or 4, or (2) intervention group: embryo transfer on day 5 and cryopreservation of supernumerary good-quality embryos on day 5 or 6. The primary outcome is the cLBR per oocyte retrieval. Secondary outcomes include LBR following fresh transfer, multiple pregnancy rate and time until pregnancy leading a live birth. We will also assess the obstetric and neonatal outcomes, costs and patients' treatment burden. Ethics and dissemination The study protocol has been approved by the Central Committee on Research involving Human Subjects in the Netherlands in June 2018 (CCMO NL 64060.000.18). The results of this trial will be submitted for publication in international peer-reviewed and in open access journals. Trial registration number Netherlands Trial Register (NL 6857)

Preimplantation genetic screening: back to the future

All agree that in hindsight the rapid adoption of preimplantation genetic screening (PGS) using cleavage stage biopsy and fluorescence in situ hybridization (FISH) in routine clinical practice without proper evaluation of (cost-)effectiveness basically resulted in couples paying more money for a less effective treatment. Now, almost 20 years later, we are on the verge of a new era of PGS. But have things really changed or are we simply going back to the future

Limitations of embryo selection methods

In in vitro fertilization (IVF), the selection of embryos for transfer is generally based on the morphology of the available embryos. However, not all embryos with good morphology implant and on average only one in four treatments are successful. This has driven a quest for alternative selection methods. The best known alternative selection method is preimplantation genetic screening (PGS), which has been used for over a decade before it was shown to be inferior to morphological selection. Now, new forms of PGS (performing biopsy at another stage of development and new methods for analysis) are emerging, just like alternative noninvasive embryo selection methods. However, the concept that better selection will lead to improved IVF results is not so certain anymore. Evidence is accumulating that all available embryos in an IVF cycle can be cryopreserved and transferred in subsequent cycles without impairing pregnancy rates or maybe even with an improvement in pregnancy rates. Embryo selection will then no longer be able to improve the live birth rate in IVF; it could even lower the live birth rate. Embryo selection will only be able to improve the time to pregnancy, if embryos with the highest implantation potential are transferred firs

Cryopreservation of human embryos and its contribution to in vitro fertilization success rates

Cryopreservation of human embryos is now a routine procedure in assisted reproductive technologies laboratories. There is no consensus on the superiority of any protocol, and substantial differences exist among centers in day of embryo cryopreservation, freezing method, selection criteria for which embryos to freeze, method of embryo thawing, and endometrial preparation for transfer of frozen-thawed embryos. In the past decade, the number of frozen-thawed embryo transfer cycles per started in vitro fertilization (IVF) cycle increased steadily, and at the same time the percentage of frozen-thawed embryo transfers that resulted in live births increased. Currently, cryopreservation of human embryos is more important than ever for the cumulative pregnancy rate after IVF. Interestingly, success rates after frozen-thawed embryo transfer are now nearing the success rates of fresh embryo transfer. This supports the hypothesis of so called freeze-all strategies in IVF, in which all embryos are frozen and no fresh transfer is conducted, to optimize success rates. High-quality randomized controlled trials should be pursued to find out which cryopreservation protocol is best and whether the time has come to completely abandon fresh transfer