Births Resulting from Oocyte Cryopreservation Using a Slow Freezing Protocol with Propanediol and Sucrose

Abstract

The human mature oocyte is particularly sensitive to cooling and low temperatures in addition to freeze-thaw damage. The efficiency of oocyte cryopreservation including the pregnancy outcome is still low. The aim of our study is to briefly introduce our preliminary clinical results achieved with oocyte cryopreservation (CP). Our work focused on the use of a slow cooling procedure using the cryoprotectants propanediol (1.5 M) and sucrose (0.3 M). Following a short incubation of 4-6 hours thawed oocytes were injected with a single sperm (ICSI) and fertilization was assessed 12-16 hours later. Laser assisted hatching (LAH) was performed on all transferred embryos and embryo transfer (ET) was carried out 48-72 hours after ICSI. One-hundred and ten eggs were thawed and a survival rate of 76% (84/110) was obtained. Of the 84 oocytes which survived, 64 subsequently fertilized (64/84; 76%) following ICSI and on the following day 55 of those had cleaved (55/64; 86%). Fifty-two embryos were transferred in 29 patients (1.8 embryo/patient), and 7 (7/29; 24%) resulted in clinical pregnancy (1 twin pregnancy). One of the pregnancies encountered first trimester abortion (1/7; 14%). Implantation rate of 15.4% per embryo transferred (8/52) and 7.3% per egg thawed (8/110) were obtained. In all cases, chorion biopsy was performed and chromosomal anomalities were not detected. Our results provide further evidence that the procedure can be applied safely and with good success in clinical assisted reproduction. However, more work is needed since the survival and implantation rate should be improved.

Implantation rates of embryos generated from slow cooled human oocytes from young women are comparable to those of fresh and frozen embryos from the same age group

“…The survival rate achieved in the present study is similar to that previously observed with the 0.3 M sucrose method [7,15,16,[31][32][33][34][35][36][37] and the 0.2 M sucrose method [17,19,21]. In some of the above studies only oocytes with optimal morphology were cryopreserved [37,38] which is not the case in the present study.…”

“…This may be a consequence of the slower embryo development [13] and organelle damage [14] reported for this method. Although parameters such as reduced time between collection and cryopreservation [15] and assisted hatching of subsequent embryos [16] may result in some improvement, the clinical efficiency has remained low (reviewed in [11]). In contrast, modifying the sucrose concentration to 0.2 M during dehydration also increased survival [17][18][19][20] but implantation did not appear to be impaired [19,21] suggesting more potential [11] with this modification, although a reduced implantation rate relative to fresh oocytes has also been reported recently [20].…”

Slow controlled-rate freezing of human in vitro matured oocytes: effects on maturation rate and kinetics and parthenogenetic activation

“…In our study, after slow controlled-rate freezing, an overall survival rate of 70% was obtained, which is consistent with other published reports (21)(22)(23)(24)(25). Because most of the donated human oocytes were immature, we compared their freezing at different developmental stages of maturation to verify the optimal stage for cryopreservation.…”

Quality and functionality of human ovarian tissue after cryopreservation using an original slow freezing procedure

“…For our freezing protocol, we chose a combination of PrOH and raffinose which are respectively penetrating and non-penetrating cryoprotectants. The use of PrOH in clinical freezing protocols gives rise to the birth of healthy babies following cryopreservation of both embryos and mature oocytes [3,26,44]. Human ovarian tissue cryopreserved with PrOH as cryoprotectant exhibits a satisfactory morphological preservation of follicles in comparison with fresh tissue [11,14].…”