Human Reproduction Update Advance Access published June 10, 2014 Human Reproduction Update, Vol.0, No.0 pp. 1– 14, 2014 doi:10.1093/humupd/dmu027
Fresh versus frozen embryo transfer: backing clinical decisions with scientific and clinical evidence
1 Uterine Biology, Prince Henry’s Institute of Medical Research, Clayton, VIC 3168, Australia 2Department of Physiology, Monash University, Clayton, VIC 3168, Australia 3Department of Obstetrics and Gynaecology, University of Melbourne, Heidelberg, VIC 3084, Australia 4Monash Health, Clayton, VIC 3168, Australia 5Monash IVF, Clayton, VIC 3168, Australia 6Department of Obstetrics and Gynaecology, Monash University, Clayton, VIC 3168, Australia
*Correspondence address. E-mail:
[email protected] Submitted on December 6, 2013; resubmitted on April 22, 2014; accepted on May 16, 2014
table of contents
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Introduction Methods The clinical arguments Embryo cryopreservation Improved outcomes The role of preimplantation genetic screening Success rates with elective FET The scientific background The endometrium in controlled ovarian stimulative cycles Human chorionic gonadotrophin Evidence from non-human models Are we there yet? The role of endometrial receptivity tests Conclusion
background: Improvements in vitrification now make frozen embryo transfers (FETs) a viable alternative to fresh embryo transfer, with reports from observational studies and randomized controlled trials suggesting that: (i) the endometrium in stimulated cycles is not optimally prepared for implantation; (ii) pregnancy rates are increased following FET and (iii) perinatal outcomes are less affected after FET. methods: This review integrates and discusses the available clinical and scientific evidence supporting embryo transfer in a natural cycle. results: Laboratory-based studies demonstrate morphological and molecular changes to the endometrium and reduced responsiveness of the endometrium to hCG, resulting from controlled ovarian stimulation. The literature demonstrates reduced endometrial receptivity in controlled ovarian stimulation cycles and supports the clinical observations that FET reduces the risk of ovarian hyperstimulation syndrome and improves outcomes for both the mother and baby.
conclusions: This review provides the basis for an evidence-based approach towards changes in routine IVF, which may ultimately result in higher delivery rates of healthier term babies. Key words: frozen embryo transfer / endometrial receptivity / human chorionic gonadotrophin / IVF success / ovarian stimulation †
J.E. and N.J.H. are equal first authors.
& The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email:
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Jemma Evans1,2,†, Natalie J. Hannan1,3,†, Tracey A. Edgell1, Beverley J. Vollenhoven 4,5,6, Peter J. Lutjen 5, Tiki Osianlis 4,5,6, Lois A. Salamonsen 1,6,*, and Luk J.F. Rombauts 4,5,6
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Introduction
Methods Literature searches were performed using PubMed, MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials and Web of Knowledge. Relevant terms and variations were used including: IVF with or without intracytoplasmic sperm injection (ICSI), fresh embryo transfer or FET, vitrification, slow freezing, cryopreservation, human chorionic gonadotrophin (hCG) and (GnRH) agonist trigger. For the scientific section, additional searching included the terms endometrium and endometrial with histology, hormonal effects, hCG and receptivity. The search was restricted to articles published in English. Further articles were identified by manually searching the references of the relevant articles and from the authors’ considerable knowledge of the literature on this topic.
The clinical arguments Embryo cryopreservation The clinical value of embryo cryopreservation has steadily increased over the decades with significant technological advances made in our ability to
Figure 1 The environments of implantation. The embryo enters the uterine cavity as a blastocyst. Within the microenvironment of the uterine cavity, it hatches, becomes apposed to and then attaches to the luminal epithelium of the endometrium. Subsequently, the trophectodermal cells penetrate between the epithelial cells and invade through the basal lamina to enter a new environment of decidualizing stromal cells. These, by their secretion of cytokines and other factors, further drive decidualization, attract macrophages (M) and uterine natural killer (NK) cells into the region, and guide the invasion of a cohort of the invading trophoblast cells (extravillous trophoblast) through the decidua. Some of these eventually penetrate and remodel the vasculature to provide blood vessels (BVs) for the increased blood flow essential for pregnancy.
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Reproductive technologies, particularly in vitro fertilization (IVF), have provided great hope for infertile couples worldwide. Indeed, more than 5 million IVF babies have been born since the birth of the first, Louise Brown, in 1978. However, in spite of considerable advances in the optimization of stimulation protocols for multi-follicular development and new technologies for the assessment of embryo quality, the success rate for IVF in fresh transfer cycles remains low worldwide. It is clear that pregnancy rates with fresh embryo transfer in IVF cycles are less than ideal. The latest results generated from European registers by ESHRE (Ferraretti et al., 2013) demonstrate clinical pregnancy rates (CPRs) per fresh transfer following IVF between 41.8% (Lithuania) and 18.9% (Iceland) with an average of 32%, making it clear that embryo implantation and ongoing pregnancy are far from guaranteed. Furthermore, a meta-analysis indicates that these differences are not due to use of different ovarian stimulation protocols as no differences in ongoing pregnancy rates were observed between GnRH antagonist and GnRH agonist facilitated cycles (Bodri et al., 2011). The CPR per transfer of frozen/thawed blastocysts in Australia and New Zealand in 2011 was 31.4% (Macaldowie et al., 2013). Across Europe, a large spread in the CPR per transfer of frozen/thawed embryos is seen, varying between 16.7 and 32.9% with an average for Europe of 22.3% (Ferraretti et al., 2012, 2013). Over the past 30 years, basic research underpinning IVF has predominantly been embryo-centric. However, successful implantation of an embryo depends not only on embryo quality, but also on endometrial receptivity and the microenvironment for embryo-maternal signalling within the uterine cavity, during the peri-implantation period (Fig. 1). The molecular and cellular mechanisms and signals underpinning
successful implantation have become better understood, with a range of models enabling the study of embryo-endometrial interactions. It is thus timely to consider how the endometrium is affected in stimulation cycles, and whether this knowledge can be applied to improve IVF outcomes. This review provides an overview of current information regarding the endometrium in stimulated cycles and provides strong evidence to suggest that IVF outcomes can be further improved with the adoption of a ‘freeze-all’ or elective frozen embryo transfer (eFET) strategy with replacement of thawed embryos in natural cycles.
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Improved outcomes A decade ago, Min and colleagues strongly argued for the adoption of a new metric, the BESST (Birth Emphasizing a Successful Singleton at Term), to measure the optimal outcome of an IVF cycle. The BESST outcome was designed to harmonize international reporting and to focus on outcomes that matter most to patients, the delivery of healthy babies and the burden of treatment to couples (Min et al., 2004). Thus, for each IVF cycle initiated, the delivery of a single, term gestation, live baby is the most relevant and important standard of success. In line with this philosophy, a ‘freeze-all’ or eFET strategy may be proposed as a safer approach for both the IVF patient and the IVF baby to achieve an optimal outcome for both.
Ovarian hyperstimulation syndrome The first strong argument for the eFET strategy is the prevention of ovarian hyperstimulation syndrome (OHSS). OHSS arguably remains a major cause of morbidity in IVF treatment. It is the single most important cause of IVF-related mortality in non-pregnant women with an estimated incidence of three deaths per 100 000 stimulated cycles (Venn et al., 2001; Braat et al., 2010). The related health-economic costs of severe OHSS are significant (Delvigne and Rozenberg, 2003). OHSS results from an increase in vascular permeability. The hCG used to trigger oocyte maturation appears to play an integral part in the aetiology of the condition and indeed, subsequent trophoblast-derived hCG dramatically worsens and prolongs the symptoms of severe OHSS (Whelan and Vlahos, 2000; Aboulghar, 2009). Thus, an important
risk-reducing strategy has been to cancel the embryo transfer and freeze all the embryos. This is necessary since our inability to reliably predict which patients will develop OHSS has severely restricted efforts to reduce its incidence. However, while elective cryopreservation in all cycles can prevent pregnancy-induced late OHSS, it cannot prevent early OHSS if hCG is used to trigger oocyte maturation (Endo et al., 2002). Currently, the stimulation protocol that all but eliminates the risk of both early and late OHSS is a GnRH antagonist protocol with a GnRH agonist trigger followed by cryopreservation of all embryos (Martinez et al., 2013; Nelson, 2013). The trade-off with this much safer protocol lies in the inadequate luteal support which all but abrogates the likelihood of pregnancy if a fresh transfer is carried out (Youssef et al., 2011). Attempts have been undertaken to re-establish appropriate luteal support with the co-administration of low doses of hCG at the time of the GnRH agonist trigger (Humaidan et al., 2013), but further evidence from randomized controlled trials (RCTs) is required to show whether such a hCG add-back protocol can restore CPRs without a concomitant rise in OHSS rates (Seyhan et al., 2013). Early evidence suggests that equivalent live birth rates following fresh embryo transfer can also be achieved when recombinant luteinizing hormone (rLH) is used instead of urinary hCG to trigger final oocyte maturation (Youssef et al., 2011) or when GnRH agonist induction of a spontaneous LH surge is followed by daily low-dose rLH for luteal support (Papanikolaou et al., 2011) or robust steroid replacement (Engmann et al., 2008). The prohibitive cost of an optimal trigger dose of rLH and the fact that OHSS rates are not reduced (Youssef et al., 2011), however, makes this an unviable proposition. The use of daily low dose of rLH or combined steroids for luteal support only, after a GnRH agonist trigger, is more promising, but the studies so far are too small to be confident that live birth rates are indeed the same. Furthermore, these attempts to rescue the fresh cycle following a GnRH agonist trigger after a GnRH antagonist protocol may be a moot point as avoidance of OHSS is not the only reason eFET should be considered. As discussed below, many of the concerns with fresh embryo transfers are linked to the effects of the supraphysiological endocrine environment created on the endometrium during ovarian stimulation.
