High Implantation Potential Research Articles

Selection of embryos for transfer in IVF: ranking embryos based on their implantation potential using morphological scoring

The selection of embryos based on morphology is still the core of daily laboratory practice in IVF/intracytoplasmic sperm injection. At present, the selection of embryos is primarily based on experience and local protocols. Since an evidence-based ranking strategy for embryos on day 3 is currently lacking, this work constructed a multivariable prediction model to rank embryos according to their implantation potential. A total of 6021 fresh embryo transfers between January 2004 and July 2009 were included, eight potential predictive factors were evaluated and a prediction model was developed using multivariable logistic regression. The model was externally validated with data from couples treated between August 2009 and September 2011 in the same clinic. Five factors were included in the final prediction model: early cleavage, number of blastomeres on days 2 and 3 and morphological score and presence of morula on day 3. With validation, the model showed moderate discriminative capacity (c-statistic 0.70) and calibrated well and was able to distinguish embryos with high ongoing implantation potential from embryos with moderate or low ongoing implantation potential. The model can be used by embryologists as an objective tool to rank embryos according to implantation potential, thereby aiding the selection of embryos for transfer.

Limitations of Embryo Selection Methods

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

Diagnóstico genético preimplantacional: alcances y límites

El diagnóstico genético preimplantacional (DGP) es el estudio de alteraciones cromosómicas y genéticas en el embrión, antes de su transferencia a la madre. Permite conocer aquellos embriones libres de anomalías cromosómicas o mutaciones genéticas. En esta revisión se describe la experiencia de un centro de reproducción asistida en Perú, realizando diagnóstico genético preimplantacional a embriones en día 3 de desarrollo. Se muestra resultados de cinco años, con 711 ciclos. El diagnóstico genético preimplantacional sería una herramienta eficaz para seleccionar los embriones con mayor potencial de implantación, que puedan dar origen a un nacido vivo sano.

Is it acceptable to destroy or include human embryos before day 5 in research programmes?

Day-3 poor-quality embryos (PQE) from IVF–embryo transfer cycles are usually destroyed or are included in research programmes. Knowing that these embryos have the ability to evolve to the blastocyst stage and yield embryonic stem cell lines, this study postulated that they could also give rise to live births. This is a prospective study including 186 IVF–embryo transfer candidates who had obtained at least one supernumerary PQE on day 3. PQE were kept for extended culture and high-quality blastocysts were frozen. A total of 620 PQE were eligible for the study, 217 (35.0%) reached the blastocyst stage and 73 (33.6%) were frozen. Blastulation rates were 7-fold higher (OR 7.29, 95% CI 5.01–10.61) in embryos compacted on day 4. Of the frozen blastocysts, 40 were thawed during 33 thawed blastocyst transfer cycles, which led to 10 clinical pregnancies. These pregnancies resulted in five miscarriages and five healthy live births at full term. PQE may achieve their development to the blastocyst stage, be frozen–thawed and harbour reasonable implantation potential. These results, thereby, raise an ethical issue regarding the fate reserved to PQE.In most IVF clinics around the world, day-3 embryos are submitted to embryo classification systems to select embryos with the higher implantation potential. This classification is usually based on the evaluation of the number of blastomeres, the degree of fragmentation, the symmetry of the cells and the presence of multinucleation. Thus, some embryos are described as poor-quality embryos (PQE) and may be destroyed or included in research programmes. Knowing that PQE have the ability to evolve to the blastocyst stage and yield embryonic stem cell lines, the authors hypothesized that PQE could also give rise to live births. They conducted a prospective study including 186 IVF–embryo transfer candidates who had obtained at least one supernumerary PQE on day 3. PQE were kept for extended culture and high quality blastocysts were frozen. Out of 620 PQE eligible for this study, 217 (35.0%) reached the blastocyst stage and 73 (33.6%) were frozen. Of these blastocysts, 40 were thawed during 33 thawed blastocyst transfer cycles, which led to 10 clinical pregnancies. These pregnancies resulted in five miscarriages and five healthy live births at full term. PQE may achieve their development to the blastocyst stage, be frozen–thawed and harbour a reasonable chance of implantation. These results, thereby, raise a ethical issue regarding the fate of PQE.

