不同获能方法对冻融塔里木马鹿精子体外获能及其蛋白酪氨酸磷酸化的影响

为探讨不同获能方法对塔里木马鹿精子体外获能及其蛋白酪氨酸磷酸化水平的影响,冻融塔里木马鹿精子随机分为4组,用钙离子载体、肝素、咖啡因和 Percool 离心4种方法进行精子获能的诱导,利用金霉素（CTC）染色法评价精子获能状态,采用 SDS-PAGE 分离精子膜蛋白,进行 Western blotting 免疫印迹分析,检测酪氨酸磷酸化蛋白的表达水平.结果显示,冻融精子经4种精子获能方法处理后,肝素诱发的精子获能率显著高于钙离子载体组和 Percool 组（P ＜0．05）.钙离子载体组、肝素组和咖啡因组精子蛋白酪氨酸磷酸化水平高于 Percool 组和对照组.另外,冻融精子随着上游处理及肝素诱导获能的进行,检测到分子量分别为14,25~30,40,47,55 ku 的酪氨酸磷酸化蛋白,这些蛋白的酪氨酸磷酸化水平在获能60~120 min 期间相对较高,而且此时精子获能率及超激活运动精子比例也显著提高（P ＜0．05）.结果提示,肝素可以较好的诱导马鹿精子获能,马鹿精子获能与蛋白酪氨酸磷酸化相关.

Effects of Different Capacitation Methods on Capacitation and Protein Tyrosine Phosphorylation of Cervus elaphus yarkandensis Frozen-thawed Sperm

In order to discuss the effects of different capacitation methods on capacitation and protein tyrosine phosphorylation of Cervus elaphus yarkandensis sperm and frozen-thawed Cervus elaphus yarkandensis sperms were randomly divided into 4 groups and treated with A23187 （IA）,Heparin,caffeine or Percool for capacitation.The sperm capacitation was evaluated by Aureomycin （CTC）method.Expres-sion of sperm protein tyrosine phosphorylation was detected by membrane-protein separation on SDS-PAGE and Western blotting respectively.The results indicated that sperm capacitation rate in treatment of heparin - induced was significantly higher than the IA treatment and Percool treatment （P 〈 0 .0 5 ）.The expression of sperm protein tyrosine phosphorylation in each group of IA,heparin and caffeine were higher than that of the Percool treatment and the control.Moreover,when freezen-thawed sperm was treated with swim-up and heparin-induced capacitation were conducted,the molecular weight of 14 ku,25 - 30 ku, 40 ku,47 ku and 55 ku protein tyrosine phosphorylation was detected respectively,these proteins tyrosine phosphorylation were increased,when the frozen-thawed sperms capacitated 60-120 mins,and the capaci-tated sperm rate and hyper-activated sperm rate were increased significantly （P 〈0.05）.The results sug-gested that sperms of Cervus elaphus yarkandensis were capacitated properly with heparin and protein tyrosine phosphorylation was associated with the capacitation of Cervus elaphus yarkandensis sperm.