Perinatal and obstetric outcomes There is a large body of evidence showing that a freeze-all strategy would lead to greatly improved birth outcomes. First, a population study in Victoria, Australia, showed that a specific group of birth defects linked to abnormal blastogenesis were increased .3-fold in fresh embryo transfers [adjusted OR (aOR) 3.65; 95% CI 2.02–6.59] but not in FETs (aOR 1.60; 95% CI 0.69–3.69) when both were compared with spontaneously conceived babies (Halliday et al., 2010). Secondly, a recent systematic review and meta-analysis (Maheshwari et al., 2012) analysed data from 11 observational studies on obstetric and perinatal outcomes. The authors concluded that singleton pregnancies following frozen versus fresh embryo transfers were significantly less likely to be complicated by perinatal mortality [relative risk (RR) 0.68; 95% CI 0.48 –0.96], small for gestational age (RR 0.45; 95% CI 0.30 – 0.66), preterm birth (RR 0.84; 95% CI 0.78 –0.90), low birthweight (RR 0.69; 95% CI 0.62 –0.76) and antepartum haemorrhage (RR 0.67; 95% CI 0.55 –0.81). Although the evidence from this meta-analysis appears to favour FET, there was heterogeneity between the 11 studies at the level of design, population and also the freezing, thawing
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freeze cleavage-stage and blastocyst-stage embryos. The first success in cryopreservation was achieved with slow-freezing methods, and for the past two decades slow freezing has been the method of choice for embryo cryopreservation. More recently, optimized vitrification methods, with the use of cryoprotectant and rapid freezing, have become more widely utilized. When implemented correctly, vitrified embryos have excellent survival rates following thawing. Indeed, a 95% survival rate has been reported in a large cohort of vitrified blastocysts (Cobo et al., 2012). Recent systematic reviews and meta-analyses have compared the outcomes of vitrification versus slow-freezing methods (Loutradi et al., 2008; AbdelHafez et al., 2010). Vitrification has higher post-thaw survival rates compared with slow freezing both for cleavagestage [OR 15.57; 95% confidence interval (95% CI) 3.68 –65.82] and blastocyst-stage embryos (OR 2.2; 95% CI 1.53 –3.16) (four studies, Loutradi et al., 2008). Furthermore, assessment of embryo vitrification by clinical outcomes (six studies, AbdelHafez et al., 2010) showed that CPRs were .50% higher in the vitrification groups compared with slow frozen embryos (OR 1.55, 95% CI 1.03–2.32), with similar results seen in ongoing pregnancy and implantation rates. This improved ability to store and thaw embryos for later use has significantly reduced our reliance on fresh embryo transfer cycles and undoubtedly has contributed to make IVF a more patient friendly and less expensive fertility option. Despite this, the prevailing view is that being able to freeze embryos is merely a welcome bonus or add-on to the assisted reproduction process rather than being the first choice option resulting in better maternal and fetal outcomes. However, below we discuss a number of clinical arguments that challenge this view and support the strategy of freezing all the viable embryos in every oocyte retrieval cycle.
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The role of preimplantation genetic screening A freeze-all strategy is also becoming more widely adopted because of the increased use of preimplantation genetic screening (PGS). This is despite a systematic review and meta-analysis (Mastenbroek et al., 2011) questioning the effectiveness of fluorescence in situ hybridization (FISH) (Rebois and Fishman, 1984) analysis on 8–11 chromosomes. The results of this review showed that PGS had a detrimental effect on live birth outcomes (RR 20.08; 95% CI 20.13 to 20.03) and suggested that technical limitations of FISH testing together with the biopsy procedure were the causal effects. Two significant developments have since led to renewed interest in PGS. First, comprehensive chromosome screening (CCS) has brought
about a significant change to PGS over the past few years. CCS allows the analysis of all 23 pairs of chromosomes thereby facilitating embryo selection based on the full karyotype (Rubio et al., 2013). However, the turn-around time is highly variable depending on the technology used and can require 4– 72 h to obtain results (Handyside, 2013). Coupled with this is evidence that the optimal time to biopsy preimplantation embryos, to avoid diminishing their developmental potential, is at the blastocyst stage (Scott et al., 2013a, b). Therefore, cryopreservation of blastocysts post-biopsy is generally recommended on Day 5 or 6 depending on the CCS technique used. Vitrification of biopsied blastocysts remains more effective than biopsied cleavage-stage embryos (Zhang et al., 2012) with better survival and pregnancy rates than with slow freezing of biopsied blastocysts (Keskintepe et al., 2009). In addition, preliminary data (Schoolcraft and Katz-Jaffe, 2013) from patients of advanced maternal age using single blastocyst transfer reinforce the argument for embryo vitrification. This study indicated that the combination of CCS with vitrification and FET in a subsequent menstrual cycle resulted in significantly higher implantation rates compared with selection of embryos based on morphological selection alone, followed by fresh or vitrified single blastocyst transfers (60% in CCS combined with vitrification versus 40.7% for morphological selection/fresh transfer and 43.8% for morphological/FET). Technological advances will very likely make it easier to obtain a PGS result from a trophectoderm biopsy within a time frame that would allow the fresh embryo transfer to proceed. Nevertheless, those same technological advances, including whole-genomic sequencing (Winand et al., 2014), will make the interpretation of the results increasingly complex. This will significantly increase the time required to provide appropriate genetic counselling (Ormond, 2013). The secondary benefit of a freeze-all strategy then is that it allows sufficient time for patients to digest and further question the information they will receive following advanced genetic analysis of their embryos.
Success rates with elective FET A stronger argument in favour of adopting eFET as the default IVF protocol is the potential for higher pregnancy rates. A recent systematic review and meta-analysis of three randomized controlled trials including 633 women assessed clinical and ongoing CPRs and miscarriage rates following eFET and fresh embryo transfers (Roque et al., 2013). The overall conclusions were that eFET resulted in an increase in the CPR (RR 1.31; 95% CI 1.10– 1.56) and the ongoing pregnancy rate (RR 1.32; 95% CI 1.10 –1.59), with no difference in the miscarriage rate (RR 0.83; 95% CI 0.43 –1.60). Randomized patients were high-responders in two studies (Aflatoonian et al., 2010; Shapiro et al., 2011a) and normal responders in the other study (Shapiro et al., 2011b), but all showed the same trend. The observed effect was also similar in those studies that transferred Day 5 (Shapiro et al., 2011a, b) and Day 2 embryos (Aflatoonian et al., 2010). Nevertheless, we must approach and interpret these data with some caution. The populations studied were generally younger, with the average age ranging between 27 and 33 years, and it remains to be seen whether these findings apply to women with a poorer prognosis. However, the preliminary study by Schoolcraft and Katz-Jaffe (2013) was conducted in women over 35 years of age, and it seems likely that these findings will bear out in a ‘poor prognosis’ population. Additionally, the RCTs did not report on live birth rates, even though a meta-analysis
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and transfer protocols: subsequent inclusions and analysis of a greater number of studies could further strengthen these conclusions. A large and more recent study from the Nordic countries (Wennerholm et al., 2013) has since largely corroborated the above findings. It also showed that compared with singletons born after fresh IVF and ICSI, singletons born after FET had a higher risk of being large for gestational age (aOR 1.45; 95% CI 1.27 –1.64) and having macrosomia (aOR 1.58; 95% CI 1.39– 1.80), which is in line with findings from other large population-based studies (Pelkonen et al., 2010; Pinborg et al., 2010; Sazonova et al., 2012). However, another large population-based study (of 56792 infants) found that fresh transfers result in low birthweight rather than FET offspring necessarily being larger (Kalra et al., 2011). It is unclear to what extent known confounding factors such as smoking, BMI and cause of infertility play a role in these associations as they were not controlled for in this study. Further recent evidence from a large observational study by Roy et al. (2014) reports on clinical pregnancy and neonatal outcomes of 1157 fresh and 645 vitrified-warmed single embryo blastocyst transfers. The blastocyst thaw survival rate was 94.4% and similar clinical outcomes were achieved for fresh and vitrified-warmed blastocyst transfers with live birth rates of 52.8 versus 55.3%, respectively, for grade I blastocysts. Babies conceived following single embryo transfer (SET) with vitrifiedwarmed blastocysts were born on average 0.3 weeks later and had a birthweight that was 145 g heavier compared with fresh transfers. Overall, the findings indicate that birth outcomes after FET are better compared with fresh embryo transfers. It should also be emphasized that these studies were all observational in nature, and that the frozen/ thawed embryos available for transfer were therefore inevitably ‘second choice’ as the best embryos would have been transferred in the preceding fresh transfer. It is thus highly conceivable that the obstetrical and neonatal outcomes following eFET, which also includes the best embryos, may be even further improved, but such studies have yet to be performed. Many of these adverse perinatal outcomes may originate in impaired early placentation with evidence of adverse effects on early placentation during stimulated IVF cycles. In particular, this may result from the associated supraphysiological estradiol levels, reported in studies showing significantly lower PAPP-A levels during first-trimester combined screening (Amor et al., 2009; Giorgetti et al., 2013). In other studies, elevated serum estradiol levels are associated with an increased risk of being small for gestational age (Griesinger et al., 2007) and the development of pre-eclampsia (Imudia et al., 2012). However, the latter risk is abrogated by implementation of FET cycles (Imudia et al., 2013).
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The scientific background The endometrium in controlled ovarian stimulative cycles Pregnancy success is dependent on a complex pre- and postimplantation dialogue between the embryo and the maternal endometrium (Fig. 1) that is initially dependent on secreted factors within the local microenvironment of the uterine cavity, but also includes subsequent interactions within the decidua. This early communication is optimal during the ‘window of implantation’ or receptive phase of the menstrual cycle, which occurs between Days 6 and 10 after ovulation (approximately) spanning the mid-secretory phase of a natural menstrual cycle (Navot and Bergh, 1991). It is clear that alterations in the timing of endometrial development during each menstrual cycle or the quality of endometrial receptivity during this window are highly implicated in failures of IVF. Indeed, a recent study has shown that, in good prognosis patients, 30% of euploid single blastocyst transfers do not result in a pregnancy (Yang et al., 2012). During the natural menstrual cycle in normo-ovulatory women, the endometrium repairs post-menses, and then regenerates (or ‘thickens’) during the proliferative phase under the influence of rising estrogen concentrations. Following ovulation and formation of the corpus luteum, the endometrium undergoes secretory transformation, which is driven by progesterone in the presence of estrogen, progressively achieving a state of receptivity over the course of around 6–10 days (in an average 28-day menstrual cycle). Progesterone induces differentiation of both endometrial epithelial and stromal cells (decidualization), along with changes in the vasculature, extracellular matrix and leucocyte content
of the tissue (Salamonsen et al., 2009). However, in an IVF cycle, ovarian multi-follicular development with exogenous hormones exposes the endometrium to supraphysiological concentrations of estrogen and progesterone, which can dramatically impact the timing of endometrial development and/or the achievement of receptivity (Fauser and Devroey, 2003). It has been clinically demonstrated that patients with high estradiol concentrations produce significantly more oocytes and also have elevated progesterone concentrations (Kyrou et al., 2009). If pregnancy is achieved, the effect of elevated estradiol can be far-reaching: for example, the risk for antepartum haemorrhage increases with increasing oocyte numbers and estradiol concentrations (Healy et al., 2010). Abnormal steroid hormone concentrations have been known for some time to detrimentally affect endometrial morphology and therefore receptivity (Thomas et al., 2002). A recent systematic review and meta-analysis of pregnancy outcomes following precocious progesterone elevation on the day of hCG administration (0.08 ng/ml) confirmed that premature elevation in progesterone is associated with a reduced probability of clinical pregnancy in fresh embryo transfer cycles, but not in frozen/thawed or donor –recipient IVF cycles (Venetis et al., 2013), implying that both appropriate concentrations and timing of steroid hormones are critical to receptivity. Alterations in the endometrium at a molecular level have been revealed in genomic analyses of stimulated endometria from women with elevated progesterone concentrations (versus those with normal progesterone concentrations), sampled between hCG+7-8 (Labarta et al., 2011; Li et al., 2011). These studies revealed alterations in both miRNA and mRNA expression, with 4 miRNAs and up to 140 mRNAs being dysregulated in an elevated progesterone group (Labarta et al., 2011; Li et al., 2011).
