Department of Animal Health and Anatomy, Faculty of Veterinary Medicine, Universitat Autònoma de Barcelona, Bellaterra, Spain.
Oncology Area, Center for Biomedical Research of La Rioja (CIBIR), Logroño, Spain.
Theriogenology. 2022 Jun;185:16-23. doi: 10.1016/j.theriogenology.2022.03.010. Epub 2022 Mar 18.
The cold-inducible RNA-binding protein (CIRBP) assists cells in adapting to new environmental conditions stabilizing specific mRNAs and promoting their translation. CIRBP participates in anti-apoptotic and anti-senescence processes, and its expression is induced by mild hypothermia, which may be advantageous to oocytes during vitrification. Several newly discovered small molecules, like zr17-2, mimic the effects of cold temperatures by increasing the expression of CIRBP at normothermia. This study aimed to evaluate the mRNA changes of a group of cold-inducible protein-encoding and apoptotic genes in response to exogenous supplementation of zr17-2 (experiment 1) or CIRBP (experiment 2) in vitro matured bovine oocytes and their cumulus cells. In experiment 1, cumulus-oocyte complexes (COCs) were randomly exposed to three concentrations of zr17-2 (1, 10, and 100 μM) during 24 h of in vitro maturation (IVM) under normothermia (38.5 °C) or mild hypothermia (34 °C) culture conditions. In experiment 2, COCs were randomly exposed to three concentrations of CIRBP (2, 4, and 6 μg/mL) or subjected to mild hypothermia (34 °C), followed by oocyte vitrification/warming after 20 h of IVM. The quantification of the selected gene transcript expression was performed separately in oocytes and cumulus cells by quantitative real-time PCR. We show that oocytes and cumulus cells exhibited similar mRNA expression responses to mild hypothermia and vitrification. However, minor differences were observed when COCs were exposed to exogenous supplementation with zr17-2 and CIRBP. In conclusion, the alterations observed in the mRNA expression in these experimental conditions may impact the quality of the cumulus-oocyte complexes in terms of vitrification and sublethal hypothermia challenges. In this sense, our results should complement other oocyte quality assessments for its application in future assisted reproductive techniques in the bovine species, including improving oocyte cryotolerance to vitrification.
冷诱导 RNA 结合蛋白 (CIRBP) 帮助细胞适应新的环境条件，稳定特定的 mRNA 并促进其翻译。CIRBP 参与抗凋亡和抗衰老过程，其表达可被轻度低温诱导，这对玻璃化过程中的卵母细胞可能有利。一些新发现的小分子，如 zr17-2，通过在常温下增加 CIRBP 的表达来模拟低温的作用。本研究旨在评估一组冷诱导蛋白编码基因和凋亡基因的 mRNA 变化，这些基因在外源补充 zr17-2（实验 1）或 CIRBP（实验 2）后，在体外成熟的牛卵母细胞及其卵丘细胞中的反应。在实验 1 中，将卵丘-卵母细胞复合物 (COC) 在常温（38.5°C）或轻度低温（34°C）培养条件下，于体外成熟（IVM）的 24 小时内随机暴露于 zr17-2 的三种浓度（1、10 和 100μM）下。在实验 2 中，COC 随机暴露于三种浓度的 CIRBP（2、4 和 6μg/mL）或接受轻度低温（34°C）处理，然后在 IVM 20 小时后进行卵母细胞玻璃化/复温。通过实时定量 PCR 分别在卵母细胞和卵丘细胞中定量所选基因转录物的表达。我们表明，卵母细胞和卵丘细胞对轻度低温和玻璃化表现出相似的 mRNA 表达反应。然而，当 COC 暴露于外源性 zr17-2 和 CIRBP 补充时，观察到 COC 表现出轻微的差异。总之，在这些实验条件下观察到的 mRNA 表达变化可能会影响卵丘-卵母细胞复合物在玻璃化和亚致死低温挑战方面的质量。在这方面，我们的结果应该补充其他卵母细胞质量评估，以将其应用于牛种的未来辅助生殖技术，包括提高卵母细胞对玻璃化的耐冻性。
