Biology of Reproduction Group (IREC), UCLM-CSIC-JCCM, Albacete, Spain.
Anim Reprod Sci. 2011 Nov;129(1-2):56-66. doi: 10.1016/j.anireprosci.2011.09.011. Epub 2011 Oct 1.
Viable epididymal sperm can be obtained in the Spanish ibex during 24h after death, but it has been observed a significant effect of the post-mortem time on fertility success, so only goats inseminated with semen recovery during the first 8h became pregnant. The aim of this study was to determine the effect of post-mortem time on epididymal semen samples from of Spanish ibex. For this purpose, sperm samples from 36 males were collected at different post-mortem times, from 2 to 24h, and cryopreserved. Thawed samples were incubated for 2h at 37°C without dilution or after dilution in a modified Tyrode medium, in order to study the sperm resistance to dilution. Moreover, flow cytometry was used to assess the sperm viability (PI), phospolipid disorder of the plasma membrane (M540), mitochondrial membrane potential (Mitotracker Deep Red), indirect apoptosis markers (YOPRO-1) and sperm chromatin stability (SCSA(®)). Sperm motility was evaluated by computer-assisted sperm analysis (CASA). Our results have shown that post-mortem time caused a reduction in mitochondrial membrane potential. In this regard, the loss of energy could be responsible for the loss of maintenance of the membrane with a consequent increase in permeability leading to a decrease in sperm viability and motility, losing linearity and speed. Moreover, the loss of maintenance of the membrane influence the extent to which sperm will survive the cryopreservation process, as it shows the results obtained from the dilution-incubation resistance test. Finally, one important finding of this study is the demonstration of no effect of post-mortem time on post-thaw DNA integrity, giving us the possibility of using sperm samples from valuable males, even if it was not possible to process during the first 8h.
在西班牙野山羊死后 24 小时内，可以获得有活力的附睾精子，但已观察到死后时间对生育成功有显著影响，因此只有在最初 8 小时内通过精液回收进行授精的山羊才能怀孕。本研究的目的是确定死后时间对西班牙野山羊附睾精液样本的影响。为此，从 36 只雄性山羊在不同的死后时间（2 至 24 小时）采集精子样本，并进行冷冻保存。解冻后的样本在 37°C 下不稀释或在改良的 Tyrode 培养基中稀释 2 小时，以研究精子对稀释的抵抗力。此外，还使用流式细胞术评估精子活力（PI）、质膜磷脂紊乱（M540）、线粒体膜电位（Mitotracker Deep Red）、间接凋亡标志物（YOPRO-1）和精子染色质稳定性（SCSA(®)）。通过计算机辅助精子分析（CASA）评估精子运动能力。我们的结果表明，死后时间导致线粒体膜电位降低。在这方面，能量的丧失可能是导致膜维持丧失的原因，随之而来的是通透性增加，导致精子活力和运动能力下降，失去直线性和速度。此外，膜的维持丧失会影响精子在冷冻保存过程中的存活率，因为这反映了稀释孵育抗性试验的结果。最后，本研究的一个重要发现是，死后时间对解冻后 DNA 完整性没有影响，这使我们有可能使用有价值的雄性的精子样本，即使在最初 8 小时内无法处理。