Histological, transcriptomic and proteomic studies A number of histological, transcriptomic and proteomic studies of endometrial tissues and of the intrauterine environment have demonstrated dramatic differences between the normal cycling endometrium and the IVF endometrium. Taken together, these data provide strong evidence that endometrial receptivity is disturbed in IVF cycles and is likely to significantly limit the success of fresh embryo transfer. Histological studies of the IVF/ICSI endometrium stimulated with the GnRH agonist or GnRH antagonist protocol have demonstrated that there is a complete failure to achieve pregnancy/implantation when an advancement (as assessed by highly experienced pathologists using Noyes’ criteria for endometrial dating) in the endometrial development of ≥3 days is observed (Chetkowski et al., 1997; Ubaldi et al., 1997; Kolibianakis and Devroey, 2002; Van Vaerenbergh et al., 2009). However, a recent more comprehensive histological/immunohistochemical study suggests that the absence of receptivity is more complicated than mere developmental advancement, and that a complex lack of synchrony of development between the different cellular and structural compartments of the endometrium results from ovarian stimulation protocols (Evans et al., 2012). This comprehensive study of endometrial histology was performed in five groups of women at LH/hCG+2. These were: GnRH agonist-treated oocyte donors; GnRH agonist-treated women who failed to become pregnant following fresh embryo transfer; GnRH agonist-treated women who became pregnant following fresh embryo transfer; GnRH antagonist-treated women with unsuccessful fresh embryo transfer and normal fertile women (LH+2) as the comparison group. The study demonstrated dramatic alterations in endometrial
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has found that conclusions on the effectiveness of a treatment based on either clinical pregnancy or live birth as end-points are comparable (Clarke et al., 2010). It would certainly be more reassuring to find that the cumulative live birth rates from one oocyte retrieval are equivalent. Also, none of the RCTs presented data on perinatal outcomes. Finally, the cost implications of the two strategies (fresh versus frozen transfers) remain unknown as no health-economic data were available. In the three RCTs, all the patients who were randomized to eFET received hormonal preparation; it is unclear why hormonal stimulation of the endometrium was offered, given that there is little clinical evidence to support this other than for women with irregular or anovulatory cycles (Groenewoud et al., 2013). Conversely, some clinical studies suggest that hormonal intervention can be detrimental to endometrial receptivity and pregnancy success (Fatemi et al., 2010; Chang et al., 2011; El Bahja et al., 2012; Xiao et al., 2012), and it is therefore plausible that the true effect size of eFET in natural cycles may well be higher in the absence of hormonal intervention. Importantly, over 1000 patients have already been recruited for a new multicentre randomized trial (The ANTARCTICA trial) to assess the effect of natural and artificial FET cycles on numerous outcomes such as live birth rate, health costs, adverse events and patient burden (Groenewoud et al., 2013) and its outcomes are keenly awaited. Studies validating these positive outcomes in women who are older or poor responders and in natural cycle eFET are certainly necessary, but the evidence so far further underpins the principle that better and safer pregnancy outcomes can be achieved with eFET and, as such, the proof of burden now seems to have shifted to those who advocate fresh embryo transfers.
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histology following stimulation for multi-follicular development (Fig. 2). As the ‘gold standard’ Noyes criteria (Noyes et al., 1975) have been shown to be highly subjective in terms of endometrial dating (Murray et al., 2004) and since the major changes in the above study were observed in individual cellular compartments with the endometrium not necessarily ‘in phase’, a ‘normality score’ was developed to assess endometrial alterations in IVF cycles. This ‘normality’ scoring system individually assessed key features of endometrial glandular epithelium, luminal epithelium, stroma and vasculature in IVF endometrium at hCG+2, in comparison with the same parameters in normal cycling women at LH+2. Women treated with GnRH agonist or GnRH antagonist protocols and who did not become pregnant displayed significantly altered endometrial histology based on eight different parameters (Fig. 3). These alterations in endometrial histology did not appear to be related to their infertility as fertile oocyte donors receiving the GnRH agonist protocol displayed a similar degree of endometrial changes. Most importantly, women who did become pregnant following ovarian stimulation displayed significantly fewer histological alterations from a perfect ‘normal score’ than did the women who did not become pregnant, emphasizing that the extent of endometrial disturbance impacted receptivity and subsequent implantation (Evans et al., 2012). Other data support endometrial advancement as a major reason for failure to establish a viable pregnancy. In normal pregnancies,
Figure 3 Endometrial changes in stimulation cycles on hCG+2. Semi-quantification of the full data represented in Fig. 2. Eight features of endometrial morphology were assessed for each tissue and normalized to that for the control samples (score of 2). Red bar: fertile women, LH+2; orange bar: fertile donor; violet bar: infertile, antagonist; purple bar: infertile, agonist (no pregnancy); blue bar: infertile, agonist (pregnant) (data derived from Evans et al., 2012).
implantation which has occurred late in the menstrual cycle (as determined by ultrasensitive hCG detection in urine) is associated with early pregnancy loss (Wilcox et al., 1999). ‘Advanced’ endometrial maturation on the day of oocyte retrieval has also been correlated with altered gene
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Figure 2 Histology of the endometrium on day hCG +2, following controlled ovarian hyperstimulation. Open arrowhead: endometrial glandular epithelium, closed arrowhead: blood vessels, arrow: secretions within endometrial glands, asterisks: oedematous/expanded stroma. Representative micrographs are shown for tissue from: (A) control women of known fertility on LH+2 of an unstimulated cycle, with open arrowheads indicating the expected morphology of endometrial glands and closed arrowheads indicating the expected appearance of blood vessels; (B) donor women treated with GnRH agonist, with open arrowheads indicating the presence of vacuoles within the endometrial glands and closed arrowheads indicating the presence of enlarged blood vessels; (C) infertile women treated with GnRH antagonist, with arrows indicating the presence of accumulated secretions within the endometrial glands and asterisks indicating oedematous stroma; (D and E) infertile women treated with GnRH agonist and who did not become pregnant in the treatment cycle, with closed arrowhead indicating the presence of enlarged blood vessels close to the endometrial luminal surface (D), arrows indicating the presence of accumulated secretions within the endometrial glands (E) and asterisks indicating the presence of oedematous stroma; and (F) GnRH agonisttreated women who did become pregnant in the treatment cycle, with open arrowheads indicating a more normal endometrial glandular morphology and an asterisk indicating limited evidence of expanded stroma (modified from Evans et al., 2012).
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growth factor at hCG+6, compared with concentrations in normally fertile women at LH+6. Such changes in inflammatory chemokines production by the stimulated endometrium may reflect a more inflamed tissue, and in the case of elevated DKK-1, also a more advanced endometrium (as discussed above). Recent data suggest that the presence of an inflamed environment within the endometrium is detrimental to implantation (Yoshii et al., 2013). Indeed, Evans et al. (2012) demonstrated not only elevated leucocyte numbers within the stimulated endometrium but, more importantly, the presence of activated leucocytes with abundant degranulating neutrophils in the stimulated endometrium of women who did not become pregnant. Thus, elevated inflammation in combination with endometrial advancement is likely to contribute to the deficient receptivity and hence failure of implantation in these women. Dramatic enlargement of blood vessels is also a clear feature of endometrium in both agonist- and antagonist-treated women on hCG+2 (Evans et al., 2012) (Fig. 4). In particular, very large vessels are clearly visible immediately below the endometrial surface in those agonisttreated women who did not become pregnant following fresh embryo transfer (Fig. 2, closed arrowhead). Although the molecular mechanisms underpinning these vascular changes are not yet defined, they are in accordance with the increase in vascular permeability seen in OHSS (as discussed above). The hCG, administered to stimulate ovulation, is a likely protagonist. While blastocyst-secreted hCG can act acutely on the endometrium in conception cycles, stimulating epithelial cell production of a number of mediators including vascular endothelial growth factor A and fibroblast growth factor 2 (FGF2; both potent angiogenic factors; Licht et al., 2007; Paiva et al., 2011), its use for ovarian stimulation and therefore longer-term actions in an IVF cycle are likely considerably modified as discussed below.
Figure 4 Immunohistology for CD34-positive blood vessels (brown colour) in the endometrium of women undergoing controlled ovarian hyperstimulation. Representative micrographs are shown for tissue from: (A) control women of known fertility on LH+2 of an unstimulated cycle, with the expected appearance of small spiral arteries at this stage of the menstrual cycle; (B) donor women treated with GnRH agonist, with the presence of enlarged blood vessels; (C) infertile women treated with GnRH antagonist, with the presence of enlarged blood vessels; (D) infertile women treated with GnRH agonist and who did not become pregnant in the treatment cycle, with the presence of enlarged blood vessels and (E) GnRH agonist-treated women who did become pregnant in the treatment cycle, with the presence of small spiral arteries (modified from Evans et al., 2012).
