Reproductive BioMedicine Online - Articles in Press

Chromosomal aberrations in 2000 couples of Indian ethnicity with reproductive failure - Accepted Manuscript

S. Gada Saxena, K. Desai, L. Shewale, P. Ranjan, D. Saranath — 2012-05-01

Abstract: Constitutional chromosomal aberrations contribute to infertility and repeated miscarriage leading to reproductive failure in couples. These aberrations may show no obvious clinical manifestations and remain undetected across multiple generations. However, infertility or recurrent spontaneous pregnancy loss, and/or genotypic/phenotypic aberrations may be manifested in the progeny during gametogenesis. The current study was a retrospective analysis to examine the chromosomal aberrations and prevalence in 2000 couples of Indian ethnicity with reproductive failure. Cytogenetic analysis via conventional G-band karyotyping analysis was carried out on phytohaemagglutinin stimulated peripheral blood lymphocytes, cultured in RPMI1640 medium. The chromosomes were enumerated as per International System for Human Cytogenetic Nomenclature at 500-550 band resolution, and recorded in the screening sheets. Chromosomal aberrations were detected in a total of 110 (2.78%) couples, with structural chromosomal aberrations in 88 cases including reciprocal translocations in 56 cases, Robertsonian translocations in 16 cases, inversions in eight cases, deletions in three cases, derivative chromosomes in five cases and numerical chromosome aberrations in 23 cases. The study emphasizes the importance of cytogenetic work up in both the partners associated with a history of reproductive failure. Genetic counselling with an option of prenatal diagnosis should be offered to couples with chromosomal aberrations.

The clinical benefit and safety of current and future assisted reproductive technology - Accepted Manuscript

Rachel Brown, Joyce Harper — 2012-05-01

Abstract: Since the first birth achieved by IVF was achieved in 1978, the techniques involved in assisted reproductive technology have grown at an enormous rate. However, new technology has rarely been robustly validated before clinical use and developing scientific understanding of the available techniques has done little to alter their use. Furthermore, there are inconsistencies in the available clinical studies and endpoints. The benefits of some technologies already established for routine use are currently dubious and there are clear ethical concerns with providing them to patients when their scientific basis is not clear. As the uptake of assisted reproductive technology increases and newer technologies continue to push the boundaries of science, it is important to consider the clinical benefits and safety of all assisted reproductive technologies. This review will discuss aspects of some of the more recent techniques, including sperm DNA-damage tests, intracytoplasmic morphologically selected sperm injection, amino acid and metabolomics profiling, preimplantation genetic screening and time-lapse imaging, and those that may have substantial impacts on the field of reproductive medicine in the future including artificial gametes, ovarian transplantation and gene therapy.

PAX2 in 192 Chinese women with Müllerian duct abnormalities: mutation analysis - Accepted Manuscript

Peng Wang, Han Zhao, Mei Sun, Yuan Li, Zi-Jiang Chen — 2012-05-01

Abstract: The paired box gene 2 (PAX2) has been proven to be a crucial gene during organogenesis of the urogenital system in mice models. This study was aimed to explore the relationship between PAX2 mutations and human Müllerian duct abnormalities (MDA). A total of 192 Chinese MDA patients (15 cases of uterine aplasia and 177 of incomplete Müllerian fusion) and 192 ethnic-matched controls were recruited from 2009 to 2011. Coding regions of PAX2 of MDA cases were amplified and sequenced. One rare novel synonymous variant (c.320G>A) was discovered in one patient with uterus didelphys, whereas this variant was not found in the control group. Mutations in PAX2 may be not a common cause of MDA.

IVF and embryo transfer: historical origin and development - Accepted Manuscript

John D. Biggers — 2012-05-01

Abstract: IVF and embryo transfer for the treatment of human infertility has now resulted in the birth of over 4 million babies. The technique did not arise as a quantum event but was built on the efforts of many earlier workers in the fields of reproductive endocrinology and development. One should remember the famous saying of Isaac Newton: ‘If I have seen further than most, it is because I have stood on the shoulder’s of giants’. Ethical and moral issues have always arisen when investigators study early mammalian development, particularly human development. This paper documents these earlier studies and also draws attention to the ethical and moral arguments that inevitably arose.

Hype or hope? Ethical and practical considerations with clinical research in women with diminished ovarian reserve - Accepted Manuscript

Norbert Gleicher, David H. Barad — 2012-04-23

Abstract: This communication suggests that investigations of treatments for women with diminished functional ovarian reserve (DOR) call for specific practical and ethical considerations, as women with DOR, because of limited remaining reproductive life spans, appropriately feel under time constraints. Another medical journal recently published an opinion piece on the use of dehydroepiandrosterone in women with DOR, raising important questions about what approaches should be taken to develop best available evidence in such patients. Their manuscript offers an excellent opportunity to consider ethical and clinical aspects of study design in clinical circumstances where patients have little to lose but face the promise of considerable gains in clinical pregnancy chances if effective treatments can be developed. This commentary concludes that, in such circumstances, common sense as well as ethical considerations support the introduction of new treatments into the clinical mainstream even in absence of prospectively randomized studies if lower levels of evidence are supportive of positive treatment effects.

Progesterone support in IVF: is evidence-based medicine translated to clinical practice? A worldwide web-based survey - Accepted Manuscript

Edi Vaisbuch, Milton Leong, Zeev Shoham — 2012-04-23

Abstract: This worldwide web-based survey compared the clinical practice for luteal-phase supplementation (LPS) in stimulated IVF cycles to the current evidence-based literature. Eighty-four treatment centres in 35 countries, representing a total of 51,155 IVF cycles/year, responded. Vaginal progesterone alone was used for LPS in 64% of cycles and in another 16% of cycles in combination with either i.m. (15%) or oral progesterone (1%). As a single agent, i.m. progesterone was used in 13% of cycles, oral progesterone in another 2% and human chorionic gonadotrophin (HCG) was still used in 5% of cycles. Progesterone was administered until 10–12 weeks’ gestation in 67% of cycles and in 22% and 12% it was discontinued when fetal heart pulsations are recognized or until β HCG was positive, respectively. In conclusion, in almost two-thirds of the assisted cycles represented in this survey, vaginal administration of progesterone is preferred for LPS. Nevertheless, despite the available literature on the disadvantages of oral progesterone, i.m. progesterone and HCG for LPS, these agents are still used routinely by many practitioners. Furthermore, although there is no firm evidence to support the continuation of LPS until 10–12 weeks’ gestation, this practice is used in the majority of IVF cycles worldwide.

Impact of an educational intervention and insurance coverage on patients’ preferences to transfer multiple embryos - Accepted Manuscript

2012-04-23

Abstract: Multiple gestations resulting from IVF continue to be a major problem associated with maternal/neonatal morbidity and mortality including preterm labour/delivery, pre-eclampsia and post-partum haemorrhage. A prospective survey at a university IVF clinic evaluated the effect of education and insurance coverage on patients’ preferences for single-embryo transfer (SET) versus double-embryo transfer (DET). Patients undergoing IVF treatment from September 2008 to October 2009 were included. The main outcome measure was patients’ preference of SET versus DET. Patients were sent an educational handout describing maternal and fetal risks of twin gestation. A total of 163 patients (32.6% response rate) returned the pre- and post-education surveys regarding preferences for SET versus DET based on three different IVF insurance coverage scenarios (no coverage, two cycles covered and unlimited coverage). There were statistically significant differences in the preference for SET before and after education across all insurance scenarios (scenario 1, 42.0% versus 61.1%; scenario 2, 50.6% versus 71.0%; and scenario 3, 61.7% versus 79.6%; P < 0.001 for all scenarios). Before education, patients preferred SET more in the unlimited coverage scenario (61.7%) versus no coverage (42.0%; P < 0.001). An educational handout and increasing the amount of insurance coverage significantly increased a patient’s preference for SET.

Fighting ‘personhood’ initiatives in the United States - Corrected Proof

Lee Rubin Collins, Susan L. Crockin — 2012-04-13

Abstract: ‘Personhood’ initiatives filed in many states within the United States threaten to impose potentially significant restrictions on infertility treatment, embryo disposition, pre-natal care, abortion, contraception, and stem-cell research, all through attempts to redefine a ‘person’ or ‘human being’ as existing from the moment of fertilization or conception, and endowed with the full legal and Constitutional rights of personhood. Virginia’s recent, unsuccessful attempt to pass such legislation provides both a dramatic example of these efforts and valuable lessons in the fight against them by infertility advocates and others. Arguments over loss of infertility treatment seemed more persuasive to legislatures than did restrictions on abortion or stem cell research. Indeed, persuading legislators or voters that they could be ‘pro-life’ and still anti-personhood initiatives was a key strategy, and consumer efforts and media attention were instrumental. The most central lessons, however, may be the degree of intensity and coordinated strategy to shift public perception that lie behind these numerous state efforts, regardless of whether the actual initiatives are won or lost.

