Human Failed-matured Oocytes Vitrification Followed By In Vitro Maturation

PartI Artificial Oocyte Activation and Human Failed-matured Oocyte Vitrification Followed by In Vitro MaturationObjective⑴To evaluate the effect of in vitro maturation(IVM) on human failed- matured oocytes from controlled ovarian hyperstimulation(COH)cycles;⑵To investigate the effect of oocytes activation in fresh or frozen -thawed human immature oocytes followed by IVM.Methods A Total of 500 failed-matured oocytes (GV and MI stages)derived from 154 patients who underwent ICSI cycles were randomly divided into two groups:fresh groupand vitrification group (GVgroup and MI group). fresh group and vitrification group were randomly divided into two groups: the controlled group and artificial oocyte activation group(AOA). Matured oocytes from fresh and freeze-thawed failed-matured oocytes followed by IVM were inseminated by ICSI. The injected oocytes in the control group were cultured in Cleavage. The AOA group oocytes were activated by exposure to 7% anhydrous alcohol for 6 minutes, then were cultured in Cleavage. The rates of fertilization and early embryonic development were compared in different groups.Result⑴In 226 fresh failed-matured oocytes followed by IVM, the maturation rate , the fertilization rate, the cleavage rate ,the high-quality embryo formation rate, the blastocyst formation rate and high-quality blastocyst formation rate were83.2% (188/ 226) ,78.7% (148 /188), 95.9% (142 /148), 19.7% (28 /142), 12.7% (18 /142) and55.6 % (10 /18), respectively. Cleavage and high-quality blastocyst formation rates were significantly higher in the AOA group than in the control group ( P <0.05).The maturation rates ,fertilization rates, high-quality embryo formation rates and blastocyst formation rates had no significant differences between the two groups (p > 0.05)⑵In MI vitrification group, the high-quality embryo formation rates and blastocyst formation rates were significantly higher in the AOA group than in the control group ( P < 0.01). The maturation rates , fertilization rates and cleavage rates had no significant differences between the two groups ( P > 0.05). In GV vitrification group,the high-quality embryo formation rate was significantly higher in the AOA group than in the control group ( P < 0.05). The maturation rate , fertilization rates and cleavage rates had no significant differences between the two groups ( P > 0.05).⑶Regardless of the maturation stage (GV+MI) ,vitrification group without AOA gave significant lower cleavage rate and high-quality embryo formation rate than fresh group without AOA(P < 0.01);Vitrification group with AOA gave significant lower cleavage rate than fresh group without AOA( P < 0.01) and gave significant higher high-quality embryo formation rate than fresh group without AOA( P < 0.05).Conclusions⑴immature human oocytes derived from COH cycles by IVM could havegood fertilization and early embryonic development ,so these immature oocytes should be fully utilized.⑵AOA on fresh and Freeze-thawed human immature oocytes may beadvantage to early embryonic development.⑶The cryodamage in vitrification on human immature oocytes influence early embryonic development, which may be partially repaired by AOA. PartⅡStudy on the Effect of Trehalose on the Vitrification of Failed-matured OocytesObjective To determine the effectiveness of trehalose as an intracellular cryoprotectant for the vitrification of immature human oocytes.Methods A Total of 296 failed-matured oocytes (GVandMI stages)derived from COH cycleswere randomly divided into two groups:control group (no trehalose) and trehalose vitrification group. Immature oocytes in control group were vitrified without trehalose. Trehalose was introduced into oocytes by extracellular penetration in trehalose vitrification group and vitrified with the Cryoleaf . Freeze-thawed immature oocytes followed by IVM in both groups were inseminated by ICSI . The survival rates ,maturation rates ,fertilization rates and early embryonic development were compared in two groups.Result⑴Cleavage and high-quality embryo formation rates were significantly higher in the MI trehalose vitrification group than in the control group ( P <0.05). The maturation rates ,fertilization rates were significantly lower in the MI trehalose vitrification group than in the control group ( P <0.05) .The survival rates had no significant differences between the two groups (p >0.05)⑵The maturation rates were significantly lower in the GV trehalose vitrification group than in the control group ( P <0.01) . The survival rates had no significant differences between the two groups (p >0.05). Mature oocytes from GV trehalose vitrification group get no normal fertilization and embryos . Conclusions⑴Trehalose as intracellular cryoprotectants will help to improve early embryonic development of freeze-thawed MI oocytes , However, process of extracellularpenetration to make trehalose into MI oocytes would produce damaging to MI oocytes ;Vitrification method in this experiment is not suitable for the GV oocytes⑵We should explore more suitable way to make trehalose into immature oocytes to play a protective role in vitrification in order to enhance freezing efficiency of immature oocytes.