No evidence that embryo selection by near-infrared spectroscopy in addition to morphology is able to improve live birth rates: results from an individual patient data meta-analysis

What is the value of embryo selection by metabolomic profiling of culture medium with near-infrared (NIR) spectroscopy as an adjunct to morphology, compared with embryo selection by morphology alone, based on an individual patient data meta-analysis (IPD MA)? The IPD MA indicates that the live birth rate after embryo selection by NIR spectroscopy and morphology is not significantly different compared with the live birth rate after embryo selection by morphology alone. Retrospective proof of principle studies has consistently shown that high NIR viability scores are correlated with a high implantation potential of embryos. However, randomized controlled trials (RCTs) have generally shown no benefit of the NIR technology over embryo morphology, although there have been some conflicting results between pregnancy outcomes on different days of embryo transfer. This IPD MA included all existing RCTs (n = 4) in which embryo selection by morphology was compared with embryo selection by morphology and the use of NIR spectroscopy of spent embryo culture medium by the Viametrics-E(™). Searches of PubMed, the Cochrane Library and the WHO International Clinical Trials Registry were conducted and the sole manufacturer of the Viametrics-E(™) was consulted to identify clinics where an RCT comparing embryo selection by morphology to embryo selection by morphology and the use of the Viametrics-E(™) (NIR viability score) was performed. A total of 20 citations were potentially eligible for inclusion, two of which met the inclusion criteria. The manufacturer of the Viametrics-E(™) provided two additional clinical sites of use. In total, four RCTs were identified as eligible for inclusion. The IPD MA was based on a fixed effect model due to the lack of heterogeneity between included studies. Differences between study groups were tested and reported using logistic regression models adjusted for significant confounders. The pooled analysis of the primary outcome led to a total sample size of 924 patients: 484 patients in the control group (embryo selection by morphology alone) and 440 patients in the treatment group (embryo selection by morphology plus NIR spectroscopy). The live birth rates in the control group and the NIR group were 34.7% (168 of 484) and 33.2% (146 of 440), respectively. The pooled odds ratio (OR) was 0.98 [95% confidence interval (CI) 0.74-1.29], indicating no difference in live birth rates between the two study groups. The data of the four studies showed no significant heterogeneity (I(2) = 26.2% P = 0.26). The multivariate regression analysis including all confounders show that maternal age (OR 0.90, 95% CI 0.87-0.94) and the number of previous IVF cycles (OR 0.83, 95% CI 0.71-0.96) were significantly related to live birth. The study group (i.e. embryo selection by morphology or embryo selection by morphology plus NIR) was not related to live birth (OR 0.97, 95% CI 0.73-1.29). The availability of at least two similar best quality embryos as an inclusion criterion prior to transfer in the two largest RCTs might have caused a selection bias towards a better prognosis patient group. There is at present no evidence that NIR spectroscopy of spent embryo culture media in its current form can be used in daily practice to improve live birth rates.

Embryo selection: the role of time-lapse monitoring.