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expression (Van Vaerenbergh et al., 2009) with an elevation of 1337 genes and a down-regulation of 1213 genes in significantly advanced (≥3 days) endometria. Indeed, transcriptomic profiling of the IVF endometrium has revealed alterations in gene expression in the endometrium taken between hCG+2 and hCG+7 compared with the natural cycle (Horcajadas et al., 2005, 2008; Liu et al., 2008; Macklon et al., 2008; Haouzi et al., 2009, 2010). Furthermore, in a tightly controlled cohort using matched samples, taken from the same women at LH+7 in nonstimulated cycles and hCG+7 in a subsequent stimulated cycle, 281 genes were found to be elevated and 277 genes were down-regulated in the endometrium at hCG+7 (Horcajadas et al., 2005). Of the identified genes, those with known roles in implantation, including leukaemia inhibitory factor (LIF) and glycodelin, were down-regulated at hCG+7. In addition, Macklon’s team (Macklon et al., 2008) demonstrated a significant elevation in Dickkopf-related protein 1 (DKK-1) in the endometrium of women treated GnRH antagonist. As DKK-1 is produced by decidualized endometrial stromal cells (Tulac et al., 2006), this strongly suggests the presence of an advanced endometrium at hCG+2 as confirmed by the presence of decidualized stromal cells (a normal feature of late secretory endometrium), in some stimulated tissues (Evans et al., 2012). Genomic analysis (Haouzi et al., 2010) has demonstrated significant alterations in a host of CCL- and CXCL-chemokines during the prereceptive to receptive transition and in stimulated endometrial tissues. These alterations are reinforced by a secretomic analysis of uterine cavity aspirates (Boomsma et al., 2010) from a mixed GnRH agonist/GnRH antagonist population, which showed significant elevation in concentrations of CCL- and CXCL-chemokines secreted from the endometrium into the uterine cavity, including interleukin (IL)-1b, IL-5, IL-10, IL-12, eotaxin, DKK-1 and heparin-binding epidermal
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Human chorionic gonadotrophin
on embryonic –endometrial crosstalk for implantation. Following preexposure to hCG, mimicking the timing of exposure in an IVF cycle (Fig. 5A), endometrial epithelial cells could no longer mount a functional implantation response to a blastocyst. The LHCG-R was downregulated, intracellular signalling (ERK 1/2 phosphorylation) could not be activated, the cells could not adhere to trophoblast-like extracellular matrices and could not relax intercellular tight junctions (Evans and Salamonsen, 2013), all of which are endometrial functions normally regulated by hCG. These data suggest that precocious exposure of the endometrium to hCG, outside the timing of normal endometrial–embryonic communication, down-regulates the endometrial epithelial LHCG-R, thus increasing the risk that the endometrium may be less responsive to peri-implantation embryonic hCG that promotes receptivity (Licht et al., 2001a, b; Paiva et al., 2011). These effects are summarized diagrammatically in Fig. 5. It is unclear how best to combine the clinical experience with the evidence from all these scientific studies on the adverse endometrial effects of early hCG exposure. Equivalent live birth rates are achieved when rLH is used instead of urinary hCG to trigger final oocyte maturation (OR 0.94; 95% CI 0.50 –1.76), but only two published RCTs were included in this analysis with a total of 280 patients randomized (Youssef et al., 2011). Along the same lines, the use of a GnRH agonist trigger to induce a spontaneous LH surge followed by daily rLH for luteal support (Papanikolaou et al., 2011) or robust steroid replacement (Engmann et al., 2008) also leads to live birth rates that are comparable to those where rhCG is used as a trigger. However, the Engmann study (Engmann et al., 2008) compared a GnRH agonist protocol/hCG trigger with a GnRH antagonist protocol/GnRH trigger. It is thus difficult to directly compare such groups. Additionally, both studies (Engmann et al., 2008; Papanikolaou et al., 2011) were again too small to be confident that no difference exists. In contrast, two studies examining the use of hCG for ovulation induction versus monitoring of the natural LH surge in natural cycle FET or IUI demonstrated a negative effect of hCG on ongoing pregnancy rates (Fatemi et al., 2010; Kyrou et al., 2012). Indeed, the study examining the use of hCG in natural cycle FET was discontinued prematurely due to the profoundly different pregnancy rates in the LH monitor group (31.1%) versus the hCG group (14.3%; Fatemi et al., 2010). These clinical studies suggest that hCG may not confer any special clinical benefits, but further studies using larger sample sizes are needed before it can be concluded that hCG either provides clinical benefit or is detrimental. It should be noted that the LHCG-R is also down-regulated after the spontaneous LH surge (Menon and Menon, 2012). It may be possible that the observed LHCG-R down-regulation in the endometrium on hCG+2 relative to LH+2 is only transient, and that the receptor recovers by the time implantation occurs 5 days later. However, it is not possible to test in vivo in humans what happens to receptor levels or activation of down-stream pathways, when a blastocyst is also present. In pseudo-pregnant rats, ovarian LHCG-R mRNA was markedly suppressed for 48 h after a large bolus of hCG, but recovered to that measured in control rats after 53 h (Harada et al., 2010). However, this study did not investigate hCG-R protein nor its functionality: it must also be remembered that rodents have very short oestrous cycles (4– 5 days) and different endometrial response times to stimuli compared with the much longer 28 day cycles in women. In baboons, a primate species with menstrual cycles similar to women, administration
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The hCG gene is transcribed by the embryo from the eight-cell stage (Bonduelle et al., 1988), and hCG protein is secreted by the embryo from the late blastocyst stage, 7–8 days after fertilization (Lopata and Hay, 1989; Lopata and Oliva, 1993). The ‘classic’ role of hCG is considered to be its luteotrophic function in maintaining the corpus luteum, and thus progesterone production in the early stages of pregnancy. Indeed, this action is essential for pregnancy establishment. Due to its structural similarities to LH and its ability to occupy the same receptor (Kessler et al., 1979), hCG has traditionally been used in IVF to mimic the mid-cycle LH surge. However, significant differences exist in the half-lives of these ligands; while that of LH is short, 60 min (Yen et al., 1968), hCG has a much longer half-life of .24 h (Damewood et al., 1989). It has been estimated that following a bolus of 10 000 IU hCG, a dose which is commonly used to trigger ovulation, hCG remains ‘bioavailable’ for receptor stimulation for .5 days (Chan et al., 2003). It has thus been proposed that prolonged stimulation of the LH/hCG receptor will have deleterious effects on both ovarian function and the luteal phase (Fauser et al., 2002; Humaidan et al., 2005; Kolibianakis et al., 2005; Griesinger et al., 2006, 2007). In elegant experiments conducted by Licht et al., (1998), hCG was infused directly into the uterine cavity of women. For the first time, a nongonadotropic role for hCG was clearly demonstrated; the infused hCG promoted endometrial secretion of vascular endothelial growth factor (VEGF) and leukaemia inhibitory factor (Kalra et al., 2011). Subsequent studies in which hCG was similarly infused into the uterine cavity of baboons (Sherwin et al., 2007), and in which human endometrial epithelial cells were treated in vitro with hCG (Paiva et al., 2011), confirmed and extended these findings, demonstrating that hCG elevates gene expression and secretion of a highly selected cohort of ‘pro-implantation’ factors including LIF, IL-11, VEGF, FGF2 and prokineticin (PROK)-1 (Perrier d’Hauterive et al., 2004; Berndt et al., 2006; Sherwin et al., 2007; Evans et al., 2009; Paiva et al., 2011). Hence, it is likely that during the normal peri-implantation period in a conception cycle, blastocyst-derived hCG can enhance endometrial receptivity. These ‘pro-implantation’ factors were induced under ‘acute’ conditions in culture, with the endometrial epithelial cells exposed to hCG for only a short duration. In the baboon model, however, when hCG was continuously infused into the uterine cavity for 5 days, the LH-CG receptor (R), through which hCG exerts its action, was down-regulated (Cameo et al., 2006; Sherwin et al., 2007). This suggested that the endometrium becomes less responsive to hCG upon prolonged exposure, due to receptor down-regulation. Indeed, this phenomenon was first described in gonadal tissues more than 30 years ago (Conti et al., 1976; Sharpe, 1976; Rebois and Fishman, 1984; Lakkakorpi et al., 1993; Peegel et al., 2005). LH-CGR down-regulation has recently been confirmed in the human endometrium retrieved 2 days after the hCG trigger (hCG+2) in patients undergoing IVF (Evans and Salamonsen, 2013). Endometrial samples from women undergoing either GnRH agonist or GnRH antagonist stimulation protocols followed by triggering with a single bolus of recombinant hCG (rhCG; 250 mg) were collected at hCG+2. In comparison with tissue at LH+2 from normally cycling women of known fertility, the endometrial epithelial LH-CGR was significantly down-regulated in the IVF endometria exposed to hCG. Subsequent functional cell culture studies confirmed the importance of receptor down-regulation
Evans et al.
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Scientific evidence supports frozen embryo transfer
of hCG 5–6 days after ovulation followed by endometrial sampling 6 days later, demonstrated a down-regulation of endometrial epithelial LHCG-R protein (Cameo et al., 2006). In confirmation, there is recent in vivo evidence that hCG can cause longer lasting changes in the human endometrium. A study comparing a traditional hCG ovulation trigger followed by luteal phase progesterone versus a GnRH agonist trigger followed by either (i) estrogen and progesterone or (ii) recombinant LH combined with estrogen and progesterone found that the endometrial gene expression profiles are significantly different 7 days after the hCG trigger, which coincides with the window of implantation (Bermejo et al., 2014). Interestingly, ADAMTS8, a proteinase with potent antiangiogenic properties, was highly significantly up-regulated in the hCG ovulation trigger group at hCG+7 compared with the GnRH trigger groups, demonstrating that the effects of hCG used for ovulation are likely perpetuated, mediating detrimental effects on the endometrium later in the stimulation cycle. It is tempting then to speculate that the 6-fold higher expression of ADAMTS8 in endometria exposed to rhCG plays a role in initiating abnormal placentation, ultimately resulting in some of the adverse perinatal outcomes associated with fresh embryo transfers.
