Number of embryos for transfer following in vitro fertilisation or intra-cytoplasmic sperm injection.

BACKGROUND: Transfer of more than one embryo during in vitro fertilisation (IVF) or intracytoplasmic sperm injection (ICSI) increases multiple pregnancy rates resulting in an increased risk of maternal and perinatal morbidity. Elective single embryo transfer offers a means of minimising this risk, but this potential gain needs to be balanced against the possibility of jeopardising the overall live birth rate (LBR). OBJECTIVES: To evaluate the effectiveness and safety of different policies for the number of embryos transferred in infertile couples undergoing assisted reproductive technology cycles. SEARCH METHODS: We searched the Cochrane Gynaecology and Fertility Group specialised register of controlled trials, CENTRAL, MEDLINE, Embase, ClinicalTrials.gov, and the World Health Organization International Clinical Trials Registry Platform from inception to March 2020. We handsearched reference lists of articles and relevant conference proceedings. We also communicated with experts in the field regarding any additional studies. SELECTION CRITERIA: We included randomised controlled trials (RCTs) comparing different policies for the number of embryos transferred following IVF or ICSI in infertile women. Studies of fresh or frozen and thawed transfer of one to four embryos at cleavage or blastocyst stage were eligible. DATA COLLECTION AND ANALYSIS: Two review authors independently extracted data and assessed trial eligibility and risk of bias. The primary outcomes were LBR and multiple pregnancy rate. The secondary outcomes were clinical pregnancy and miscarriage rates. We analysed data using risk ratios (RR), Peto odds ratio (Peto OR) and a fixed effect model. MAIN RESULTS: We included 17 RCTs in the review (2505 women). The main limitation was inadequate reporting of study methods and moderate to high risk of performance bias due to lack of blinding. A majority of the studies had low numbers of participants. None of the trials compared repeated single embryo transfer (SET) with multiple embryo transfer. Reported results of multiple embryo transfer below refer to double embryo transfer. Repeated single embryo transfer versus multiple embryo transfer in a single cycle Repeated SET was compared with double embryo transfer (DET) in four studies of cleavage-stage transfer. In these studies the SET group received either two cycles of fresh SET (one study) or one cycle of fresh SET followed by one frozen SET (three studies). The cumulative live birth rate after repeated SET may be little or no different from the rate after one cycle of DET (RR 0.95, 95% CI (confidence interval) 0.82 to 1.10; I² = 0%; 4 studies, 985 participants; low-quality evidence). This suggests that for a woman with a 42% chance of live birth following a single cycle of DET, the repeated SET would yield pregnancy rates between 34% and 46%. The multiple pregnancy rate associated with repeated SET is probably reduced compared to a single cycle of DET (Peto OR 0.13, 95% CI 0.08 to 0.21; I² = 0%; 4 studies, 985 participants; moderate-quality evidence). This suggests that for a woman with a 13% risk of multiple pregnancy following a single cycle of DET, the risk following repeated SET would be between 0% and 3%. The clinical pregnancy rate (RR 0.99, 95% CI 0.87 to 1.12; I² = 47%; 3 studies, 943 participants; low-quality evidence) after repeated SET may be little or no different from the rate after one cycle of DET. There may be little or no difference in the miscarriage rate between the two groups. Single versus multiple embryo transfer in a single cycle A single cycle of SET was compared with a single cycle of DET in 13 studies, 11 comparing cleavage-stage transfers and three comparing blastocyst-stage transfers.One study reported both cleavage and blastocyst stage transfers. Low-quality evidence suggests that the live birth rate per woman may be reduced in women who have SET in comparison with those who have DET (RR 0.67, 95% CI 0.59 to 0.75; I² = 0%; 12 studies, 1904 participants; low-quality evidence). Thus, for a woman with a 46% chance of live birth following a single cycle of DET, the chance following a single cycle of SET would be between 27% and 35%. The multiple pregnancy rate per woman is probably lower in those who have SET than those who have DET (Peto OR 0.16, 95% CI 0.12 to 0.22; I² = 0%; 13 studies, 1952 participants; moderate-quality evidence). This suggests that for a woman with a 15% risk of multiple pregnancy following a single cycle of DET, the risk following a single cycle of SET would be between 2% and 4%. Low-quality evidence suggests that the clinical pregnancy rate may be lower in women who have SET than in those who have DET (RR 0.70, 95% CI 0.64 to 0.77; I² = 0%; 10 studies, 1860 participants; low-quality evidence). There may be little or no difference in the miscarriage rate between the two groups. AUTHORS' CONCLUSIONS: Although DET achieves higher live birth and clinical pregnancy rates per fresh cycle, the evidence suggests that the difference in effectiveness may be substantially offset when elective SET is followed by a further transfer of a single embryo in fresh or frozen cycle, while simultaneously reducing multiple pregnancies, at least among women with a good prognosis. The quality of evidence was low to moderate primarily due to inadequate reporting of study methods and absence of masking those delivering, as well as receiving the interventions.

Morula transfer achieves better clinical outcomes than post-thawed cleavage embryos after overnight culture in frozen embryo transfer (FET) cycles.