In vitro fertilization has been available for over 3 decades. Its use is becoming more widespread worldwide, and in the developed world, up to 5% of children have been born following IVF. It is estimated that over 5 million children have been conceived in vitro. In addition to giving hope to infertile couples to have their own family, in vitro fertilization has also introduced risks as well. The risk of multiple gestation and the associated maternal and neonatal morbidity/mortality has increased significantly over the past few decades. While stricter transfer policies have eliminated the majority of the high-order multiples, these changes have not yet had much of an impact on the incidence of twins. A twin pregnancy can be avoided by the transfer of a single embryo only. However, the traditionally used method of morphologic embryo selection is not predictive enough to allow routine single embryo transfer; therefore, new screening tools are needed. Time-lapse embryo monitoring allows continuous, non-invasive embryo observation without the need to remove the embryo from optimal culturing conditions. The extra information on the cleavage pattern, morphologic changes and embryo development dynamics could help us identify embryos with a higher implantation potential. These technologic improvements enable us to objectively select the embryo(s) for transfer based on certain algorithms. In the past 5-6 years, numerous studies have been published that confirmed the safety of time-lapse technology. In addition, various markers have already been identified that are associated with the minimal likelihood of implantation and others that are predictive of blastocyst development, implantation potential, genetic health and pregnancy. Various groups have proposed different algorithms for embryo selection based on mostly retrospective data analysis. However, large prospective trials are needed to study the full benefit of these (and potentially new) algorithms before their introduction into daily practice can be recommended.

The combination of embryo selection with high implantation potential and the best moment in term of endometrial receptivity for embryo replacement improves pregnancy outcomes

The aim of this study was to optimize pregnancy outcome using personalized care management by combining the embryo competence and endometrial receptivity assessments for infertile couples refereed for IVF/ICSI under controlled ovarian stimulation (COS).

Time-lapse embryo selection for single blastocyst transfer – results of a multicenter, prospective, randomized clinical trial

A multiple pregnancy is an undesired outcome of assisted reproduction. Current selection methods are inefficient at identifying the embryo with the highest implantation potential. Time lapse (TL) embryo monitoring provides additional information about embryo development and therefore may aid embryo selection. Our aim is to study whether TL monitoring combined with morphological scoring is superior to traditional embryo observation when a single blastocyst (BC) is selected for transfer (ET).

Multiple Pregnancies After ART and How to Minimize their Occurrence

In the past few decades, the number of multiple pregnancies has increased significantly. Advanced maternal age, use of ovarian stimulation and use of IVF are all responsible for this. A multiple pregnancy carries excess maternal and neonatal risks and is associated with undesired extra health care expenses as well. Up to 1/3 of the pregnancies following IVF are multiple gestations; most being twins. The best way to minimize the risk of twins is to transfer a single embryo only. The majority of the embryos do not implant however and most clinics routinely transfer 2 or even more embryos to maintain high IVF success rates (pregnancy rates). A high success rate despite the transfer of fewer, preferably a single embryo could be maintained if we had better tools to identify the embryo with the highest implantation potential. Current selection criteria based on the cleavage rate and morphology of the embryo are not particularly effective for this. The past decade has seen a lot of research aiming to screen the embryos based on various properties. This review will discuss the perinatal outcome of IVF multiples, available screening tools for embryo selection and the current status of the development of new tools.

Correlation between embryo morphology and development and chromosomal complement

To analyze the correlation between embryo morphology and the chromosomal status using the comparative genomic hybridization (CGH) technique for screening 46 chromosomes in a single blastomere biopsy from Day 3 embryos. Retrospective study in an in vitro fertilization laboratory. Four hundred fifty two embryos were included from 63 cycle ICSI patients undergoing preimplantation genetic screening. The 46 chromosomes were analyzed by blastomere biopsy from day 3 embryos using CGH array method. Data was analyzed by Chi-Square analysis for embryo development rate, fragmentation rate and chromosome status of the analyzed blastomeres. From 452 embryos biopsied, 26 samples were not able to analyze. A total of 263/426 (61.7%) embryos tested on day 3 were abnormal. 175 blastocysts developed from biopsied embryos. 51/175 (29.1%) were analyzed as abnormal vs 124/175 (70.9%) from normal a-CGH results. The incidence of chromosomal abnormalities was significantly lower in a day 3 embryo with more than 6 cells compared with embryos with 6 cells or less (59.3% versus 74.3%, respectively). The numerical analysis of chromosomes was related to the percentage of fragmentation. In the total of 426 embryos tested, the total incidence of chromosomal abnormalities was 23.6% with 0 to 15% fragmentation and 66.1% with 15% or more fragmentation. Embryo development rate and morphological parameter such as degree of fragmentation to a large extent reflect the cytogenetic status of the embryo and thus are important in the selection of embryos with the highest implantation potential.