Evidence from non-human models Despite substantial differences in implantation between species (Carson et al., 2000), rodents can provide models for functional studies not possible in women. For example in embryo transfer studies in rodents (Ertzeid and Storeng, 2001), when normal embryos were transferred to recipients which had undergone ovarian stimulation, there was a highly significant decrease in the implantation rate compared with embryo transfer to control recipients, again indicative of an effect of ovarian stimulation protocols on endometrial receptivity. More recent molecular studies in a rat ovarian stimulation model have further defined these changes including: (i) altered uterine expression of TGF-b1 and -b2 (Jovanovic and Kramer, 2010) and (ii) decreased b3 integrin and VEGF (Biyiksiz et al., 2011), the latter being in accordance with human data (Thomas et al., 2003a, b). A novel recent finding is that following ovarian stimulation in the rat model, there is an increase in the fluid-transporting molecule aquaporin 5 and an accompanying decrease in claudin 4 at the time of implantation. These findings strongly indicate disturbance of the osmotic gradient for water across tight junctions (Lindsay and Murphy, 2013). In turn, these regulate the uterine fluid dynamics that contributes to a favourable implantation environment. Aquaporin 2, the human equivalent to rat
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Figure 5 A schematic summarizing experimentally proven alterations in the response of the endometrium to either blastocyst hCG within a normal pregnancy or within a cycle in which hCG has been used to induce ovulation. (A) Endometrial epithelial cells were treated with a low dose of hCG (0.5– 5 IU) each day for 5 days (arrows). Both cells pre-exposed to hCG and naı¨ve cells (never been exposed to hCG) were then treated with 20 IU hCG to mimic that secreted by the incoming blastocyst in a conception cycle and functional outcomes were examined. (B) In cells pre-exposed to low doses of hCG, 20 IU hCG could not mediate signalling, relaxation of tight junctions or adhesiveness. In these pre-exposed cells, the LHCG-R was downregulated suggesting the cells were refractory to hCG. In naı¨ve cells exposed to the dose of hCG mimicking endometrial exposure to blastocyst-secreted hCG for the first time as experienced in a normal conception cycle, 20 IU hCG enhanced ERK 1/2 phosphorylation, relaxation of intercellular tight junctions and adhesiveness of endometrial epithelial cells to trophoblast-like extracellular matrices, all of which are important features of the epithelium associated with successful implantation.
10 aquaporin 5, has been identified in human endometrial epithelium and appears to be regulated for implantation (Hildenbrand et al., 2006); whether it changes in women following ovarian stimulation should be examined.
Are we there yet?
The role of endometrial receptivity tests It is of course hoped that further evidence from robust RCTs will help overcome the significant hurdles discussed above. However, since biomarker tests for receptivity are close to becoming a reality, the future
application of such tests will undoubtedly assist the patient and the clinician in deciding whether the transfer of a fresh embryo should even be contemplated. In recent years, considerable effort has been applied to the discovery of biomarkers for endometrial receptivity, based on either mRNA, protein or lipids, that could identify a receptive endometrium and hence an appropriate environment for embryo implantation. These have been well summarized in recent reviews (Edgell et al., 2013; Lessey et al., 2013) and include tests that can be performed on endometrial biopsies or on lavage or aspirates of the uterine cavity (Cheong et al., 2013; Garrido-Gomez et al., 2013; Vilella et al., 2013). It is also important to consider how such tests might be used in a clinical environment to provide the information needed to guide clinical practice. Commercial tests using endometrial tissue biopsies are already available (Kliman et al., 2006; Garrido-Gomez et al., 2013; Vilella et al., 2013). Because of the tissue damage to the endometrium, these tests can only be applied in a natural cycle during the investigation of infertility, but not in the stimulation cycle during which the embryo will be replaced. More relevant in the context of this review, they could be performed in a cycle preceding the embryo transfer. The test results can then be used to ‘time’ the optimal window of implantation or to optimize endometrial receptivity with hormonal therapy during the transfer cycle. However, as discussed extensively above, the endometrium in an IVF cycle presents a dramatically altered environment compared with the normal cycle and the relevance of these biopsy-based diagnoses in a natural cycle preceding the stimulated one is therefore not immediately apparent. Caution is also necessary when interpreting these tests as their clinical value remains to be fully determined. Neither test has been validated in large independent cohorts using the STARD guidelines (Bossuyt and Reitsma, 2003). In view of this and the potential for cycle-to-cycle variability of endometrial receptivity (Ruiz-Alonso et al., 2013), the ideal test would be relatively non-invasive and performed within the embryo transfer cycle itself. Uterine lavage or aspiration is feasible within a stimulated IVF cycle, enabling the measurement of potential biomarkers that predict the attainment of full receptivity later in the cycle (Boomsma et al., 2010; Edgell et al., 2013). Given the significant alterations to the endometrium caused by the ovarian stimulation during IVF (Evans et al., 2012), these tests could be useful in determining whether it may be better to proceed with eFET than to continue with a fresh transfer; but again full validation of such diagnostic tests is still awaited.
Conclusion The evidence supporting eFET, when taken together from large observational studies and RCTs, is growing, not only in terms of achieving higher pregnancy rates but, more importantly, also in terms of lower maternal and infant morbidity and mortality. Additional RCTs in less selected populations are welcome, but they will have to prove that the reported health risks of fresh transfer cycles are significantly outweighed by higher cumulative pregnancy rates or lower costs. In the absence of such RCTs, the major pressures against eFET remain the required upskilling of IVF units, patient preference, restrictive healthcare funding models and government regulation. It is likely that the implementation of eFET will be gradual. The addition of endometrial receptivity tests to the repertoire of tools for clinical decision making will provide further guidance about when to abandon fresh embryo transfers in individual cases.
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Given that the clinical and scientific evidence in favour of eFET is growing, why is it that we have not already seen a greater adoption of this strategy? Major reasons currently given for continuing with fresh embryo transfers are patient preference and governmental funding models that discourage a freeze-all strategy. There are currently no studies that have investigated patient attitudes towards eFET, and it is therefore presumptuous to assume that the majority of patients would prefer a fresh embryo transfer if given access to objective information. In a very similar policy debate, studies investigating patient attitudes towards elective SET(eSET) clearly show that the majority of informed patients are sufficiently concerned about adverse outcomes to make deliberate choices regarding their IVF treatment (Hope and Rombauts, 2010; Martini et al., 2011; Stillman et al., 2013). It is also true that patients become less risk averse when faced with multiple IVF failures and if the personal cost of IVF is high (Maheshwari and Bhattacharya, 2013) and the same may be true when deciding between a fresh embryo transfer or eFET. Nevertheless, the available evidence on eSET has persuaded countries including Belgium and Sweden to overrule patient autonomy by legislating the number of embryos that can be transferred even though the scientific debate continues (Stillman et al., 2013; Gleicher et al., 2014). The key considerations bringing about these legislative initiatives were health-economic factors, in particular the increased health burden of multiple pregnancies (Stillman et al., 2013), leading Chambers and colleagues to argue that ‘for the sake of the health of children born following ART, we should be asking: “Can we afford not to fund it?”’ (Chambers et al., 2011). A similar argument, of course, holds for eFET given the 60% increased risk of blastogenesis defects, the 55% increase in babies born small for gestational age and the more than 30% increased risk for perinatal mortality, low birthweight and antepartum haemorrhage with fresh embryo transfers (Maheshwari et al., 2012). Future RCTs comparing fresh transfers versus eFET will therefore also have to measure health savings as one of their important end-points. As it stands, legislative change lifting the restrictions on the ability to freeze embryos will be required in some countries before patients there can even consider eFET (Beier and Beckman, 1991; Levi Setti et al., 2011). The implementation of eFET may also require changes to reporting rules, given that embryo-banking cycles can have the potential to distort publicly available ‘league’ tables (Kushnir et al., 2013a, b; Meldrum, 2013). Clearly, much more work needs to be done to educate both patients and government regulators of the short- and long-term benefits of eFET.
Evans et al.
Scientific evidence supports frozen embryo transfer
Authors’ roles The review was planned and finalized by J.E., L.A.S. and L.J.F.R., while written sections and critical discussions were contributed by all authors. J.E., N.J.H. and T.A.E. performed much of the cited experimental work on endometrial receptivity. L.J.R. and B.J.V. contributed to the collections of uterine tissue used in the experimental work.
Funding
Conflict of interest L.J.F.R., B.J.V. and P.J.L. are minority shareholders of Monash IVF. There are no other conflicts of interest to report.
References AbdelHafez FF, Desai N, Abou-Setta AM, Falcone T, Goldfarb J. Slow freezing, vitrification and ultra-rapid freezing of human embryos: a systematic review and meta-analysis. Reprod Biomed Online 2010;20:209 –222. Aboulghar M. Symposium: update on prediction and management of OHSS. Prevention of OHSS. Reprod Biomed Online 2009;19:33–42. Aflatoonian A, Oskouian H, Ahmadi S, Oskouian L. Can fresh embryo transfers be replaced by cryopreserved-thawed embryo transfers in assisted reproductive cycles? A randomized controlled trial. J Assist Reprod Genet 2010;27:357 –363. Amor DJ, Xu JX, Halliday JL, Francis I, Healy DL, Breheny S, Baker HW, Jaques AM. Pregnancies conceived using assisted reproductive technologies (ART) have low levels of pregnancy-associated plasma protein-A (PAPP-A) leading to a high rate of false-positive results in first trimester screening for Down syndrome. Hum Reprod 2009;24:1330 –1338. Beier HM, Beckman JO. Implications and consequences of the German Embryo Protection Act. Hum Reprod 1991;6:607– 608. Bermejo A, Cerrillo M, Ruiz-Alonso M, Blesa D, Simo´n C, Pellicer A, Garcia-Velasco JA. Impact of final oocyte maturation using gonadotropin-releasing hormone agonist triggering and different luteal support protocols on endometrial gene expression. Fertil Steril 2014;101:138– 146. Berndt S, Perrier d’Hauterive S, Blacher S, Pequeux C, Lorquet S, Munaut C, Applanat M, Herve MA, Lamande N, Corvol P et al. Angiogenic activity of human chorionic gonadotropin through LH receptor activation on endothelial and epithelial cells of the endometrium. FASEB J 2006;20:2630– 2632. Biyiksiz PC, Filiz S, Vural B. Is sildenafil citrate affect endometrial receptivity? An immunohistochemical study. Gynecol Endocrinol 2011;27:767 –774. Bodri D, Sunkara SK, Coomarasamy A. Gonadotropin-releasing hormone agonists versus antagonists for controlled ovarian hyperstimulation in oocyte donors: a systematic review and meta-analysis. Fertil Steril 2011;95:164 –169. Bonduelle ML, Dodd R, Liebaers I, Van Steirteghem A, Williamson R, Akhurst R. Chorionic gonadotrophin-beta mRNA, a trophoblast marker, is expressed in human 8-cell embryos derived from tripronucleate zygotes. Hum Reprod 1988; 3:909– 914. Boomsma CM, Kavelaars A, Eijkemans MJ, Fauser BC, Heijnen CJ, Macklon NS. Ovarian stimulation for in vitro fertilization alters the intrauterine cytokine, chemokine, and growth factor milieu encountered by the embryo. Fertil Steril 2010;94:1764 –1768. Bossuyt PM, Reitsma JB. The STARD initiative. Standards for reporting of diagnostic accuracy. Lancet 2003;361:71. Braat DD, Schutte JM, Bernardus RE, Mooij TM, van Leeuwen FE. Maternal death related to IVF in the Netherlands 1984 –2008. Hum Reprod 2010;25:1782– 1786.