Long-term duration of function of ovarian tissue transplants: case reports - Accepted Manuscript

2012-04-09

Abstract: These three case reports describe the long-term duration of function of ovarian cortical tissue grafts among patients in a university fertility preservation programme in Europe and in a private practice programme in the USA. One woman underwent sterilizing cancer treatment and had frozen ovarian tissue transplanted, and two women underwent fresh ovarian tissue transplants. The function of ovarian cortical strips has continued for more than 7 years in these three women, with the birth of eight healthy babies following a single graft per patient. In addition to these three cases, transplantation (repeatedly in some cases) of cryopreserved ovarian tissue has restored reproductive function to all other women in the study centres’ programmes for some years. The sustained longevity of function of the transplanted tissue suggests that it may also be possible to postpone the normal time of menopause or to alleviate its symptoms.

Estradiol valerate pretreatment in GnRH-antagonist cycles - Accepted Manuscript

C. Blockeel — 2012-04-09

NADPH oxidase as an important source of reactive oxygen species at the mouse maternal–fetal interface: putative biological roles - Corrected Proof

2012-04-09

Abstract: Oxygen derivatives that comprise the large family of reactive oxygen species (ROS) are actively involved in placental biology. They are generated at the maternal–fetal interface at the level of decidual, trophoblast and mesenchymal components. In normal conditions, ROS produced in low concentrations participate in different functions as signalling molecules, regulating activation of redox-sensitive transcription factors and protein kinases involved in cell survival, proliferation and apoptosis, hence much of cell functioning. Physiological ROS generation is also associated with such defence mechanisms as phagocytosis and microbiocidal activities. In mice, particularly but not exclusively, trophoblast cells phagocytose intensively during implantation and post-implantation periods and express enzymic machinery to address a ROS-producing response to changes in the environment. The cells directly associated with ROS production are trophoblast giant cells, which mediate each and every relationship with the maternal organism. In this review, the production of ROS by the implanting mouse trophoblast is discussed, focusing on NADPH oxidase expression, regulatory mechanisms and similarities with NOX2 from phagocytes. Some of the current controversies are assessed by attempting to integrate data from studies in human trophoblast and mouse models.Although oxygen is vital for life, in specific conditions it can also give rise to unstable molecules, the reactive oxygen species (ROS). In physiological conditions, ROS are used as a defence mechanism by immune cells and as messengers for cell survival, proliferation and death among neighbouring cells. However, due to their high capability to react with other organic molecules changing their structure and function, such as proteins, lipids and DNA, ROS can also cause biological damage, which in general occurs when natural antioxidant mechanisms are unable to counterbalance the reactivity of these molecules. NAD(P)H oxidase is an enzymic complex able to generate a large amount of ROS. Production of ROS has been described at the maternal–fetal interface in many species and is associated with an unbalance and gestational diseases such as pre-eclampsia and hypertension. This review discusses the physiological NADPH oxidase expression by the fetal component of the mouse placenta, the trophoblast, which mediates each and every relationship with the maternal organism.

Placental adaptations to the maternal–fetal environment: implications for fetal growth and developmental programming - Corrected Proof

Ionel Sandovici, Katharina Hoelle, Emily Angiolini, Miguel Constância — 2012-04-09

Abstract: The placenta is a transient organ found in eutherian mammals that evolved primarily to provide nutrients for the developing fetus. The placenta exchanges a wide array of nutrients, endocrine signals, cytokines and growth factors with the mother and the fetus, thereby regulating intrauterine development. Recent studies show that the placenta is not just a passive organ mediating maternal–fetal exchange. It can adapt its capacity to supply nutrients in response to intrinsic and extrinsic variations in the maternal–fetal environment. These dynamic adaptations are thought to occur to maximize fetal growth and viability at birth in the prevailing conditions in utero. However, some of these adaptations may also affect the development of individual fetal tissues, with patho-physiological consequences long after birth. Here, this review summarizes current knowledge on the causes, possible mechanisms and consequences of placental adaptive responses, with a focus on the regulation of transporter-mediated processes for nutrients. This review also highlights the emerging roles that imprinted genes and epigenetic mechanisms of gene regulation may play in placental adaptations to the maternal–fetal environment.The placenta regulates intrauterine development in mammals by performing the function of several adult organs for the growing fetus. These tasks are achieved by a multitude of specialized cell types. Recent work suggests the placenta has the ability to ‘sense’ the maternal and fetal environment and respond to changes in a dynamic fashion. These placental adaptations are particularly important when dealing with suboptimal conditions for growth and development. In this paper, we review the current knowledge on what triggers these adaptations, how the placenta responds to maternal and fetal signals, what these signals might be and what the long-term consequences are for the function of adult organs if the placenta fails to fully adapt. We argue that a special class of genes, so-called imprinted genes, are key regulators of placental adaptive responses to physiological stressors. These genes are different from the rest because they retain information about their parental origin and because they have evolved to be ‘selfish’. They play special roles in the placenta, as we will describe, which include controlling the amounts of maternal nutrients that go across to the fetus and the manipulation of maternal physiology, sometimes in conflict with the mother’s own interests.

A placenta for life - Corrected Proof

Rosalind John, Myriam Hemberger — 2012-04-09

Abstract: The chorioallantoic placenta is the defining organ of eutherians that has enabled prolonged intrauterine gestation. As such, normal placental development and function are essential for mammalian reproductive success. Reflecting the key role of this organ in providing nutrients to the embryo, the characteristic cell type that forms substantial parts of the placenta is called ‘trophoblast’ (from Greek trephein ‘to feed’ and blastos ‘germinator’). However, in addition to regulating nutrient supply, the placenta also exerts a number of other pivotal functions that highlight the importance of normal trophoblast differentiation for a successful pregnancy. In this guest symposium, ‘Trophoblast Development’, several contributors summarize insights gained from recent studies in the mouse that have advanced our understanding of trophoblast biology. This includes how the earliest trophoblast cells are set aside to expand in a stem- or progenitor-cell compartment under tight genetic and epigenetic control and how subsequent differentiation into the various placental cell types is controlled to ensure normal placentation. The relevance of these contributions range from early developmental cell fate decisions, stem cell biology and placental development for healthy pregnancy to the impact of placental failures on long-term health, with important clinical implications for assisted reproductive technology procedures and pregnancy-associated complications.Mammals give birth to live young which have been nurtured in utero by the mother via an organ called the placenta. The placenta is derived primarily from a specialized cell lineage provided by the embryo, called the trophoblast. This trophoblast lineage is responsible for communicating with the mother, supplying nutrients to the growing embryo and removing waste products. While the appearance and structure of the placenta varies widely between different mammalian species, these key functions are conserved. The genetically manipulatable mouse has provided a tool to complement studies on human placental material to help us understand both the genetic and epigenetic processes required to generate a functional placenta essential for both short-term wellbeing and long-term health. In this symposium, several contributors highlight novel insights in our understanding of how development of this critical organ is genetically and epigenetically controlled.

The placental imprintome and imprinted gene function in the trophoblast glycogen cell lineage - Corrected Proof

Louis Lefebvre — 2012-04-06

Abstract: Imprinted genes represent a unique class of autosomal genes expressed from only one of the parental alleles during development. The choice of the expressed allele is not random but rather is determined by the parental origin of the allele. Consequently, the mouse genome contains more than 100 genes expressed preferentially or exclusively from the maternally or the paternally inherited allele. Current research efforts are focused on understanding the molecular mechanism of this epigenetic phenomenon as well as the biological functions of the genes under its regulation. Both theoretical considerations and experimental results support a role for genomic imprinting in the regulation of embryonic growth and placental biology. In this review, recent efforts to establish the complete set of genes showing imprinted expression in the mouse placenta are first discussed. Then, the evidence suggesting that imprinted genes might be implicated in the emergence, maintenance and function of trophoblast glycogen cells is presented. Although the origin and functions of this trophoblast cell lineage are currently unknown, the analysis of mutations in imprinted genes in the mouse are providing new insights into these issues. The implications of this work for placental pathologies in human are also discussed.