Timing of cell division in human cleavage-stage embryos is linked with blastocyst formation and quality

Noninvasive markers of embryo quality are being sought to improve IVF success. The present study aimed to discover possible associations between embryo division kinetics in the cleavage stage, the subsequent ability of human embryos to reach the blastocyst stage and the resulting blastocyst morphology. A retrospective cohort study analysed 834 embryos from 165 oocyte donation couples using a time-lapse monitoring system that allowed the recording of the exact timings for key events related to embryo development. Timing parameters were categorized into four quartiles. The probability of an embryo developing to a blastocyst was linked to a strict chronology of development. To further evaluate the relationships between these morphokinetic parameters and subsequent blastocyst formation, the ensuing blastocyst morphology was compared with a viability score based on a hierarchical classification of the cleavage-stage morphokinetic parameters. It is concluded that the kinetics of early embryo development and the potential for human embryos to develop to the blastocyst stage on day 5 are closely related and that time-lapse-based evaluation of the exact timing of early events in embryo development is a promising tool for the prediction of blastocyst formation and quality according to the proposed model.Assisted reproduction treatment has helped several thousand infertile couples to start a family. Even though treatment is presently very successful, there is still scope for improvement in the process of selecting the embryos for transfer that have the highest implantation potential. An improved selection procedure may be able to increase the pregnancy rates, or alternatively, to reduce the number of embryos transferred while maintaining high pregnancy rates.We report the results after using an incubator with time-lapse video recording and image analysis to determine the timing of key events during embryo development. The events were assessed as potential markers of embryo morphology and development. One of the major factors that has already helped to increase success rates is extended embryo culture and blastocyst transfer on day 5 of development, because at this stage the embryo has stronger developmental competence. Also embryo/uterine synchrony is better achieved by a blastocyst-stage than a cleavage-stage embryo. However, extended culture in vitro is time consuming and extended exposure to artificial culture conditions in vitro may stress the embryo and could have negative side effects. For that reason, we sought evidence in the data obtained from the time-lapse monitoring system for a link between timing of early embryo cell divisions and the probability of subsequent development of embryos into blastocysts. The purpose of this was to avoid extended culture if blastocyst development could instead be reliably predicted from time-lapse images of the early cleavage states.

D’autres acteurs dans la croissance folliculaire et ovocytaire : rôle des microARNs dans le dialogue cumulo-ovocytaire

The good folliculogenesis evolution is fundamental for the obtaining of a competent oocyte, able to lead to pregnancy, once fertilized. During the follicular development, the oocyte is in close contact with surrounding cumulus cells (CCs) to form a cumulus-oocyte complex. The bidirectional exchange between oocyte and contiguous CCs via gap junction communications and paracrine signaling is important for oocyte competence and CCs development. These reciprocal regulations are controlled by some key genes. Recently, it has been demonstrated that these genes are themselves regulated by short RNAs fragments (approximately 22 nucleotides), called microRNAs. The identification and the quantification in the CCs of the microRNAs regulating these genes could promote the development of non invasive tests in order to assess the oocyte quality and its ability to provide embryo with a high implantation potential. This approach could be decisive in the embryo selection to transfer and could avoid the risk of multiple pregnancies by the replacement of a single embryo.

