Department of Animal Biochemistry and Biotechnology, Faculty of Animal Bioengineering, University of Warmia and Mazury in Olsztyn, 10-719 Olsztyn, Poland.
Laboratory of Mass Spectrometry Research Centre EIT+, Stabłowicka 147, 54-066 Wrocław, Poland.
Genes (Basel). 2021 Oct 2;12(10):1574. doi: 10.3390/genes12101574.
This study aimed to characterize the protein composition of fractionated seminal plasma (SP) by liquid chromatography mass spectrometry (LC-MS/MS) analysis and investigate its effects on survival of frozen-thaw (FT) boar spermatozoa following storage. Seminal plasma (SP) was fractionated by gel filtration chromatography to give two fractions, SP1 with more than 40 kDa (>40 kDa) and SP2 with less than 40 kDa (<40 kDa). SP1 and SP2 were subjected to LC-MS/MS and bioinformatics analysis. Following cryopreservation, FT boar semen ( = 7) was thawed in Beltsville Thawing Solution (BTS), BTS + SP1 or BTS + SP2, stored at different periods and subjected to post-thaw (PT) quality assessment. A total of 52 and 22 abundant proteins were detected in SP1 and SP2, respectively. FN1, ANGPTL1, and KIF15 proteins were more abundance in SP1, whereas a high abundance of spermadhesins (PSP-I and PSP-II) was detected in SP2. Proteins of the fractionated SP were involved in various biological processes, such as cell motility and signal transduction. The dominant pathway of SP1 proteins was the apelin signaling pathway (GNA13, MEF2D, SPHK2, and MEF2C), whereas a pathway related to lysosome (CTSH, CTSB, and NPC2) was mainly represented by SP2 proteins. In most of the boars, significantly higher motility characteristics, membrane integrity, and viability were observed in FT spermatozoa exposed to SP1 or SP2 compared with BTS. The results of our study confirm that a combination of several proteins from the fractionated SP exerted beneficial effects on the sperm membrane, resulting in improved quality characteristics following PT storage.
本研究旨在通过液相色谱-质谱联用（LC-MS/MS）分析对精液进行蛋白质组成特征分析，并研究其对冷冻-解冻（FT）后猪精子存活的影响。采用凝胶过滤色谱法将精液分为两个部分，SP1 部分大于 40 kDa (>40 kDa)，SP2 部分小于 40 kDa(<40 kDa)。对 SP1 和 SP2 进行 LC-MS/MS 和生物信息学分析。冷冻保存后，FT 公猪精液（n = 7）在贝尔茨维尔解冻液（BTS）、BTS+SP1 或 BTS+SP2 中解冻，在不同时间段储存并进行解冻后（PT）质量评估。SP1 和 SP2 中分别检测到 52 种和 22 种丰富蛋白。FN1、ANGPTL1 和 KIF15 蛋白在 SP1 中含量较高，而 SP2 中检测到高丰度的精子结合蛋白（PSP-I 和 PSP-II）。分离的 SP 蛋白参与多种生物学过程，如细胞运动和信号转导。SP1 蛋白的主要途径是阿片素信号通路（GNA13、MEF2D、SPHK2 和 MEF2C），而 SP2 蛋白主要代表与溶酶体相关的途径（CTSH、CTSB 和 NPC2）。在大多数公猪中，与 BTS 相比，暴露于 SP1 或 SP2 的 FT 精子的运动特性、膜完整性和活力明显更高。我们的研究结果证实，来自分离 SP 的几种蛋白质的组合对精子膜产生有益影响，导致解冻后储存的质量特征得到改善。
