The optimization of sperm cryopreservation protocols in ram is a feasible tool to reinforce artificial insemination technologies considering the desirable application of sperm by vaginal/cervical or transcervical deposition. Cryopreservation provokes different types of damage on spermatozoa and many of these detrimental effects are triggered by redox deregulation. For this reason, the antioxidant supplementation in sperm cryopreservation protocols to decrease reactive oxygen species (ROS) levels and to equilibrate redox status has been widely employed in different species. Despite this, more fertility trials are necessary to provide the definitive tool to ensure the antioxidant effectiveness on sperm quality. For this reason, in this work, we performed a multiparametric analysis of some previously tested antioxidants (crocin, GSH and Trolox) on ram sperm cryopreservation from field trials to sperm quality analyses focused on new strategies to measure redox balance. Attending to fertility trial, Trolox supplementation registered an improvement concerning to fertility (when we considered high fertility males) and multiple lambing frequency and other complementary and descriptive data related to lambing performance such as prolificacy and fecundity. This positive effect was more evident in multiple lambing frequency when we considered low fertility males than in global male analysis. In vitro analyses of sperm quality confirmed in vivo trials registering a positive effect on sperm viability and redox balance. In this study, we provided the definitive evidence that the role of trolox on redox balance maintenance has a direct effect on fertility parameters, such as prolificacy. The effectiveness of antioxidant treatments was tested, for the first time in ovine species, using an integrative and multiparametric approach combining in vivo and in vitro analyses and novel approaches, such as RedoxSYS. These types of strategies should be applied to improve sperm conservation methods and optimize AI technologies upgrading the correlation between in vitro and in vivo analyses.

中文翻译：

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抗氧化作用对精子冷冻保存的多参数研究-从田间试验到研究工作

考虑到通过阴道/子宫颈或经子宫颈沉积的精子的理想应用，优化ram中精子冷冻方案的方法是增强人工授精技术的可行工具。冷冻保存会对精子造成不同类型的损害，其中许多有害作用是由氧化还原调节失调引起的。因此，在精子冷冻保存方案中添加抗氧化剂以降低活性氧（ROS）水平并平衡氧化还原状态已在不同物种中广泛使用。尽管如此，还需要进行更多的生育力试验以提供确定的工具，以确保抗氧化剂对精子质量的有效性。因此，在这项工作中，我们对一些先前测试过的抗氧化剂（crocin，从实地试验到精子质量分析的ram精子冷冻保存的GSH和Trolox）重点是测量氧化还原平衡的新策略。参加生育力试验时，Trolox补充剂在生育力（当我们考虑高生育力的男性）和多个产羔频率以及与产羔性能有关的其他补充性和描述性数据（例如多产和繁殖力）方面取得了进步。当我们考虑低生育力的雄性时，这种积极作用在多重产程频率下比在全球雄性分析中更为明显。精子质量的体外分析证实了体内试验对精子活力和氧化还原平衡具有积极作用。在这项研究中，我们提供了明确的证据，表明trolox在维持氧化还原平衡方面的作用对生育参数有直接影响，例如多产。抗氧化剂治疗的有效性首次在绵羊物种中使用结合体内和体外分析以及新颖方法（例如RedoxSYS）的集成和多参数方法进行了测试。这些类型的策略应应用于改善精子保存方法和优化AI技术，从而提高体外和体内分析之间的相关性。