The type of protocol used for art induces significant differences in embryo developmental kinetics – A time-lapse study

ObjectiveTo explore whether differences in embryo kinetics exist between protocols employing GnRH agonist (GnRH-a) down-regulation and hCG for triggering versus GnRH antagonist (GnRH- ant) and GnRH agonist for triggering.DesignRetrospective cohort study.Materials and MethodsWe studied 748 embryos from oocyte donation. Donors were stimulated with recombinant FSH and down-regulated with either GnRHa or GnRH-ant. Triggering was performed with hCG in the GnRHa group and with GnRH-a in the GnRH-ant one. Through a time-lapse system (Embryoscope™ Unisense Fertilitech, Aarhus, Denmark), we obtained a monitoring of the embryos. Timing values of cleavage to 2-cell embryos (T2), to 3-cell embryos (T3), to 4-cells embryos (T4), time of compaction (M), appearance of a blastocoel cavity (B), length of the second cell cycle (CC2) and the synchrony from 2-cell to 4-cell embryos (S2) were recorded. The proportion of optimal embryos was determined from our previous observations, identifying embryos in a best time range with a higher implantation potential. We applied a T-test with P<0.05 statistically significant.ResultsTable I.GnRH-a/hCG (n = 44)CI 95%GnRH-ant/GnRH-a (n = 66)CI 95%pT2 (h)27.026.5-27.527.527.1-27.8NST3 (h)38.838.2-39.440.039.5-40.5<0.05T4 (h)40.039.2-40.841.040.0-42.0NSS2 (h)1.30.8-1.81.91.0-2.8NSCC2 (h)11.911.4-12.312.612.3-13.0<0.05M (h)89.086.6-91.491.590.3-92.7NSB (h)106.0104.0-108.0103.4102.2-104.6NS%optimal embryos%optimal embryosT2 (h) 24.6-28.24438.3-49.747.542.9-52.1NST3 (h) 35.2-40.543.738.0-49.442.638.0-47.2NSS2 (h) <0.7584.480.0-88.885.079.1-90.9NSCC2 (h) <1237.832.2-43.430.626.3-34.9NS Open table in a new tab ConclusionThese new data deriving from a time-lapse system, suggest that the type of protocol used for ovarian stimulation seems to impact embryonic kinetics. ObjectiveTo explore whether differences in embryo kinetics exist between protocols employing GnRH agonist (GnRH-a) down-regulation and hCG for triggering versus GnRH antagonist (GnRH- ant) and GnRH agonist for triggering. To explore whether differences in embryo kinetics exist between protocols employing GnRH agonist (GnRH-a) down-regulation and hCG for triggering versus GnRH antagonist (GnRH- ant) and GnRH agonist for triggering. DesignRetrospective cohort study. Retrospective cohort study. Materials and MethodsWe studied 748 embryos from oocyte donation. Donors were stimulated with recombinant FSH and down-regulated with either GnRHa or GnRH-ant. Triggering was performed with hCG in the GnRHa group and with GnRH-a in the GnRH-ant one. Through a time-lapse system (Embryoscope™ Unisense Fertilitech, Aarhus, Denmark), we obtained a monitoring of the embryos. Timing values of cleavage to 2-cell embryos (T2), to 3-cell embryos (T3), to 4-cells embryos (T4), time of compaction (M), appearance of a blastocoel cavity (B), length of the second cell cycle (CC2) and the synchrony from 2-cell to 4-cell embryos (S2) were recorded. The proportion of optimal embryos was determined from our previous observations, identifying embryos in a best time range with a higher implantation potential. We applied a T-test with P<0.05 statistically significant. We studied 748 embryos from oocyte donation. Donors were stimulated with recombinant FSH and down-regulated with either GnRHa or GnRH-ant. Triggering was performed with hCG in the GnRHa group and with GnRH-a in the GnRH-ant one. Through a time-lapse system (Embryoscope™ Unisense Fertilitech, Aarhus, Denmark), we obtained a monitoring of the embryos. Timing values of cleavage to 2-cell embryos (T2), to 3-cell embryos (T3), to 4-cells embryos (T4), time of compaction (M), appearance of a blastocoel cavity (B), length of the second cell cycle (CC2) and the synchrony from 2-cell to 4-cell embryos (S2) were recorded. The proportion of optimal embryos was determined from our previous observations, identifying embryos in a best time range with a higher implantation potential. We applied a T-test with P<0.05 statistically significant. ResultsTable I.GnRH-a/hCG (n = 44)CI 95%GnRH-ant/GnRH-a (n = 66)CI 95%pT2 (h)27.026.5-27.527.527.1-27.8NST3 (h)38.838.2-39.440.039.5-40.5<0.05T4 (h)40.039.2-40.841.040.0-42.0NSS2 (h)1.30.8-1.81.91.0-2.8NSCC2 (h)11.911.4-12.312.612.3-13.0<0.05M (h)89.086.6-91.491.590.3-92.7NSB (h)106.0104.0-108.0103.4102.2-104.6NS%optimal embryos%optimal embryosT2 (h) 24.6-28.24438.3-49.747.542.9-52.1NST3 (h) 35.2-40.543.738.0-49.442.638.0-47.2NSS2 (h) <0.7584.480.0-88.885.079.1-90.9NSCC2 (h) <1237.832.2-43.430.626.3-34.9NS Open table in a new tab ConclusionThese new data deriving from a time-lapse system, suggest that the type of protocol used for ovarian stimulation seems to impact embryonic kinetics. These new data deriving from a time-lapse system, suggest that the type of protocol used for ovarian stimulation seems to impact embryonic kinetics.