Cameo P, Szmidt M, Strakova Z, Mavrogianis P, Sharpe-Timms KL, Fazleabas AT. Decidualization regulates the expression of the endometrial chorionic gonadotropin receptor in the primate. Biol Reprod 2006;75:681–689. Carson DD, Bagchi I, Dey SK, Enders AC, Fazleabas AT, Lessey BA, Yoshinaga K. Embryo implantation. Dev Biol 2000;223:217– 237. Chambers GM, Illingworth PJ, Sullivan EA. Assisted reproductive technology: public funding and the voluntary shift to single embryo transfer in Australia. Med J Aust 2011;195:594– 598. Chan CC, Ng EH, Chan MM, Tang OS, Lau EY, Yeung WS, Ho PC. Bioavailability of hCG after intramuscular or subcutaneous injection in obese and non-obese women. Hum Reprod 2003;18:2294 –2297. Chang EM, Han JE, Won HJ, Kim YS, Yoon TK, Lee WS. Effect of estrogen priming through luteal phase and stimulation phase in poor responders in in-vitro fertilization. J Assist Reprod Genet 2011;29:225 –230. Cheong Y, Boomsma C, Heijnen C, Macklon N. Uterine secretomics: a window on the maternal-embryo interface. Fertil Steril 2013;99:1093– 1099. Chetkowski RJ, Kiltz RJ, Salyer WR. In premature luteinization, progesterone induces secretory transformation of the endometrium without impairment of embryo viability. Fertil Steril 1997;68:292–297. Clarke JF, van Rumste MM, Farquhar CM, Johnson NP, Mol BW, Herbison P. Measuring outcomes in fertility trials: can we rely on clinical pregnancy rates? Fertil Steril 2010; 94:1647– 1651. Cobo A, de los Santos MJ, Castello D, Gamiz P, Campos P, Remohi J. Outcomes of vitrified early cleavage-stage and blastocyst-stage embryos in a cryopreservation program: evaluation of 3,150 warming cycles. Fertil Steril 2012;98:1138–1146. e1131. Conti M, Harwood JP, Hsueh AJ, Dufau ML, Catt KJ. Gonadotropin-induced loss of hormone receptors and desensitization of adenylate cyclase in the ovary. J Biol Chem 1976;251:7729 –7731. Damewood MD, Shen W, Zacur HA, Schlaff WD, Rock JA, Wallach EE. Disappearance of exogenously administered human chorionic gonadotropin. Fertil Steril 1989; 52:398– 400. Delvigne A, Rozenberg S. Review of clinical course and treatment of ovarian hyperstimulation syndrome (OHSS). Hum Reprod Update 2003;9:77– 96. Edgell TA, Rombauts LJ, Salamonsen LA. Assessing receptivity in the endometrium: the need for a rapid, non-invasive test. Reprod Biomed Online 2013;27:486–496. El Bahja D, Hertz P, Schweitzer T, Lestrade F, Ragage JP. Frozen embryo transfer protocol: does spontaneous cycle give good results?. Gynecol Obstet Fertil 2013; 41:648– 652. Endo T, Honnma H, Hayashi T, Chida M, Yamazaki K, Kitajima Y, Azumaguchi A, Kamiya H, Kudo R. Continuation of GnRH agonist administration for 1 week, after hCG injection, prevents ovarian hyperstimulation syndrome following elective cryopreservation of all pronucleate embryos. Hum Reprod 2002;17:2548– 2551. Engmann L, DiLuigi A, Schmidt D, Benadiva C, Maier D, Nulsen J. The effect of luteal phase vaginal estradiol supplementation on the success of in vitro fertilization treatment: a prospective randomized study. Fertil Steril 2008;89:554 –561. Ertzeid G, Storeng R. The impact of ovarian stimulation on implantation and fetal development in mice. Hum Reprod 2001;16:221–225. Evans J, Salamonsen LA. Too much of a good thing? Experimental evidence suggests prolonged exposure to hCG is detrimental to endometrial receptivity. Hum Reprod 2013;28:1610 –1619. Evans J, Catalano RD, Brown P, Sherwin R, Critchley HO, Fazleabas AT, Jabbour HN. Prokineticin 1 mediates fetal-maternal dialogue regulating endometrial leukemia inhibitory factor. FASEB J 2009;23:2165– 2175. Evans J, Hannan NJ, Hincks C, Rombauts LJ, Salamonsen LA. Defective soil for a fertile seed? Altered endometrial development is detrimental to pregnancy success. PLoS ONE 2012;7:e53098. Fatemi HM, Kyrou D, Bourgain C, Van den Abbeel E, Griesinger G, Devroey P. Cryopreserved-thawed human embryo transfer: spontaneous natural cycle is superior to human chorionic gonadotropin-induced natural cycle. Fertil Steril 2010; 94:2054– 2058. Fauser BC, Devroey P. Reproductive biology and IVF: ovarian stimulation and luteal phase consequences. Trends Endocrinol Metab 2003;14:236–242. Fauser BC, Bouchard P, Coelingh Bennink HJ, Collins JA, Devroey P, Evers JL, van Steirteghem A. Alternative approaches in IVF. Hum Reprod Update 2002;8:1 –9. Ferraretti AP, Goossens V, de Mouzon J, Bhattacharya S, Castilla JA, Korsak V, Kupka M, Nygren KG, Nyboe Andersen A, European IVF monitoringet al. Assisted reproductive technology in Europe, 2008: results generated from European registers by ESHRE. Hum Reprod 2012;27:2571 –2584.
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Funding for work in the authors’ laboratories has been provided by the NHMRC of Australia [programme grant #494802, project grants #1047056 and #047756 as well as fellowships grants #1002028 (L.A.S.) and #629927 (N.J.H)], the Monash IVF Research and Education Foundation, a Merck-Serono Grant for Fertility Innovation and the Victorian Government’s Operational Infrastructure Support Program.
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Imudia AN, Awonuga AO, Kaimal AJ, Wright DL, Styer AK, Toth TL. Elective cryopreservation of all embryos with subsequent cryothaw embryo transfer in patients at risk for ovarian hyperstimulation syndrome reduces the risk of adverse obstetric outcomes: a preliminary study. Fertil Steril 2013;99:168–173. Jovanovic A, Kramer B. The effect of hyperstimulation on transforming growth factor beta(1) and beta(2) in the rat uterus: possible consequences for embryo implantation. Fertil Steril 2010;93:1509 –1517. Kalra SK, Ratcliffe SJ, Coutifaris C, Molinaro T, Barnhart KT. Ovarian stimulation and low birth weight in newborns conceived through in vitro fertilization. Obstet Gynecol 2011;118:863– 871. Keskintepe L, Sher G, Machnicka A, Tortoriello D, Bayrak A, Fisch J, Agca Y. Vitrification of human embryos subjected to blastomere biopsy for pre-implantation genetic screening produces higher survival and pregnancy rates than slow freezing. J Assist Reprod Genet 2009;26:629– 635. Kessler MJ, Reddy MS, Shah RH, Bahl OP. Structures of N-glycosidic carbohydrate units of human chorionic gonadotropin. J Biol Chem 1979;254:7901 –7908. Kliman HJ, Honig S, Walls D, Luna M, McSweet JC, Copperman AB. Optimization of endometrial preparation results in a normal endometrial function test (EFT) and good reproductive outcome in donor ovum recipients. J Assist Reprod Genet 2006; 23:299– 303. Kolibianakis EM, Devroey P. The luteal phase after ovarian stimulation. Reprod Biomed Online 2002;5(Suppl 1):26 –35. Kolibianakis EM, Bourgain C, Papanikolaou EG, Camus M, Tournaye H, Van Steirteghem AC, Devroey P. Prolongation of follicular phase by delaying hCG administration results in a higher incidence of endometrial advancement on the day of oocyte retrieval in GnRH antagonist cycles. Hum Reprod 2005;20: 2453 – 2456. Kushnir VA, Barad DH, Gleicher N. Defining assisted reproductive technology success. Fertil Steril 2013a;100:e30. Kushnir VA, Vidali A, Barad DH, Gleicher N. The status of public reporting of clinical outcomes in assisted reproductive technology. Fertil Steril 2013b; 100:736 –741. Kyrou D, Kolibianakis EM, Venetis CA, Papanikolaou EG, Bontis J, Tarlatzis BC. How to improve the probability of pregnancy in poor responders undergoing in vitro fertilization: a systematic review and meta-analysis. Fertil Steril 2009; 91:749– 766. Kyrou D, Kolibianakis EM, Fatemi HM, Grimbizis GF, Theodoridis TD, Camus M, Tournaye H, Tarlatzis BC, Devroey P. Spontaneous triggering of ovulation versus HCG administration in patients undergoing IUI: a prospective randomized study. Reprod Biomed Online 2012;25:278– 283. Labarta E, Martinez-Conejero JA, Alama P, Horcajadas JA, Pellicer A, Simon C, Bosch E. Endometrial receptivity is affected in women with high circulating progesterone levels at the end of the follicular phase: a functional genomics analysis. Hum Reprod 2011;26:1813 –1825. Lakkakorpi JT, Pietila EM, Aatsinki JT, Rajaniemi HJ. Human chorionic gonadotrophin (CG)-induced down-regulation of the rat luteal LH/CG receptor results in part from the down-regulation of its synthesis, involving increased alternative processing of the primary transcript. J Mol Endocrinol 1993;10:153 –162. Lessey BA, Lebovic DI, Taylor RN. Eutopic endometrium in women with endometriosis: ground zero for the study of implantation defects. Semin Reproduct Med 2013;31:109–124. Levi Setti PE, Albani E, Cesana A, Novara PV, Zannoni E, Baggiani AM, Morenghi E, Arfuso V, Scaravelli G. Italian Constitutional Court modifications of a restrictive assisted reproduction technology law significantly improve pregnancy rate. Hum Reprod 2011;26:376 –381. Li R, Qiao J, Wang L, Li L, Zhen X, Liu P, Zheng X. MicroRNA array and microarray evaluation of endometrial receptivity in patients with high serum progesterone levels on the day of hCG administration. Reproduct Biol Endocrinol 2011;9:29. Licht P, Losch A, Dittrich R, Neuwinger J, Siebzehnrubl E, Wildt L. Novel insights into human endometrial paracrinology and embryo-maternal communication by intrauterine microdialysis. Hum Reprod Update 1998;4:532– 538. Licht P, Russu V, Lehmeyer S, Wildt L. Molecular aspects of direct LH/hCG effects on human endometrium—lessons from intrauterine microdialysis in the human female in vivo. Reprod Biol 2001a;1:10– 19. Licht P, Russu V, Wildt L. On the role of human chorionic gonadotropin (hCG) in the embryo-endometrial microenvironment: implications for differentiation and implantation. Semin Reprod Med 2001b;19:37– 47.
