To summarize, this study described the correlations between blastocyst morphologic grades and their implantation rates. Our findings indicated that good-quality euploid blastocysts had higher implantation rates than poor-quality blastocysts, although this increase was only apparent for patients under 35 years old. On the same day as TE biopsy, morphologic grading was associated with the implantation rates of euploid blastocysts. Day 5 euploid blastocysts showed no significant difference in implantation potential compared with similarly graded day 6 euploid blastocysts. Unlike the implantation rates, however, the early spontaneous abortion rate was not correlated with blastocyst grading.
In this study, TE biopsy was performed at the blastocyst stage and NGS-based PGT-A was conducted to select euploid blastocysts for single vitrified-warmed blastocyst transfer. NGS has grown in popularity due to its ability to identify unbalanced translocations, segmental aneuploidies, some triploidies[6] and lower levels of mosaicism than other techniques[13]. Some research has found improved pregnancy outcomes and reduced spontaneous abortion rates with NGS due to the exclusion of these abnormal embryos[14]. However, the findings are not unanimous. In a multicenter randomized clinical trial[15] that performed PGT-A versus morphology as a selection criterion for single frozen-thawed embryo transfer in good-prognosis patients, PGT-A did not improve overall pregnancy outcomes for the cohort of women, as analyzed per embryo transfer or per intention to treat. It is possible that the detrimental effect of the biopsy pre-vitrification on the embryo viability may outweigh the benefit of PGT-A[16].
Despite the advances made with PGT-A, a considerable proportion of euploid embryos still fail to implant due to as yet unknown etiologies. In our study, the priority for euploid blastocyst selection for transfer was based on blastocyst morphology. Our data confirm that good-quality euploid blastocysts were correlated with higher implantation rates than poor-quality blastocysts, which partially explains some of the failed euploid cycles. Our data are consistent with previously reported data by Irani et al., who demonstrated that excellent-quality embryos yielded a statistically significantly higher ongoing pregnancy rate than poor-quality embryos (84.2% vs. 35.8%; adjusted OR, 11.0; 95% CI, 3.8–32.1) and average-quality embryos (84.2% vs. 55.8%; adjusted OR, 4.8; 95% CI, 1.7–13.3)[17]. The difference is that whereas the present study excluded the potential effects of embryo‑embryo interactions by only analyzing single‑blastocyst cases, Irani et al. analyzed cycles in which either one or two blastocysts of the same overall quality were transferred. Blastocyst quality is assessed based on blastocoel expansion, appearance of TE and appearance of ICM. TE grade has been shown to be a useful predictor of ongoing pregnancy rate for euploid embryos, and the critical role of the TE in mediating correct embryo implantation has been well established[1].
Preventing miscarriage and maximizing the chance of implantation by SET are of utmost importance in euploid blastocysts. Here, we provide evidence that the early spontaneous abortion rate is not correlated with blastocyst grading. Barash et al. demonstrated an association of euploidy rates with morphologic characteristics of blastocysts. Embryos with an advanced degree of blastocyst expansion and well formed ICM/TE had higher chances of being euploid (χ2 = 10.73, P < 0.05 and χ2 = 4.34, P < 0.05, respectively)[18]. Elsewhere, poor-quality blastocysts showed a higher miscarriage rate per clinical pregnancy (36.4%) than non-poor-quality blastocysts (13.9%) after euploid SET[19]. However, once the risk of miscarriage is overcome, poor-quality blastocysts go to term apparently without greater obstetrical or perinatal risks[20].
Although aneuploidy is the most significant determinant of cycle outcome in the older population, an age-related decline in implantation occurs in euploid embryos, supporting the view that factors other than chromosome segregation errors play a role in age-related fertility decline[21]. During a 5-year study period, Cimadomo collected various data to outline the clinical significance of poor-quality blastocysts during an ICSI cycle with PGT-A. The only feature that showed an association with the prevalence of poor-quality blastocysts was the maternal age at oocyte retrieval[19]. These results contradict previous findings in which the differences in outcomes based on age were nonsignificant after adjusting for embryo morphology[22]. After controlling for maternal age, here we provide evidence that good-quality euploid blastocysts have a higher chance of implanting than poor-quality blastocysts in women under 35 years; however, we found no differences in implantation rate based on blastocyst morphology in women over 35. The most likely explanation is that for women aged 35 or older, the most important feature associated with the euploid blastocyst implantation rate is the maternal age at oocyte retrieval rather than blastocyst morphology, which highlights the competence of poor-quality euploid embryos in women of advanced maternal age. This conjecture is supported by Gonzalez’s finding that in women of advanced maternal age following euploid SET after PGT-A via NGS, clinical outcomes were not significantly affected by the embryo morphology[18].
Our study demonstrated that good-quality euploid blastocysts yield higher implantation rates than same-day poor-quality blastocysts. Both day 5 and day 6 euploid blastocysts with good quality showed increased implantation ability, and no significant difference in implantation potential was found between similarly graded blastocysts from days 5 and 6. Shapiro et al.[23] found that day 5 blastocysts were associated with higher clinical pregnancy rates than their day 6 counterparts in fresh cycles, but had similar outcomes in FET cycles. While day 5 and day 6 embryos may have similar implantation potentials, the difference in the success rates observed in fresh cycles is essentially related to the suboptimal embryo-endometrial synchrony of day 6 blastocysts. Contradicting our findings, Irani et al., in a study correlating blastocyst development rate with pregnancy outcomes after euploid SET, showed that day 5 euploid blastocysts yielded higher implantation rates and live birth rates than similarly graded day 6 blastocysts[9]. Blastocyst grading and the speed of embryo development to the blastocyst stage may reflect the metabolic health of the embryo. Irani et al. confirmed that day 5 blastocysts that were transferred were more likely to be of good quality and less likely to be classified as poor quality, while the higher implantation rate of day 5 blastocysts was mainly due to there being more good-quality blastocysts on day 5 than day 6. In any event, morphologic grading is correlated with implantation potential, which is why it was chosen as the main criterion used to select the embryos for transfer in this study.
In this regard, it is vital to provide patients with specific counseling focused on the evidence reported in the literature so they can make an informed choice.
This study has several strengths. First, it was specifically designed to find a strategy for selecting the best euploid embryo in patients who have multiple euploid embryos. Second, the chosen age range allowed the results to be clearly stratified by age. Third, we evaluated the role of the blastocyst morphology along with blastocyst development in euploid embryo selection. Fourth, as this study involved single euploid blastocyst transfer, all of the embryos were subjected to the same NGS platform protocols with uniform analysis of the PGT-A results. Our embryologists also used a consistent scoring system consisting of a number of standardized transfer parameters.
This study also has several limitations. First, being a retrospective study with a relatively small sample size and data collection from a single center, some bias was inevitable. Second, implantation rate, not live birth rate, was the primary outcome. Third, patients over 40 years of age were not included because of the risk of not having access to euploid blastocyst transplantation. This may affect the applicability of the clinical outcomes to older patients with different quality blastocysts. Fourth, although blastocyst morphology and blastocyst development were both explored, the study was not capable of identifying specifically which embryos were most able to be implanted successfully.