PDF(Pubmed)
Abstract：
Kaempferol (KAE) is a natural flavonoid with powerful reactive oxygen species (ROS) scavenging properties and beneficial effects on ex vivo sperm functionality. In this paper, we studied the ability of KAE to prevent or ameliorate structural, functional or oxidative damage to frozen-thawed bovine spermatozoa. The analysis focused on conventional sperm quality characteristics prior to or following thermoresistance tests, namely the oxidative profile of semen alongside sperm capacitation patterns, and the levels of key proteins involved in capacitation signaling. Semen samples obtained from 30 stud bulls were frozen in the presence of 12.5, 25 or 50 μM KAE and compared to native ejaculates (negative control-CtrlN) as well as semen samples cryopreserved in the absence of KAE (positive control-CtrlC). A significant post-thermoresistance test maintenance of the sperm motility (p < 0.001), membrane (p < 0.001) and acrosome integrity (p < 0.001), mitochondrial activity (p < 0.001) and DNA integrity (p < 0.001) was observed following supplementation with all KAE doses in comparison to CtrlC. Experimental groups supplemented with all KAE doses presented a significantly lower proportion of prematurely capacitated spermatozoa (p < 0.001) when compared with CtrlC. A significant decrease in the levels of the superoxide radical was recorded following administration of 12.5 (p < 0.05) and 25 μM KAE (p < 0.01). At the same time, supplementation with 25 μM KAE in the cryopreservation medium led to a significant stabilization of the activity of Mg2+-ATPase (p < 0.05) and Na+/K+-ATPase (p < 0.0001) in comparison to CtrlC. Western blot analysis revealed that supplementation with 25 μM KAE in the cryopreservation medium prevented the loss of the protein kinase A (PKA) and protein kinase C (PKC), which are intricately involved in the process of sperm activation. In conclusion, we may speculate that KAE is particularly efficient in the protection of sperm metabolism during the cryopreservation process through its ability to promote energy synthesis while quenching excessive ROS and to protect enzymes involved in the process of sperm capacitation and hyperactivation. These properties may provide supplementary protection to spermatozoa undergoing the freeze-thaw process.
摘要：
山奈酚（KAE）是一种天然类黄酮，具有强大的活性氧（ROS）清除特性和对离体精子功能的有益作用。在本文中,我们研究了KAE预防或改善结构的能力，对冻融牛精子的功能性或氧化性损伤。该分析集中在耐热性测试之前或之后的常规精子质量特征。即精液的氧化谱和精子获能模式，以及参与获能信号传导的关键蛋白质的水平。在12.5、25或50μM的KAE存在下冷冻从30头公牛获得的精液样品,并与天然射精(阴性对照-CtrlN)以及在不存在KAE的情况下冷冻保存的精液样品(阳性对照-CtrlC)进行比较。一个显著的后热阻试验维持精子运动(p<0.001),膜（p<0.001）和顶体完整性（p<0.001），与CtrlC相比,在补充所有KAE剂量后观察到线粒体活性(p<0.001)和DNA完整性(p<0.001)。当与CtrlC相比时,补充有所有KAE剂量的实验组呈现显著较低比例的过早获能精子(p<0.001)。在施用12.5(p<0.05)和25μMKAE(p<0.01)后记录到超氧自由基水平的显著降低。同时,与CtrlC相比,在冷冻保存培养基中补充25μM的KAE导致Mg2+-ATP酶(p<0.05)和Na+/K+-ATP酶(p<0.0001)的活性显著稳定。蛋白质印迹分析显示，在冷冻保存培养基中补充25μM的KAE可防止蛋白激酶A（PKA）和蛋白激酶C（PKC）的丢失，这些都与精子激活过程密切相关。总之,我们可能推测KAE在冷冻保存过程中通过促进能量合成同时抑制过量的ROS以及保护参与精子获能和过度激活过程的酶的能力，在保护精子代谢方面特别有效。这些性质可以为经历冻融过程的精子提供补充保护。
