利用孤雌激活评估ICSI后未成熟卵母细胞的利用价值
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摘要
目的本研究以ICSI后未成熟卵母细胞为研究对象,分析比较卵母细胞不同发育阶段冷冻对其后续效果的影响,评估ICSI后未成熟卵母细胞的利用价值。方法未成熟卵母细胞直接成熟培养(新鲜组)与玻璃化冷冻后成熟培养(冷冻组)的成熟率,并利用孤雌激活的方法比较卵母细胞的发育潜力。结果发现新鲜组与冷冻组体外培养卵母细胞成熟率、受精率、卵裂率、优质胚胎率和囊胚率均无差异(P>0.05)。但两组的GV期卵母细胞成熟率低于MI期(P<0.05),且冷冻组的GV期卵母细胞受精率低于MI期(P<0.05)。不过裸卵体外成熟培养效果欠佳,特别是对GV期卵母细胞,体外成熟培养后的卵母细胞发育潜力低下,无囊胚形成。结论ICSI后未成熟卵母细胞的冷冻对卵母细胞的发育潜力没有明显影响,但体外成熟培养的卵母细胞发育潜力低下,有待进一步提高体外成熟培养技术。
Objective In this study, the immature oocytes after ICSI were used to analyze the effects of freezing on the subsequent development of oocytes at different developmental stages, and to evaluate the utilization value of immature oocytes after ICSI(intracytoplasmic sperm injection). Methods The immature oocytes was directly cultured(fresh group) and matured after vitrification(frozen group), and the development potential of the oocytes was compared by parthenogenetic activation. Results There was no significant difference in the oocyte maturation rate, fertilization rate, embryo cleavage rate, high quality embryo rate and blastocyst rate between the fresh and frozen groups(P>0.05). However, the maturation rate of GV oocytes in the two groups was lower than MI oocytes(P<0.05), and the fertilization rate of GV oocytes in the frozen group was lower than MI oocytes(P<0.05). However, the in vitro maturation of naked oocytes was not effective, especially for GV oocytes, the oocyte development potential after in vitro maturation was low, there was no blastocyst formation. Conclusion The freezing of immature oocytes after ICSI has no significant effect on the development potential, but the development potential of naked oocytes matured in vitro was low, and the in vitro maturation culture technology of naked oocyte needs to be further improved.
Objective In this study, the immature oocytes after ICSI were used to analyze the effects of freezing on the subsequent development of oocytes at different developmental stages, and to evaluate the utilization value of immature oocytes after ICSI(intracytoplasmic sperm injection). Methods The immature oocytes was directly cultured(fresh group) and matured after vitrification(frozen group), and the development potential of the oocytes was compared by parthenogenetic activation. Results There was no significant difference in the oocyte maturation rate, fertilization rate, embryo cleavage rate, high quality embryo rate and blastocyst rate between the fresh and frozen groups(P>0.05). However, the maturation rate of GV oocytes in the two groups was lower than MI oocytes(P<0.05), and the fertilization rate of GV oocytes in the frozen group was lower than MI oocytes(P<0.05). However, the in vitro maturation of naked oocytes was not effective, especially for GV oocytes, the oocyte development potential after in vitro maturation was low, there was no blastocyst formation. Conclusion The freezing of immature oocytes after ICSI has no significant effect on the development potential, but the development potential of naked oocytes matured in vitro was low, and the in vitro maturation culture technology of naked oocyte needs to be further improved.
引文
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[23] NIKIFORAKI D,VANDEN MEERSCHAUT F,QIAN C,et al.Oocyte cryopreservation and in vitro culture affect calcium signalling during human fertilization [J].Hum Reprod,2014,29(1):29- 40.
[2] YOON T K,CHUNG H M,LIM J M,et al.Pregnancy and delivery of healthy infants developed from vitrified oocytes in a stimulated in vitro fertilization-embryo transfer program [J].Fertil Steril,2000,74(1):180-181.
[3] CAO Y X,XING Q,LI L,et al.Comparison of survival and embryonic development in human oocytes cryopreserved by slow-freezing and vitrification [J].Fertil Steril,2009,92(4):1306-1311.
