Zona pellucida gene mRNA expression in human oocytes is related to oocyte maturity, zona inner layer retardance and fertilization competence
Zona pellucida gene mRNA expression in human oocytes is related to oocyte maturity, zona inner layer retardance and fertilization competence
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DOI：
10.1093/molehr/gax008
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发表时间：
2017-05-01
影响因子：
4
通讯作者：
Smitz, J.
中科院分区：
文献类型：
--
作者：
Canosa, S.;Adriaenssens, T.;Smitz, J.
研究问题：卵母细胞和卵丘细胞中透明质酸（ZP）基因ZP 1、2、3和4的mRNA表达水平是否揭示了卵母细胞的相关信息？简要答复：ZP mRNA在人类卵母细胞中的表达与卵母细胞成熟度、卵内层发育迟缓和受精能力有关。已知：ZP结构和双折射为卵母细胞胞质成熟、胚胎生长发育能力、囊胚形成和妊娠提供了有用的信息。为了理解ZP形态学变化的分子基础，在本研究中，偏振光显微镜（PLM）方法与卵母细胞和周围CC中编码ZP 1、2、3和4的基因的表达分析相结合。这是一项回顾性研究，包括98个额外的人卵丘卵母细胞复合体（COC）[80中期II（MII），10个中期I（MI）和8个生殖囊泡（GV）]从39名患者（中位年龄33.4岁，范围22-42）在控制性卵巢刺激后获得。使用PLM检查卵母细胞，并对这些个体卵母细胞及其CC中的ZP 1、2、3和4进行定量RT-PCR。在CC中测量Ephrin-B2（EFNB 2）mRNA作为对照。ZP 3蛋白在CC和卵母细胞中的存在进行了研究，使用免疫细胞化学。数据进行了分析，使用单参数和多变量分析，并进行了校正的潜在影响的病人和周期characteristics.Main结果和角色的机会：卵母细胞中含有ZP 1/2/3和4 mRNA，而在CC只有ZP 3是可量化的。在人CC中也检测到ZP 3蛋白。在比较成熟卵母细胞（MII）和未成熟卵母细胞（MI/GV）及其相应的CC时，ZP 1/2和4在成熟卵母细胞中的表达低于未成熟卵母细胞中的表达（均P < 0.05），ZP 3在成熟卵母细胞CC中的表达低于未成熟卵母细胞CC中的表达（P < 0.05）。成熟卵母细胞IL-ZP面积和厚度均显著小于未成熟卵母细胞（均P <0. 05），成熟卵母细胞IL-ZP延迟与4种ZP mRNA表达显著相关（均P <0. 05）。卵母细胞ZP 3表达是受精能力的主要预测因子，其次是IL-延迟和IL-厚度。使用逐步回归分析，IL-厚度结合CC中EFNB 2表达和患者的卵巢反应导致非侵入性卵母细胞受精预测模型。大规模数据：不适用。局限性，解释原因：这是一项回顾性研究，卵母细胞mRNA水平与受精能力的关系是间接的，因为卵母细胞基因表达分析需要卵母细胞裂解。成功地记录了PLM观察结果、mRNA表达变化和内在卵母细胞能力之间的总体关系。因此，PLM和CC基因表达被证实是有价值的非侵入性技术，以评估卵母细胞的能力。
STUDY QUESTION: Do the mRNA expression levels of zona pellucida (ZP) genes, ZP1, 2, 3 and 4 in oocyte and cumulus cells (CC) reveal relevant information on the oocyte?SUMMARY ANSWER: The ZP mRNA expression in human oocytes is related to oocyte maturity, zona inner layer (IL) retardance and fertilization capacity.WHAT IS KNOWN ALREADY: ZP structure and birefringence provide useful information on oocyte cytoplasmic maturation, developmental competence for embryonic growth, blastocyst formation and pregnancy. In order to understand the molecular basis of morphological changes in the ZP, in the current study, the polarized light microscopy (PLM) approach was combined with analysis of the expression of the genes encoding ZP1, 2, 3 and 4, both in the oocytes and in the surrounding CC.STUDY DESIGN, SIZE, DURATION: This is a retrospective study comprising 98 supernumerary human cumulus oocyte complexes (COC) [80 Metaphase II (MII), 10 Metaphase I (MI) and 8 germinal vesicle (GV)] obtained from 39 patients (median age 33.4 years, range 22-42) after controlled ovarian stimulation.PARTICIPANTS/MATERIALS, SETTING, METHODS: Single oocytes and their corresponding CC were analysed. Oocytes were examined using PLM, and quantitative RT-PCR was performed for ZP1, 2, 3 and 4 in these individual oocytes and their CC. Ephrin-B2 (EFNB2) mRNA was measured in CC as a control. Presence of ZP3 protein in CC and oocytes was investigated using immunocytochemistry. Data were analysed using one-parametric and multivariate analysis and were corrected for the potential impact of patient and cycle characteristics.MAIN RESULTS AND THE ROLE OF CHANCE: Oocytes contained ZP1/2/3 and 4 mRNA while in CC only ZP3 was quantifiable. Also ZP3 protein was detected in human CC. When comparing mature (MII) and immature oocytes (MI/GV) or their corresponding CC, ZP1/2 and 4 expression was lower in mature oocytes compared to the expression in immature oocytes (all P < 0.05) and ZP3 expression was lower in the CC of mature oocytes compared to the expression in CC of immature oocytes (P < 0.05). This coincided with a significantly smaller IL-ZP area and thickness in mature oocytes than in immature oocytes (all P < 0.05).In mature oocytes, IL-ZP retardance was significantly correlated with the expression of all four ZP mRNAs (all P < 0.05). The oocyte ZP3 expression was the main predictor of the fertilization capacity, next to IL-retardance and IL-thickness. Using stepwise regression analysis, IL-thickness combined with EFNB2 expression in CC and the patient's ovarian response resulted in a noninvasive oocyte fertilization prediction model.LARGE SCALE DATA: Not applicable.LIMITATIONS, REASONS FOR CAUTION: This is a retrospective study and the relation of oocyte mRNA levels to fertilization capacity is indirect as oocyte gene expression analysis required lysis of the oocyte.WIDER IMPLICATIONS OF THE FINDINGS: Overall relations between PLM observations, mRNA expression changes and intrinsic oocyte competence were successfully documented. As such PLM and CC gene expression are confirmed as valuable noninvasive techniques to evaluate oocyte competence.