PURPOSE: This study aimed to investigate the clinical outcomes of morula stage transfer derived from post-thawed cleavage embryos undergoing overnight culture in frozen embryo transfer (FET) cycles. METHODS: We performed a retrospective study that included 392 FET cycles with 784 thawed embryos undergoing overnight culture between January 2014 and December 2018. Embryos were divided into three groups in terms of their status: 8-16 cells without morula (group I), one morula (group II), and two morulae (group III). The clinical outcomes of these cycles were then compared between the three groups. Logistic regression analysis was performed to control for confounders. RESULTS: Group III was associated with a significantly higher clinical pregnancy rate (odds ratio [OR] 2.35; 95% confidence interval [CI] 1.29-4.27; P = 0.005), implantation rate (OR 3.00; CI 1.75-5.16; P < 0.001), multiple pregnancy rate (OR 4.91; CI 2.11-11.40; P < 0.001), and live birth rate (OR 1.96; CI 1.10-3.49; P = 0.022) than group I. Group II had a higher live birth rate than group I after adjustment (OR 1.70; CI 1.04-2.79; P = 0.035). There was no difference in the rate of premature delivery when compared across the three groups after adjustment. CONCLUSION: The transfer of morula stage embryos following the overnight culture of post-thawed cleavage embryos led to an improvement in the clinical outcomes of FET cycles. It is important to reduce the number of morula embryos transferred in order to achieve a singleton pregnancy.

Laser-assisted hatching and clinical outcomes in frozen-thawed cleavage-embryo transfers of patients with previous repeated failure.

Assisted hatching (AH) is initially developed to provide an artificial manipulation of the zona pellucida (ZP) to help embryos hatch and improve the capacity of the embryos to implant. However, these effects remain unclear and controversial because of variation in patient characteristics, and it is critical to ascertain the indications for AH and to identify those patients who might benefit from AH. Here, this study aimed to assess the effect of laser-assisted zona thinning hatching technology (LAH) during the frozen-thawed D3 embryos on pregnancy outcomes in patients with previous repeated failures in vitro fertilization-embryo transfer (IVF-ET). To the best of our knowledge, these relationships have not been previously investigated. A retrospective cohort analysis was carried out. Infertility patients with previous repeated failure who underwent assisted reproductive therapy at our in vitro fertilization (IVF) center from May 2014 to May 2016 were enrolled. A total of 415 cleavage FET cycles (225 in the LAH group and 190 in the control group) were analyzed. Clinical outcomes including clinical pregnancy, implantation, live birth, miscarriage, and multiple gestation rates after transfer were compared between the LAH and control groups. The clinical pregnancy (49.3% versus 38.9%) and implantation rates (31.2% versus 24.6%) were significantly higher for the LAH group than the control group (P < 0.05). The live birth (44.8% versus 35.8%), multiple pregnancy (32.4% versus 31.0%), preterm birth (22.8% versus 17.1%), miscarriage (7.2% versus 5.4%), and ectopic rates (1.9% versus 0%) did not differ significantly between the two groups (P > 0.05). This study showed that LAH via zona pellucida (ZP) thinning significantly improves clinical outcomes, particularly clinical pregnancy and implantation rates, associated with FET cycles among patients with previous repeated failure.

Birthweight of singletons born after cleavage-stage or blastocyst transfer in fresh and warming cycles.

STUDY QUESTION: Does extended culture to the blastocyst stage affect singleton birthweight after either fresh or vitrified-warmed embryo transfer? SUMMARY ANSWER: Singleton birthweight z-scores did not vary significantly after a fresh blastocyst transfer, whereas the additional effect of vitrification remains inconclusive. WHAT IS KNOWN ALREADY: Observational studies have associated extended culture with an increased risk of preterm birth and low birthweight. On the contrary, in terms of birthweight and gestational age, singletons born after vitrification have been associated with a better perinatal outcome when compared to those born following a fresh transfer. STUDY DESIGN, SIZE, DURATION: Our post-hoc cohort analysis on neonatal outcomes included 447 liveborn singletons was derived from a recent retrospective analysis on cumulative live birth rates after cleavage-stage and blastocyst transfers. These babies were born following a fresh single cleavage-stage transfer (FCT Day 3, n = 113), fresh single blastocyst transfer (FBT Day 5, n = 218), vitrified-warmed cleavage-stage transfer (VCT Day 3, n = 58) or vitrified-warmed blastocyst transfer (VBT Day 5, n = 58). PARTICIPANTS/MATERIALS, SETTING, METHODS: Singleton birthweight was the primary outcome measure. Gestational age and gender of the newborn were accounted for by using birthweight z-scores in a multivariable linear regression analysis, adjusting for other confounders (maternal age, BMI, parity and smoking behaviour). Vanishing twins were excluded from the analysis. MAIN RESULTS AND THE ROLE OF CHANCE: A significantly lower z-score was observed after blastocyst transfer compared to cleavage-stage transfer in the vitrified-warmed Day 5 group (P = 0.013), a difference not observed in the fresh transfer groups (P = 0.32). Following multivariable regression analysis [adjusted regression coefficient (95% confidence interval)], the FCT and FBT groups showed no significant influence on the birthweight z-scores after fresh transfer [-0.19 (-0.44; 0.05)], but the transfer of vitrified blastocysts (VBT) was associated with a lower birthweight [-0.52 (-0.90; -0.15)] compared with the transfer of vitrified cleavage-stage embryos (VCT). LIMITATIONS, REASONS FOR CAUTION: The present cohort was relatively small, especially in the vitrified-warmed subgroups. Pregnancy-associated factors possibly influencing birthweight (such as diabetes, hypertension, pre-eclampsia) were also not accounted for in the analysis. WIDER IMPLICATIONS OF THE FINDINGS: Different ART procedures, including extended culture and vitrification, may hold potential safety issues. These results require further confirmation in future larger studies. STUDY FUNDING/COMPETING INTEREST(S): None. TRIAL REGISTRATION NUMBER: N/A.

Clinical analysis of the patients with single fair cleavage-stage embryo on day 3.

