影响马精液离心后精子回收率和存活率的因素。
Factors affecting sperm recovery rates and survival after centrifugation of equine semen.
机构信息
Equine Health Studies Program, Department of Veterinary Clinical Sciences, Louisiana State University, Baton Rouge, Louisiana, USA.
出版信息
Theriogenology. 2012 Nov;78(8):1814-23. doi: 10.1016/j.theriogenology.2012.07.011. Epub 2012 Sep 10.
Conventional centrifugation protocols result in important sperm losses during removal of the supernatant. In this study, the effect of centrifugation force (400 or 900 × g), duration (5 or 10 min), and column height (20 or 40 mL; Experiment 1); sperm concentration (25, 50, and 100 × 10(6)/mL; Experiment 2), and centrifugation medium (EZ-Mixin CST [Animal Reproduction Systems, Chino, CA, USA], INRA96 [IMV Technologies, Maple Grove, MN, USA], or VMDZ [Partnar Animal Health, Port Huron, MI, USA]; Experiment 3) on sperm recovery and survival after centrifugation and cooling and storage were evaluated. Overall, sperm survival was not affected by the combination of centrifugation protocol and cooling. Total sperm yield was highest after centrifugation for 10 min at 400 × g in 20-mL columns (95.6 ± 5%, mean ± SD) or 900 × g in 20-mL (99.2 ± 0.8%) or 40-mL (91.4 ± 4.5%) columns, and at 900 × g for 5 min in 20-mL columns (93.8 ± 8.9%; P < 0.0001). Total (TMY) and progressively motile sperm yield followed a similar pattern (P < 0.0001). Sperm yields were not significantly different among samples centrifuged at various sperm concentrations. However, centrifugation at 100 × 10(6)/mL resulted in significantly lower total sperm yield (83.8 ± 10.7%) and TMY (81.7 ± 6.8%) compared with noncentrifuged semen. Centrifugation in VMDZ resulted in significantly lower TMY (69.3 ± 22.6%), progressively motile sperm yield (63.5 ± 18.2%), viable yield (60.9 ± 36.5%), and survival of progressively motile sperm after cooling (21 ± 10.8%) compared with noncentrifuged semen. In conclusion, centrifuging volumes of ≤ 20 mL minimized sperm losses with conventional protocols. With 40-mL columns, it may be recommended to increase the centrifugal force to 900 × g for 10 min and dilute the semen to a sperm concentration of 25 to 50 × 10(6)/mL in a milk- or fractionated milk-based medium. The semen extender VMDZ did not seem well suited for centrifugation of equine semen.
传统的离心方案会导致在去除上清液的过程中丢失大量精子。在这项研究中，我们研究了离心力（400 或 900×g）、持续时间（5 或 10 分钟）和柱高（20 或 40 mL；实验 1）；精子浓度（25、50 和 100×10(6)/mL；实验 2）和离心介质（EZ-Mixin CST[Animal Reproduction Systems，Chino，CA，USA]、INRA96[IMV Technologies，Maple Grove，MN，USA]或 VMDZ[Partnar Animal Health，Port Huron，MI，USA]；实验 3）对离心冷却和储存后精子回收和活力的影响。总体而言，离心方案和冷却对精子活力没有影响。在 20-mL 柱中以 400×g 离心 10 分钟（95.6±5%，平均值±标准差）或 900×g 离心 20-mL 柱（99.2±0.8%）或 40-mL 柱（91.4±4.5%），或在 20-mL 柱中以 900×g 离心 5 分钟，精子总回收率最高（93.8±8.9%；P<0.0001）。总（TMY）和前向运动精子的产量也遵循类似的模式（P<0.0001）。在不同精子浓度下离心的精子产量之间没有显著差异。然而，与未经离心的精液相比，以 100×10(6)/mL 离心会导致总精子产量（83.8±10.7%）和 TMY（81.7±6.8%）显著降低。在 VMDZ 中离心会导致 TMY（69.3±22.6%）、前向运动精子产量（63.5±18.2%）、活力产量（60.9±36.5%）和冷却后前向运动精子活力（21±10.8%）显著降低，与未经离心的精液相比。总之，使用传统方案时，将离心体积控制在 20 mL 以下可最大程度地减少精子损失。对于 40-mL 柱，建议增加离心力至 900×g 离心 10 分钟，并将精液稀释至 25 至 50×10(6)/mL 的精子浓度，使用牛奶或分馏牛奶基介质。精液稀释剂 VMDZ 似乎不适合马精液的离心。
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