Gutiérrez-Cepeda Luna, Crespo Francisco, Blazquez Juan Carlos, Serres Consuelo
Animal Medicine and Surgery Department, Veterinary Faculty, UCM, Avda. Puerta de Hierro s/n, 28040 Madrid, Spain.
Centro Militar de Cría Caballar (CCFAA), C/Arsenio Gutiérrez Palacios s/n, 05005 Ávila, Spain.
Animals (Basel). 2023 Jan 22;13(3):382. doi: 10.3390/ani13030382.
The Purebred Spanish Horse, according to our clinical experience, is characterized by having a high number of stallions that do not meet the international commercial recommendations for equine-sperm cryopreservation. This means that artificial insemination with frozen semen from these stallions is less widespread than in other breeds. In this study, we investigated if the incorporation of single-layer colloidal centrifugation prior to cryopreservation in clinical conditions could increase the number of ejaculates of Purebred Spanish stallions suitable for this processing, observing the influence of centrifugation and freezing extender protocol on post-thawed sperm motility. Using colloidal centrifugation, the percentage of ejaculates available to be frozen was increased from 35% (6/17) to 71% (12/17), doubling the number of samples that could have been subjected to cryopreservation. We only found significant differences in linearity (LIN) and lateral head displacement (ALH) after 5 min of incubation at 37 °C between colloidal and simple centrifugation processing techniques. No significant differences were found between the two different colloidal protocols in any of the variables considered. Colloidal centrifugation allowed us to obtain, from worse fresh-quality ejaculates, thawed sperm doses with similar quality to that of good-quality ejaculates. BotuCrio produced, in general, higher motility parameters and its characteristics than the other extenders analyzed, with significant differences found in comparison to Inra-Freeze and Lac-Edta in both total (MOT) and progressive motility (PMOT) when using colloidal centrifugation and only in PMOT when applying simple centrifugation. Colloidal centrifugation optimized the efficiency of cryopreservation, as it allowed us to increase the number of ejaculates of Purebred Spanish Horses suitable to be frozen. Including these semen processing techniques in the freeze test could help to optimize equine-sperm cryopreservation protocols, especially when dealing with individuals or breeds for which initially low sperm quality prevents or limits their inclusion in sperm cryopreservation programs.
根据我们的临床经验，纯种西班牙马的特点是，有大量种公马不符合马精液冷冻保存的国际商业建议。这意味着，用这些种公马的冷冻精液进行人工授精的情况不如其他品种普遍。在本研究中，我们调查了在临床条件下，冷冻保存前采用单层胶体离心法是否能增加适合该处理的纯种西班牙种公马的射精量，并观察离心和冷冻稀释液方案对解冻后精子活力的影响。使用胶体离心法，可用于冷冻的射精量百分比从35%（6/17）增加到71%（12/17），使可进行冷冻保存的样本数量增加了一倍。我们仅发现，在37℃孵育5分钟后，胶体离心和简单离心处理技术之间在线性度（LIN）和头部侧向位移（ALH）方面存在显著差异。在所考虑的任何变量中，两种不同的胶体方案之间均未发现显著差异。胶体离心法使我们能够从质量较差的新鲜射精样本中获得解冻后精子剂量，其质量与优质射精样本相似。总体而言，BotuCrio产生的活力参数及其特性高于所分析的其他稀释液，在使用胶体离心法时，与Inra - Freeze和Lac - Edta相比，在总活力（MOT）和渐进性活力（PMOT）方面均存在显著差异，而在应用简单离心法时，仅在PMOT方面存在显著差异。胶体离心法优化了冷冻保存的效率，因为它使我们能够增加适合冷冻的纯种西班牙马的射精量。在冷冻测试中纳入这些精液处理技术有助于优化马精液冷冻保存方案，特别是在处理个体或品种时，其初始精子质量较低会阻碍或限制它们纳入精子冷冻保存计划。