Downloaded from http://humupd.oxfordjournals.org/ at Memorial University of Newfoundland on July 3, 2014
Ferraretti AP, Goossens V, Kupka M, Bhattacharya S, de Mouzon J, Castilla JA, Erb K, Korsak V, Nyboe Andersen A. Assisted reproductive technology in Europe, 2009: results generated from European registers by ESHRE. Hum Reprod 2013; 28:2318– 2331. Garrido-Gomez T, Ruiz-Alonso M, Blesa D, Diaz-Gimeno P, Vilella F, Simon C. Profiling the gene signature of endometrial receptivity: clinical results. Fertil Steril 2013; 99:1078– 1085. Giorgetti C, Vanden Meerschaut F, De Roo C, Saunier O, Quarello E, Hairion D, Penaranda G, Chabert-Orsini V, De Sutter P. Multivariate analysis identifies the estradiol level at ovulation triggering as an independent predictor of the first trimester pregnancy-associated plasma protein-A level in IVF/ICSI pregnancies. Hum Reprod 2013;28:2636 –2642. Gleicher N, Kushnir VA, Barad DH. What ‘misguided campaign’ against single embryo transfer? Hum Reprod 2014;29:380 –381. Griesinger G, Dafopoulos K, Schultze-Mosgau A, Jelkmann W, von Otte S, Diesing D, Diedrich K. Vascular endothelial growth factor response to exogenous chorionic gonadotropic hormone in the luteal phase of women with a history of severe ovarian hyperstimulation syndrome. Arch Gynecol Obstet 2006;274:29– 33. Griesinger G, Kolibianakis EM, Papanikolaou EG, Diedrich K, Van Steirteghem A, Devroey P, Ejdrup Bredkjaer H, Humaidan P. Triggering of final oocyte maturation with gonadotropin-releasing hormone agonist or human chorionic gonadotropin. Live birth after frozen-thawed embryo replacement cycles. Fertil Steril 2007;88:616–621. Groenewoud ER, Cantineau AE, Kollen BJ, Macklon NS, Cohlen BJ. What is the optimal means of preparing the endometrium in frozen-thawed embryo transfer cycles? A systematic review and meta-analysis. Hum Reprod Update 2013;19:458– 470. Halliday JL, Ukoumunne OC, Baker HW, Breheny S, Jaques AM, Garrett C, Healy D, Amor D. Increased risk of blastogenesis birth defects, arising in the first 4 weeks of pregnancy, after assisted reproductive technologies. Hum Reprod 2010; 25:59 –65. Handyside AH. 24-chromosome copy number analysis: a comparison of available technologies. Fertil Steril 2013;100:595– 602. Haouzi D, Assou S, Mahmoud K, Tondeur S, Reme T, Hedon B, De Vos J, Hamamah S. Gene expression profile of human endometrial receptivity: comparison between natural and stimulated cycles for the same patients. Hum Reprod 2009;24: 1436 – 1445. Haouzi D, Assou S, Dechanet C, Anahory T, Dechaud H, De Vos J, Hamamah S. Controlled ovarian hyperstimulation for in vitro fertilization alters endometrial receptivity in humans: protocol effects. Biol Reprod 2010;82:679 –686. Harada M, Peegel H, Menon KM. Expression of vascular endothelial growth factor A during ligand-induced down-regulation of luteinizing hormone receptor in the ovary. Mol Cell Endocrinol 2010;328:28– 33. Healy DL, Breheny S, Halliday J, Jaques A, Rushford D, Garrett C, Talbot JM, Baker HW. Prevalence and risk factors for obstetric haemorrhage in 6730 singleton births after assisted reproductive technology in Victoria Australia. Hum Reprod 2010;25:265–274. Hildenbrand A, Lalitkumar L, Nielsen S, Gemzell-Danielsson K, Stavreus-Evers A. Expression of aquaporin 2 in human endometrium. Fertil Steril 2006;86:1452 –1458. Hope N, Rombauts L. Can an educational DVD improve the acceptability of elective single embryo transfer? A randomized controlled study. Fertil Steril 2010;94:489–495. Horcajadas JA, Riesewijk A, Polman J, van Os R, Pellicer A, Mosselman S, Simon C. Effect of controlled ovarian hyperstimulation in IVF on endometrial gene expression profiles. Mol Hum Reprod 2005;11:195–205. Horcajadas JA, Minguez P, Dopazo J, Esteban FJ, Dominguez F, Giudice LC, Pellicer A, Simon C. Controlled ovarian stimulation induces a functional genomic delay of the endometrium with potential clinical implications. J Clin Endocrinol Metab 2008; 93:4500– 4510. Humaidan P, Bredkjaer HE, Bungum L, Bungum M, Grondahl ML, Westergaard L, Andersen CY. GnRH agonist (buserelin) or hCG for ovulation induction in GnRH antagonist IVF/ICSI cycles: a prospective randomized study. Hum Reprod 2005; 20:1213– 1220. Humaidan P, Polyzos NP, Alsberg B, Erb K, Mikkelsen AL, Elbaek HO, Papanikolaou EG, Andersen CY. GnRHa trigger and individualized luteal phase hCG support according to ovarian response to stimulation: two prospective randomized controlled multi-centre studies in IVF patients. Hum Reprod 2013; 28:2511– 2521. Imudia AN, Awonuga AO, Doyle JO, Kaimal AJ, Wright DL, Toth TL, Styer AK. Peak serum estradiol level during controlled ovarian hyperstimulation is associated with increased risk of small for gestational age and preeclampsia in singleton pregnancies after in vitro fertilization. Fertil Steril 2012;97:1374– 1379.
Evans et al.
Scientific evidence supports frozen embryo transfer
Peegel H, Towns R, Nair A, Menon KM. A novel mechanism for the modulation of luteinizing hormone receptor mRNA expression in the rat ovary. Mol Cell Endocrinol 2005;233:65 –72. Pelkonen S, Koivunen R, Gissler M, Nuojua-Huttunen S, Suikkari AM, Hyden-Granskog C, Martikainen H, Tiitinen A, Hartikainen AL. Perinatal outcome of children born after frozen and fresh embryo transfer: the Finnish cohort study 1995 – 2006. Hum Reprod 2010;25:914– 923. Perrier d’Hauterive S, Charlet-Renard C, Berndt S, Dubois M, Munaut C, Goffin F, Hagelstein MT, Noel A, Hazout A, Foidart JM et al. Human chorionic gonadotropin and growth factors at the embryonic-endometrial interface control leukemia inhibitory factor (LIF) and interleukin 6 (IL-6) secretion by human endometrial epithelium. Hum Reprod 2004;19:2633 –2643. Pinborg A, Wennerholm UB, Romundstad LB, Loft A, Aittomaki K, SoderstromAnttila V, Nygren KG, Hazekamp J, Bergh C. Why do singletons conceived after assisted reproduction technology have adverse perinatal outcome? Systematic review and meta-analysis. Hum Reprod Update 2010;19:87– 104. Rebois RV, Fishman PH. Down-regulation of gonadotropin receptors in a murine Leydig tumor cell line. J Biol Chem 1984;259:3096– 3101. Roque M, Lattes K, Serra S, Sola I, Geber S, Carreras R, Checa MA. Fresh embryo transfer versus frozen embryo transfer in in vitro fertilization cycles: a systematic review and meta-analysis. Fertil Steril 2013;99:156– 162. Roy TK, Bradley CK, Bowman MC, McArthur SJ. Single-embryo transfer of vitrified-warmed blastocysts yields equivalent live-birth rates and improved neonatal outcomes compared with fresh transfers. Fertil Steril 2014;101:1294–1301. Rubio C, Rodrigo L, Mir P, Mateu E, Peinado V, Milan M, Al-Asmar N, Campos-Galindo I, Garcia S, Simon C. Use of array comparative genomic hybridization (array-CGH) for embryo assessment: clinical results. Fertil Steril 2013;99:1044 –1048. Ruiz-Alonso M, Blesa D, Diaz-Gimeno P, Go´mez E, Ferna´ndez-Sa´nchez M, Carranza F, Carrera J, Vilella F, Pellicer A, Simo´n C. The endometrial receptivity array for diagnosis and personalized embryo transfer as a treatment for patients with repeated implantation failure. Fertil Steril 2013;100:818 –824. Salamonsen LA, Nie G, Hannan NJ, Dimitriadis E. Society for Reproductive Biology Founders’ Lecture 2009. Preparing fertile soil: the importance of endometrial receptivity. Reprod Fertil Dev 2009;21:923– 934. Sazonova A, Kallen K, Thurin-Kjellberg A, Wennerholm UB, Bergh C. Neonatal and maternal outcomes comparing women undergoing two in vitro fertilization (IVF) singleton pregnancies and women undergoing one IVF twin pregnancy. Fertil Steril 2012;99:731 –737. Schoolcraft WB, Katz-Jaffe MG. Comprehensive chromosome screening of trophectoderm with vitrification facilitates elective single-embryo transfer for infertile women with advanced maternal age. Fertil Steril 2013;100:615 –619. Scott KL, Hong KH, Scott RT Jr. Selecting the optimal time to perform biopsy for preimplantation genetic testing. Fertil Steril 2013a;100:608– 614. Scott RT Jr, Upham KM, Forman EJ, Zhao T, Treff NR. Cleavage-stage biopsy significantly impairs human embryonic implantation potential while blastocyst biopsy does not: a randomized and paired clinical trial. Fertil Steril 2013b;100:624–630. Seyhan A, Ata B, Polat M, Son WY, Yarali H, Dahan MH. Severe early ovarian hyperstimulation syndrome following GnRH agonist trigger with the addition of 1500 IU hCG. Hum Reprod 2013;28:2522 –2528. Shapiro BS, Daneshmand ST, Garner FC, Aguirre M, Hudson C, Thomas S. Evidence of impaired endometrial receptivity after ovarian stimulation for in vitro fertilization: a prospective randomized trial comparing fresh and frozenthawed embryo transfer in normal responders. Fertil Steril 2011a;96: 344 – 348. Shapiro BS, Daneshmand ST, Garner FC, Aguirre M, Hudson C, Thomas S. Evidence of impaired endometrial receptivity after ovarian stimulation for in vitro fertilization: a prospective randomized trial comparing fresh and frozen-thawed embryo transfers in high responders. Fertil Steril 2011b;96:516– 518. Sharpe RM. hCG-induced decrease in availability of rat testis receptors. Nature 1976; 264:644 –646. Sherwin JR, Sharkey AM, Cameo P, Mavrogianis PM, Catalano RD, Edassery S, Fazleabas AT. Identification of novel genes regulated by chorionic gonadotropin in baboon endometrium during the window of implantation. Endocrinology 2007; 148:618 –626. Stillman RJ, Richter KS, Jones HWJ. Refuting a misguided campaign against the goal of single-embryo transfer and singleton birth in assisted reproduction. Hum Reprod 2013;28:2599 –2607.