OBJECTIVE: This study aimed to explore an appropriate selection for the patients with single fair cleavage-stage embryo on day 3. METHODS: This study included 469 fresh transfers and 220 frozen-thawed transfers from January 2014 to June 2016. Furthermore, in 72 patients who have only 4-6 fair embryos (4-5 blastomeres) on day 3, the blastocysts were cultured to day 5 for transfer. RESULTS: In the fresh transfers, the clinical pregnancy rate of 4-5 blastomeres group was significantly lower than 6-7 and 8-10 blastomeres group (5.88 vs. 30.13%, p<.001and 5.88 vs. 26.09%, p < .001). In the frozen-thawed transfers, the clinical pregnancy rate of 4-5 blastomeres group was also significantly lower than 6-7 and 8-10 blastomeres group (10.00 vs. 28.57%, p = .040 and 10.00 vs. 33.33%, p = .005). For the blastocyst transfers derived from fair embryos with 4-5 blastomeres, the clinical pregnancy rate was significantly higher than single and double fair embryo transfers of similar quality (44.44 vs. 7.04%, p < .001 and 44.44 vs. 28.09%, p = .013). CONCLUSIONS: For the patients with single fair embryo (6-7 blastomeres or 8-10 blastomeres), transfer at the cleavage stage is feasible. For the patients with single fair embryo (4-5 blastomeres), transfer of single fair embryo at the blastocyst stage or accumulating two fair embryos might be worthy of consideration.

Blastocyst vs cleavage-stage embryo transfer: systematic review and meta-analysis of reproductive outcomes.

OBJECTIVES: Blastocyst transfer in assisted reproduction techniques could be advantageous because the timing of exposure of the embryo to the uterine environment is more analogous to a natural cycle and permits embryo self-selection after activation of the embryonic genome on day 3. Conversely, the in-vitro environment is likely to be inferior to that in vivo, and in-vitro culture beyond embryonic genomic activation could potentially harm the embryo. Our objective was to identify, appraise and summarize the available evidence comparing the effectiveness of blastocyst vs cleavage-stage embryo transfer. METHODS: This was a systematic review and meta-analysis of randomized controlled trials (RCTs) comparing the transfer of blastocysts (days 5-6) with the transfer of cleavage-stage embryos (days 2-3) in women undergoing in-vitro fertilization or intracytoplasmic sperm injection. The last electronic searches were run on 1 August 2016. Abstracts and studies with a mean difference between the two study groups of > 0.5 for the number of embryos transferred were excluded. RESULTS: We screened 1187 records and assessed 33 potentially eligible studies. Twelve studies were included, comprising a total of 1200 women undergoing blastocyst transfer and 1218 undergoing cleavage-stage embryo transfer. We observed low-quality evidence of no significant difference of blastocyst transfer on live birth/ongoing pregnancy (relative risk (RR), 1.11 (95% CI, 0.92-1.35), 10 RCTs, 1940 women, I2 = 54%), clinical pregnancy (RR, 1.10 (95% CI, 0.93-1.31), 12 RCTs, 2418 women, I2 = 64%), cumulative pregnancy (RR, 0.89 (95% CI, 0.67-1.16), four RCTs, 524 women, I2 = 63%) and miscarriage (RR, 1.08 (95% CI, 0.74-1.56), 10 RCTs, 763 pregnancies, I2 = 0%). There was moderate-quality evidence of a decrease in the number of women with surplus embryos after the blastocyst-stage embryo transfer (RR, 0.78 (95% CI, 0.66-0.91)). Overall, the quality of the evidence was limited by the quality of the included studies and by unexplained inconsistency across studies. CONCLUSIONS: Current evidence shows no superiority of blastocyst compared with cleavage-stage embryo transfer in clinical practice. As the quality of the evidence for the primary outcomes is low, additional well-designed RCTs are still needed before robust conclusions can be drawn. Copyright © 2016 ISUOG. Published by John Wiley & Sons Ltd.

Production of calves by the transfer of cryopreserved bovine elongating conceptuses and possible application for preimplantation genomic selection.

Preimplantation genomic selection based on single nucleotide polymorphism (SNP) genotypes is expected to accelerate genetic improvement in cattle. However, genome-wide genotyping at the early embryonic stage has several limitations, such as the technical difficulty of embryonic biopsy and low accuracy of genotyping resulting from a limited number of biopsied cells. After hatching from the zona pellucida, the morphology of the bovine embryo changes from spherical to filamentous, in a process known as elongation. The bovine nonsurgical elongating conceptus transfer technique was recently developed and applied for sexing without requiring specialized skills for biopsy. In order to develop a bovine preimplantation genomic selection system combined with the elongating conceptus transfer technique, we examined the accuracy of genotyping by SNP chip analysis using the DNA from elongating conceptuses (Experiment 1) and optimal cryopreservation methods for elongating conceptuses (Experiment 2). In Experiment 1, the call rates of SNP chip analysis following whole genome amplification in biopsied cells from two elongating conceptuses were 95.14% and 99.32%, which were sufficient for estimating genomic breeding value. In Experiment 2, the rates of dead cells in elongating conceptuses cryopreserved by slow freezing were comparable to those in fresh elongating conceptuses. In addition, we obtained healthy calves by the transfer of elongating conceptuses cryopreserved by slow freezing. Our findings indicate that the elongating conceptus transfer technology enables preimplantation genomic selection in cattle based on SNP chip analysis. Further studies on the optimization of cryopreservation methods for elongating conceptuses are required for practical application of the selection system.

Effects of high three pro-nuclei (3PN) proportion incidence on clinical outcomes in the fresh cleavage-stage and blastocyst-stage embryo transfer (ET) cycles.

