Department of Life Sciences, Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal.
PLoS One. 2011 Mar 23;6(3):e18112. doi: 10.1371/journal.pone.0018112.
Human sperm samples are very heterogeneous and include a low amount of truly functional gametes. Distinct strategies have been developed to characterize and isolate this specific subpopulation. In this study we have used fluorescence microscopy and fluorescence-activated cell sorting to determine if mitochondrial function, as assessed using mitochondrial-sensitive probes, could be employed as a criterion to obtain more functional sperm from a given ejaculate. We first determined that mitochondrial activity correlated with the quality of distinct human samples, from healthy donors to patients with decreased semen quality. Furthermore, using fluorescence-activated cell sorting to separate sperm with active and inactive mitochondria we found that this was also true within samples. Indeed, sperm with active mitochondria defined a more functional subpopulation, which contained more capacitated and acrosome intact cells, sperm with lower chromatin damage, and, crucially, sperm more able to decondense and participate in early development using both chemical induction and injection into mature bovine oocytes. Furthermore, cell sorting using mitochondrial activity produced a more functional sperm subpopulation than classic swim-up, both in terms of improvement in a variety of functional sperm parameters and in statistical significance. In conclusion, whatever the true biological role of sperm mitochondria in fertilization, mitochondrial activity is a clear hallmark of human sperm functionality.
人类精子样本非常不均匀，其中真正有功能的配子数量很少。人们已经开发出了不同的策略来对这一特定的亚群进行特征描述和分离。在本研究中，我们使用荧光显微镜和荧光激活细胞分选来确定使用线粒体敏感探针评估的线粒体功能是否可以作为从特定精液中获得更多有功能精子的标准。我们首先确定线粒体活性与不同人类样本的质量相关，从健康供体到精液质量下降的患者。此外，使用荧光激活细胞分选分离具有活性和非活性线粒体的精子，我们发现这在样本内也是如此。事实上，具有活性线粒体的精子定义了一个更有功能的亚群，其中包含更多获能和顶体完整的细胞、具有较低染色质损伤的精子，而且至关重要的是，使用化学诱导和注入成熟牛卵母细胞，精子更有能力去浓缩和参与早期发育。此外，与经典的泳动法相比，使用线粒体活性进行细胞分选可以产生更有功能的精子亚群，无论是在改善各种功能精子参数方面，还是在统计学意义方面。总之，无论精子线粒体在受精中的真正生物学作用如何，线粒体活性都是人类精子功能的一个明显标志。
