Evaluation of DNA damage in rainbow trout (Oncorhynchus mykiss) and gilthead sea bream (Sparus aurata) cryopreserved sperm
Evaluation of DNA damage in rainbow trout (Oncorhynchus mykiss) and gilthead sea bream (Sparus aurata) cryopreserved sperm
复制标题
DOI：
10.1016/j.cryobiol.2004.12.003
复制
发表时间：
2005-04-01
期刊：
影响因子：
2.7
通讯作者：
Herráez, MP
中科院分区：
文献类型：
--
作者：
Cabrita, E;Robles, V;Herráez, MP
低温保存对精子造成多种类型的损伤，如质膜完整性和功能的丧失，运动性和ATP含量的丧失，导致生育率下降。这种精子损害已经在几种海洋和淡水鱼中得到了广泛的研究。然而，对核DNA的研究却很少。本研究的目的是确定冷冻保存对虹鳟（Oncorhynchus tnykiss）和金头鲷（Sparus aurata）两种商业养殖物种精子DNA损伤的程度，这两种物种都可以从大规模冷冻保存策略的发展中受益。我们使用单细胞凝胶电泳，通常被称为彗星试验来检测DNA中的链断裂。这项技术是在两个物种的新鲜精子和冷冻精子上进行的。在虹鳟鱼中，冷冻保存后的DNA片段和橄榄尾矩的平均值显著增加（新鲜精子和冷冻保存后的精子尾部DNA的平均值分别为11.19- 30.29%和13.4-53.48%），并且DNA片段率较高的细胞比例也显著增加。在对照样品和1:6稀释和冷冻保存的精子中，鳙鱼尾部DNA的百分比差异不显著（分别为28.23%和31.3%）。然而，精子稀释率的增加导致DNA碎片率的增加（41.4%）。我们的研究表明，冷冻保存可以诱导这些物种的DNA损伤，这一事实应该在评估冷冻/解冻方案时考虑到，特别是当精子冷冻保存将用于基因库目的时。(c) 2005爱思唯尔公司版权所有。
Cryopreservation causes several types of damage to spermatozoa, such as loss of plasma membrane integrity and functionality, loss of motility, and ATP content, resulting in decrease of fertility rates. This spermatozoal damage has been widely investigated for several marine and freshwater fish species. However, not much attention has been paid to the nuclear DNA. The objective of this study was to determine the degree to which cryopreservation induces spermatozoal DNA damage in two commercially cultured species, rainbow trout (Oncorhynchus tnykiss) and gilthead sea bream (Sparus aurata), both of which could benefit from the development of cryopreservation strategies on a large scale. We have used the single-cell gel electrophoresis, commonly known as Comet assay to detect strand breaks in DNA. This technique was performed on fresh and cryopreserved sperm from both species. In rainbow trout there was a significant increase in the averages of fragmented DNA and Olive tail moment after cryopreservation (11.19-30.29 % tail DNA and 13.4-53.48% Olive tail moment in fresh and cryopreserved sperm, respectively), as well as in the proportion of cells with a high percentage of DNA fragmentation. For gilthead sea bream there were no significant differences in the percentage of tail DNA between the control samples and sperm diluted 1:6 and cryopreserved (28.23 and 31.3% DNA,, respectively). However, an increase in the sperm dilution rate produced an increase in the percentage of DNA fragmentation (41.4 %). Our study demonstrates that cryopreservation can induce DNA damage in these species, and that this fact should be taken into account in the evaluation of freezing/thawing protocols, especially when sperm cryopreservation will be used for gene bank purposes. (c) 2005 Elsevier Inc. All rights reserved.