Downloaded from http://humupd.oxfordjournals.org/ at Memorial University of Newfoundland on July 3, 2014
Licht P, Fluhr H, Neuwinger J, Wallwiener D, Wildt L. Is human chorionic gonadotropin directly involved in the regulation of human implantation? Mol Cell Endocrinol 2007; 269:85 –92. Lindsay LA, Murphy CR. Ovarian hyperstimulation affects fluid transporters in the uterus: a potential mechanism in uterine receptivity. Reprod Fertil Develop 2013; doi:10.1071/RD12396. Liu Y, Lee KF, Ng EH, Yeung WS, Ho PC. Gene expression profiling of human peri-implantation endometria between natural and stimulated cycles. Fertil Steril 2008;90:2152 –2164. Lopata A, Hay DL. The potential of early human embryos to form blastocysts, hatch from their zona and secrete HCG in culture. Hum Reprod 1989;4:87 –94. Lopata A, Oliva K. Chorionic gonadotrophin secretion by human blastocysts. Hum Reprod 1993;8:932–938. Loutradi KE, Kolibianakis EM, Venetis CA, Papanikolaou EG, Pados G, Bontis I, Tarlatzis BC. Cryopreservation of human embryos by vitrification or slow freezing: a systematic review and meta-analysis. Fertil Steril 2008;90:186– 193. Macaldowie A, Wang YA, Chambers GM, Sullivan EA. Assisted Reproductive Technology in Australia and New Zealand 2011. Sydney: National Perinatal Epidemiology and Statistics Unit, the University of New South Wales, 2013. Macklon NS, van der Gaast MH, Hamilton A, Fauser BC, Giudice LC. The impact of ovarian stimulation with recombinant FSH in combination with GnRH antagonist on the endometrial transcriptome in the window of implantation. Reprod Sci 2008; 15:357– 365. Maheshwari A, Bhattacharya S. Elective frozen replacement cycles for all: ready for prime time? Hum Reprod 2013;28:6 –9. Maheshwari A, Pandey S, Shetty A, Hamilton M, Bhattacharya S. Obstetric and perinatal outcomes in singleton pregnancies resulting from the transfer of frozen thawed versus fresh embryos generated through in vitro fertilization treatment: a systematic review and meta-analysis. Fertil Steril 2012;98:368 –377. Martinez MC, Ruiz FJ, Garcia-Velasco JA. GnRH-agonist triggering to avoid ovarian hyperstimulation syndrome: a review of the evidence. Curr Drug Targets 2013; 14:843– 849. Martini S, Van Voorhis BJ, Stegmann BJ, Sparks AE, Shochet T, Zimmerman MB, Ryan GL. In vitro fertilization patients support a single blastocyst transfer policy. Fertil Steril 2011;96:993– 997. Mastenbroek S, Twisk M, van der Veen F, Repping S. Preimplantation genetic screening: a systematic review and meta-analysis of RCTs. Hum Reprod Update 2011; 17:454– 466. Meldrum DR. Pregnancies and deliveries per fresh cycle are no longer adequate indicators of in vitro fertilization program quality: how should registries adapt? Fertil Steril 2013;100:620– 621. Menon KM, Menon B. Structure, function and regulation of gonadotropin receptors—a perspective. Mol Cell Endocrinol 2012;356:88–97. Min JK, Breheny SA, MacLachlan V, Healy DL. What is the most relevant standard of success in assisted reproduction? The singleton, term gestation, live birth rate per cycle initiated: the BESST endpoint for assisted reproduction. Hum Reprod 2004;19:3 – 7. Murray MJ, Meyer WR, Zaino RJ, Lessey BA, Novotny DB, Ireland K, Zeng D, Fritz MA. A critical analysis of the accuracy, reproducibility, and clinical utility of histologic endometrial dating in fertile women. Fertil Steril 2004;81: 1333 – 1343. Navot D, Bergh P. Preparation of the human endometrium for implantation. Ann N Y Acad Sci 1991;622:212– 219. Nelson SM. Venous thrombosis during assisted reproduction: novel risk reduction strategies. Thromb Res 2013;131(Suppl 1):S1 –S3. Noyes RW, Hertig AT, Rock J. Dating the endometrial biopsy. Am J Obstet Gynecol 1975; 122:262 –263. Ormond KE. From genetic counseling to ‘genomic counseling’. Mol Genet Genomic Med 2013;1:189–193. Paiva P, Hannan NJ, Hincks C, Meehan KL, Pruysers E, Dimitriadis E, Salamonsen LA. Human chorionic gonadotrophin regulates FGF2 and other cytokines produced by human endometrial epithelial cells, providing a mechanism for enhancing endometrial receptivity. Hum Reprod 2011;26:1153 –1162. Papanikolaou EG, Verpoest W, Fatemi H, Tarlatzis B, Devroey P, Tournaye H. A novel method of luteal supplementation with recombinant luteinizing hormone when a gonadotropin-releasing hormone agonist is used instead of human chorionic gonadotropin for ovulation triggering: a randomized prospective proof of concept study. Fertil Steril 2011;95:1174 –1177.
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after frozen-thawed embryo transfer: a Nordic cohort study from the CoNARTaS group. Hum Reprod 2013;28:2545 –2553. Whelan JG III, Vlahos NF. The ovarian hyperstimulation syndrome. Fertil Steril 2000; 73:883– 896. Wilcox AJ, Baird DD, Weinberg CR. Time of implantation of the conceptus and loss of pregnancy. N Engl J Med 1999;340:1796 – 1799. Winand R, Hens K, Dondorp W, de Wert G, Moreau Y, Vermeesch JR, Liebaers I, Aerts J. In vitro screening of embryos by whole-genome sequencing: now, in the future or never? Hum Reprod 2014;29:842 –851. Xiao Z, Zhou X, Xu W, Yang J, Xie Q. Natural cycle is superior to hormone replacement therapy cycle for vitrificated-preserved frozen-thawed embryo transfer. Syst Biol Reprod Med 2012;58:107– 112. Yang Z, Liu J, Collins GS, Salem SA, Liu X, Lyle SS, Peck AC, Sills ES, Salem RD. Selection of single blastocysts for fresh transfer via standard morphology assessment alone and with array CGH for good prognosis IVF patients: results from a randomized pilot study. Mol Cytogenet 2012;5:24. Yen SS, Llerena O, Little B, Pearson OH. Disappearance rates of endogenous luteinizing hormone and chorionic gonadotropin in man. J Clin Endocrinol Metab 1968;28:1763 –1767. Yoshii N, Hamatani T, Inagaki N, Hosaka T, Inoue O, Yamada M, Machiya R, Yoshimura Y, Odawara Y. Successful implantation after reducing matrix metalloproteinase activity in the uterine cavity. Reprod Biol Endocrinol 2013;11:37. Youssef MA, Van der Veen F, Al-Inany HG, Griesinger G, Mochtar MH, Aboulfoutouh I, Khattab SM, van Wely M. Gonadotropin-releasing hormone agonist versus HCG for oocyte triggering in antagonist assisted reproductive technology cycles. Cochrane Database Syst Rev 2011;13:CD003719. Zhang XJ, Yang YZ, Lv Q, Wang Y, Cao XH, Li XJ, Liao MX, Guo C. The impact of two different thaw protocols on outcomes of vitrified cleavage-stage embryos transfer. Cryo Lett 2012;33:411– 417.
Downloaded from http://humupd.oxfordjournals.org/ at Memorial University of Newfoundland on July 3, 2014
Thomas K, Thomson AJ, Sephton V, Cowan C, Wood S, Vince G, Kingsland CR, Lewis-Jones DI. The effect of gonadotrophic stimulation on integrin expression in the endometrium. Hum Reprod 2002;17:63– 68. Thomas K, Thomson A, Wood S, Kingsland C, Vince G, Lewis-Jones I. Endometrial integrin expression in women undergoing in vitro fertilization and the association with subsequent treatment outcome. Fertil Steril 2003a;80:502 –507. Thomas K, Thomson AJ, Wood SJ, Kingsland CR, Vince G, Lewis-Jones DI. Endometrial integrin expression in women undergoing IVF and ICSI: a comparison of the two groups and fertile controls. Hum Reprod 2003b;18:364– 369. Tulac S, Overgaard MT, Hamilton AE, Jumbe NL, Suchanek E, Giudice LC. Dickkopf-1, an inhibitor of Wnt signaling, is regulated by progesterone in human endometrial stromal cells. J Clin Endocrinol Metab 2006;91:1453 – 1461. Ubaldi F, Bourgain C, Tournaye H, Smitz J, Van Steirteghem A, Devroey P. Endometrial evaluation by aspiration biopsy on the day of oocyte retrieval in the embryo transfer cycles in patients with serum progesterone rise during the follicular phase. Fertil Steril 1997;67:521 –526. Van Vaerenbergh I, Van Lommel L, Ghislain V, In’t Veld P, Schuit F, Fatemi HM, Devroey P, Bourgain C. In GnRH antagonist/rec-FSH stimulated cycles, advanced endometrial maturation on the day of oocyte retrieval correlates with altered gene expression. Hum Reprod 2009;24:1085– 1091. Venetis CA, Kolibianakis EM, Bosdou JK, Tarlatzis BC. Progesterone elevation and probability of pregnancy after IVF: a systematic review and meta-analysis of over 60 000 cycles. Hum Reprod Update 2013;19:433 –457. Venn A, Hemminki E, Watson L, Bruinsma F, Healy D. Mortality in a cohort of IVF patients. Hum Reprod 2001;16:2691 – 2696. Vilella F, Ramirez LB, Simon C. Lipidomics as an emerging tool to predict endometrial receptivity. Fertil Steril 2013;99:1100– 1106. Wennerholm UB, Henningsen AK, Romundstad LB, Bergh C, Pinborg A, Skjaerven R, Forman J, Gissler M, Nygren KG, Tiitinen A. Perinatal outcomes of children born
Evans et al.