Single FBT has been a widely accepted by increasing number of reproductive centers, and it is important to choose the blastocyst with best development potential to transfer. As known, endometrium receptivity is critical for both implantation and gestation [24]. In our study, only the FBT cycles using hormone replacement treatment for endometrial preparation were included, which enhanced embryo-endometrium synchronization. We conducted a matched-cohort study to investigate the embryonic factor of blastocyst development potential, including the day of blastocyst expansion and embryo quality. To the best of knowledge, this is the first matched-cohort analysis to compare the pregnancy potential of frozen-thawed blastocyst cryopreserved on day 5 versus day 6.
The comparison of pregnancy outcomes of FBT cycles showed that the CPR and LBR of day 5 were significantly higher than day 6, which consisted with most previous study [17–20, 25, 26]. Furthermore, the FBT cycles of day 5 and day 6 were divided into two subgroups according to the embryo quality (good or poor). The significant advantage of FBT cycles on day 5 was still maintained after accounting the embryo quality in our present study. However, whether the blastocyst with good quality cryopreserving on day 5 had superior pregnancy potential was still under debate. Some original articles showed higher pregnancy potential of day 5 blastocyst than day 6 [17, 19], while others exhibited equal pregnancy potential between day 5 and day 6 [22, 26]. Only a few research have performed a comparison of the pregnancy outcomes of FBT cycles with poor embryo quality, and both suggested the superiority of blastocyst on day 5 [19, 26].
Besides the original articles, two systematic reviews also analyzed the different outcomes of FBT cycles on day 5 and day 6 [20, 25]. On the whole, both of them approved of the superiority of blastocyst on day 5. However, controversy on the pregnancy outcome of blastocyst cryopreserved on day 5 versus day 6 with the same quality still exited. This inconsistent result partly caused by the cryopreservation method of the studies they included. Slow-freezing method was more widely used than vitrification ten years ago or even earlier. This hypothesis was also confirmed by Bourdon et al., who suggested that calculated subgroup relative risk was1.27 (95% CI: 1.16–1.39, P < 0.001) for vitrified/warmed blastocyst transfer, and 1.15 (95% CI: 0.93–1.41, P = 0.20) for slow frozen/thawed blastocyst transfer [20]. Our results seemed more consistent with the review of Bourdon et al., which included more newly published articles.
Based on the literatures and our present results, we conclude that the FBT cycles of day 5 showed higher pregnancy potential than day 6 with the same embryo quality. However, an interesting result was also found in our study. The good-quality blastocyst of day 6 showed higher pregnancy potential than poor-quality blastocyst of day 5, which indicate embryo quality was an important factor for successful gestation. Limited studies have reported the specific data we concerning, even showed conflict results. The study of Yang et al. showed higher CPR of blastocyst on day 6 with good quality than day5 with poor quality [26], which was similar with our results. However, the study of Ferreux et al. supported the superiority of blastocyst on day 5 even with poor quality [19]. These conflict results may attribute to the heterogeneity of the number in FBT cycles. We only analyzed the single FBT cycles, while some studies also included the frozen-thawed cycles of transferring two blastocysts. In the routine operation, clinicians tended to transfer two blastocysts if the embryo quality were poor. This may partly increase the successful gestation of blastocyst with poor quality. Additionally, the embryo culture strategy such as the number of cleavage embryo transfer, whether cleavage embryo cryopreservation and blastocyst transfer were performed in fresh cycles may also affect the clinical outcomes of following FBT cycles.
Besides the day of blastocyst expansion, the results of multivariate logistic regression analysis showed that maternal age at retrieval, AMH, endometrial thickness and embryo quality was independently correlated with CPR and LBR. It was indisputable that maternal age and embryo quality were closely related to successful gestation. Age had a negative effect on pregnancy chance [27–30], while morphological embryo quality was positively associated with the pregnancy outcomes [6, 7], and our data also reinforced it (Fig. 1). AMH is also an independent predictor of live birth outcome [31, 32]. In addition, some studies reported that increased endometrial thickness is associated with better pregnancy outcome [33, 34].
Furthermore, the neonatal outcomes were also analyzed in our study. No significant difference was observed in the birth weight, gestational age or gender ratio between the FBT cycles of day 5 and day 6. Our results were consistent with those reported by Wang et al. [35].
As a retrospective analysis, there some unavoidable limitations in our study. Such as the difference of operators, culture system (Vitrolife or Cook) may produce biases. But the operators and culture medium were fixed during the same period, which were exactly the same in the FBT cycles of day 5 and day 6.