Secciani Federica, Bianchi Laura, Ermini Leonardo, Cianti Riccardo, Armini Alessandro, La Sala Giovan Battista, Focarelli Riccardo, Bini Luca, Rosati Floriana
Laboratory of Cell Biology, Department of Evolutionary Biology, Siena University, Siena, Italy.
J Proteome Res. 2009 Jul;8(7):3377-89. doi: 10.1021/pr900031r.
Freshly ejaculated sperm acquire the fertilizing potential by a continuing process that occurs during sperm transport through the female genital tract, and it is physiologically not complete until the spermatozoon reaches the oocyte. The process termed capacitation can be mimicked in vitro by using appropriate capacitation media. Despite its importance, the molecular mechanisms underlying capacitation are poorly understood. This work deals with a proteomic approach to the analysis of protein profile variations in human normospermic samples as a consequence of three hours in vitro capacitation. 2DE gels were produced per freshly ejaculated sperm and per capacitated sperm and several quantitative and qualitative significant variations were found. Among the MS obtained identifications, proteins with a significant decrease after capacitation were found to be involved in protein fate, metabolism, and flagellar organization; on the contrary, increasing proteins were found to be related to cellular stress. Interestingly, the detected flagellar organization proteins decreased during capacitation whereas their corresponding fragments increased. A swim-up selected and three-hour capacitated sperm subpopulation has also been resolved by 2DE, and its synthetic gel has been analyzed for the variations observed in the entire sperm population. An immunofluorescence analysis of this sperm typology was carried out with antiactin and antitubulin antibodies.
刚射出的精子通过在雌性生殖道内运输过程中持续发生的一个过程获得受精潜能，并且在精子到达卵母细胞之前，其生理过程并不完整。通过使用合适的获能培养基，可以在体外模拟称为获能的过程。尽管其很重要，但获能背后的分子机制仍知之甚少。这项工作采用蛋白质组学方法来分析人正常精子样本在体外获能三小时后蛋白质谱的变化。针对刚射出的精子和获能后的精子分别制备了二维凝胶，发现了一些定量和定性的显著差异。在质谱鉴定结果中，获能后显著减少的蛋白质参与蛋白质命运、代谢和鞭毛组织；相反，增加的蛋白质与细胞应激有关。有趣的是，检测到的鞭毛组织蛋白在获能过程中减少，而其相应片段增加。还通过二维凝胶电泳分离了上浮选择并获能三小时的精子亚群，并对其合成凝胶进行了分析，以观察整个精子群体中观察到的变化。用抗肌动蛋白和抗微管蛋白抗体对这种精子类型进行了免疫荧光分析。
