Giaretta E, Damato A, Zennaro L, Bonfatti V, Mislei B, Vigolo V, Falomo M E, Bertuzzo F, Gabai G, Bucci D
Department of Comparative Biomedicine and Food Science, University of Padova, Via Dell'Università 6, 35020, Legnaro, PD, Italy.
Institute of Biomolecular Chemistry, CNR, Padova Unit, Department of Chemistry University of Padova, Via Marzolo 1, 35131, Padova, Italy.
Theriogenology. 2025 Mar 15;235:262-274. doi: 10.1016/j.theriogenology.2025.01.015. Epub 2025 Jan 17.
Seminal plasma composition has important role in sperm functionality and its freezability. The objective of this study was to test the hypothesis that seminal plasma (SP) oxidative status and metabolome are associated with fresh semen characteristics and freezability of bull sperm. To accomplish this objective, oxidative status markers and metabolome of SP of ejaculates obtained from 20 Holstein bulls (3 for each bull) were analyzed using spectrophotometry and nuclear magnetic resonance (H NMR). The ejaculates were classified into higher motility fresh semen (HMF) and lower motility fresh semen (LMF), according to total motility (TM) and progressive motility (PM) values of fresh semen. Then the ejaculates was cryopreserved and assigned to higher motility thawed group (HMT) or lower motility thawed group (LMT) according to TM and PM at 0 h post-thawing. Multivariate analyses were performed to identify the association between the functional characteristics of fresh and thawed semen and the SP parameters, in terms of the oxidative status and the metabolomic composition. According to our results, the advanced oxidative protein products (AOPP) and thiol concentrations in SP are significantly related to some physiological characteristics of the thawed sperm, such as higher viability, TM, PM and LIN and lower mitochondrial and cytoplasmic superoxide production in viable thawed cells. In contrast, a higher amount of C in the SP was negatively related to TM and PM of thawed semen and was associated with higher mitochondrial and cytoplasmic superoxide production. In addition, partial least squares-discriminant analysis (PLS-DA) performed on the H NMR spectra indicated a discrete separation between HMF and LMF groups, and good discrimination between HMT and LMT groups. Higher levels of formic acid, lactate, glycerol and phosphocholine, were found in the SP of the HMF group than in the LMF group. On the other hand, alanine, phenylalanine, and tyrosine were higher in the SP of the LMF group than in the HMF group. GABA, glutamate, histidine and glycerol were found in higher concentrations in the HMT group than in the LMT group, while fructose decreased in the HMT group. Our results showed that the oxidative and metabolomic status of SP is related to the physiological properties of semen and its freezability and open new fields in research of SP biomarkers of bull semen preservation and fertility.
精浆成分在精子功能及其冷冻保存能力方面起着重要作用。本研究的目的是验证以下假设：精浆（SP）的氧化状态和代谢组与公牛精子的新鲜精液特征及冷冻保存能力相关。为实现这一目标，使用分光光度法和核磁共振（¹H NMR）对从20头荷斯坦公牛采集的射精样本（每头公牛3份）的精浆氧化状态标志物和代谢组进行了分析。根据新鲜精液的总活力（TM）和前向运动率（PM）值，将射精样本分为高活力新鲜精液（HMF）组和低活力新鲜精液（LMF）组。然后将射精样本进行冷冻保存，并根据解冻后0小时的TM和PM分为高活力解冻组（HMT）或低活力解冻组（LMT）。进行多变量分析以确定新鲜和解冻精液的功能特征与精浆参数在氧化状态和代谢组组成方面的关联。根据我们的结果，精浆中的晚期氧化蛋白产物（AOPP）和硫醇浓度与解冻精子的一些生理特征显著相关，例如解冻后活力更高、TM、PM和LIN更高，以及存活解冻细胞中线粒体和细胞质超氧化物产生更低。相反，精浆中较高含量的C与解冻精液的TM和PM呈负相关，并与较高的线粒体和细胞质超氧化物产生相关。此外，对¹H NMR光谱进行的偏最小二乘判别分析（PLS - DA）表明HMF组和LMF组之间有明显分离，HMT组和LMT组之间有良好区分。HMF组精浆中的甲酸、乳酸、甘油和磷酸胆碱水平高于LMF组。另一方面，LMF组精浆中的丙氨酸、苯丙氨酸和酪氨酸高于HMF组。HMT组中的γ - 氨基丁酸（GABA）、谷氨酸、组氨酸和甘油浓度高于LMT组，而HMT组中的果糖减少。我们的结果表明，精浆的氧化和代谢组状态与精液的生理特性及其冷冻保存能力相关，并为公牛精液保存和生育力的精浆生物标志物研究开辟了新领域。