Trophoblast tales - Corrected Proof

Martin H. Johnson, Jacques Cohen, Gedis Grudzinskas — 2012-04-06

The placenta has to be the most complex organ in the body – genetically, functionally and structurally. It is uniquely complex genetically, comprising tissues of both maternal and fetal genotype in close apposition, as well as having a complex gene expression profile according to the parental epigenetic imprint inherited on selected genes. It is complex functionally in its sheer range of hormonal products, combining roles assigned to many or most other endocrine glands, and also in its transport systems, its physical anchoring role, and in its immunological properties. Structural complexity is evident in the placenta’s location between, and mixture of, maternal and fetal tissues, as well as in its cellular diversity, unique tissue organisation and the release of its cells into the maternal circulation from whence they can be harvested for genetic testing. Moreover, each of these complexities shows marked and rapid changes as pregnancy progresses, from the early delineation of placental lineages among the zygote-derived cells in the pre-embryo, through contained invasiveness at implantation, to the proliferation of cell types and tissue functions during embryogenesis. The transition to the fetal stage marks a major shift in most eutherians, in which the haemotrophic chorio-allantoic placenta supplants the previously dominant histiotrophic chorio-vitelline or yolk sac placenta – a transition neither observed nor required in marsupials, with their early delivery of young to the pouch. In eutherians, this transition ensures that the delicate and seminal events of embryogenesis occur in an environment protected from potentially damaging reactive oxygen species. Given the complexity, and relative inaccessibility, of this extra-ordinary organ – or should one more properly say organs such are its remarkable transitions? – it is perhaps not surprising that until recently our understanding of it was so rudimentary. There can be few organs of such central significance to our very existence that have been less studied – indeed, usually it is either discarded after birth or transformed into ‘rejuvenating’ skin creams! The relatively recent emergence of the placenta from this shadowy ignorance has been stimulated in part by the creation five years ago of the Centre for Trophoblast Research (CTR). This virtual centre is based in the University of Cambridge, but has global affiliations. However, it does build upon strong local Cambridge traditions in placental and trophoblast research, as represented especially by Barcroft, Boyd, Loke and Steven (). It also draws on and stimulates local current research and expertise in the University and the adjacent Gurdon and Babraham Institutes. By providing resources and a community of interests, the CTR encourages cooperation within and well beyond Cambridge. This wide-sprung cooperativity is reflected in the Symposium in this issue edited by Myriam Hemberger (Cambridge) and Rosalind John (Cardiff) to mark the fifth anniversary of the CTR. Graham Burton, the driving force behind the formation of the CTR and its first director, writes about the Centre itself, and we have six further contributions from Brazil, Canada, and Germany as well as Cambridge: an exciting set of papers with a thoughtful introduction by our two editors. Our warmest thanks to Myriam and Ros for doing an excellent editorial job and being such a joy to work with. We hope you enjoy and are informed!

Anti-Müllerian hormone in women with hypopituitarism diagnosed before or during adolescence - Accepted Manuscript

B. Sonntag, F. Nawroth, M. Ludwig, C. Bullmann — 2012-04-05

Abstract: In women, anti-Müllerian hormone (AMH) is exclusively expressed in granulosa cells and an established marker of ovarian reserve. In menstrual cycle disorders, low AMH is usually interpreted as an indicator of primary ovarian insufficiency. This study is a case series of 11 patients with hypopituitarism. AMH concentrations were on or below the age-specific 25th percentile in three of the four patients diagnosed in infancy, but not in the remaining seven patients, who were diagnosed during adolescence or later. In patients with hypopituitarism, the detection of low AMH serum concentrations can present a diagnostic pitfall and its value in the interpretation of ovarian reserve in these patients is challenging.

Intracytoplasmic morphologically selected sperm injection improves development and quality of preimplantation embryos in teratozoospermia patients - Accepted Manuscript

Katja Knez, Tomaz Tomazevic, Branko Zorn, Eda Vrtacnik-Bokal, Irma Virant-Klun — 2012-04-05

Abstract: This prospective randomized study investigated whether intracytoplasmic sperm injection (ICSI) outcome can be improved with sperm preselection under magnification × 6000 and intracytoplasmic morphologically selected sperm injection (IMSI) in patients with teratozoospermia and characterized embryo development and quality regarding sperm morphology and presence of head vacuoles. Couples with isolated teratozoospermia were divided into two groups: IMSI group (n = 52) and ICSI group (n = 70) and fertilization, blastocyst and clinical pregnancy rates were compared. Oocytes from 30 randomly chosen patients from the IMSI group were injected with spermatozoa that had been previously classified under × 6000 magnification into four classes according to number and size of vacuoles in the head and then cultured separately. Pronuclear morphology, embryo development and blastomere viability were estimated to investigate the influence of sperm morphology, especially vacuoles, on embryo developmental capacity. A significantly higher clinical pregnancy rate was achieved in the IMSI group compared with the ICSI group (48% versus 24%, P < 0.05). Fertilization with spermatozoa without head vacuoles yielded higher number of morphologically normal zygotes, higher blastocyst rate and smaller proportion of arrested embryos than spermatozoa with vacuoles and other head defects. IMSI is a method of choice in patients with teratozoospermia.

Relationship between psychological stress and recurrent miscarriage - Accepted Manuscript

W. Li, J. Newell-Price, G.L. Jones, W.L. Ledger, T.C. Li — 2012-04-05

Abstract: Stress arousal may compromise the feedback regulation of the hypothalamo–pituitary–adrenal axis, releasing stress-related biomarkers and thereby affecting establishment of pregnancy. This study examined the relationship between stress and recurrent miscarriage (RM) and the impact of stress on establishment of pregnancy. The stress status of 45 patients with unexplained RM and 40 fertile women was investigated with the Fertility Problem Inventory (FPI), Perceived Stress Scale (PSS), Positive and Negative Affect Schedule, peripheral natural killer (NK) cells and cortisol. Patients with unexplained RM had significantly higher scores on the FPI (P < 0.05, adjusted OR 1.02), PSS (P < 0.05, adjusted OR 1.13) and Negative Affect scale (P < 0.05, adjusted OR 1.12) and lower scores on the Positive Affect scale (P < 0.05, adjusted OR 0.89) than fertile controls. Patients who had live births (n = 20) during the study period had significantly lower scores in the Positive Affect scale (P < 0.05, adjusted OR 1.17) than those who miscarried (n = 10). There was little association between psychological stress measurements and biochemical stress measurements. These results suggest that stress is a risk factor of RM. Within women with RM, moderate stress appears to be associated with improved pregnancy outcome.

The use of vaginal natural progesterone for prevention of preterm birth in IVF/ICSI pregnancies - Accepted Manuscript

2012-04-05

Abstract: The aim of this study was to evaluate the effect of vaginal natural progesterone on the prevention of preterm birth in IVF/intracytoplasmic sperm injection (ICSI) pregnancies. A single-centre prospective placebo-controlled randomized study was performed. A total of 313 IVF/ICSI pregnant patients were randomized into two groups for either treatment with daily 400 mg vaginal natural progesterone or placebo, starting from mid-trimester up to 37 weeks or delivery. Amongst the patients, there were 215 singleton and 91 twin pregnancies. There was no significant difference in risk of preterm birth among all patients (OR 0.672, 95% CI 0.42–1.0. There was a significantly lower preterm birth rate in singleton pregnancies in the natural progesterone arm (OR 0.53, 95% CI 0.28–0.97) and no significant difference between both arms in twin pregnancies (OR 0.735, 95% CI 0.36–2). In conclusion, the administration of 400 mg vaginal natural progesterone from mid trimester reduced the incidence of preterm birth in singleton, but not in twin, IVF/ICSI pregnancies.

Intensive luteal-phase support with oestradiol and progesterone after GnRH-agonist triggering: does it help? - Corrected Proof

Raoul Orvieto — 2012-03-19

We read with interest the recent review by which discussed comprehensively the literature concerning the luteal phase after GnRH-agonist (GnRHa) triggering of ovulation, including suggested modification of luteal-phase support. We agree with the authors’ conclusion that the most plausible reason for the luteal-phase defect seen in cycles triggered with GnRHa seems to be a lack of endogenous LH activity during the early to mid-luteal phase, an assumption that might be seen to be supported by the beneficial effect of adding HCG () or recombinant LH () to the luteal-phase support. However, we are more sceptical regarding the concept of overcoming the luteal-phase insufficiency seen after GnRHa triggering via intensive luteal support using oestradiol and progesterone, as described by .

Response: Intensive luteal phase support with oestradiol and progesterone after GnRH agonist triggering: does it help? - Corrected Proof

Peter Humaidan — 2012-03-19

I would like to thank Professor Orvieto for his interest in our latest review () in which we discuss the luteal phase insufficiency seen after GnRH agonist (GnRHa) trigger and the various trials that have been performed to assess the most optimal luteal support in relation to GnRHa trigger. We conclude that although more research is needed, GnRHa trigger is now an alternative to HCG trigger, combining a significant reduction in, or total elimination of, ovarian hyperstimulation syndrome (OHSS) with high ongoing pregnancy rates.

Mutations in WNT4 are not responsible for Müllerian duct abnormalities in Chinese women - Corrected Proof

2012-03-19

Abstract: The WNT4 gene plays a crucial role in sexual differentiation and female genital tract development. This study screened WNT4 for mutation in 189 Chinese women with Müllerian duct abnormalities (10 Mayer–Rokitansky–Küster–Hauser syndrome, five Müllerian aplasia and 174 incomplete Müllerian fusion) and detected no perturbation that would indicate a major role for WNT4. Only one novel synonymous mutation (c.1091G>A) in exon 5 and one known single-nucleotide polymorphism (rs16826648) in exon 2 were found. The results suggest that WNT4 might not contribute to the aetiology of Müllerian duct abnormalities in Chinese women.A genetic factor is considered to be one of the major causes for Müllerian duct abnormalities (MDA), and previous studies have showed that the WNT4 gene plays a crucial role in mammalian uterus development. We conducted a large cohort of well-characterized patients with MDA and screened variants of WNT4, as far as is known for the first time, but no causative mutations in the coding region of WNT4 were detected except for one known single-nucleotide polymorphism (rs16826648) and a novel synonymous mutation (c.1091G>A).

