Oxidative stress is one of the major issues associated with cryopreservation because it causes a marked reduction in the post-thaw quality of semen. This study investigated the ability of kinetin to preserve the structural and functional integrity of dog sperm during cryopreservation. Pooled ejaculates were divided into 5 equal aliquots, diluted with buffer 2 supplemented with different concentrations of kinetin (0, 25, 50, 100, and 200 μM), and finally cryopreserved. The optimal concentration of kinetin was 50 μM based on the significantly improved (P < 0.05) motion characteristics and viability of post-thaw sperm samples. Moreover, kinetin-supplemented samples exhibited significantly higher (P < 0.05) sperm counts with the intact plasma membrane, normal acrosomes, mitochondria, and chromatin than control. The beneficial effects of kinetin were also reflected by the significant increase in the expression levels of anti-apoptotic (B-cell lymphoma, BCL2) and protamine-related genes (protamine 2, PRM2; protamine 3, PRM3), and decrease in the expression of pro-apoptotic (BCL2-associated X, BAX) and mitochondrial reactive oxygen species-modulating genes (ROS modulator 1, ROMO1) in kinetin-supplemented sperm samples than in control. The results demonstrated that supplementation of buffer 2 with 50 μM kinetin is ideal for reducing the magnitude of oxidative damage during semen cryoprocessing and improving the post-thaw quality of dog semen.

中文翻译：

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激动素补充剂对狗精子解冻后活力、活力和结构完整性的影响

氧化应激是与冷冻保存相关的主要问题之一，因为它会导致精液解冻后质量显着降低。本研究调查了激动素在冷冻保存过程中保持狗精子结构和功能完整性的能力。汇集的精液被分成 5 等份，用补充有不同浓度激动素（0、25、50、100 和 200 μM）的缓冲液 2 稀释，最后冷冻保存。基于显着改善 (P < 0.05) 运动特性和解冻后精子样本的活力，激动素的最佳浓度为 50 μM。此外，与对照相比，补充激动素的样本显示出具有完整质膜、正常顶体、线粒体和染色质的精子计数显着更高（P < 0.05）。激动素的有益作用还体现在抗凋亡（B 细胞淋巴瘤，BCL2）和鱼精蛋白相关基因（鱼精蛋白 2，PRM2；鱼精蛋白 3，PRM3）的表达水平显着增加，以及表达水平的降低与对照相比，补充激动素的精子样本中促凋亡（BCL2 相关的 X，BAX）和线粒体活性氧调节基因（ROS 调节剂 1，ROMO1）的含量。结果表明，用 50 μM 激动素补充缓冲液 2 是降低精液冷冻处理过程中氧化损伤程度和提高狗精液解冻后质量的理想选择。与对照相比，补充激动素的精子样本中促凋亡（BCL2 相关 X，BAX）和线粒体活性氧调节基因（ROS 调节剂 1，ROMO1）的表达降低。结果表明，用 50 μM 激动素补充缓冲液 2 是降低精液冷冻处理过程中氧化损伤程度和提高狗精液解冻后质量的理想选择。与对照相比，补充激动素的精子样本中促凋亡（BCL2 相关 X，BAX）和线粒体活性氧调节基因（ROS 调节剂 1，ROMO1）的表达降低。结果表明，用 50 μM 激动素补充缓冲液 2 是降低精液冷冻处理过程中氧化损伤程度和提高狗精液解冻后质量的理想选择。