Highly differentiated mature spermatozoa carry not only genetic but also epigenetic information that is to be transmitted to the embryo. DNA methylation is one epigenetic actor associated with sperm nucleus compaction, gene silencing, and prepatterning of embryonic gene expression. Therefore, the stability of this mark toward reproductive biotechnologies is a major issue in animal production. The present work explored the impact of hormonal induction of spermiation and sperm cryopreservation in two cyprinids, the goldfish (Carassius auratus) and the zebrafish (Danio rerio), using LUminometric Methylation Assay (LUMA). We showed that while goldfish hormonal treatment did increase sperm production, it did not alter global DNA methylation of spermatozoa. Different sperm samples repeatedly collected from the same males for 2 months also showed the same global DNA methylation level. Similarly, global DNA methylation was not affected after cryopreservation of goldfish spermatozoa with methanol, whereas less efficient cryoprotectants (dimethylsulfoxide and 1,2-propanediol) decreased DNA methylation. In contrast, cryopreservation of zebrafish spermatozoa with methanol induced a slight, but significant, increase in global DNA methylation. In the less compact nuclei, that is, goldfish fin somatic cells, cryopreservation did not change global DNA methylation regardless of the choice of cryoprotectant. To conclude, global DNA methylation is a robust parameter with respect to biotechnologies such as hormonal induction of spermiation and sperm cryopreservation, but it can be altered when the best sperm manipulation conditions are not met.

中文翻译：

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鱼精子中DNA甲基化稳定性受外部限制：鱼激素刺激和精子冷冻保存的影响。

高分化的成熟精子不仅携带遗传信息，而且还携带表观遗传信息，这些信息将传递给胚胎。DNA甲基化是一种与精子细胞核的紧实，基因沉默和胚胎基因表达的预模式有关的表观遗传因子。因此，该标记对生殖生物技术的稳定性是动物生产中的主要问题。本研究使用光度甲基化测定法（LUMA）探索了激素诱导的两个精子中金鱼（Carassius auratus）和斑马鱼（Danio rerio）的精子的精子化和精子冷冻的影响。我们发现，尽管金鱼激素治疗确实增加了精子的产量，但它并没有改变精子的整体DNA甲基化。从相同的雄性中重复采集两个月的不同精子样本也显示出相同的总体DNA甲基化水平。同样，用甲醇冷冻保存金鱼精子后，总体DNA甲基化作用不受影响，而效率较低的冷冻保护剂（二甲基亚砜和1,2-丙二醇）则降低了DNA甲基化作用。相比之下，用甲醇冷冻保存斑马鱼的精子可引起总体DNA甲基化的轻微但显着的增加。在较不紧密的核中，即金鱼鳍体细胞中，无论选择何种冷冻保护剂，冷冻保存都不会改变整体DNA甲基化。总而言之，相对于生物技术，例如激素诱导的精子和精子冷冻保存，总体DNA甲基化是一个可靠的参数。