Boar spermatozoa are extremely sensitive to low temperatures and the cryopreservation causes dramatic changes in sperm survivability, but it is not clear which part of the cryopreservation process affects the most. The aim of this work was to assess early events of apoptotic changes as damage indicators in boar sperm cooled to 5 °C and exposed to different glycerol (GLY) concentrations. For this purpose, progressive sperm motility (CASA), plasmatic and acrosome membranes integrity (CFDA/PI; phase contrast), plasma membrane functionality (HOS), phosphatidylserine translocation (Annexin-V/FITC) and reduction of mitochondrial membrane potential (Ψm) (JC-10) were carried out at 37 °C, 17 °C and 5 °C in eight boar sperm pools. Afterwards, three aliquots were diluted in different freezing extenders (control: 0% GLY; A: 2% GLY and B: 3% GLY); sperm quality and early apoptotic changes were assessed. Motility was negatively affected during cooling to 5 °C. Furthermore, plasma membrane functionality was the most affected by cooling. The number of necrotic cells was higher at 5 °C. However, no differences were observed in phosphatidylserine translocation. The extender with 3% GLY at 5 °C presented better Ψm than 0 and 2% GLY. Based on this analysis, boar sperm cooling to 5 °C does not modify the rate of early apoptotic changes, although alterations in the Ѱm were evident.

中文翻译：

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不同甘油浓度对冷冻公猪精子磷脂酰丝氨酸易位和线粒体膜电位的影响

公猪精子对低温极为敏感，冷冻保存会导致精子存活率发生巨大变化，但目前尚不清楚冷冻保存过程的哪一部分影响最大。这项工作的目的是评估细胞凋亡变化的早期事件，因为公猪精子中的损伤指标冷却至 5°C 并暴露于不同的甘油 (GLY) 浓度。为此，进行性精子活力 (CASA)、质膜和顶体膜完整性 (CFDA/PI；相差)、质膜功能 (HOS)、磷脂酰丝氨酸易位 (Annexin-V/FITC) 和线粒体膜电位 (Ψm) 降低(JC-10) 在 8 个公猪精子池中在 37°C、17°C 和 5°C 下进行。然后，将三份等分试样在不同的冷冻稀释剂中稀释（对照：0% GLY；A：2% GLY 和 B：3% GLY）；评估精子质量和早期凋亡变化。在冷却至 5 °C 期间，运动性受到负面影响。此外，质膜功能受冷却的影响最大。坏死细胞的数量在 5°C 时更高。然而，在磷脂酰丝氨酸易位中没有观察到差异。在 5°C 下含有 3% GLY 的增量剂表现出比 0 和 2% GLY 更好的 Ψm。根据该分析，公猪精子冷却至 5 °C 不会改变早期凋亡变化的速率，尽管 Ѱm 的变化很明显。在 5°C 下含有 3% GLY 的增量剂表现出比 0 和 2% GLY 更好的 Ψm。根据该分析，公猪精子冷却至 5 °C 不会改变早期凋亡变化的速率，尽管 Ѱm 的变化很明显。在 5°C 下含有 3% GLY 的增量剂表现出比 0 和 2% GLY 更好的 Ψm。根据该分析，公猪精子冷却至 5 °C 不会改变早期凋亡变化的速度，尽管 Ѱm 的变化很明显。