OBJECTIVES: The aim of this study was to explore the effect of three pro-nuclei (3PN) incidence on clinical outcomes in the fresh cleavage-stage embryo transfer (CSET) and blastocyst-stage embryo transfer (BSET) cycles. METHODS: This retrospective cohort study included 1427 CSET cycles, 632 BSET cycles, and 313 elective single BSET cycles from January 2013 to June 2015. The patients were divided into two groups as follows: Group 1 included patients with no 3PN zygotes and Group 2 included patients with >20% 3PN zygotes. RESULTS: We observed that the fertilization rate was significantly lower in 3PN = 0% than 3PN > 20% group (p < 0.05), but the day-3 grade I + II embryo and day-3 grade I + II + III embryo rates were not significantly different between 3PN = 0% and 3PN > 20% group (p > 0.05). Interestingly, in the CSET, the implantation (42.87% and 41.76%, p = 0.585) and clinical pregnancy (59.94% and 58.25%, p = 0.538) rates were not significantly different between two groups. In the BSET, the implantation (61.93% and 49.62%, p < 0.001) and clinical pregnancy rates (69.45% and 61.02%, p = 0.043) were significantly higher in 3PN = 0% than 3PN > 20% group. In the elective single BSET, the implantation (68.91% and 61.33%, p = 0.223) and clinical pregnancy rates (68.48% and 61.33%, p = 0.251) were higher in 3PN = 0% than 3PN > 20% group, but there was no significant difference. CONCLUSIONS: We concluded that a high 3PN incidence may predict poor outcomes in BSET but not CSET cleavage-stage.

Obstetrical and perinatal outcomes following blastocyst transfer compared to cleavage transfer: a systematic review and meta-analysis.

STUDY QUESTION: Is blastocyst transfer safe when compared to cleavage stage embryo transfer regarding obstetric and perinatal outcomes? SUMMARY ANSWER: The clinical equipoise between blastocyst and cleavage stage embryo transfer remains as the evidence associating blastocyst transfer with some adverse perinatal outcomes is of low/very low quality. WHAT IS KNOWN ALREADY: Extended embryo culture to the blastocyst stage provides some theoretical advantages and disadvantages. While it permits embryo self-selection, it also exposes those embryos to possible harm due to the in vitro environment. Both effectiveness and safety should be weighed to permit evidence-based decisions in clinical practice. STUDY DESIGN, SIZE, DURATION: This is a systematic review and meta-analysis of randomized controlled trials (RCTs) and observational studies reporting perinatal outcomes for singletons comparing the deliveries resulting from blastocyst and cleavage stage embryo transfer. Observational studies were included because the primary outcomes, perinatal mortality and birth defects, are rare and require a large number of participants (>50 000) to be properly assessed. The last electronic searches were last run on 11 March 2016. PARTICIPANTS/MATERIALS, SETTING, METHOD: There were 12 observational studies encompassing 195 325 singleton pregnancies included in the study. No RCT reported the studied outcomes. The quality of the included studies was evaluated according to the Newcastle-Ottawa Scale and the quality of the evidence was evaluated according to GRADE criteria. MAIN RESULTS AND THE ROLE OF CHANCE: Blastocyst stage transfer was associated with increased risks of preterm birth (<37 weeks), very preterm birth (<32 weeks), large for gestational age and perinatal mortality, although the latter was only identified from one study. Conversely, blastocyst stage transfer was associated with a decrease in the risks of small for gestational age and vanishing twins, although the latter was reported by only one study. LIMITATIONS, REASONS FOR CAUTION: The observational nature of the included studies and some inconsistency and imprecision in the analysis contributed to decreasing our confidence in the estimates. WIDER IMPLICATIONS OF THE FINDINGS: Due to the overall low quality of available evidence, the clinical equipoise between cleavage stage and blastocyst transfer remains. More large well-conducted studies are needed to clarify the potential risks and benefits of blastocyst transfer. As this review was initiated to support global recommendations on best practice, and in light of the challenges in lower resource settings to offer extended culture to blastocyst stage, it is critical to take into consideration these obstetric and neonatal outcomes in order to ensure any recommendation will not result in the overburdening of existing maternal and child health care systems and services. STUDY FUNDING/COMPETING INTERESTS: No external funding was either sought or obtained for this study. The authors have no competing interests to declare. PROSPERO REGISTRATION NUMBER: CRD42015023910.

Cleavage stage versus blastocyst stage embryo transfer in assisted reproductive technology.

