A total of 2531 blastocysts from 839 PGT-A cycles were obtained in the present study. There were 269 PGT-A cycles with both day 5 and day 6 blastocysts, 422 cycles with day 5 blastocysts alone, and 148 cycles with day 6 blastocysts only. The euploid rate of day 5 blastocysts seemed to be significantly higher than that of day 6 blastocysts, either from the same OS cycles (49.9% vs 35.7%, P < 0.001) or from different cycles (48.2% vs 27.8%, P < 0.001). However, after single euploid embryo transfer, the clinical outcomes, including clinical pregnancy rate, miscarriage rate, ongoing pregnancy rate and live birth rate, were comparable between day 5 and day 6 blastocysts, no matter whether they were from the same OS cycles or not.
Baseline characteristics
The baseline characteristics of PGT-A cycles are shown in Table 1. The ovarian reserve varied significantly among the three groups. Among them, female patients in cycles with day 6 blastocysts only seemed to have the poorest ovarian reserve, with the average age of 36.63 ± 4.81 years and the AMH concentration of 2.33 ± 1.64 ng/mL (P < 0.001). Accordingly, the percentage of AMA as the PGT-A indication (35.8%) was the greatest in those cycles, compared with those in the other two groups (26.4% for both day 5 and day 6 blastocysts group, and 26.3% for day 5 blastocysts alone group, P = 0.02). Besides, in cycles with only day 6 blastocysts, the average male age was the oldest (38.54 ± 6.30 years), compared with that of 35.11 ± 5.67 years in cycles with both day 5 and day 6 blastocysts, and 36.52 ± 6.15 years in cycles with only day 5 blastocysts (P < 0.001). The other basic characteristics regarding female body mass index (BMI), basal FSH, LH, E2 concentration, semen volume, sperm count, total motility, and percentages of progressive and slow progressive spermatozoa were comparable among the three study groups.
As confirmed by the multivariate logistic regression model, women with a higher AMH concentration inclined to have blastocyst formation by day 5 (adjusted OR = 1.130, 95% CI: 1.080-1.182, P<0.001), after adjusting for female age and male age (Table 4).
OS characteristics
As for OS cycles, those with day 6 blastocysts alone mostly received the mild stimulation protocol (43.2%), compared with the other two groups (20.8% for both day 5 and day 6 blastocysts group, 37.9% for only day 5 blastocysts group, P < 0.001). Meanwhile, the average stimulation duration was the shortest in those cycles (7.86 ± 1.92 days) relative to the other two groups (8.31 ± 1.13 days for both day 5 and day 6 blastocysts group, 7.97 ± 1.67 days for only day 5 blastocysts group, P < 0.01). The initial Gn dose and total Gn dose were comparable among the three groups. On the stimulation day, cycles with only day 6 blastocysts had the lowest E2 concentration (1647.97 ± 1165.22 pg/mL) compared with the other two groups (2737.98 ± 1709.63 pg/mL for both day 5 and day 6 blastocysts group, 2062.09 ± 1444.06 pg/mL for only day 5 blastocysts group, P < 0.001). But the LH concentration (5.23 ± 4.22 IU/L) was the highest in cycles with only day 6 blastocysts, compared with the other two groups (3.87 ± 3.17 IU/L for both day 5 and day 6 blastocysts group, 4.75 ± 3.74 IU/L for only day 5 blastocysts group, P < 0.001). Finally, in cycles with both day 5 and day 6 blastocysts, the average numbers of oocytes retrieved, MII oocytes, oocytes fertilized (2PN), fertilized oocytes that cleaved, and blastocysts for biopsy were 12.20 ± 6.69, 10.57 ± 5.98, 8.29 ± 4.79, 8.07 ± 4.64 and 4.31 ± 2.14, respectively; those in cycles with only day 5 blastocysts were 8.81 ± 6.61, 7.41 ± 5.79, 5.72 ± 4.69, 5.62 ± 4.59 and 2.77 ± 2.30, respectively; and those in cycles with only day 6 blastocysts were 6.95 ± 5.95, 5.66 ± 4.50, 4.34 ± 3.48, 4.23 ± 3.36 and 1.41 ± 0.61, respectively. Differences among the three groups were statistically significant (P < 0.001) (Table 2).
Outcome comparisons
The comparisons between day 5 and day 6 blastocysts regarding PGT-A outcomes and clinical outcomes are presented in Table 3. It was found that day 5 blastocysts had better euploid status than day 6 blastocysts, no matter whether they were from the same OS cycles or from different cycles (P < 0.001). Specifically, the euploid rate, aneuploid rate and mosaic rate of day 5 blastocysts were 49.9%, 31.1% and 19.0%, respectively; while those of day 6 blastocysts from the same OS cycles were 35.7%, 41.3% and 23.0%, respectively. By contrast, the euploid rate, aneuploid rate and mosaic rate of day 5 blastocysts were 48.2%, 37.6% and 14.2%, respectively; while those of day 6 blastocysts from different OS cycles were 27.8%, 51.2% and 21.0%, respectively (Figure 2A). The clinical outcomes were comparable between single day 5 blastocyst transfer and single day 6 blastocyst transfer, either from the same OS cycles or different OS cycles. For single day 5 blastocyst transfer, the clinical pregnancy rate, biochemical pregnancy rate, miscarriage rate, ongoing pregnancy rate and live birth rate were 59%,7.7%, 10.4%, 73.9% and 15.7%, respectively; while those for single day 6 blastocyst transfer from the same OS cycles were 44.2%, 7.0%, 21.0%, 63.2% and 15.8%, respectively. In comparison, the clinical pregnancy rate, biochemical pregnancy rate, miscarriage rate, ongoing pregnancy rate and live birth rate were 62%, 8.1%, 14.4%, 72.5% and 13.1%, respectively, for single day 5 blastocyst transfer; while those for single day 6 blastocyst transfer from different OS cycles were 43.9%, 7.3%, 11.1%, 66.7% and 22.2%, respectively (Figure 2B).
Further, multivariate logistic regression analysis confirmed the higher euploid rate for younger female age (adjusted OR = 0.917, 95% CI: 0.892-0.944, P < 0.001) and day 5 blastocysts (adjusted OR = 1.735, 95% CI: 1.415-2.127, P < 0.001), after adjusting for the significantly different basal characteristics (Table 4).