Donor insemination: a follow-up study of disclosure decisions, family relationships and child adjustment at adolescence - Accepted Manuscript

T. Freeman, S. Golombok — 2012-03-19

Abstract: The call for greater openness about gamete donation highlights the need to assess the long-term implications of telling donor-conceived children about their origins. This longitudinal study examined the consequences of secrecy versus openness about donor insemination (DI) for family relationships and child adjustment at adolescence. Thirty heterosexual families with an adolescent (aged 10–14 years) conceived by anonymous DI were assessed using standardized measures of parent–child and marital relationships, and parents’ and adolescents’ psychological wellbeing. Ten (33%) adolescents had been told about their donor conception. The only differences found between disclosed and non-disclosed families concerned parent–child relationships. In particular, whilst disclosure was associated with lower levels of conflict between mothers and sons, adolescents who were aware of their donor origins reported less warm father–child relationships. This is of interest given that identity issues and a fuller understanding of donor conception are likely to arise at adolescence. However, differences between disclosing and non-disclosing families cannot be directly attributed to parents’ disclosure decisions. Overall, these findings suggest that openness about DI does not create significant difficulties for family functioning or child adjustment and that a child’s age and sex may be important in assessing the impact of secrecy and disclosure.

Consecutive repeat miscarriages are likely to occur in the same gestational period - Corrected Proof

2012-03-16

Abstract: This is a retrospective, observational study to determine how often repeated consecutive miscarriages occur consistently in the same gestational period, including 1589 miscarriages among 543 women with recurrent miscarriage. In women who had two miscarriages only, 49.17% of both miscarriages occurred in the same gestational period, which was significantly higher than the expected probability of 34.54% (P<0.01). The proportions of all miscarriages occurring in the same gestational period in women with three, four and five or more miscarriages were 28.72%, 19.44% and 18.60%, compared with the expected probabilities of 14.36% (P<0.01), 6.57% (P<0.05) and <3.15% (P<0.05). The proportions of miscarriages occurring consistently in the same gestational period are higher than the theoretical probabilities calculated for the whole population.To determine how often repeated consecutive miscarriages occur consistently in the same gestational period, we did a retrospective, observational study including 1589 miscarriages among 543 women with recurrent miscarriage. It was found that in women who had two miscarriages only, 49.17% of both miscarriages occurred in the same gestational period, which was significantly higher than the expected probability of 34.54%. The proportions of all miscarriages occurring in the same gestational period in women with three, four or five or more miscarriages were 28.72%, 19.44% and 18.60%, compared with the expected probabilities of 14.36%, 6.57% and <3.15%. So we draw the conclusion that the proportions of the miscarriages occurring consistently in the same gestational period are higher than the theoretical probabilities calculated for the whole population.

Glycodelin suppresses endometrial cell migration and invasion but stimulates spheroid attachment - Corrected Proof

Kam-Hei So, Cheuk-Lun Lee, William S.B. Yeung, Kai-Fai Lee — 2012-03-16

Abstract: Glycodelin contains four isoforms with diverse biological functions. Glycodelin-A is potentially a diagnostic marker for cancer patients and receptivity marker of the secretory endometrium. Yet, direct evidence for the role of glycodelin in the regulation of endometrial epithelial cell migration, invasion and attachment of trophoblastic spheroids (blastocyst surrogate) is lacking. In this study, the human glycodelin gene was stably transfected into human endometrial (HEC1-B) cells. Forced expression of glycodelin in HEC1-B cells did not affect cell proliferation, cell viability or cell-cycle progression, but significantly reduced migration and invasion of the stably transfected cells (both P<0.05). The migration rate returned to normal levels when the glycodelin-forced-expressing HEC1-B cells were treated with glycodelin RNAi. Furthermore, forced expression of glycodelin in HEC1-B cells significantly increased the attachment of trophoblastic spheroids onto the endometrial epithelial cells (P<0.05). In summary, glycodelin suppressed endometrial cell migration and invasion but enhanced spheroid attachment.Glycodelin contains four similar forms with different biological functions. Glycodelin-A is a potential diagnostic marker for cancer patients and receptivity marker of the uterus. Yet, direct evidence for the role of glycodelin on the regulation of uterine epithelial cell migration, invasion and attachment of trophoblastic spheroids (embryo surrogate) is lacking. In this study, the human glycodelin gene was stably expressed in the human endometrial (HEC1-B) cells. Forced expression of glycodelin in HEC1-B cells did not affect cell proliferation, cell viability or cell-cycle progression, but significantly reduced migration and invasion of the stably expressed cells. The migration rate returned to normal levels when the glycodelin-forced-expressing HEC1-B cells were treated with glycodelin RNAi. Furthermore, forced expression of glycodelin in HEC1-B cells significantly increased the attachment of trophoblastic spheroids onto the endometrial epithelial cells. In summary, glycodelin has dual roles on cell migration/invasion and may be involved in embryo attachment.

Successful delivery following ICSI with macrocephalic sperm head syndrome: a case report - Corrected Proof

2012-03-14

Abstract: This article reports a case of macrocephalic sperm head syndrome, which is defined as the presence of a very high percentage of spermatozoa with enlarged heads and multiple flagellae, together with detailed morphological analysis. After a couple presented with infertility, sperm analysis showed severe teratozoospermia and almost all of the spermatozoa had macrocephaly with multiple tails. The morphological analysis revealed that most of the sperm heads contained several nuclei and had a similar number of tails as that of nuclei. However, detailed analysis revealed that there were a very few spermatozoa with an almost normal morphology. After genetic counselling, intracytoplasmic sperm injection was performed using a few spermatozoa that had an almost normal morphology, resulting in pregnancy and successful delivery. Even in macrocephalic sperm head syndrome, which may be caused by meiotic division failure, pregnancy is possible if some spermatozoa with almost normal morphology can be utilized, although there may be genetic risks.

Intra-cycle fluctuations of anti-Müllerian hormone in normal women with a regular cycle: a re-analysis - Corrected Proof

2012-03-14

Abstract: Anti-Müllerian hormone (AMH) has emerged as an important marker of ovarian reserve. Its variation throughout the cycle, however, may still be a matter of debate. The objective of this study was to re-evaluate the intra-cycle fluctuations of AMH in individuals in a prospective clinical study with focus on the age-related effects on these fluctuations. Frequent blood samples were obtained from the mid-luteal phase of the first cycle to the mid-luteal phase of the second cycle in 44 healthy, regularly menstruating Caucasian women. Main outcome measures were individual fluctuations of AMH concentrations during the natural menstrual cycle. AMH concentrations exhibited large fluctuations throughout the cycle and did not follow a defined pattern. Female age was negatively correlated with mean AMH concentrations. The absolute intra-individual variation was also negatively associated with age, whereas the relative intra-individual variation was positively associated with age. Although the fluctuation in relative intra-individual variation was higher in the older group, the absolute variation is very low and these fluctuations might therefore be of limited clinical relevance in this age group. These data show that in younger women caution should be exerted with the interpretation of a single randomly taken AMH measurement as a representative of ovarian reserve.Anti-Müllerian hormone (AMH) is currently under investigation to evaluate its use as a marker for the number of eggs remaining in the ovary, also known as the ovarian reserve. Ovarian reserve is used for decision making in infertility treatment. Some say it is a reliable marker that can be measured once at any time in the cycle; others claim that AMH fluctuates throughout the menstrual cycle. The aim of the current study was to evaluate whether AMH fluctuates during the cycle and whether the amount by which it fluctuates is dependent on the age of the individual. To investigate this, we took frequent blood samples during two cycles in 44 healthy, regularly menstruating women. In conclusion, AMH concentrations showed large fluctuations throughout the cycle but did not follow a defined pattern. The AMH concentration decreased with age. The amount of fluctuation was larger in younger women. These data show that, in younger women, caution should be exerted if a single randomly taken AMH measurement is used for clinical decision making.