BACKGROUND: Advances in cell culture media have led to a shift in in vitro fertilisation (IVF) practice from cleavage stage embryo transfer to blastocyst stage transfer. The rationale for blastocyst transfer is to improve both uterine and embryonic synchronicity and enable self selection of viable embryos, thus resulting in better live birth rates. OBJECTIVES: To determine whether blastocyst stage (day 5 to 6) embryo transfers improve the live birth rate, and other associated outcomes, compared with cleavage stage (day 2 to 3) embryo transfers. SEARCH METHODS: We searched the Cochrane Gynaecology and Fertility Group Specialised Register of controlled trials, Cochrane Central Register of Controlled Trials (CENTRAL; the Cochrane Library; 2016, Issue 4), MEDLINE, EMBASE, PsycINFO, CINAHL, and Bio extracts from inception to 4th April 2016. We also searched registers of ongoing trials and the reference lists of studies retrieved. SELECTION CRITERIA: We included randomised controlled trials (RCTs) which compared the effectiveness of blastocyst versus cleavage stage transfers. DATA COLLECTION AND ANALYSIS: We used standard methodological procedures recommended by Cochrane. Our primary outcomes were live birth and cumulative clinical pregnancy rates. Secondary outcomes were clinical pregnancy, multiple pregnancy, high order pregnancy, miscarriage, failure to transfer embryos, and embryo freezing. We assessed the overall quality of the evidence for the main comparisons using GRADE methods. MAIN RESULTS: We included 27 RCTs (4031 couples or women).The live birth rate following fresh transfer was higher in the blastocyst transfer group (odds ratio (OR) 1.48, 95% confidence interval (CI) 1.20 to 1.82; 13 RCTs, 1630 women, I(2) = 45%, low quality evidence) following fresh transfer. This suggests that if 29% of women achieve live birth after fresh cleavage stage transfer, between 32% and 42% would do so after fresh blastocyst stage transfer.There was no evidence of a difference between the groups in rates per couple of cumulative pregnancy following fresh and frozen-thawed transfer after one oocyte retrieval (OR 0.89, 95% CI 0.64 to 1.22; 5 RCTs, 632 women, I(2) = 71%, very low quality evidence).The clinical pregnancy rate was also higher in the blastocyst transfer group, following fresh transfer (OR 1.30, 95% CI 1.14 to 1.47; 27 RCTs, 4031 women, I(2) = 56%, moderate quality evidence). This suggests that if 36% of women achieve clinical pregnancy after fresh cleavage stage transfer, between 39% and 46% would do so after fresh blastocyst stage transfer.There was no evidence of a difference between the groups in rates of multiple pregnancy (OR 1.05, 95% CI 0.83 to 1.33; 19 RCTs, 3019 women, I(2) = 30%, low quality evidence), or miscarriage (OR 1.15, 95% CI 0.88 to 1.50; 18 RCTs, 2917 women, I(2) = 0%, low quality evidence). These data are incomplete as under 70% of studies reported these outcomes.Embryo freezing rates were lower in the blastocyst transfer group (OR 0.48, 95% CI 0.40 to 0.57; 14 RCTs, 2292 women, I(2) = 84%, low quality evidence). This suggests that if 60% of women have embryos frozen after cleavage stage transfer, between 37% and 46% would do so after blastocyst stage transfer. Failure to transfer any embryos was higher in the blastocyst transfer group (OR 2.50, 95% CI 1.76 to 3.55; 17 RCTs, 2577 women, I(2) = 36%, moderate quality evidence). This suggests that if 1% of women have no embryos transferred in (planned) fresh cleavage stage transfer, between 2% and 4% will have no embryos transferred in (planned) fresh blastocyst stage transfer.The evidence was of low quality for most outcomes. The main limitation was serious risk of bias, associated with failure to describe acceptable methods of randomisation, and unclear or high risk of attrition bias. AUTHORS' CONCLUSIONS: There is low quality evidence for live birth and moderate quality evidence for clinical pregnancy that fresh blastocyst stage transfer is associated with higher rates than fresh cleavage stage transfer. There was no evidence of a difference between the groups in cumulative pregnancy rates derived from fresh and frozen-thawed cycles following a single oocyte retrieval, but the evidence for this outcome was very low quality. Thus, although there is a benefit favouring blastocyst transfer in fresh cycles, it remains unclear whether the day of transfer impacts on cumulative live birth and pregnancy rates. Future RCTs should report rates of live birth, cumulative live birth, and miscarriage to enable couples or women undergoing assisted reproductive technology (ART) and service providers to make well informed decisions on the best treatment option available.

Effects of three pro-nuclei (3PN) proportion incidence on clinical outcomes of patients with lower retrieved oocytes in the fresh cleavage-stage embryo transfer (ET) cycles.

PURPOSE: To analyze the three pro-nuclei (3PN) incidence on clinical outcomes of patients with lower retrieved oocytes in the fresh cleavage-stage embryo transfer (ET) cycles. METHODS: This study included 1200 fresh cleavage-stage ET cycles from January 2013 to June 2015. The patients were divided into 3PN = 0% (773 cycles) and 3PN > 0% (427 cycles) group. Main outcomes compared were fertilization, cleavage, normal fertilization, good quality embryo, implantation, clinical pregnancy, and early abortion rate. RESULTS: We observed that there was no significant difference in female's age, the number of retrieved oocytes, the number of transferred embryos, the number of good quality embryos, endometrial thickness, infertile time, basal serum follicle-stimulating hormone, and E2 value between two groups (p > 0.05). The fertilization (89.43 versus 83.90%, p < 0.001) and cleavage (98.34 versus 97.19%, p = 0.048) rates were significantly higher in 3PN > 0% than 3PN = 0% group. However, the normal fertilization (70.05 versus 50.67%, p < 0.001), good quality embryos (37.11 versus 26.47%, p < 0.001), and clinical pregnancy (49.81 versus 43.79%, p = 0.046) rates were significantly higher in 3PN = 0% than 3PN > 0% group. The implantation (35.88 versus 33.78%, p = 0.333) and early abortion (8.83 versus 10.70%, p = 0.474) rates were not significantly different between two groups. CONCLUSION: 3PN incidence might make a negative effect on clinical outcomes for patients with lower retrieved oocytes in the fresh cleavage-stage ET cycles.

Pregnancy outcome after blastocyst stage transfer comparing to early cleavage stage embryo transfer.

Blastocyst transfer has been introduced as an alternative for improving the chance for in vitro fertilizations (IVF) implantation. The present study was to evaluate pregnancy rates when embryo transfer was performed either on day 2-3 (cleavage stage) or on day 4-5 (blastocyst stage). This randomized clinical trial included 118 infertile women. All the study subjects underwent controlled ovarian stimulation using a long protocol and randomized into two groups. BS group (n = 57), the culture was extended to day 5 (blastocyst stage) and in the CS-group (n = 61), embryo culture was continued to day 3 (cleavage stage). Ongoing pregnancies, abortion, implantation rate were evaluated. No significant differences were seen in the pregnancy rate between the two groups (33.3% in the BS group versus 27.9% in the CS group; p = 0.519). Abortion, implantation rate in two groups are not significant. Despite the lack of statistical difference between the two study groups, our data suggest that blastocyst transfer may be associated with a higher pregnancy and an overall better implantation rates. However, further studies with larger sample size are mandatory to confirm these findings.

