Department of Animal and Fish Production, College of Agricultural and Food Sciences, King Faisal University, Al-Ahsa, Saudi Arabia.
Department of Poultry Diseases, Faculty of Veterinary Medicine, Cairo University, Giza, Egypt.
PLoS One. 2021 Feb 2;16(2):e0246307. doi: 10.1371/journal.pone.0246307. eCollection 2021.
Vitrification is an economically effective method for embryo cryopreservation in human and livestock animals; however, it carries the risk of damage by the exposure to severe oxidative stress. The present study was conducted to evaluate the effect of leptin at different levels on the in vitro development of fresh and vitrified preimplantation embryos in a rabbit model. Normal embryos at morulae stage were randomly cultured for 2 h with 0, 10, 20 or 100 ng/mL of leptin, then were cultured for further 48 h as freshly or after vitrification. Thereafter, developed blastocysts form the best leptin level in fresh and vitrified embryos along with their controls were allocated to analyze the pro-oxidant (malondialdehyde, MDA; nitric oxide, NO), antioxidant (total antioxidant capacity, TAC; superoxide dismutase, SOD; glutathione peroxidase, GPx), apoptotic (Bcl-2 associated X protein, BAX; heat shock 60kD protein member 1, HSP60; tumor necrosis factor alpha, TNFα) and developmental (sex determining region Y box protein 2, SOX2; Nanog homeobox protein, NANOG; Octamer-binding protein 4, OCT4) biomarkers. Results indicate that expanding and hatching rates of embryos were significantly higher at 20 ng/mL leptin than the other levels, while vitrification had an independent suppression effect on the in vitro development rates. The MDA and NO were significantly higher, while TAC, SOD and GPx were significantly lower in the vitrified than fresh embryos. In contrast, leptin treatment significantly decreased the pro-oxidant biomarkers and increased the antioxidant biomarkers in both fresh and vitrified embryos. Vitrification significantly increased the antiapoptotic biomarkers, and decreased the developmental biomarkers in embryos. In contrast, leptin decreased the BAX and TNFα, increased the HSP60, and moreover, ameliorated the reduction of developmental biomarkers in the vitrified embryos. These results conclude that leptin could be used as antiapoptotic and antioxidant promotor to support the in vitro embryonic development, particularly under oxidative stress emerged from cryopreservation programs.
玻璃化是一种经济有效的人类和家畜胚胎冷冻保存方法；然而，它存在因暴露于严重氧化应激而受损的风险。本研究旨在评估不同水平的瘦素对兔模型中新鲜和玻璃化前植入胚胎体外发育的影响。处于桑葚胚阶段的正常胚胎被随机培养 2 小时，培养液中含有 0、10、20 或 100ng/mL 的瘦素，然后在新鲜或玻璃化后再培养 48 小时。此后，将在新鲜和玻璃化胚胎中最佳瘦素水平下发育的囊胚与各自的对照组分离出来，分析其促氧化剂（丙二醛 MDA；一氧化氮 NO）、抗氧化剂（总抗氧化能力 TAC；超氧化物歧化酶 SOD；谷胱甘肽过氧化物酶 GPx）、凋亡（Bcl-2 相关 X 蛋白 BAX；热休克 60kD 蛋白 1 成员 HSP60；肿瘤坏死因子 α TNFα）和发育（性别决定区 Y 框蛋白 2 SOX2；Nanog 同源盒蛋白 NANOG；八聚体结合蛋白 4 OCT4）生物标志物。结果表明，20ng/mL 瘦素组胚胎的扩展和孵化率明显高于其他水平，而玻璃化处理对体外发育率有独立的抑制作用。MDA 和 NO 在玻璃化胚胎中明显升高，而 TAC、SOD 和 GPx 在新鲜胚胎中明显降低。相比之下，瘦素处理在新鲜和玻璃化胚胎中均显著降低了促氧化剂生物标志物，增加了抗氧化生物标志物。玻璃化处理显著增加了胚胎的抗凋亡生物标志物，降低了发育生物标志物。相比之下，瘦素降低了 BAX 和 TNFα，增加了 HSP60，并且改善了玻璃化胚胎中发育生物标志物的减少。这些结果表明，瘦素可用作抗凋亡和抗氧化促进剂，支持体外胚胎发育，特别是在冷冻保存方案引起的氧化应激下。
