人IVF周期中卵母细胞透明带及发育潜力相关影响因素的研究
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摘要
在体外受精(IVF)和单精注射(ICSI)中,选择发育潜能较高(能发育成有附植潜力的胚胎)的卵母细胞对于改善辅助生殖治疗结局至关重要。近年来,人们发现透明带特性可作为一项预测卵母细胞质量的重要指标,偏光显微系统的应用使得通过非损伤研究透明带特性来评价卵母细胞／胚胎的质量成为了可能。然而,以往的研究结果都是集中于ICSI周期。本研究通过IVF周期的追溯性分析,系统研究了透明带双折射率(ZB)与卵母细胞、胚胎发育潜力以及怀孕率的关系,同时探讨了IVF周期中精卵共孵育时间、病人病因、超促排卵处理方案等对卵母细胞透明带、受精率、胚胎发育和妊娠结果的影响。
1.利用OCTAX系统研究了卵母细胞ZB值与病人年龄、卵母细胞成熟度、原核、胚胎质量以及怀孕率的关系。结果显示,卵母细胞ZB值与病人年龄、卵母细胞成熟度以及原核密切相关,但与随后的胚胎发育潜力和怀孕率关系不大。由此表明,卵母细胞的ZB值能在一定程度上预测IVF周期中卵母细胞的质量,可作为辅助生殖卵母细胞和胚胎早期选择的参考指标。
2.为进一步了解IVF周期中卵母细胞透明带在预测卵母细胞以及胚胎发育潜力中的作用,研究了精卵孵育时间对受精率、卵裂率、胚胎优质率以及ZB值和透明带厚度(ZT)等指标的影响。结果发现,适当缩短精卵共孵育时间对于提高受精率、卵裂率、附植率和临床妊娠率具有一定作用,ZB值随精卵共孵育时间的延长而呈增大趋势,但D1胚胎ZT基本不受精卵孵育时间长短的影响。
3.研究了临床上诸多因素如输卵管因素、多囊卵巢综合症、子宫内膜异位症、原因不明性不孕等对卵母细胞透明带、受精率、胚胎发育和妊娠结果等的影响。结果发现,不同病因对获卵数、受精率、卵裂率和随后的附植和妊娠率以及ZB和ZT值有影响,且因病因不同其影响作用亦不同。其中,子宫内膜异位症的影响最明显,尤其是ZB值显著低于其它各组,ZT值显著高于其它各组。ZB和ZT值可在一定程度上反映不同病因对卵母细胞发育潜力的影响。
4.研究了标准长方案、短方案、拮抗剂方案和微刺激方案四种超排处理方法对卵母细胞发育潜力、临床效果、妊娠结果以及卵母细胞透明带的影响。发现在病人年龄、病因相当的情况下,不同超排方案的成熟率、受精率、多精受精率、卵裂率、优质胚胎率、附植率、妊娠率和周期取消率之间并无统计学差异。不过进一步研究卵母细胞ZB和ZT值时发现,不同超排卵方案对ZB和ZT有明显影响。尤其是卵母细胞ZB值在一定程度上能反映超促排卵方案对卵母细胞质量的影响。
5.探讨了超促排卵时使用hMG对获卵数、受精率、优质胚胎率、妊娠率、以及卵母细胞的ZB和ZT值的影响。结果显示,使用部分hMG对IVF周期病人的受精率、妊娠率以及附植率等无显著影响,但在降调节病人中适当添加hMG,成熟率、受精率、卵裂率、优质胚胎率和附植率较单纯使用FSH组略有增加,但获卵数略有减少。添加hMG组的卵母细胞ZB值低于单纯使用r-FSH组,说明添加促黄体生成素(LH)不仅能促进卵母细胞减数分裂的恢复,还能促进透明带由不成熟向成熟转化。LH (hMG)对卵母细胞ZT的影响存在双向作用,LH过高或过低都不利于透明带的正常发育。
In in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) cycles of human assisted reproduction, the precise evaluating quality of oocyte with the potential to develop to implantation-competent embryos is crucial for the success of clinical treatment. In recent years, zona pellucida (ZP) has been found to be a valuable predictive marker for evaluation of oocyte and embryo quality. Currently, polarized light microscopy has been introduced to facilitate the non-invasive visualization of the oocyte ZP. Through this technique, the zona birefringence (ZB) and light retardance of oocytes as new prognostic factors of the oocyte quality were proposed. However, the previously reported results mainly focused on the ICSI cycles. The aim of the present study was to explore the relationship between ZB and oocyte quality and the factors affecting the developmental potential of oocyte in human IVF cycles.