Changes in sex ratio from fertilization to birth in assisted-reproductive-treatment cycles.

BACKGROUND: In Western gender-neutral countries, the sex ratio at birth is estimated to be approximately 1.06. This ratio is lower than the estimated sex ratio at fertilization which ranges from 1.07 to 1.70 depending on the figures of sex ratio at birth and differential embryo/fetal mortality rates taken into account to perform these estimations. Likewise, little is known about the sex ratio at implantation in natural and assisted-reproduction-treatment (ART) cycles. In this bioessay, we aim to estimate the sex ratio at fertilization and implantation using data from embryos generated by standard in-vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) in preimplantation genetic diagnosis cycles. Thereafter, we compare sex ratios at implantation and birth in cleavage- and blastocyst-stage-transfer cycles to propose molecular mechanisms accounting for differences in post-implantation male and female mortality and thereby variations in sex ratios at birth in ART cycles. METHODS: A literature review based on publications up to December 2013 identified by PubMed database searches. RESULTS: Sex ratio at both fertilization and implantation is estimated to be between 1.29 and 1.50 in IVF cycles and 1.07 in ICSI cycles. Compared with the estimated sex ratio at implantation, sex ratio at birth is lower in IVF cycles (1.03 after cleavage-stage transfer and 1.25 after blastocyst-stage transfer) but similar and close to unity in ICSI cycles (0.95 after cleavage-stage transfer and 1.04 after blastocyst-stage transfer). CONCLUSIONS: In-vitro-culture-induced precocious X-chromosome inactivation together with ICSI-induced decrease in number of trophectoderm cells in female blastocysts may account for preferential female mortality at early post-implantation stages and thereby variations in sex ratios at birth in ART cycles.

Time-lapse embryo imaging for improving reproductive outcomes: systematic review and meta-analysis.

OBJECTIVE: To identify, appraise and summarize the available evidence regarding the effectiveness and safety of time-lapse embryo monitoring on the main outcomes of assisted reproductive techniques. METHODS: In this systematic review and meta-analysis, we included only randomized controlled trials (RCTs) comparing time-lapse embryo imaging with standard embryo monitoring. Our primary outcomes were live births (efficacy) and congenital abnormalities (safety). The secondary outcomes were clinical pregnancy, ongoing pregnancy and miscarriage. RESULTS: Two RCTs were considered eligible, and their data were extracted and included in a meta-analysis. In both studies embryos were transferred at the blastocyst stage. No studies reported rates of live birth or congenital abnormalities. Our estimates were not sufficiently precise to identify whether time-lapse monitoring provided a small benefit, no effect or minor harm on rates of clinical pregnancy (relative risk (RR), 1.05 (95% CI, 0.80-1.38)) or ongoing pregnancy (RR, 1.05 (95% CI, 0.76-1.45)), based on two studies involving 138 women with moderate-quality evidence. Considering the available data, we were unable to determine whether the intervention poses substantial benefit, no effect or substantial harm in the risk of miscarriage (RR, 0.95 (95% CI, 0.30-2.99)), based on two studies involving 76 clinical pregnancies with low-quality evidence. CONCLUSIONS: Time-lapse embryo imaging is unlikely to have a large effect on the chance of achieving clinical and/or ongoing pregnancy when transferring embryos at the blastocyst stage. More studies are required to improve the quality of the current evidence and also to examine whether this intervention is useful when transferring embryos at the cleavage stage.

The importance of the cleavage stage morphology evaluation for blastocyst transfer in patients with good prognosis.

PURPOSE: To evaluate: (i) the influence of morphology at cleavage stage on blastocyst formation and implantation, and (ii) whether the transfer of low-quality embryos on day-three would be a better approach than the transfer at blastocyst stage. METHODS: This study included 8,444 embryos obtained from 1,125 patients undergoing ICSI cycles between January/2011 and September/2013. The influence of the quality of the embryo on days-two and -three on blastocyst formation and implantation success was evaluated. Moreover, the implantation potential of low-quality embryos, at cleavage stage, transferred on day-three was compared with the implantation potential of low-quality embryos, at cleavage stage, transferred on day-five. RESULTS: Low-quality embryos on day-two had an approximate 20 % decreased chance of achieving the blastocyst stage, and blastocysts derived from low-quality embryos on day-two had a nearly 40 % decrease in the implantation chance. Low-quality embryos on day-three had a 30 % decreased chance of achieving the blastocyst stage, and blastocysts derived from low-quality embryos on day-three had an almost 40 % decreased implantation chance. The implantation rate didn't differ when low-quality embryos on the cleavage stage were transferred on day-three or left in culture and transferred on day-five. CONCLUSIONS: The transfer of low-quality embryos on day-three is a better approach than transfer at the blastocyst stage. In addition, the embryo morphology evaluation at the cleavage stage is still needed for the selection of the embryo with the best implantation potential in extended embryo culture programmes.

Number of embryos for transfer following in vitro fertilisation or intra-cytoplasmic sperm injection.