The type of gonadotropins used for controlled ovarian stimulation affects embryo developmental kinetics

We aimed to explore whether the type of gonadotropin used for COS has any influence on embryo kinetics, using a time-lapse system. Retrospective cohort study. We examined 751 embryos from our oocyte donation program. Donors were stimulated with either HMG or rFSH using a GnRH agonist long protocol and triggering with hCG. For monitoring the embryos an incubator with a built-in camera (Embryoscope™, Unisense Fertilitech, Aarhus, Denmark) was used. We determined timing values of cleavage to 2-cell embryos (T2), to 3-cell embryos (T3), to 4-cells embryos (T4), time of morulae (M) and the appearance of blastocoel cavity (B), length of the second cell cycle (CC2) and the synchrony from 2-cell to 4-cell embryos (S2). “Optimal embryos”, pointing out those embryos included in a real interval time to be of high quality and with a high implantation potential. A T-test with P<0.05 was considered to be statistically significant. Embryos obtained after stimulation with rFSH divided significantly earlier (T2, T3) as compared to those after HMG stimulation. From T3 and onwards these kinetic differences disappeared. We noticed a significant increase in embryo quality in cycles stimulated with rFSH.Tabled 1Embryo development kinetics Vs gonadotropinsrFSH (n = 43)CI 95%HMG (n = 58)CI 95%pT2 (h)27.026.5-27.528.327.9-28.7<0.05T3 (h)38.838.1-39.540.139.6-40.6<0.05T4 (h)40.039.2-40.841.240.5-41.9NSS2 (h)1.30.8-1.81.81.3-2.3NSCC2 (h)11.811.4-12.212.011.5-12.5NSM(h)88.286.1-90.391.689.8-93.4NSB (h)103.8101.8-105.8103.3101.8-104.8NS%optimal embryos%optimal embryosT2 (h) 24.6-28.244.037.9-50.141.336.8-45.8NST3 (h) 35.2-40.543.738.0-49.433.028.6-37.7<0.05S2 (h) <0.7584.480.0-88.882.778.6-86.8NSSS2 (h) <1237.832.2-43.430.025.7-34.3NS Open table in a new tab We observed differences in T2 and T3 between preparations with or without LH-like activity. Gonadotropin used for stimulation might have an impact downstream on embryo development and implantation potential.