[4] BAKA S G,TOTH T L,VEECK L L,et al.Evaluation of the spindle apparatus of in-vitro matured human oocytes following cryopreservation [J].Hum Reprod,1995,10(7):1816-1820.
[5] BOISO I,MARTI M,SANTALO J,et al.A confocal microscopy analysis of the spindle and chromosome configurations of human oocytes cryopreserved at the germinal vesicle and metaphase II stage [J].Hum Reprod,2002,17(7):1885-1891.
[6] MICHAEL J,TUCKER,GRAHAM WRIGHT,et al.Birth after cryopreservation of immature oocytes with subsequent in vitro maturation [J].Fertil Sterilt,1998,70(3):578-579.
[7] CHIAN R C,GILBERT L,JACK Y J,et al.Live birth after vitrification of in vitro matured human Oocytes [J].Fertil Steril,2009,91(2):372-376.
[8] EPPIG J J,SCHULTZ R M,O’BRIEN M,et al.Relationship between the developmental programs controlling nuclear and cytoplasmic maturation of mouse oocytes [J].Dev Biol,1994,164(1):1-9.
[9] EPPIG J J.Coordination of nuclear and cytoplasmic oocyte maturation in eutherian mammals [J].Reprod Fertil Dev,1996,8(4):485- 489.
[10] KRISHER R L.The effect of oocyte quality on development [J].J Anim Sci,2004,82(E.Suppl.):E14-23.
[11] WATSON A J.Oocyte cytoplasmic maturation:a key mediator of oocyte and embryo developmental [J].J Anim Sci,2007,85(E.Suppl.):e1-e3.
[12] STERN S,RAYYIS A,KENNEDY J F.Incorporation of amino acids during maturation in vitro by the mouse oocyte:effect of puromycin on protein synthesis [J].Biol Reprod,1972,7(3):341-346.
[13] SCHULTZ R M,WASSARMAN P M.Specific changes in the pattern of protein synthesis during meiotic maturation of mammalian oocytes in vitro [J].Proc Natl Acad Sci USA,1977,74(2):538-541.
[14] SIRARD M A,FIRST N L.In vitro inhibition of oocyte nuclear maturation in the bovine [J].Biol Reprod,1988,39(2):229-234.
[15] SIRARD M A,FLORMAN H M,LEIBFRIED-RUTLEDGE M L,et al.Timing of nuclear progression and protein synthesis necessary for meiotic maturation of bovine oocytes [J].Biol Reprod,1989,40(6):1257-1263.
[16] CLARKE H J,MASUI Y.The induction of reversible and irreversible chromosome decondensation by protein synthesis inhibition during meiotic maturation of mouse oocytes [J].Dev Biol,1983,97(2):291-301.
[17] GERHART J,WU M,KIRSCHNER M.Cell cycle dynamics of an M-phase-specific cytoplasmic factor in Xenopus laevis oocytes and eggs [J].J Cell Biol,1984,98(4):1247-1255.
[18] GIFFORD D J,FLEETHAM J A,MAHADEVAN M M,et al.Protein synthesis in mature human oocytes [J].Gamete Res,1987,18(2):97-107.
[19] SCHULTZ G A,GIFFORD D J,MAHADEVAN M M,et al.Protein synthetic patterns in immature and mature human oocytes [J].Ann NYAcad Sci,1988,541(1):237-247.
[20] GOUD P T,GOUD A P,QIAN C,et al.In-vitro maturation of human germinal vesicle stage oocytes:role of cumulus cells and epidermal growth factor in the culture medium [J],1998,13(6):1638-1644.
[21] JOHNSON J E,LEE HIGDON III H,BOONE W R.Effect of human granulosa cell co-culture using standard culture media on the maturation and fertilization potential of immature human oocytes [J].Fertil Steril,2008,90(5):1674-1679.
[22] DEL MASTRO L,VENTURINI M,SERTOLI M R,et al.Amenorrhea induced by adjuvant chemotherapy in early breast cancer patients:prognostic role and clinical implications [J].Breast Cancer Res Treat,1997,43(2):183-190.
[23] NIKIFORAKI D,VANDEN MEERSCHAUT F,QIAN C,et al.Oocyte cryopreservation and in vitro culture affect calcium signalling during human fertilization [J].Hum Reprod,2014,29(1):29- 40.