Ghafari Fataneh, Salavati Mazdak, Piercy Richard J, Fouladi-Nashta Ali A
Reproduction. 2025 Mar 14;169(4). doi: 10.1530/REP-24-0388. Print 2025 Apr 1.
Oxidative stress due to high-fat content damages canine oocytes during long period of in vitro maturation and compromises their developmental competence. This paper shows how melatonin protects the oocytes and increases development to metaphase II stage. Results from this study would be applicable to conservation of endangered and other animal species.
Unlike other domesticated animals, in vitro maturation of canine oocytes results in poor nuclear maturation to the metaphase II stage and high oocyte degeneration. The high-fat content of canine oocytes is the likely cause; it predisposes them to oxidative stress and deleterious reactive oxygen species (ROS). Melatonin (MTN) (potentially acting as a powerful antioxidant) was reported to support in vitro cultured oocytes in other species. In this work, canine cumulus oocyte complexes (COCs) were collected after routine ovariohysterectomy. Immunocytochemistry for expression of melatonin receptors (MTNR-1A and 1B) revealed that both receptors were highly expressed in canine oocytes and there was lower expression in the cumulus cells. Canine COCs matured in vitro in melatonin-supplemented culture at 100 nM concentration in low (5%) O2 incubator had a lower percentage at GV stage (6.7% ± 4.2 vs 19.8% ± 3), a higher MII stage (32.3% ± 6.4 vs 15.81% ± 8.1), lower degeneration (20.5% ± 3.2 vs 45.2% ± 5.15) and higher meiotic resumption (GVBD-MII; 56.2% ± 8.6 vs 19% ± 3), and produced lower ROS (determined after DCHFDA staining) (all P < 0.005) than oocytes cultured in high O2 (20%) incubator. The expression of ROS-regulating genes GPX-1 and catalase was significantly reduced in oocytes cultured with melatonin in low O2 compared with high O2 (P < 0.05). Importantly, the oocyte maturation rate increased significantly when G-IVF PLUS, a commercial culture medium used in human oocyte culture, was supplemented with MTN (P < 0.05). These results support the main objective of this study to analyze the beneficial effects of melatonin and low oxygen tension on both health and the developmental competence of in vitro matured canine oocytes.
高脂肪含量导致的氧化应激在犬类卵母细胞长时间体外成熟过程中对其造成损害，并损害其发育能力。本文展示了褪黑素如何保护卵母细胞并增加其发育至中期II阶段的能力。本研究结果将适用于濒危及其他动物物种的保护。
与其他家养动物不同，犬类卵母细胞的体外成熟导致核成熟至中期II阶段的情况较差，且卵母细胞退化率较高。犬类卵母细胞的高脂肪含量可能是原因所在；这使其易受氧化应激和有害活性氧（ROS）的影响。据报道，褪黑素（MTN）（可能作为一种强大的抗氧化剂）可支持其他物种体外培养的卵母细胞。在本研究中，在常规卵巢子宫切除术后收集犬类卵丘卵母细胞复合体（COC）。对褪黑素受体（MTNR - 1A和1B）表达进行免疫细胞化学分析显示，两种受体在犬类卵母细胞中高表达，而在卵丘细胞中表达较低。在低氧（5%）培养箱中，在添加100 nM褪黑素的培养液中体外成熟的犬类COC，处于生发泡（GV）期的比例较低（6.7%±4.2对比19.8%±3），处于中期II（MII）期的比例较高（32.3%±6.4对比15.81%±8.1），退化率较低（20.5%±3.2对比45.2%±5.15），减数分裂恢复率（GVBD - MII）较高（56.2%±8.6对比19%±3），并且与在高氧（20%）培养箱中培养的卵母细胞相比，产生的ROS较低（经二氯二氢荧光素二乙酸酯（DCHFDA）染色后测定）（所有P<0.005）。与高氧环境相比，在低氧环境中用褪黑素培养的卵母细胞中，ROS调节基因GPX - 1和过氧化氢酶的表达显著降低（P<0.05）。重要的是，当在人类卵母细胞培养中使用的商业培养基G - IVF PLUS添加MTN时，卵母细胞成熟率显著提高（P<0.05）。这些结果支持了本研究的主要目的，即分析褪黑素和低氧张力对体外成熟犬类卵母细胞的健康和发育能力的有益影响。