Embryo selection in IVF

To optimize success rates of IVF, selection of the most viable embryo(s) for transfer has always been essential, as embryos that are cryopreserved are thought to have a reduced chance of implanting after thawing. Recent developments challenge this concept. Evidence is accumulating that all embryos can now be cryopreserved and transferred in subsequent cycles without impairing pregnancy rates or maybe even with an improvement in pregnancy rates. In such a scenario no selection method will ever lead to improved live birth rates, as, by definition, the live birth rate per stimulated IVF cycle can never be improved when all embryos are serially transferred. In fact, selection could then only lower the live birth rate after IVF. The only parameter that could possibly be improved by embryo selection would be time to pregnancy, if embryos with the highest implantation potential are transferred first.

Cytogenetic analysis of human blastocysts with the use of FISH, CGH and aCGH: scientific data and technical evaluation

Recent studies have suggested that biopsy of several trophectoderm (TE) cells from blastocysts followed by comparative genomic hybridization (CGH) analysis might represent an optimal strategy for aneuploidy detection, but few data on accuracy are available. The main question concerns the rate of mosaicism at the blastocyst stage, and to what extent this might cause misdiagnoses. We assessed blastocyst aneuploidy and mosaicism rates and evaluated the accuracy and efficiency of CGH and microarray-CGH (aCGH) for TE analysis. A total of 52 blastocysts, from 20 couples, were biopsied and their chromosomes examined by CGH. The remaining cells were spread and tested by fluorescent in situ hybridization (FISH). Of the 52 blastocysts, 20 underwent a second TE biopsy and were tested using aCGH. CGH and aCGH produced results for 98% of TE samples. 42.3% of blastocysts were uniformly euploid, 30% were uniformly aneuploid and 32.4% were mosaic. Of the mosaic embryos, 15.4% were found to be composed of a mixture of different aneuploid cell lines, while 17% contained both normal and aneuploid cells. Mosaic diploid-aneuploid blastocysts with >30% normal cells accounted for <6% of analysed embryos. Comprehensive chromosome screening and follow-up assessment of large numbers of cells provided a unique insight into the cytogenetics of human blastocysts. Meiotic and post-zygotic errors leading to mosaicism were common. However, most mosaic blastocysts contained no normal cells. Hence, CGH or aCGH TE analysis is an accurate aneuploidy detection tool and may assist in identifying viable euploid embryos with higher implantation potential.

The types of chromosome abnormality in blastocysts generated by infertile patients

OBJECTIVE: While much is known about cleavage stage embryo aneuploidy, the chromosomal status of blastocysts has been little investigated. Blastocysts can activate their own genome, undergo the first cellular differentiation into trophectoderm (TE) and inner cell mass, and are usually considered to have high implantation potential. However, growth to the blastocyst stage does not eliminate aneuploid embryos. This study aimed to catalogue all chromosome errors seen in blastocysts generated by IVF couples for the following indications, recurrent implantation failure (RIF), recurrent pregnancy loss (RPL), or advanced maternal age (AMA). DESIGN: 845 blastocysts from 127 couples were examined (mean maternal age: 39 years). MATERIALS AND METHODS: Combination of TE biopsy and comparative genomic hybridization (CGH) was used. RESULTS: The total aneuploidy rate was 58%. The rates per indication were 54% for RIF, 56% for RPL, and 61% for AMA. In total, 784 anomalies were scored and all chromosomes were affected. The distribution of monosomies and trisomies showed significant variation between the groups (p<0.01). Trisomies were more frequent for RPL and RIF, whilst monosomies dominated in AMA. A strong relationship between maternal age and aneuploidy was observed. Women over 34 years of age had twice the aneuploidy rate as younger patients (p<0.01). CONCLUSION: The rate of aneuploidy in blastocysts was significantly lower than the published rates at the cleavage stage. Hence, the final stage of preimplantation development is not an absolute selective barrier to most forms of error. Differences in the types of abnormality seen for RIF, RPL and AMA groups indicate that different mechanisms may lead to aneuploidy in these groups of patients. The high aneuploidy rates detected suggest that chromosome screening at the blastocyst stage may be beneficial and may prove particularly effective for increasing the success rates following elective single embryo transfer.