1. To examine if the age of the female patients, oocyte maturation stage, pronuclei, embryonic quality, and cycle outcome are related to the zona birefringence (ZB) intensity of oocytes in IVF cycles, a retrospective study on214IVF cycles with a polarization imaging system was conducted. The score of ZB intensity was closely correlated with the age of female patients and the maturation stage of oocytes, but not with the embryonic quality and gestation rate. These results indicate that the score of ZB intensity is related to the quality of oocyte and can be used as a reference for early selection of reproductive oocyte and embryo in IVF cycles.
2. Effects of sperm-oocyte coincubation time on the fertility, cleavage, embryo quality, ZB and zona thickness (ZT) were investigated. It was found that properly shortening the sperm-oocyte coincubation time could increase the fertiliy rate, cleavage, implantation and pregnancy rate, moreover, ZB values were increased with the increase of sperm-oocyte incubation time. However, the time of sperm-oocyte coincubation did not effect ZT on day1.
3. Effects of clinical diseases such as tubal factor infertile、polycystic ovarian syndrome (PCOS)、endometriosis and unexplained infertility on the ZP of oocyte, embryo development and clinical outcome in IVF cycles were analyzed. It was found that different diseases had different effects on the retrieved oocyte number, the rate of fertility, implant and pregnancy, ZB and ZT. Endometriosis had the most obvious effects, particularly with significantly lower ZB and significantly higher ZT than those effects caused by other diseases. Therefore, ZB and ZT of oocyte could indicate the effects of different diseases on developmental potential of oocytes to some extent.
4. The efficiency of different superovulation protocols, including standard GnRH agonist long protocol, short protocol, GnRH antagonist protocol and minimal stimulation protocol on developmental potential of oocytes, clinical effects, pregnancy, and ZB of oocytes were compared. No statistic difference was found in maturation, fertilization, mulfertilization, cleavage, embryo quality, implantation, pregnancy and cycle concel with the different superovulation protocols when the patient age and etiology were comparative. However, the different superovulation protocols had distinct influence on the ZB and ZT of oocytes. Particularly, ZB could reflect the effects of different superovulation protocols on oocyte quality to some extent.
5. Effects of human menopausal gonadotropin (hMG) in superovulation treatment on follicle stimulating hormone (FSH) dosage, the number of oocytes retrieved, fertilization rate, cleavage rate, embryo quality, pregnancy rate, embryo implantation and the ZB and ZT of oocyte were investigated. The maturation rate, fertilization rate, cleavage rate, embryo quality, pregnancy and embryo implantation rate were higher in rFSH+hMG group than those in rFSH group, whereas, the number of oocytes retrieved was lower in the former. The ZB was lower in rFSH+hMG group than that in the rFSH group, which indicates that luteinizing hormone (LH) can not only promote the recover of oocyte meiosis but also facilitate the maturation of ZP. LH played the two-way role for ZT of oocytes, which implies that exceptionally high or low level of LH is detrimental for the development of oocyte ZP.
1.利用OCTAX系统研究了卵母细胞ZB值与病人年龄、卵母细胞成熟度、原核、胚胎质量以及怀孕率的关系。结果显示,卵母细胞ZB值与病人年龄、卵母细胞成熟度以及原核密切相关,但与随后的胚胎发育潜力和怀孕率关系不大。由此表明,卵母细胞的ZB值能在一定程度上预测IVF周期中卵母细胞的质量,可作为辅助生殖卵母细胞和胚胎早期选择的参考指标。
2.为进一步了解IVF周期中卵母细胞透明带在预测卵母细胞以及胚胎发育潜力中的作用,研究了精卵孵育时间对受精率、卵裂率、胚胎优质率以及ZB值和透明带厚度(ZT)等指标的影响。结果发现,适当缩短精卵共孵育时间对于提高受精率、卵裂率、附植率和临床妊娠率具有一定作用,ZB值随精卵共孵育时间的延长而呈增大趋势,但D1胚胎ZT基本不受精卵孵育时间长短的影响。
3.研究了临床上诸多因素如输卵管因素、多囊卵巢综合症、子宫内膜异位症、原因不明性不孕等对卵母细胞透明带、受精率、胚胎发育和妊娠结果等的影响。结果发现,不同病因对获卵数、受精率、卵裂率和随后的附植和妊娠率以及ZB和ZT值有影响,且因病因不同其影响作用亦不同。其中,子宫内膜异位症的影响最明显,尤其是ZB值显著低于其它各组,ZT值显著高于其它各组。ZB和ZT值可在一定程度上反映不同病因对卵母细胞发育潜力的影响。
4.研究了标准长方案、短方案、拮抗剂方案和微刺激方案四种超排处理方法对卵母细胞发育潜力、临床效果、妊娠结果以及卵母细胞透明带的影响。发现在病人年龄、病因相当的情况下,不同超排方案的成熟率、受精率、多精受精率、卵裂率、优质胚胎率、附植率、妊娠率和周期取消率之间并无统计学差异。不过进一步研究卵母细胞ZB和ZT值时发现,不同超排卵方案对ZB和ZT有明显影响。尤其是卵母细胞ZB值在一定程度上能反映超促排卵方案对卵母细胞质量的影响。
5.探讨了超促排卵时使用hMG对获卵数、受精率、优质胚胎率、妊娠率、以及卵母细胞的ZB和ZT值的影响。结果显示,使用部分hMG对IVF周期病人的受精率、妊娠率以及附植率等无显著影响,但在降调节病人中适当添加hMG,成熟率、受精率、卵裂率、优质胚胎率和附植率较单纯使用FSH组略有增加,但获卵数略有减少。添加hMG组的卵母细胞ZB值低于单纯使用r-FSH组,说明添加促黄体生成素(LH)不仅能促进卵母细胞减数分裂的恢复,还能促进透明带由不成熟向成熟转化。LH (hMG)对卵母细胞ZT的影响存在双向作用,LH过高或过低都不利于透明带的正常发育。
