Castillo-Martín Miriam, Yeste Marc, Soler Albert, Morató Roser, Bonet Sergi
Biotechnology of Animal and Human Reproduction (TechnoSperm), Department of Biology, Institute of Food and Agricultural Technology, University of Girona, Campus Montilivi, E-17071 Girona, Spain.
Unit of Animal Reproduction, Department of Animal Medicine and Surgery, Faculty of Veterinary Medicine, Autonomous University of Barcelona, E-08193 Bellaterra, Spain.
Reprod Fertil Dev. 2015 Sep;27(7):1115-23. doi: 10.1071/RD14078.
The aim of the present study was to determine the effect of L-ascorbic acid on embryo quality and gene expression of porcine blastocysts after supplementations of in vitro culture medium and/or vitrification-warming media. Embryo quality, in terms of total cell number (TCN), DNA fragmentation and peroxide levels, together with the relative transcript abundance of BCL-2 associated X protein (BAX), BCL2-like 1 (BCL2L1), POU class 5 homeobox 1 (POU5F1) and heat shock protein 70 (HSPA1A), was analysed. In Experiment 1, gene expression and embryo quality of fresh blastocysts were evaluated after culture with or without L-ascorbic acid; no significant differences were observed between the groups. In Experiment 2, blastocysts cultured with or without L-ascorbic acid were vitrified using two different vitrification solutions, supplemented or not with L-ascorbic acid. Supplementation of culture and vitrification media significantly enhanced survival rates and reduced peroxide levels. No significant differences in TCN, DNA fragmentation and BAX, BCL2L1 and POU5F1 expression were found in vitrified blastocysts among experimental groups. Vitrification procedures increase HSPA1A transcript abundance, but this increase was significantly lower in embryos cultured and/or vitrified with L-ascorbic acid. Thus, supplementing culture and/or vitrification media with L-ascorbic acid enhances survival rates of porcine blastocysts, suggesting a relationship with HSPA1A expression.
本研究的目的是确定在体外培养基和/或玻璃化冷冻复苏培养基中添加L-抗坏血酸对猪囊胚胚胎质量和基因表达的影响。分析了胚胎质量，包括总细胞数（TCN）、DNA片段化和过氧化物水平，以及BCL-2相关X蛋白（BAX）、BCL2样1蛋白（BCL2L1）、POU5类同源盒1蛋白（POU5F1）和热休克蛋白70（HSPA1A）的相对转录丰度。在实验1中，评估了添加或不添加L-抗坏血酸培养后新鲜囊胚的基因表达和胚胎质量；两组之间未观察到显著差异。在实验2中，使用两种不同的玻璃化溶液对添加或不添加L-抗坏血酸培养的囊胚进行玻璃化冷冻，玻璃化溶液添加或不添加L-抗坏血酸。培养基和玻璃化冷冻复苏培养基的添加显著提高了存活率并降低了过氧化物水平。在实验组的玻璃化囊胚中，TCN、DNA片段化以及BAX、BCL2L1和POU5F1表达未发现显著差异。玻璃化冷冻程序会增加HSPA1A转录丰度，但在添加L-抗坏血酸培养和/或玻璃化冷冻的胚胎中，这种增加显著降低。因此，在培养基和/或玻璃化冷冻复苏培养基中添加L-抗坏血酸可提高猪囊胚的存活率，提示其与HSPA1A表达有关。