Soluble BAFF-R produced by decidual stromal cells plays an inhibitory role in monocytes and macrophages - Corrected Proof

2012-03-14

Abstract: A sophisticated immunological regulation between decidual stromal cells (DSC) and monocytes and macrophages is essential for the successful symbiosis of the mother and her fetus, but the mechanisms remain incompletely understood. The mRNA and proteins of B lymphocyte stimulator (BAFF, also known as BLys) and its receptor, BAFF-R (also known as BR3, CD268 or TNFRSF17), have been detected in both first-trimester and term placentas, but whether BAFF or BAFF-R participates in the cross-talk between DSC and monocytes and macrophages in the first-trimester pregnancy has not been described. This study found that purified DSC extensively shed BAFF-R and that polyinosinic:polycytidylic acid (poly(I:C); a synthetic toll-like receptor (TLR) 3 agonist) dramatically up-regulated BAFF-R secretion, suggesting that release of these soluble proteins was an inherent property of DSC and its induction might have relevance to TLR-3-mediated signal transduction. When monocytes were cultured with the supernatants of resting DSC or poly(I:C)-treated DSC, the proliferation of CD14+HLA-DR+ monocytes (P=0.025 and 0.045) and the secretion levels of tumour necrosis factor α (P=0.035 and 0.031) and interleukin 6 (P=0.021 and 0.035) were significantly increased after the BAFF-R was blocked. Soluble BAFF-R may play inhibitory roles in monocytes and macrophages.The immunological regulation between decidual stromal cells (DSC) and monocytes and macrophages is essential and complex for the successful pregnancy, but the mechanisms remain indistinct. The mRNA and proteins of B lymphocyte stimulator (BAFF) and its receptor, BAFF-R, have been detected in both first-trimester and term placentas, but whether BAFF or BAFF-R participates in the cross-talk between DSC and monocytes and macrophages in the first-trimester pregnancy has not been described. In this study, we found purified DSC extensively shed BAFF-R and this cytokine was further up-regulated by polyinosinic:polycytidylic acid (poly(I:C); a synthetic toll-like receptor 3 agonist). When monocytes and macrophages were co-cultured with the supernatants of resting DSC or poly(I:C)-treated DSC, the proliferation of CD14+HLADR+ monocytes and the secretion levels of tumour necrosis factor α and interleukin 6 were significantly increased after the BAFF-R was blocked. These results show soluble BAFF-R may play inhibitory roles in monocytes and macrophages.

Effects of HMG on revascularization and follicular survival in heterotopic autotransplants of mouse ovarian tissue - Corrected Proof

2012-03-14

Abstract: Ovarian tissue transplantation is now considered as a procedure to preserve the fertility of young women patients undergoing cancer therapy. The present study investigated the effects and mechanism of human menopausal gonadotrophin (HMG) intervention on vascular remoulding in ovarian heterotopic autotransplantation. Ovaries of 8-week-old mice were cultured in vitro with different concentrations of HMG for 3h for measuring the expression of vascular endothelial growth factor (VEGF). The cultured ovaries were implanted under the kidney capsule and removed 24, 36, 48h or 1month after transplantation. Revascularization, fluid exudation and the number of surviving ovarian follicles were observed. The results showed that VEGF was increased 1.6–6.5 times in the HMG intervention groups. Revascularization appeared 24–36h after transplantation and was earlier than that of the control. Fluid exudation increased incrementally with increasing HMG concentrations. The total number of surviving ovarian follicles was increased by 1.2–1.5 times in the HMG 0.15IU/ml group as compared with the other groups 1month after transplantation. It is concluded that intervention with HMG in vitro before transplantation could improve the blood supply reconstruction and survival of the autotransplanted ovarian follicles, which might be associated with increased VEGF expression.Early diagnosis and effective therapy have been improving the long-term survival rate of children, adolescents and young women suffering from malignant tumours. Chemotherapy and/or radiation therapy commonly compromise the ovary functions. Cryopreservation of cortical fragments of ovarian tissue before cancer therapy, followed by later transplantation, is an emerging technique, but ischaemia after transplantation appears to be one of the main obstacles to successful transplantation. The aim of the present study was to investigate the effects of human menopausal gonadotrophin (HMG) on revascularization and follicular survival of autotransplanted ovary. Half ovaries of 8-week-old mice were cultured in vitro with different concentrations of HMG (0.00, 0.15, 0.30 and 0.60IU/ml) for 3h and transplanted back to the mice. The expression of vascular endothelial growth factor (VEGF), blood vessel repatency and survival ovarian follicles were observed. We found that the in vitro intervention of HMG could improve expression of VEGF in ovarian tissue, shorten the blood supply reconstruction time and increase the number of survival ovarian follicles. The starting date of the oestrous cycle was significantly earlier in the HMG 0.60IU/ml treatment group than the HMG 0.00IU/ml group. We also found that treatment with HMG 0.15IU/ml was better in preserving ovarian follicle numbers than HMG 0.30 or 0.60IU/ml. We concluded the intervention of HMG in vitro before transplantation could improve the blood supply reconstruction and survival of the autotransplanted ovarian follicles, which might be associated with increased VEGF expression. This method may have the potential for clinical usage.

Perception of control, coping and psychological stress of infertile women undergoing IVF - Corrected Proof

2012-03-14

Abstract: The study aimed to examine: (i) the association between perception of infertility controllability and coping strategies; and (ii) the association between perception of infertility controllability and coping strategies to psychological distress, applying multivariate statistical techniques to control for the effects of demographic variables. This cross-sectional study included 137 women with fertility problems undergoing IVF in a public hospital. All participants completed questionnaires that measured fertility-related stress, state anxiety, depressive symptomatology, perception of control and coping strategies. Pearson’s correlation coefficients were calculated between all study variables, followed by hierarchical multiple linear regression. Low perception of personal and treatment controllability was associated with frequent use of avoidance coping and high perception of treatment controllability was positively associated with problem-focused coping. Multivariate analysis showed that, when controlling for demographic factors, low perception of personal control and avoidance coping were positively associated with fertility-related stress and state anxiety, and problem-appraisal coping was negatively and significantly associated with fertility-related stress and depressive symptomatology scores. The findings of this study merit the understanding of the role of control perception and coping in psychological stress of infertile women to identify beforehand those women who might be at risk of experiencing high stress and in need of support.Infertility and its treatment are severe stressors and many women undergoing fertility treatment experience significant emotional distress. The existing literature indicates that several factors may influence emotional reactions to infertility and fertility treatment, including coping strategies and sense of controllability. This study showed that low perception of controllability was associated with avoidance coping. In addition, the study demonstrated that perception of infertility controllability and coping strategies were related to psychological stress. Therefore, healthcare professionals should enhance the implementation of interventions and support services for infertile women undergoing fertility treatment in order to enhance sense of control and alter coping skills of those women believed to be at risk.

The Centre for Trophoblast Research: improving health through placental research - Corrected Proof

Graham J. Burton — 2012-03-14

Abstract: The placenta is an essential but widely neglected organ. As the interface between the mother and her fetus, the placenta represents the platform for a healthy life. The majority of the major complications of pregnancy, including miscarriage, pre-eclampsia, intrauterine growth restriction and stillbirth, have their pathophysiological roots in poor placentation. In addition, there is now irrefutable evidence that low birthweight predisposes to a higher risk of cardiovascular and other disorders in later life. The Centre for Trophoblast Research was established in the University of Cambridge with the aim of generating new impetus and a fresh approach to address these problems. Placentation involves many different cell biological processes, some of which are unique to the trophoblast, as well as complex interactions with the maternal immune system. The Centre brings together academic clinicians and basic scientists working in diverse disciplines and provides a rich intellectual environment that facilitates novel collaborative links. The Centre also encourages new investigators into the field and fosters their careers through a number of initiatives, including support for studentships and fellowships, developing research resources, hosting an annual scientific meeting and running a training course in placental biology. Full details can be found at www.trophoblast.cam.ac.uk.The Centre for Trophoblast Research was established in the University of Cambridge to promote research into the placenta. The placenta (afterbirth) is the interface between the mother and her baby. All nutrients and waste products pass across the placenta and so it plays a key role in determining the growth of the baby. Many adverse complications of pregnancy are caused by defects in placental development or function. The placenta carries the same genes as the baby; these are related to those of the mother but are not identical. Hence, there are complex interactions with the cells of the maternal immune system that must be carefully negotiated if the pregnancy is not to be rejected. Many of the functions of the placenta and of the immune interactions depend on a unique cell type, the trophoblast. The Centre brings together basic scientists and academic clinicians to generate a unique interdisciplinary approach to problems of pregnancy. The Centre also provides research training and acts as an international resource centre for placental research.

Oocyte cryopreservation following failed testicular sperm extraction: a French case report with implications for the management of non-obstructive azoospermia - Corrected Proof

2012-03-09

Abstract: The French law regulating assisted reproductive technologies forbids donor spermatozoa to be available in case of failed testicular sperm extraction (TESE) performed on the day of oocyte retrieval. This article reports the first French live birth after intracytoplasmic sperm injection of donated spermatozoa into frozen–thawed oocytes cryopreserved following failure of TESE. By reinforcing the relevance of TESE performed on the day of oocyte retrieval, oocyte cryopreservation in couples having beforehand consented to go to sperm donation will avoid cycle cancellation and potentially lead to successful live birth. Therefore, it could modify the French policy of management of patients suffering from non-obstructive azoospermia.

