The present study analyzed the relationship between bovine oocytes developmental competence and mRNA expression of apoptotic and mitochondrial genes following the change of vitrification temperatures (VTs) and cryoprotectant agent concentrations (CPAs). Cumulus oocyte complexes were randomly divided into five groups: control, vitrified in liquid nitrogen (LN; -196 °C) with 5.6 M CPAs (LN 5.6 M), LN with 6.6 M CPAs (LN 6.6 M), liquid helium (LHe; -269 °C) with 5.6 M CPAs (LHe 5.6 M), and LHe with 6.6 M CPAs (LHe 6.6 M). After vitrification and warming, oocytes of vitrified and control groups were subjected to in vitro maturation (IVM), in vitro fertilization and in vitro culture. The blastocyst rate in LHe 5.6 M group was the highest among the four vitrified groups (13.7% vs. 9.4%, 1.3%, and 8.4%; P < 0.05). The mRNA expression level of 8 apoptotic- and 12 mitochondria-related genes were detected through qRT-PCR after IVM. Lower VT (LHe, -269 °C) positively affected the mRNA expression levels of apoptotic genes (BAD, BID, BTK, TP53, and TP53I3) and mitochondrial genes (COX6B1, DERA, FIS1, NDUFA1, NDUFA4, PRDX2, SLC25A5, TFB1M, and UQCRB), and reduced oxidative stress from freezing. Decreased CPAs (5.6 M) positively affected mRNA expression levels of apoptotic genes (BAD, BCL2A1, BID, and CASP3) in LHe vitrification but negatively affected apoptotic genes (BAD, BAX, BID, BTK, and BCL2A1) in LN vitrification. In conclusion, decreased VTs and CPAs in LHe vitrification may increase the blastocyst rate by changing the mRNA expression levels of these apoptotic and mitochondrial genes for the vitrified oocytes.

中文翻译：

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玻璃化温度和冷冻保护剂浓度变化后牛卵母细胞发育能力与凋亡和线粒体基因mRNA表达的关系

本研究分析了随着玻璃化温度 (VT) 和冷冻保护剂浓度 (CPA) 的变化，牛卵母细胞发育能力与凋亡和线粒体基因 mRNA 表达之间的关系。卵丘卵母细胞复合物随机分为五组：对照，在液氮（LN；-196°C）中玻璃化，含 5.6 M CPA（LN 5.6 M），LN 含 6.6 M CPA（LN 6.6 M），液氦（LHe； -269 °C) 使用 5.6 M CPA (LHe 5.6 M)，LHe 使用 6.6 M CPA (LHe 6.6 M)。玻璃化和加温后，玻璃化组和对照组的卵母细胞进行体外成熟（IVM）、体外受精和体外培养。LHe 5.6 M 组的囊胚率在四个玻璃化组中最高（13.7% vs. 9.4%、1.3% 和 8.4%；P < 0.05）。IVM后通过qRT-PCR检测8个凋亡相关基因和12个线粒体相关基因的mRNA表达水平。较低的 VT (LHe, -269 °C) 对凋亡基因（BAD、BID、BTK、TP53 和 TP53I3）和线粒体基因（COX6B1、DERA、FIS1、NDUFA1、NDUFA4、PRDX2、SLC25A5、TFB1MD）的 mRNA 表达水平产生积极影响, 和 UQCRB)，并减少冷冻引起的氧化应激。CPA 的减少 (5.6 M) 对 LHe 玻璃化中凋亡基因（BAD、BCL2A1、BID 和 CASP3）的 mRNA 表达水平产生积极影响，但对 LN 玻璃化中的凋亡基因（BAD、BAX、BID、BTK 和 BCL2A1）产生负面影响。总之，LHe玻璃化过程中VT和CPA的降低可能通过改变玻璃化卵母细胞凋亡和线粒体基因的mRNA表达水平来提高囊胚率。