Ectopic pregnancy after assisted reproductive technology: what are the risk factors?

This review discusses recent publications that investigate risk factors associated with ectopic pregnancy after IVF. Data on the risk factors for developing ectopic pregnancy after IVF are still inconsistent. Between fresh nondonor IVF and embryo transfer cycles, the significant risk factor for ectopic pregnancy was tubal factor infertility, and endometriosis, rather than male factor infertility. Higher ectopic pregnancy rate could be associated with zygote intrafallopian transfer, assisted hatching, large embryo transfer volume, deep fundal transfer, and frozen embryo transfer. The supraphysiologic progesterone level may decrease uterine contractility and enhance implantation in the uterine cavity in fresh embryo transfer compared with frozen embryo transfer cycles. Although recent results suggest reassurance in risk of ectopic pregnancy with frozen transfer, clinicians should be remembering this possibility while performing a frozen embryo transfer. Higher implantation potential per embryo at the blastocyst stage may increase the risk of ectopic pregnancy than cleavage stage. Especially, according to numbers of embryos transferred, different risk of ectopic pregnancy after IVF was noted. Different hormonal milieu, the reproductive health characteristics of infertile women such as distorted tubal function, technical issues of IVF procedures, and the estimated embryo implantation potential are possible risk factors. How each factor contributes to the risk of occurring ectopic pregnancy after assisted reproductive technology is uncertain and needs further investigation.

Human cumulus cells as biomarkers for embryo and pregnancy outcomes

Single-embryo transfer is becoming increasingly common in in vitro fertilization (IVF) treatment as a means of reducing multiple pregnancy rates leading to a higher incidence of medical, perinatal and neonatal complications. Consequently, selecting embryos with the highest implantation potential is of great importance in assisted reproductive technology. To date, the choice of the best embryos to transfer is based on subjective morphological parameters. However, as judged by their subjective aspect, movement towards more sophisticated technologies to select the most competent oocytes and/or embryos with the greatest implantation potential have become available, including emerging 'omics' sciences, such as genomics, transcriptomics, proteomics and metabolomics. In this way, the study of the cumulus cells (CCs) transcriptomic profile offers the opportunity, by a non-invasive method, to predict oocyte and embryo competence because bidirectional traffic between CCs and the oocyte is very important for the acquisition of this competence. Using either RT-PCR or DNA microarrays, some studies have provided evidence for the genes expressed in CCs presenting potential biomarkers to predict embryo quality and pregnancy outcomes. This review provides an overview of the current knowledge about CCs as biomarkers for oocyte and embryo selection under an IVF program.

A cycle-based model to predict blastocyst transfer cancellation

Extended culture for blastocyst transfer is thought to result in embryos with high implantation potential, enabling the transfer of fewer embryos and the reduction of multiple pregnancies after in vitro fertilization (IVF). However, one major drawback of extended culture is the risk of transfer cancellation if no blastocyst develops by Day 5, despite the observation of adequate early embryo development. We set out to develop a model to predict blastocyst transfer cancellation. The model was built from 562 consecutive first IVF cycles and tested on an independent validation cohort. Multivariable logistic regression analysis was used to test the association of patient and cycle characteristics with the presence of cultured blastocysts on Day 5 and to create a nomogram. The model's performance was quantified by discrimination and calibration and clinical utility was evaluated for various thresholds. Fertilization technique, number and quality ratio of the Day 3 embryos and female age were independently associated with blastocyst development. The final prediction model showed an area under the curve (AUC) of 0.75 in the training set (95% confidence interval (CI): 0.73-0.77) and was well calibrated. The AUC for the validation data set was 0.80 (95% CI: 0.78-0.83) and calibration was acceptable. Using a decision threshold of 0.55, the model showed a negative predictive value of 80.5% and a false positive rate of 17%. A cycle-based model could prove clinically-relevant for reducing the incidence of cancelled Day 5 transfers.