In in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) cycles of human assisted reproduction, the precise evaluating quality of oocyte with the potential to develop to implantation-competent embryos is crucial for the success of clinical treatment. In recent years, zona pellucida (ZP) has been found to be a valuable predictive marker for evaluation of oocyte and embryo quality. Currently, polarized light microscopy has been introduced to facilitate the non-invasive visualization of the oocyte ZP. Through this technique, the zona birefringence (ZB) and light retardance of oocytes as new prognostic factors of the oocyte quality were proposed. However, the previously reported results mainly focused on the ICSI cycles. The aim of the present study was to explore the relationship between ZB and oocyte quality and the factors affecting the developmental potential of oocyte in human IVF cycles.
1. To examine if the age of the female patients, oocyte maturation stage, pronuclei, embryonic quality, and cycle outcome are related to the zona birefringence (ZB) intensity of oocytes in IVF cycles, a retrospective study on214IVF cycles with a polarization imaging system was conducted. The score of ZB intensity was closely correlated with the age of female patients and the maturation stage of oocytes, but not with the embryonic quality and gestation rate. These results indicate that the score of ZB intensity is related to the quality of oocyte and can be used as a reference for early selection of reproductive oocyte and embryo in IVF cycles.
2. Effects of sperm-oocyte coincubation time on the fertility, cleavage, embryo quality, ZB and zona thickness (ZT) were investigated. It was found that properly shortening the sperm-oocyte coincubation time could increase the fertiliy rate, cleavage, implantation and pregnancy rate, moreover, ZB values were increased with the increase of sperm-oocyte incubation time. However, the time of sperm-oocyte coincubation did not effect ZT on day1.
3. Effects of clinical diseases such as tubal factor infertile、polycystic ovarian syndrome (PCOS)、endometriosis and unexplained infertility on the ZP of oocyte, embryo development and clinical outcome in IVF cycles were analyzed. It was found that different diseases had different effects on the retrieved oocyte number, the rate of fertility, implant and pregnancy, ZB and ZT. Endometriosis had the most obvious effects, particularly with significantly lower ZB and significantly higher ZT than those effects caused by other diseases. Therefore, ZB and ZT of oocyte could indicate the effects of different diseases on developmental potential of oocytes to some extent.
4. The efficiency of different superovulation protocols, including standard GnRH agonist long protocol, short protocol, GnRH antagonist protocol and minimal stimulation protocol on developmental potential of oocytes, clinical effects, pregnancy, and ZB of oocytes were compared. No statistic difference was found in maturation, fertilization, mulfertilization, cleavage, embryo quality, implantation, pregnancy and cycle concel with the different superovulation protocols when the patient age and etiology were comparative. However, the different superovulation protocols had distinct influence on the ZB and ZT of oocytes. Particularly, ZB could reflect the effects of different superovulation protocols on oocyte quality to some extent.
5. Effects of human menopausal gonadotropin (hMG) in superovulation treatment on follicle stimulating hormone (FSH) dosage, the number of oocytes retrieved, fertilization rate, cleavage rate, embryo quality, pregnancy rate, embryo implantation and the ZB and ZT of oocyte were investigated. The maturation rate, fertilization rate, cleavage rate, embryo quality, pregnancy and embryo implantation rate were higher in rFSH+hMG group than those in rFSH group, whereas, the number of oocytes retrieved was lower in the former. The ZB was lower in rFSH+hMG group than that in the rFSH group, which indicates that luteinizing hormone (LH) can not only promote the recover of oocyte meiosis but also facilitate the maturation of ZP. LH played the two-way role for ZT of oocytes, which implies that exceptionally high or low level of LH is detrimental for the development of oocyte ZP.
引文
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