BACKGROUND: Multiple embryo transfer during in vitro fertilisation (IVF) increases multiple pregnancy rates causing maternal and perinatal morbidity. Single embryo transfer is now being seriously considered as a means of minimising the risk of multiple pregnancy. However, this needs to be balanced against the risk of jeopardising the overall live birth rate. OBJECTIVES: To evaluate the effectiveness and safety of different policies for the number of embryos transferred in couples who undergo assisted reproductive technology (ART). SEARCH METHODS: We searched the Cochrane Menstrual Disorders and Subfertility Group Trials Register, the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE and EMBASE, from inception to July 2013. We handsearched reference lists of articles, trial registers and relevant conference proceedings and contacted researchers in the field. SELECTION CRITERIA: We included randomised controlled trials (RCTs) comparing different policies for the number of embryos transferred following IVF or intra-cytoplasmic sperm injection (ICSI) in subfertile women. Studies of fresh or frozen and thawed transfer of one, two, three or four embryos at cleavage or blastocyst stage were eligible. DATA COLLECTION AND ANALYSIS: Two review authors independently assessed trial eligibility and risk of bias and extracted the data. The overall quality of the evidence was graded in a summary of findings table. MAIN RESULTS: Fourteen RCTs were included in the review (2165 women). Thirteen compared cleavage-stage transfers (2017 women) and two compared blastocyst transfers (148 women): one study compared both. No studies compared repeated multiple versus repeated single embryo transfer (SET). DET versus repeated SETDET was compared with repeated SET in three studies of cleavage-stage transfer. In these studies the SET group received either two cycles of fresh SET (one study) or one cycle of fresh SET followed by one frozen SET in a natural or hormone-stimulated cycle (two studies). When these three studies were pooled, the cumulative live birth rate after one cycle of DET was not significantly different from the rate after repeated SET (OR 1.22, 95% CI 0.92 to 1.62, three studies, n=811, I(2)=0%, low quality evidence). This suggests that for a woman with a 40% chance of live birth following a single cycle of DET, the chance following repeated SET would be between 30% and 42%. The multiple pregnancy rate was significantly higher in the DET group (OR 30.54, 95% CI 7.46 to 124.95, three RCTs, n = 811, I(2) = 23%, low quality evidence), suggesting that for a woman with a 15% risk of multiple pregnancy following a single cycle of DET, the risk following repeated SET would be between 0% and 2%. Single-cycle DET versus single-cycle SETA single cycle of DET was compared with a single cycle of SET in 10 studies, nine comparing cleavage-stage transfers and two comparing blastocyst-stage transfers. When all studies were pooled the live birth rate was significantly higher in the DET group (OR 2.07, 95% CI 1.68 to 2.57, nine studies, n = 1564, I(2) = 0%, high quality evidence). This suggests that for a woman with a 40% chance of live birth following a single cycle of DET, the chance following a single cycle of SET would be between 22% and 30%. The multiple pregnancy rate was also significantly higher in the DET group (OR 8.47, 95% CI 4.97 to 14.43, 10 studies, n = 1612, I(2) = 45%, high quality evidence), suggesting that for a woman with a 15% risk of multiple pregnancy following a single cycle of DET, the risk following a single cycle of SET would be between 1% and 4%. The heterogeneity for this analysis was attributable to a study with a high rate of cross-over between treatment arms. Other comparisons Other fresh cycle comparisons were evaluated in three studies which compared DET versus transfer of three or four embryos. Live birth rates did not differ significantly between the groups for any comparison, but there was a significantly lower multiple pregnancy rate in the DET group than in the three embryo transfer (TET) group (OR 0.36, 95% CI 0.13 to 0.99, two studies, n = 343, I(2) = 0%). AUTHORS' CONCLUSIONS: In a single fresh IVF cycle, single embryo transfer is associated with a lower live birth rate than double embryo transfer. However, there is no evidence of a significant difference in the cumulative live birth rate when a single cycle of double embryo transfer is compared with repeated SET (either two cycles of fresh SET or one cycle of fresh SET followed by one frozen SET in a natural or hormone-stimulated cycle). Single embryo transfer is associated with much lower rates of multiple pregnancy than other embryo transfer policies. A policy of repeated SET may minimise the risk of multiple pregnancy in couples undergoing ART without substantially reducing the likelihood of achieving a live birth. Most of the evidence currently available concerns younger women with a good prognosis.

Cleavage stage versus blastocyst stage embryo transfer in assisted reproductive technology.

BACKGROUND: Advances in cell culture media have led to a shift in in vitro fertilization (IVF) practice from early cleavage embryo transfer to blastocyst stage transfer. The rationale for blastocyst culture is to improve both uterine and embryonic synchronicity and enable self selection of viable embryos thus resulting in higher implantation rates. OBJECTIVES: To determine if blastocyst stage (Day 5 to 6) embryo transfers (ETs) improve live birth rate and other associated outcomes compared with cleavage stage (Day 2 to 3) ETs. SEARCH METHODS: Cochrane Menstrual Disorders and Subfertility Group Specialised Register of controlled trials, Cochrane Central Register of Controlled Trials (CENTRAL) (The Cochrane Library), MEDLINE, EMBASE and Bio extracts. The last search date was 21 February 2012. SELECTION CRITERIA: Trials were included if they were randomised and compared the effectiveness of early cleavage versus blastocyst stage transfers. DATA COLLECTION AND ANALYSIS: Of the 50 trials that were identified, 23 randomised controlled trials (RCTs) met the inclusion criteria and were reviewed (five new studies were added in this update). The primary outcome was rate of live birth. Secondary outcomes were rates per couple of clinical pregnancy, cumulative clinical pregnancy, multiple pregnancy, high order pregnancy, miscarriage, failure to transfer embryos and cryopreservation. Quality assessment, data extraction and meta-analysis were performed following Cochrane guidelines. MAIN RESULTS: Twelve RCTs reported live birth rates and there was evidence of a significant difference in live birth rate per couple favouring blastocyst culture (1510 women, Peto OR 1.40, 95% CI 1.13 to 1.74) (Day 2 to 3: 31%; Day 5 to 6: 38.8%, I(2) = 40%). This means that for a typical rate of 31% in clinics that use early cleavage stage cycles, the rate of live births would increase to 32% to 42% if clinics used blastocyst transfer.There was no difference in clinical pregnancy rate between early cleavage and blastocyst transfer in the 23 RCTs (Peto OR 1.14, 95% CI 0.99 to 1.32) (Day 2 to 3: 38.6%; Day 5 to 6: 41.6%) and no difference in miscarriage rate (13 RCTs, Peto OR 1.18, 95% CI 0.86 to 1.60). The four RCTs that reported cumulative pregnancy rates (266 women, Peto OR 1.58, 95% CI 1.11 to 2.25) (Day 2 to 3: 56.8%; Day 5 to 6: 46.3%) significantly favoured early cleavage. Embryo freezing rates (11 RCTs, 1729 women, Peto OR 2.88, 95% CI 2.35 to 3.51) and failure to transfer embryos (16 RCTs, 2459 women, OR 0.35, 95% CI 0.24 to 0.51) (Day 2 to 3: 3.4%; Day 5 to 6: 8.9%) favoured cleavage stage transfer. AUTHORS' CONCLUSIONS: This review provides evidence that there is a small significant difference in live birth rates in favour of blastocyst transfer (Day 5 to 6) compared to cleavage stage transfer (Day 2 to 3). However, cumulative clinical pregnancy rates from cleavage stage (derived from fresh and thaw cycles) resulted in higher clinical pregnancy rates than from blastocyst cycles. The most likely explanation for this is the higher rates of frozen embryos and lower failure to transfer rates per couple obtained from cleavage stage protocols. Future RCTs should report miscarriage, live birth and cumulative live birth rates to enable ART consumers and service providers to make well informed decisions on the best treatment option available.

