Mills Kayla M, Minton Amanda M, Ferreira Christina R
US Department of Agriculture, Agricultural Research Service, Beltsville Agricultural Research Center (BARC), Beltsville, MD 20705, USA.
AcuFast, Breese, IL 62230, USA.
Transl Anim Sci. 2024 Sep 19;8:txae139. doi: 10.1093/tas/txae139. eCollection 2024.
Multiple reaction monitoring (MRM) profiling is a sensitive method of lipid screening that has the capability to distinguish between different fertility phenotypes in gilts. However, MRM profiling has not yet been utilized to evaluate fertility phenotypes in boars. Markers indicative of fertility status in boars would be valuable as inclusion of subfertile boars in breeding programs results in a loss of efficiency and negative economic consequences. In addition, semen samples for lipidomic analysis are transported in liquid nitrogen or on dry ice to suspend metabolic activity within the sperm cells, however, these cryopreservation techniques are not commonly available at commercial boar studs. Therefore, the objective of this study was to develop a method of sample processing for MRM profiling that suspends metabolic activity within semen without freezing the sample. Five, sexually mature boars of similar genetics enrolled in a commercial breeding program were collected for the study. Following collection, ejaculates were aliquoted into methanol to suspend metabolic activity and shipped to Purdue University overnight for lipid extraction. Lipids were extracted using the Bligh and Dyer method and MRM profiling was used for lipid screening. A total of 329 ion transitions (MRMs) related to lipids were detected with most lipids being characterized as plasma membrane lipids (74%) which were comprised of phosphatidylcholines (40%), ceramides (16%), phosphatidylethanolamines (11%), and phosphatidylserines (7%). acylcarnitines (AC) represented approximately 8% of the ejaculate lipidome. Hierarchical cluster and principal component analysis revealed that boars have a distinct ejaculate lipidome profile based on major plasma membrane lipid classes. In addition, we observed that one boar was unique in his abundance of AC which are related to progressive motility and sperm cell metabolism. These results indicate that this method of sample processing for MRM profiling is suitable to be used to evaluate the lipidome of ejaculates from commercial boars and has the potential for broader applications across different livestock species in commercial environments.
多反应监测（MRM）分析是一种灵敏的脂质筛查方法，能够区分后备母猪的不同繁殖力表型。然而，MRM分析尚未用于评估公猪的繁殖力表型。由于繁殖计划中纳入亚生育力公猪会导致效率损失和负面经济后果，因此指示公猪繁殖力状态的标志物将具有重要价值。此外，用于脂质组学分析的精液样本需在液氮或干冰中运输，以暂停精子细胞内的代谢活动，然而，这些冷冻保存技术在商业公猪站并不常用。因此，本研究的目的是开发一种用于MRM分析的样本处理方法，该方法可在不冷冻样本的情况下暂停精液中的代谢活动。本研究收集了参与商业育种计划的5头遗传背景相似的性成熟公猪。采集后，将射精精液分装到甲醇中以暂停代谢活动，并连夜运往普渡大学进行脂质提取。使用布莱和戴尔方法提取脂质，并使用MRM分析进行脂质筛查。共检测到329个与脂质相关的离子跃迁（MRM），大多数脂质被鉴定为质膜脂质（74%），其中包括磷脂酰胆碱（40%）、神经酰胺（16%）、磷脂酰乙醇胺（11%）和磷脂酰丝氨酸（7%）。酰基肉碱（AC）约占射精脂质组的8%。层次聚类和主成分分析表明，基于主要质膜脂质类别，公猪具有独特的射精脂质组谱。此外，我们观察到一头公猪的AC丰度独特，AC与精子的前进运动和代谢有关。这些结果表明，这种用于MRM分析的样本处理方法适用于评估商业公猪射精精液的脂质组，并且在商业环境中具有在不同家畜物种中更广泛应用的潜力。