The effects of letrozole on ovarian stimulation for fertility preservation in cancer-affected women - Corrected Proof

2012-03-09

Abstract: Survival rates for fertile women with cancer have increased significantly, lending importance to considering the possibility of motherhood after cancer. This study was a retrospective analysis of a prospective database comparing two groups of patients who underwent fertility preservation after being diagnosed with either breast cancer or a non-hormone-dependent cancer between 2009 and 2011. Nineteen oncology patients were included in the study. The objective was to assess the efficacy of ovarian stimulation with aromatase inhibitors versus a standard antagonist protocol. This study sought to quantify oestradiol concentrations in patients receiving letrozole and to determine the length of time between diagnosis of malignancy and onset of fertility preservation. Number of mature oocytes retrieved in the non-hormone-dependent cancer group was comparable to that in the breast cancer group (15.4±8.19 versus 16.3±7.31). Oestradiol concentrations were higher for patients with non-hormone-dependent cancer (1666.4±739.42pg/ml versus 829±551.11pg/ml, P=0.006). There were no differences between the groups in the length of time between diagnosis and fertility preservation (17.4±4.93 versus 16.4±1.74days). Oestradiol concentrations of breast cancer patients on the letrozole protocol remained much lower than those of patients on the antagonist protocol.The survival rates for fertile women with cancer have increased significantly, lending importance to considering the possibility of motherhood after cancer. In our study, we compared two groups of patients with cancer: a group with breast cancer, which is hormone dependent and is the most common type of cancer in women, and a group of non-hormone-dependent cancer patients. We compared the efficacy of two protocols in a retrospective analysis of a prospective database. In the patients with breast cancer, we used the protocol from the Controlled Ovarian Stimulation Treatment with Letrozole Supplementation Study (COST-LESS), modified to avoid the high concentrations of oestradiol to prevent the risks of the increase in hormone-dependent cancer. In the other group, we used the usual antagonist protocol. No difference was observed between the two groups in terms of the numbers of mature oocytes retrieved or in the time taken to complete the stimulation. The oestradiol concentrations of the breast cancer patients on the modified COST-LESS protocol remained much lower than those of the patients on the antagonist protocol, which is beneficial for preventing tumour growth in patients with hormone-dependent breast cancer. In addition, healthcare providers should inform their oncology patients of their options at the time of diagnosis and urge them not to delay initiating fertility preservation.

Unexplained infertility: an update and review of practice - Corrected Proof

Arpita Ray, Amit Shah, Anil Gudi, Roy Homburg — 2012-03-09

Abstract: Of the couples unable to conceive without any identifiable cause, 30% are defined as having unexplained infertility. Management depends on duration of infertility and age of female partner. This review describes and comments on the definition and evidence for the management of unexplained infertility. A literature search was conducted in EMBASE, Medline, Ovid and Cochrane Database of Systematic reviews using the terms ‘infertility’, ‘unexplained infertility’, ‘idiopathic infertility’, ‘definition of infertility’, ‘treatment options’, ‘intrauterine insemination’, ‘ovulation induction’, ‘Fallopian tube sperm’, ‘GIFT’ and ‘IVF’. There is no uniform definition for unexplained infertility. This varies in the literature depending on the duration of infertility and the age of the female partner. The treatment of unexplained infertility is empirical and many different regimens have been used. Among these are expectant management, ovulation stimulation with clomiphene citrate, gonadotrophins and aromatase inhibitors, Fallopian tube sperm perfusion, tubal flushing, intrauterine insemination, gamete intra-Fallopian transfer and IVF. The standard protocol is to progress from low-technology to high-technology treatment options. On the best available evidence, an algorithm for management is suggested. There is a definite need for multicentre randomized controlled trials to identify the best treatment option in unexplained infertility using a standard definition.Of the couples unable to conceive without any identifiable cause, 30% are defined as having unexplained infertility. The management for unexplained infertility is empirical. There is no uniform definition for unexplained infertility. This varies in the literature depending on the duration of infertility and the age of the female partner. It may arise in two ways: in some couples it may be due to some undetectable factor and in others it may be due to reduced fecundity. The treatment of unexplained infertility is empirical and many different regimens have been used. Among these various treatment options are expectant management, ovulation stimulation with clomiphene citrate, gonadotrophins and other agents, Fallopian tube sperm perfusion, tubal flushing, intrauterine insemination, gamete intra-Fallopian transfer and IVF. There is no definite evidence to show that any one treatment is better than the other. Treatment of unexplained infertility is very much dependent on availability of resources and patients’ age and duration of infertility. The standard protocol is to proceed from low-tech to high-tech treatment options. There is a definite need for multicentre randomized controlled trials to identify the best treatment option in unexplained infertility.

Response: Adjustment of cryoprotectant exposure time to counteract reduced cooling rates - Corrected Proof

Dominic Stoop — 2012-03-05

We acknowledge that the closed vitrification approach described by is different from the approach described in our paper (). We did not intend to claim similarity by citing their work, as we were well aware of the differences at the time of writing the manuscript. Therefore, the citation should not be interpreted as if we believe that the conclusions by Vanderzwalmen et al. are in line or supportive of our conclusions. Indeed, we cited the work by Vanderzwalmen et al. for exactly the opposite reason. As mentioned in their work, and cited in their recent correspondence (), they adjusted the exposure time to the cryoprotectant solution in order to minimize the risk of intracellular ice crystal formation. However, our study demonstrates that excellent results can be achieved without these changes.

Adjustment of cryoprotectant exposure time to counteract reduced cooling rates - Corrected Proof

Pierre Vanderzwalmen — 2012-03-05

In the article by ‘Clinical validation of a closed vitrification system in an oocyte-donation programme’, the authors state that “successful vitrification can be achieved without the need to modify cryoprotectant exposure to counteract reduced cooling rates” by citing our work (). However, we do not agree with this statement. The vitrification protocols of , both in closed carrier conditions, are derived from vitrification protocols in open cooling conditions but with two different strategies regarding exposure to cryoprotectant (CP) solutions.

Implication of MEK1 and MEK2 in the establishment of the blood–placenta barrier during placentogenesis in mouse - Corrected Proof

Jean Charron, Vickram Bissonauth, Valérie Nadeau — 2012-03-02

Abstract: The ERK/MAPK signalling cascade is involved in many cellular functions. In mice, the targeted ablation of genes coding for members of this pathway is often associated with embryonic death due to the abnormal development of the placenta. The placenta is essential for nutritional and gaseous exchanges between maternal and embryonic circulations, as well as for the elimination of metabolic waste. These exchanges occur without direct contact between the two circulations. In mice, the blood–placenta barrier consists of a triple layer of trophoblast cells adjacent to endothelial cells from the embryo. In the ERK/MAPK cascade, MEK1 and MEK2 are dual-specificity kinases responsible for the activation of the ERK1 and ERK2 kinases. Inactivation of Mek1 causes placental malformations resulting from defective proliferation and differentiation of the labyrinthine trophoblast cells and leading to a severe delay in the development and the vascularization of the placenta, which explains the embryonic death. Although Mek2−/− mutants survive without any apparent phenotype, a large proportion of Mek1+/− Mek2+/− double heterozygous mutants die during gestation from placenta anomalies affecting the establishment of the blood–placenta barrier. Together, these data reveal how crucial is the role of the ERK/MAPK pathway during the formation of the placenta.In normal cells, various cellular processes are tightly controlled via a cascade of protein kinases, which transduce various signals from the cell surface to the nucleus to activate programmes of cell growth and differentiation. This network of sequential protein kinases referred to as the mitogen-activated protein kinase (MAPK) signalling cascade has been conserved during evolution. Our studies on the role of Mek1 and Mek2, components of this cascade, during mouse development have shown that both Mek1 and Mek2 genes contribute to normal morphogenesis and vascularization of the placenta. Mek1 mutant mice die during gestation of abnormal development of the labyrinthine region of the placenta. In contrast, Mek2 mutant mice are viable without any obvious anomaly. However, a majority of double heterozygous Mek1+/− Mek2+/− embryos die during late gestation from placental defects. Thus, the lack of one allele of each Mek gene is deleterious to embryo survival suggesting a potential contribution of Mek2 to placenta development. Despite similarities in their biochemical properties, MEK1 and MEK2 appear to have specific biological functions. Our long-term objective is to unravel the molecular mechanisms responsible for the specificity of action of MEK1 and MEK2 as well as to identify in which cellular processes the ERK/MAPK cascade is implicated. Characterization of the mechanisms involved in signal transduction is essential for our comprehension of the role of the ERK/MAPK signalling cascade in placenta diseases.

