Berthelot F, Venturi E, Cognié J, Furstoss V, Martinat-Botté F
INRA, UMR 6175 INRA-CNRS, Université de TOURS-Haras Nationaux, PRC, 37380 Nouzilly, France.
Theriogenology. 2007 Jul 15;68(2):178-85. doi: 10.1016/j.theriogenology.2007.04.050. Epub 2007 Jun 6.
Unhatched blastocysts from Large White hyperprolific gilts (n=103) were identified, measured and vitrified using the Open Pulled Straw (OPS) technique to evaluate the effects of the collected blastocyst size and cryoprotectant concentrations used for vitrification, and the number of embryos transferred per recipient. Vitrified/warmed blastocyst viability was estimated in vitro, as the percentage of embryos developing after 72h, and in vivo, on pregnancy Day 30. In the in vitro study, we compared the use of three cryoprotectant concentrations (16.5, 18, or 20% DMSO+16.5, 18, or 20% EG+0.4M sucrose). Survival rates differed significantly between the control (98.3%) and the three cryoprotectant concentrations (67, 62.3, and 57%, respectively). Blastocyst size at vitrification determined the further in vitro development of embryos (26% survival for blastocysts 126-144microm versus 100% for blastocysts >199microm). For the in vivo study, blastocysts were vitrified using cryoprotectant concentrations of 16.5 or 18% DMSO+EG and transferred surgically in groups of 20 or 30 per recipient (n=40). Recipients were slaughtered on pregnancy D30. No significant differences were detected in gestation rates (50-70%) and embryo survival rates (14.7-25%), although survival was higher (P=0.0003) when 20 blastocysts were transferred compared to 30 (24.7% versus 15.5%). Our findings indicate that best results, in terms of subsequent in vivo embryo survival, were achieved after transferring 20 embryos at the blastocyst or expanded blastocyst stage, previously vitrified using cryoprotectant concentrations of 16.5 or 18%.
识别、测量并使用开放式拉长细管（OPS）技术对来自大白高繁殖力母猪（n = 103）的未孵化囊胚进行玻璃化处理，以评估所收集囊胚大小、用于玻璃化的冷冻保护剂浓度以及每个受体移植胚胎数量的影响。在体外评估玻璃化/复温后囊胚的活力，以72小时后发育胚胎的百分比表示，在体内则在妊娠第30天进行评估。在体外研究中，我们比较了三种冷冻保护剂浓度（16.5%、18%或20%二甲基亚砜 + 16.5%、18%或20%乙二醇 + 0.4M蔗糖）的使用情况。对照组（98.3%）与三种冷冻保护剂浓度组（分别为67%、62.3%和57%）的存活率存在显著差异。玻璃化时囊胚的大小决定了胚胎的进一步体外发育（126 - 144微米的囊胚存活率为26%，而大于199微米的囊胚存活率为100%）。在体内研究中，使用16.5%或18%二甲基亚砜 + 乙二醇的冷冻保护剂浓度对囊胚进行玻璃化处理，并以每组20个或30个的数量手术移植到每个受体中（n = 40）。在妊娠第30天屠宰受体。虽然与移植30个囊胚相比，移植20个囊胚时存活率更高（P = 0.0003，分别为24.7%和15.5%），但在妊娠率（50 - 70%）和胚胎存活率（14.7 - 25%）方面未检测到显著差异。我们的研究结果表明，就随后的体内胚胎存活率而言，在囊胚或扩张囊胚阶段移植20个先前使用16.5%或18%冷冻保护剂浓度进行玻璃化处理的胚胎可获得最佳结果。
