Costa Camila Bortoliero, Silva Nathália Covre, Costa Érika Ferreira, Ferreira Christina Ramires, Sanches Tamires Korchovei, Zangirolamo Amanda Fonseca, Seneda Marcelo Marcondes, Nogueira Marcelo Fábio Gouveia
Institute of Biosciences, São Paulo State University (UNESP), Campus Botucatu, Botucatu, Brazil.
School of Sciences, Humanities and Languages, São Paulo State University (UNESP), Assis, Brazil.
Sci Rep. 2025 Jul 2;15(1):22972. doi: 10.1038/s41598-025-07453-0.
The objective of this study was to evaluate the effect of adding of C-Type Natriuretic (CNP) to the in vitro culture medium of bovine embryos on cryotolerability through modulation of lipid content and profile, as well as modulation of gene transcripts linked to embryonic metabolism. Initially, a concentration of 400 nM of CNP was used throughout the in vitro culture and blastocysts were collected for lipid content analysis by Sudan Black B. In addition, blastocysts were selected by morphological quality and developmental stage, and the samples collected were analyzed using MRM- profiling. After, blastocysts were vitrified using OPS. Subsequently the warming, hatched blastocysts were collected and evaluated for transcript abundance in a microfluidic platform. Differences of probabilities lower than P < 0.05, and/or fold change ˃1.5 were considered significant. The CNP group presented a reduction in the relative abundance of ions belonging to different lipid subclasses, such as acylcarnitine, sphingomyelin, cholesteryl esters, free fatty acids, and glycerophospholipid. Furthermore, the triacylglycerol lipids TG 52:3 NL 16:1, TG 56:3 NL 18:1, and the glycerophospholipid C22:6, were increased in the CNP group. A modulation of blastocyst transcripts was also observed by increased transcription of ATF4, and a trend statistical significance of BMP15 and GFPT2 transcripts. There was no difference in blastocyst development rates after warming of CNP-treated embryos.
本研究的目的是通过调节脂质含量和分布以及与胚胎代谢相关的基因转录本，评估向牛胚胎体外培养基中添加C型利钠肽（CNP）对冷冻耐受性的影响。最初，在整个体外培养过程中使用400 nM的CNP浓度，并收集囊胚用苏丹黑B进行脂质含量分析。此外，根据形态质量和发育阶段选择囊胚，并使用MRM分析对收集的样本进行分析。之后，使用OPS对囊胚进行玻璃化冷冻。随后进行解冻，收集孵化的囊胚并在微流控平台上评估转录本丰度。概率差异低于P < 0.05和/或倍数变化>1.5被认为具有显著性。CNP组中属于不同脂质亚类的离子相对丰度降低，如酰基肉碱、鞘磷脂、胆固醇酯、游离脂肪酸和甘油磷脂。此外，CNP组中三酰甘油脂质TG 52:3 NL 16:1、TG 56:3 NL 18:1和甘油磷脂C22:6增加。还观察到囊胚转录本的调节，即ATF4转录增加，以及BMP15和GFPT2转录本的统计学趋势显著性。CNP处理的胚胎解冻后的囊胚发育率没有差异。