The role of transcription factor Tcfap2c/TFAP2C in trophectoderm development - Corrected Proof

Peter Kuckenberg, Caroline Kubaczka, Hubert Schorle — 2012-03-02

Abstract: In recent years, knowledge regarding the genetic and epigenetic programmes governing specification, maintenance and differentiation of the extraembryonic lineage has advanced substantially. Establishment and analysis of mice deficient in genes implicated in trophoblast lineage and the option to generate and manipulate murine stem cell lines from the inner cell mass and the trophectoderm in vitro represent major advances. The activating enhancer binding protein 2 (AP2) family of transcription factors is expressed during mammalian development and in certain malignant diseases. This article summarizes the data regarding expression and function of murine Tcfap2 and human TFAP2 in extraembryonic development and differentiation. It also presents a model integrating Tcfap2c into the framework of trophoblast development and highlights the requirement of Tcfap2c to maintain trophoblast stem cells. With regard to human trophoblast cell-lineage restriction, the role of TFAP2C in lineage specification and maintenance is speculated upon. Furthermore, an overview of target genes of AP2 in mouse and human affecting placenta development and function is provided and the evidence suggesting that defects in regulating TFAP2 members might contribute to placental defects is discussed.During recent years, much has been learned about role and function of TFAP2C/Tcfap2c with regard to trophoblast development and placenta development. Using murine models and cell systems, we learned that this gene serves as an important regulator in trophoblast stem cell maintenance and have begun to decipher the molecular network TFAP2C/Tcfap2c is integrated in. However, knowledge on its function and role in the differentiating placental lineage is rather limited and needs to be extended. In this respect, transgenic systems aimed at conditional deletion of Tcfap2c in the trophectoderm once differentiation is initiated are on the way. In addition, we have evidence that elevated TCFAP2C concentrations might lead to placental defects such as pre-eclampsia. Clearly more work is needed to further substantiate this claim. However, description and analysis on upstream regulators of TFAP2C will eventually lead to a better understanding of the molecular nature underlying placental defects. Such basic research may open up the field for translational projects aiming at treatment options via patient-specific targeted therapy.

Egg-sharing, consent and exploitation: examining donors’ and recipients’ circumstances and retrospective reflections - Corrected Proof

Zeynep B. Gürtin, Kamal K. Ahuja, Susan Golombok — 2012-02-29

Abstract: Egg-sharing schemes involve a woman sharing a portion of her eggs with another in exchange for free or reduced-cost fertility treatment and have been regulated in the UK since 1998. However, while being perceived as a unique anomaly (within the UK system) such schemes have caused considerable debate. Some critics have been concerned that the benefit-in-kind (i.e. fertility treatment) offered in exchange for donating eggs might compromise the consent of potential donors, particularly those who have no other means to access treatment, and lead to exploitation and the commodification of gametes. This article reports empirical findings that engage directly with these concerns. First, data on the demographic characteristics, circumstances and treatment outcomes of donors and recipients are presented, followed by data on egg sharers’ own retrospective assessments of egg-sharing and their opinions regarding its ethics and regulation. This study shows that, contrary to some expectations, there are very few differences in the characteristics, experiences and opinions of egg-sharing donors and recipients, and also highlights the overwhelmingly positive assessment of egg-sharing by women who have taken part in such schemes.Egg-sharing schemes involve a woman sharing a portion of her eggs with another in exchange for free or reduced-cost fertility treatment and have been regulated in the UK since 1998. However, while being perceived as a unique anomaly (within the UK system) such schemes have caused considerable debate. Some critics have been concerned that the benefit-in-kind (i.e. fertility treatment) offered in exchange for donating eggs might compromise the consent of potential donors, particularly those who have no other means to access treatment, and lead to exploitation and the commodification of gametes. This article reports empirical findings that engage directly with these concerns. First, data on the demographic characteristics, circumstances and treatment outcomes of donors and recipients are presented, followed by data on egg sharers’ own retrospective assessments of egg-sharing and their opinions regarding its ethics and regulation. This study shows that contrary to some expectations, there are very few differences in the characteristics, experiences, and opinions of egg-sharing donors and recipients, and highlights the overwhelmingly positive assessment of egg-sharing by women who have taken part in such schemes.

Genes r us? Making sense of genetic and non-genetic relationships following anonymous donor insemination - Corrected Proof

Eric Blyth — 2012-02-29

Abstract: This exploratory qualitative study investigates the experiences of eight adults conceived following anonymous sperm donation who had discovered the identity both of their donor and of donor half-siblings and had established contact with each other. It focuses primarily on participants’ reflections on genetic and social kinship relationships. Data were collected from this group as well as from the son of the donor and the donor-conceived half-sister of one participant by means of semistructured interviews utilizing asynchronous email and digitalized voice recording. Participants discussed their experience of genetic disconnection resulting from learning of their donor-conceived status and of revising their personal biographies and developing new kinship networks as a result of discovering the identity of their donor and the existence of donor half-siblings. The study highlights participants’ agency expressed through their ability to draw on both genetic and non-genetic elements of their inheritance to redefine their self-identity and extend their familial/kinship networks in meaningful ways.This paper reports findings from a study investigating the experiences of eight adults who learned of their conception following anonymous donor insemination provided by the same fertility clinic, the identity of their shared donor and their relatedness to each other and who had subsequently established communications with each other. The donor-conceived sister of one participant and the son of the donor also participated. Data were collected by means of email communications using a semi-structured interview schedule. The specific focus of this paper examines participants’ experiences of genetic disconnection resulting from learning of their donor-conceived status and of revising their personal biographies and developing new kinship networks as a result of discovering the identity of their donor and the existence of donor half-siblings. It concludes that participants were able to draw on both genetic and non-genetic elements of their ‘roots’ in order to redefine their self-identity and extend their familial/kinship networks.

Array CGH analysis shows that aneuploidy is not related to the number of embryos generated - Corrected Proof

2012-02-27

Abstract: This study retrospectively analysed array comparative genomic hybridization (CGH) results of 7753 embryos from 990 patients to determine the frequency of embryonic euploidy and its relationship with the cohort size (i.e. the number of embryos available for biopsy and array CGH analysis). Linear regression analysis was performed to assess the effect of cohort size on euploidy rate adjusted for the effect of female age. While increasing female age was associated with a significant decrease in euploidy rate of day-3 and day-5 embryos (P<0.001 for both groups), cohort size was not significantly associated with euploidy rate. Logistic regression analysis was performed to assess the effect of cohort size, adjusted for maternal age, on the likelihood of having at least one euploid embryo available for transfer. The odds of having at least one euploid embryo in an assisted cycle was significantly decreased by increasing female age (P<0.01 for both day-3 and day-5 embryos) and was significantly increased by every additional embryo available for analysis (P<0.001 for both day-3 and day-5 embryos).Array comparative genomic hybridization (CGH) enables assessment of the entire chromosomal component of human embryos, which was not possible with the formerly used preimplantation genetic diagnosis (PGD) technology. We analysed array CGH PGD results of 7753 embryos from 990 women to determine the frequency of embryonic euploidy and its relationship with the number of embryos available for biopsy (hereafter called cohort size) in an assisted reproduction cycle. While increasing female age was associated with a significant decrease in euploidy rate of day-3 and day-5 embryos (P<0.001 for both groups), cohort size was not significantly associated with euploidy rate. Logistic regression analysis was performed to assess the effect of cohort size, adjusted for maternal age, on the likelihood of having at least one euploid embryo available for transfer. The odds of having at least one euploid embryo in an assisted cycle was significantly decreased by increasing female age (P<0.01 for both day-3 and day-5 embryos) and was significantly increased by every additional embryo available for analysis (P<0.001 for both day-3 and day-5 embryos). In conclusion, increasing the number of embryos available in a conventionally stimulated cycle increases the chance of having an euploid embryo available for transfer.

Validation of array comparative genome hybridization for diagnosis of translocations in preimplantation human embryos - Corrected Proof

2012-02-24

Abstract: Fluorescent in-situ hybridization (FISH) for preimplantation genetic diagnosis (PGD) of structural chromosome abnormalities has limitations, including carrier testing, inconclusive results and limited aneuploidy screening. Array comparative genome hybridization (CGH) was used in PGD cases for translocations. Unbalances could be identified if three fragments were detectable. Smallest detectable fragments were ∼6Mbp and ∼5Mbp for blastomeres and trophectoderm, respectively. Cases in which three or more fragments were detectable by array CGH underwent PGD by FISH and concordance was obtained in 53/54 (98.1%). The error rate for array CGH was 1.9% (1/54). Of 402 embryos analysed, 81 were normal or balanced, 92 unbalanced but euploid, 123 unbalanced and aneuploid and 106 balanced but aneuploid. FISH with additional probes to detect other aneuploidies would have missed 28 abnormal embryos in the reciprocal group and 10 in the Robertsonian group. PGD cases (926) were retrospectively reviewed for reciprocal translocations performed by FISH to identify which could have been analysed by array CGH. This study validates array CGH in PGD for translocations and shows that it can identify all embryos with unbalanced reciprocal and Robertsonian translocations. Array CGH is a better approach than FISH since it allows simultaneous screening of all chromosomes for aneuploidy.Fluorescent in-situ hybridization (FISH) has been used in preimplantation genetic diagnosis (PGD) for structural chromosome abnormalities. This approach has limitations such as the need for testing of carriers, poor fixation and no possibility for aneuploidy screening of all chromosomes. The use of array comparative genome hybridization (CGH) can solve these limitations. Array CGH was used in 47 PGD cases for Robertsonian or reciprocal translocations. The smallest detectable fragment size was ∼6Mbp for single blastomeres (day-3) and ∼5Mbp for trophectoderm (day-5) biopsies. Translocation cases in which three or more translocated fragments were detectable by array CGH underwent PGD. Of those, a total of 54 non-transferred embryos analysed by array CGH were reanalysed by FISH. The error rate for array CGH was 1.9% (1/54). Of the 402 embryos analysed by array CGH, 81 embryos were normal or balanced, 92 were unbalanced but euploid, 123 unbalanced and aneuploid and 106 balanced but aneuploid. We retrospectively reviewed 926 PGD cases for reciprocal translocations performed by FISH in order to identify those in which fewer than two fragments were less than ∼6Mbp. In conclusion, this study validates the use of array CGH in PGD for translocations showing a low error rate and shows that array CGH can identify all embryos with unbalanced reciprocal and Robertsonian translocations. Array CGH is a better approach than FISH or single-nucleotide polymorphism arrays because it allows simultaneous screening of all chromosomes for aneuploidy and does not require previous genetic testing of the parents.

