In the present study, we found that the usable blastocyst formation rate (derived from the MG day 3 embryos), as well as the in vivo developmental potential of MG blastocysts was reduced significantly in the L group, compared to the H group. However, transfer of the MG day 3 embryos resulted in comparable clinical outcomes in the H and L groups. Our study support the strategy of transfer of MG day 3 embryos instead of extended culture for treatment of patients with LRTD3, particularly when few MG embryos were available on day 3.
A previous study showed that the number of good quality embryos available on day 3 over 3 is a strong predictive value for both pregnancy rate and implantation rate . Different from that study, we grouped the patients/cycles by rate of MG embryos on day 3 in the present study. This is a new perspective focusing on the capacity of overall MG embryos formation on day 3 rather than the absolute counts of MG day 3 embryos. The follicles within a patient promoted by controlled ovarian hyperstimulation share a common growth environment and genetic foundation. Therefore, we assumed that rate of MG day 3 embryos may better reflect the quality of follicles, rather than the absolute counts. To clarify this hypothesis, the developmental potential of MG embryos from patients with low and high RTD3 was compared. For controlling the fluctuation of rate of MG day 3 embryos, patients underwent a fresh cycle with 2 PN-zygotes = > 5 were included for analysis. Therefore patients included in this study were normal responders with relatively good prognosis.
In the present, we found that fresh cycles with transfer of MG day 3 embryos is distinct from fresh cycles with blastocyst transfer, and only a small fracture of fresh cycles have both transfer of MG day3 embryos and blastocysts in either the L or H groups. This was due to the transfer strategy. In our reproductive center, 2–4 MG day 3 embryos (if available) will be frozen or transferred, and the remaining will be further cultured. For embryo selection, MG blastocyst has the highest priority, followed by MG day 3 embryos, the next is MNG blastocyst, and the last is MNG day 3 embryos. Patients having frozen MG day 3 embryo transfer may indicate no available MG blastocysts or no extra MG embryos for further culture. The baseline characteristics of patients and consequent ovulation protocols were comparable between the L and H groups in patients with MG blastocyst and day 3 embryo transfer. However, we noticed that patients with transfer of MG blastocysts have higher ovarian reserve and ovarian response, compared to patients with transfer of MG day 3 embryos. This is reasonable. Because with similar RTD3, more number of eggs means more chance having MG blastocysts.
For patients with day 3 embryos transfer, although similar Gn use, the number of mature follicles and oocyte retrieved were significantly lower in the H group than that in the L group, indicating relatively low response of patients in the H group, compared to patients in the L group. However, the reduced oocytes retrieved in the H group were compensated by the HRTD3, resulting in more available MG embryos on day 3 and subsequent more MG day 3 embryos for expended culture. For patients with MG blastocyst transfer, the only difference between the two groups was the RTD3. HRTD3 resulted in more available MG embryos on day 3 and more MG day 3 embryos for expended culture in the H group. Taken all together, grouping by RTD3 didn’t introduce the obvious confounding factors regarding baseline characteristics of patients and cycles.
Usable blastocyst formation rate is a clinically important parameter reflecting the developmental potential of embryos. In the present study, we observed a higher of usable blastocyst formation rate in patients with transfer of blastocyst than in patients with transfer of MG day3 embryos. It has been reported that more dose of Gn use adversely affect the quality of eggs[11, 12]. Therefore, it is possible that high dose of Gn use in patients with MG day 3 embryo transfer deteriorates the oocyte quality. However, we found that usable blastocyst formation rate from MG day 3 embryos was reduced significantly in the L group, compared to the H group in either the MG day 3 embryo transfer or blastocyst transfer group, indicating that MG day 3 embryos from patients with low RTD3 showed decreased ability of being usable blastocyst in vitro, compared to patients with HRTD3.
The golden maker reflecting the quality of embryo is whether the embryo can result in a live birth . Although a decreased ability of being usable blastocyst by in vitro culture of the MG day 3 embryos from the patients with LRTD3 was observed, how about the in vivo developmental potential of MG blastocysts from the patients with LRTD3? In the present study, we found that not only clinical pregnancy rate and livebirth rate, but also implantation rate and livebirth rate per embryo were significantly higher in the H group than the L group. It is know that female age and BMI, number of embryos transferred and semen DFI have profound influences on the clinical outcomes of in vitro fertilization [13–19]. In addition, we found that there was a trend towards decreased male age in the H group, compared with L group. Although conflicting results were reported regarding the effect of male age on the clinical outcomes of in vitro fertilization, the factor of male age should be also considered [20, 21]. After adjusted for age of male or female, number of embryos transferred, female BMI and sperm DFI, we found that the embryos from the H group was positively associated with the clinical pregnancy and livebirth. Therefore, although similar morphological appearance (they were both MG blastocyst), the MG blastocysts from patients with LRTD3 showed reduced in vivo developmental potential, compared to that from patients with HRTD3.
To our surprise, transfer of MG day 3 embryos resulted in similar clinical outcomes in the H and L groups. After adjusted for factors including male or female age, number of embryos transferred, female BMI and sperm DFI, type of embryo transfer (fresh or frozen transfer), we found that embryos from the H group were not associated with the clinical pregnancy and livebirth. This was quite different from the results from blastocyst transfer. In addition, this result was also paradox with the finding that a decreased useful blastocyst formation rate was observed in the L group for group of MG day 3 embryo transfer. One reasonable explanation is the difference of developmental potential of MG day 3 embryos resulted from in vitro and in vivo. It has been proposed that the day 3 embryos which can’t develop into blastocysts in vitro may lead to live births in vivo . Therefore, we deduced that that MG day 3 embryos from patients with LRTD3 is more sensitive to in vitro culture but direct transfer, resulting in decreased useable blastocyst formation rate and similar clinical outcomes of direct transfer of MG day 3 embryos, compared with patients with high LRTD3.
Data from direct MG day 3 embryo transfer showed that patients with LRTD3 had limited MG day 3 embryos with a decreased rate of useful blastocyst formation rate compared to patients with HRTD3. However, direct transfer of MG day 3 embryos from patients with LRTD3 may achieve similar clinical pregnancy outcomes as patients with HRTD3. Data from transfer of blastocyst showed that rate of blastocyst formation is significantly reduced in patients with LRTD3. More importantly, transfer of MG blastocyst resulted in poorer clinical pregnancy outcomes in patients with LRTD3 than patients with HRTD3. Therefore, we concluded that the quality of MG day 3 embryos or blastocysts from patients with LRTD3 was reduced compared to patients with HRTD3. However, it seems that direct transfer of MG day 3 embryos effectively avoid the flaw of poor quality of MG day 3 embryos from patients with RTD3.
The main limitation of this study is its nature of the retrospective study. We noticed that transfer of MG day 3 embryos and transfer of blastocysts represented relatively separate population in either the L group or the H group. Population of transfer of blastocyst had higher response to Gn and less Gn use, more oocytes retrieved and higher blastocyst formation rate, compared to patients with transfer of MG day 3 embryos. A concern is that whether it is appropriate that results from separate populations were integrated for analysis. This study was a comparison study, did not provide the direct evidence. Furthermore, the data included for analysis were from single center and the cases included in the present study is limited, therefore, the indications from this study need to be confirmed by randomized control trial.