Faculty of Veterinary Medicine, Utrecht University, Yalelaan 114, 3,584 CM Utrecht, the Netherlands.
Theriogenology. 2012 Mar 15;77(5):998-1007. doi: 10.1016/j.theriogenology.2011.10.007. Epub 2012 Jan 5.
The purpose of this study was to investigate the occurrence of sperm DNA fragmentation in Asian elephant (Elephas maximus) spermatozoa at various processing stages before and after cryopreservation. Five semen samples from four elephants were assessed at four different stages during processing; after (1) collection and reextension in TEST-egg yolk; (2) cooling to 5 °C; (3) equilibration for 1 h with glycerol; (4) thawing. An experimental approach was adopted that allowed comparisons of DNA fragmentation rates developed after the various processing stages. For this, spermatozoa were incubated in TEST-yolk media at 37 °C for 0, 4, 8, 24 and 48 h, and sperm DNA fragmentation rates were estimated using an elephant-specific Halosperm procedure. Incubation at 37 °C induced a rapid increase in DNA fragmentation, and significant differences between males were observed. The overall rate of increase over 4 h was estimated at about 5% per hour, and no significant changes to this rate were observed at the different processing stages, even, including the post-thaw samples. As semen quality of the five ejaculates was relatively poor, the basic semen parameter data were compared with nine different samples collected 11 mo earlier to see whether the tested samples were atypical or representative of the population, As there was no significant difference between the two sets of samples, it is believed that the samples tested for DNA stability were not unusually sensitive. These results suggest that Asian elephant spermatozoa are more susceptible to DNA fragmentation than spermatozoa of other mammals.
本研究旨在探讨亚洲象（Elephas maximus）精子在冷冻保存前后不同处理阶段的 DNA 碎片化发生情况。从四只大象中采集了五份精液样本，在处理过程中的四个不同阶段进行了评估：（1）收集后在 TEST-卵黄中重新扩展；（2）冷却至 5°C；（3）用甘油平衡 1 小时；（4）解冻。采用实验方法，允许比较在各种处理阶段后发展的 DNA 碎片化率。为此，将精子在 TEST-卵黄培养基中于 37°C 孵育 0、4、8、24 和 48 小时，并使用特定于大象的 Halosperm 程序估计精子 DNA 碎片化率。37°C 的孵育会导致 DNA 碎片化迅速增加，并且观察到雄性之间存在显著差异。在 4 小时内的总体增长率估计约为每小时 5%，即使在不同的处理阶段，包括解冻后的样本，也没有观察到这种速率的显著变化。由于五份精液的质量相对较差，因此将基本精液参数数据与 11 个月前收集的 9 个不同样本进行了比较，以确定测试样本是否异常或代表群体。由于两组样本之间没有显著差异，因此可以认为测试 DNA 稳定性的样本没有异常敏感。这些结果表明，亚洲象精子比其他哺乳动物的精子更容易发生 DNA 碎片化。