Perceptions of embryo status and embryo use in an Australian community - Corrected Proof

2012-01-30

Abstract: Increasingly, important areas of medical therapy and research rely on the donation and use of human embryos. Yet their use is commonly determined by community tolerance and ethico-legal regulation. The aim of this study was to explore the views of an Australian community about what an embryo is, how it should be used and who should make disposition decisions. The findings of a large representative population survey showed that most participants thought of an embryo as human or potentially human but that this did not affect a majority community view that embryos should be used rather than discarded. This study also found divergent views about what the community perceived to be acceptable uses of embryos. The majority perceived the couple as having the authority to make a disposition decision. Women held different views to men across all three questions. The way an embryo was perceived related significantly to how it should be used and who should decide its disposition. These differences and relationships should be considered when developing clinic practices and ethico-legal frameworks to regulate embryo use in science or treatment.Increasingly, important areas of medical therapy and research rely on the donation and use of human embryos. Yet their use is commonly determined by community tolerance and ethico-legal regulation. The aim of this study was to explore the views of an Australian community about what an embryo is, how it should be used and who should make disposition decisions. The findings of a large representative population survey showed that most participants thought of an embryo as human or potentially human but that this did not affect a majority community view that embryos should be used rather than discarded. We also found divergent views about what the community perceived to be acceptable uses of embryos. The majority perceived the couple as having the authority to make a disposition decision. Women held different views to men across all three questions. The way an embryo was perceived related significantly to how it should be used and who should decide its disposition. These differences and relationships should be considered when developing clinic practices and ethico-legal frameworks to regulate embryo use in science or treatment.

Disclosure patterns of embryo donation mothers compared with adoption and IVF - Corrected Proof

Fiona MacCallum, Sarah Keeley — 2012-01-30

Abstract: Attitudes towards information sharing in donor conception have changed in recent years in some parts of the world, with a move towards openness. This study follows up a sample of embryo donation mothers, examining their current disclosure patterns and comparing them with adoptive and IVF mothers to investigate any influence of the method of family creation. This study compared 17 mothers with an embryo donation child aged 5–9years with 24 mothers with an adopted child and 28 mothers with a genetically related IVF child. Embryo donation mothers were far less likely to share information with the child; 43% were inclined towards disclosing, compared with all adoptive mothers and nearly 90% of IVF mothers. Furthermore, embryo donation and IVF mothers who had disclosed had often only given partial explanations of the child’s conception. Differences between embryo donation and adoption in particular should be taken into account when advising embryo recipients.

Epigenetic features of the mouse trophoblast - Corrected Proof

Peter J. Rugg-Gunn — 2012-01-27

Abstract: Trophoblast cells are required for the growth and survival of the fetus during pregnancy, and failure to maintain appropriate trophoblast regulation is associated with placental insufficiencies and intrauterine growth restriction. Development of the trophoblast lineage is mediated by interactions between genetic and epigenetic factors. This review will focus on new insights that have been gained from analysis of mouse models into the epigenetic mechanisms that are required for the early establishment of the trophoblast lineage and for the development of specialized cell types of the fetal placenta. In particular, the importance of DNA methylation, 5-hydroxymethylcytosine and histone modifications in orchestrating trophoblast gene expression and functional outcome will be discussed. These insights are beginning to be extended towards human studies and initial results suggest that the causes and consequences of a variety of placental pathologies are related to epigenetic processes. Furthermore, the epigenetic landscape that regulates trophoblast cells seems to be particularly vulnerable to perturbation during development. This has major implications for diet and other environmental factors during pregnancy.The placenta is required for the growth and survival of the fetus during pregnancy. Placental insufficiencies, including pre-eclampsia and intrauterine growth restriction, occur in ∼10% of pregnancies and are associated with serious complications for mother and baby. Better understanding of how the placenta is formed is critical for the development of early diagnoses and therapies for treatment. Studies of animal models have revealed several critical pathways that are required for the proper development and function of the placenta. In particular, epigenetic processes, which determine how genes are switched on and off, are necessary for the early establishment of the placental lineage and also for the development of specialized cell types within the mature placenta. These crucial and recent insights form the basis for this review. Importantly, much of our understanding gleaned from animal models are now beginning to be extended towards human studies. Initial results suggest that the causes and consequences of a variety of placental pathologies are related to epigenetic pathways. As epigenetic processes can be susceptible to alterations by environmental factors, these studies have major implications for diet and other external influences during pregnancy.

Forming a family with sperm donation: a survey of 244 non-biological parents - Corrected Proof

Lucy Frith, Neroli Sawyer, Wendy Kramer — 2012-01-27

Abstract: There has been little research on the views and experiences of non-biological parents of sperm donor children. This paper reports the results of a survey of non-biological mothers and fathers. An online survey was designed and conducted by the Donor Sibling Registry, a US-based non-profit organization that supports those who have used donor conception. A total of 244 people responded (199 non-biological mothers and 45 non-biological fathers). The survey aimed to understand the perspectives of the respondents who had used donor spermatozoa within heterosexual or same-sex relationships, by exploring their views on a number of key issues. Certain issues and concerns associated with not being genetically related to their offspring were experienced differently by men and women. However, there were many important areas of common ground: a concern for getting a healthy donor, the importance of matching the donor to the non-biological partner, and the amount of thought that went into selecting the donor. The implications of these results for policies concerning donor spermatozoa are discussed.There has been little research on the views and experiences of non-biological parents of sperm donor children born from assisted conception. This paper reports the results of a survey of 244 non-biological mothers and fathers. An online survey designed by the Donor Sibling Registry, a US-based non-profit organization that supports those who have used donor conception, was conducted. Two hundred and forty four people responded (199 non-biological mothers and 45 non-biological fathers). The survey aimed to understand the perspectives of these parents within heterosexual and same-sex relationships who had used donor insemination in greater depth by exploring their views on a number of key issues. Certain issues and concerns that related to not being genetically related to their offspring were experienced differently by the men and women. However, there were many important areas of common ground: a concern for getting a healthy donor, the importance of matching the donor to the non-biological partner and the amount of thought that went into selecting the donor. The implications of these results for policies concerning donor insemination will be discussed.

Contraceptive counselling and factors affecting women’s contraceptive choices: results of the CHOICE study in Austria - Corrected Proof

2011-12-19

Abstract: Empirical evidence of the impact of contraceptive counselling and factors affecting women’s contraceptive choices are limited. CHOICE (Contraceptive Health Research Of Informed Choice Experience) was a large-scale study in 11 European countries. Women in Austria aged 15–40 years considering a short-acting, reversible form of combined hormonal contraceptive were eligible to participate. The choices included the combined daily pill, weekly transdermal patch, and monthly vaginal ring. This study assessed and compared 2478 women’s original preferences with their post-counselling choices and evaluated their perceptions and criteria for their choice. Women who were ‘undecided’ decreased from 18.1% pre-counselling to 3.2% post-counselling; significantly more women post-counselling chose the monthly ring (8.7% to 23.8%; difference 15.1%, 95% CI 13.3–16.8%; P<0.0001) or the weekly patch (6.2% to 7.8%; difference 1.7%, 95% CI 0.5–2.9%; P=0.0014). Women’s primary reasons for choosing a method included ‘easy to use’ (daily pill, weekly patch and monthly ring) and ‘still effective if I experience vomiting, diarrhoea’ (weekly patch and monthly ring). Structured and balanced counselling led to changes in the method chosen.The impact of contraceptive counselling and factors affecting women’s contraceptive choices are not well investigated. In the present study, women in Austria aged 15–40 years considering a short-acting, reversible form of combined hormonal contraceptive were eligible to participate. We assessed and compared women’s original preferences with their choices after extensive counselling and evaluated their perceptions and criteria for their choice. Women who were ‘undecided’ decreased from 18.1% before counselling to 3.2% after counselling; significantly more women chose the monthly ring after counselling or the weekly patch. Women’s primary reasons for choosing a method included ‘easy to use’ (pill, patch and ring) and ‘still effective if I experience vomiting, diarrhoea’ (patch and ring). A balanced counselling led to changes in the method chosen, even in women possessing profound knowledge about their preferred method.