Effect of Age and Morphology on Live Birth Rate After Cleavage Stage Embryo Transfer.

Accurate knowledge of the live birth rate for cleavage stage embryos is essential to determine an appropriate number of embryos to transfer at once. Results from previous studies lack details needed for practical use. This is a mathematical analysis and model building study of day 3 cleavage stage embryo transfers. A total of 996 embryos were transferred in 274 fresh and 83 frozen embryo transfers. Embryo morphology was divided into 4 groups based on number of cells and fragmentation percentage. Each embryo transfer was modeled as an equation equating the sum of the live birth rates of the transferred embryos to the number of live births that resulted. The least squares solution to the system of embryo transfer equations was determined using linear algebra. This analysis was repeated for ages 35 to 42 years old at oocyte retrieval. The best fit live birth rates per embryo in the age group centered on 35 years old were 29%, 13%, 10%, and 9% for embryos in the 8-cell with ≤ 5% fragmentation, 8-cell with > 5% fragmentation, 9-12 cell, and 6-7 cell groups, respectively. Cleavage stage embryos with fewer than 6 cells on day 3 had very low best fit live birth rates close to 0% at age 39 years and were excluded from the primary analysis to prevent overfitting. These live birth rates can be used with a simple embryo transfer model to predict rates of single and multiple gestation prior to a planned cleavage stage embryo transfer.

Cumulative ongoing pregnancy rate achieved with oocyte vitrification and cleavage stage transfer without embryo selection in a standard infertility program.

BACKGROUND: Recent advancement of minimum volume vitrification methods has resulted in a dramatic increase in the efficiency of the process. The aim of this study was to estimate the cumulative reproductive outcome of a cohort of infertile couples undergoing ICSI and oocyte vitrification in restrictive legal conditions, where only a limited number of oocytes could be inseminated per cycle and embryo selection and cryopreservation were forbidden. METHODS: In this prospective longitudinal cohort study, the cumulative ongoing pregnancy rates obtained by the insemination of fresh and vitrified oocytes from the same cohort were calculated as primary outcome measures. Moreover, the effect of basal and cycle characteristics on clinical outcomes were assessed. RESULTS: Between September 2008 and May 2009, 182 ICSI cycles were performed where oocyte vitrification was possible. A total of 104 first and 11 second oocyte warming cycles were then performed in non-pregnant patients of the same cohort. The overall ongoing pregnancy rates obtained in the fresh, and first and second warming cycles were 37.4, 25.0 and 27.3%, respectively. The overall cumulative ongoing clinical pregnancy rate observed per stimulation cycle was 53.3%. Maternal age was the only characteristic found to influence the reproductive outcome, with an inverse correlation between the age >40 and the ongoing pregnancy rates (P = 0.04, by Cox regression analysis). CONCLUSIONS: High cumulative ongoing pregnancy rates can be obtained with transfers of embryos derived from fresh and cryopreserved oocytes in a typical infertile population. Female age significantly affects outcomes in this system.

Successful transfer of day 10 horse embryos: influence of donor-recipient asynchrony on embryo development.

A total of 78 day 10 horse embryos were transferred non-surgically to recipient mares that had ovulated 9, 7, 6, 5, 4, 3, 2 or 1 day after (negative asynchrony), on the same day (synchronous), or 2 or 4 days before (positive asynchrony) the donor (n=6 or 8 mares per group). Pregnancy rates between 100% (6/6) and 63% (5/8) were seen in recipient mares that were between +2 and -6 days asynchronous. Embryo survival to the heartbeat stage declined in recipients that were -7 days asynchronous and no embryos survived in recipients that were -9 days asynchronous. Irrespective of uterine asynchrony, cessation of embryo mobility and fixation at the base of a uterine horn occurred when the conceptus was approximately 17 days old. Conceptus growth and development was slowed when embryos were placed in negatively asynchronous uteri. At the greatest degree of negative asynchrony at which embryos survived to the heartbeat stage, i.e. -7 and -6 days, development of the embryo proper and allantois was retarded. Luteostasis was achieved in recipient mares when day 10 embryos were transferred to recipient mares at any stage of asynchrony between -9 and +2 days with respect to the donor. These results indicate that in the horse, there is a wide window for establishment of pregnancy following embryo transfer to asynchronous recipients. Although progesterone priming of the uterus to a stage equivalent to that of the transferred embryo does not appear to be a prerequisite for embryo survival, it does nonetheless influence embryonic development.