Use of conjugated linoleic acid (trans 10, cis 12) to cultivate bovine embryos: effect on cryoresistance and lipid content

ABSTRACT This study evaluated the effect of the addition of conjugated linoleic acid (CLA) to in vitro culture on viability, lipid content, and cryoresistance of bovine embryos at different in vitro culture times. Cumulus oocyte complexes (N = 974) were maturated in vitro for 22 h. In vitro fecundation ensued for 18 h. Viable zygotes were cultivated in vitro in medium supplemented with CLA (100 mM) in the first 72 h (CLA-F), last 72 h (CLA-L), or throughout the culture period (CLA-T). Control embryos (control) were cultivated with no CLA. Embryos were cryopreserved by vitrification for subsequent analysis after devitrification. Effect of CLA on cryoresistance was assessed by cultivating embryos in synthetic oviductal fluid containing 5% fetal bovine serum. Lipid content was quantified using Nile Red staining. No significant difference was observed in cleavage rate, blastocyst:total oocyte ratio, and blastocyst:cleaved oocyte ratio. Culture in CLA-L reduced survival rate 24 h after devitrification compared with CLA-F and CLA-T, although with no statistically significant difference compared with control group. However, CLA-T improved embryo hatching rate and affected lipid content of embryos. Cultures CLA-F and CLA-L increased lipid content compared with control. Yet, lipid content values decreased in embryos treated with CLA-T, but they did not differ significantly from the values observed for oocytes at the germinal vesicle stage. Treatment of bovine embryos with CLA during in vitro cultivation did not affect the production of blastocysts, reducing lipid content and improving cryoresistance. However, the effects of CLA on cryoresistance and lipid content is significant only when embryos are exposed to the compound throughout the cultivation period.

摘要 本研究评估了在体外培养体系中添加共轭亚油酸（conjugated linoleic acid, CLA）对不同体外培养时长的牛胚胎的存活率、脂质含量及抗冻性能的影响。本研究共纳入974枚卵丘-卵母细胞复合体，先对其进行22小时的体外成熟培养，随后开展18小时的体外受精。将存活的受精卵分为四组：三组分别在培养基中添加100 mM共轭亚油酸，仅在培养的前72小时（CLA-F组）、后72小时（CLA-L组）或整个培养周期（CLA-T组）进行处理；对照组则不添加CLA进行培养。胚胎经玻璃化冷冻保存，解冻后进行后续分析。为评估CLA对胚胎抗冻性能的影响，本研究将胚胎置于添加5%胎牛血清的合成输卵管液中培养；采用尼罗红染色法对胚胎脂质含量进行定量检测。结果显示，各组在卵裂率、囊胚/总卵母细胞比值及囊胚/卵裂卵比值上均无显著差异。与CLA-F组和CLA-T组相比，CLA-L组胚胎在解冻后24小时的存活率有所降低，但与对照组相比无统计学显著性差异。然而，CLA-T组可显著提升胚胎的孵化率，并对胚胎脂质含量产生影响：CLA-F组与CLA-L组的胚胎脂质含量均高于对照组，而CLA-T组胚胎的脂质含量则有所降低，且与生发泡期卵母细胞的脂质含量无显著统计学差异。本研究表明，体外培养期间添加CLA处理牛胚胎，并不会影响囊胚的生成率，但可降低胚胎脂质含量并改善其抗冻性能；不过仅当胚胎在整个培养周期中均暴露于CLA时，其对胚胎抗冻性能与脂质含量的影响才具有统计学显著性。