不同抗氧化剂组合对蒙古马精液冷冻保存效果和抗氧化相关指标的影响

Effects of antioxidants combination on cryopreservation and antioxidant activity of equine frozen semen

关键词:  蒙古马  精液  冷冻保存  抗氧化剂  抗氧化酶

Key words:  Mongolian horse  spermatozoa  cryopreservation  antioxidants  antioxidant activity

摘要:

为研究不同抗氧化剂对蒙古马冷冻精液的品质和抗氧化相关指标的影响，选取10匹蒙古公马作为试验动物，以INRA82作为基础稀释液(对照组)，分别添加谷胱甘肽(0、5.0、7.0、10.0和20.0 mg/mL)、海藻糖(0、25.0、35.0和50.0 mg/mL)和联合添加谷胱甘肽+海藻糖((0+0)、(5.0+35.0)、(5.0+50.0)、(7.0+35.0)和(7.0+50.0)mg/mL)制备成含有不同氧化剂的冷冻保护液，将浓缩处理后的马精子分别置于稀释液中稀释、冷冻和解冻，利用精子常规品质检测方法和酶联免疫分析法，检测运动精子比例(TM)、前进运动精子比例(PM)、质膜完整率(MI)、顶体完整率(AI)及丙二醛(MDA)含量、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-PX)和谷胱甘肽还原酶(GR)的活性。结果显示:1)5.0 mg/mL谷胱甘肽对马精液TM、PM和MI均有提高，可降低精液中MDA含量，可显著提高CAT和GSH-PX活性(P<0.05)；35.0和50.0 mg/mL海藻糖可提高精子TM、PM、AI和MI，且35.0 mg/mL海藻糖可显著降低MDA含量(P<0.05)；2)联合添加5.0 mg/mL谷胱甘肽和35.0 mg/mL海藻糖可显著提升马精液的TM、PM、AI和MI(P<0.05)，还可显著降低MDA含量(P<0.05)，提高SOD活性和GSH-PX活性。综上，联合添加5.0 mg/mL谷胱甘肽和35.0 mg/mL海藻糖一定程度上可通过改变酶活性，对精子氧化应激产生保护作用，提高马精液冷冻-解冻后质量。

Abstract:

This study was conducted to evaluate the effects of antioxidants combination on stallion sperm quality during freeze-thaw process. Sperms of ten stallions(3 or 4 individuals each time)were diluted into different levels of glutathione(0, 5. 0, 7. 0, 10. 0, 20. 0 mg/mL), trehalose(0, 25. 0, 35. 0, 50. 0 mg/mL), and the combination of glutathione and trehalose((0+0), (5. 0+35. 0), (5. 0+50. 0), (7. 0+35. 0), (7. 0+50. 0)mg/mL), which were supplemented with INRA82 extender and then subjected to cryopreservation. Sperm motility(TM), progressive motility(PM), plasma membrane integrity(MI), acrosome integrity(AI)and its malondialdehyde(MDA)content, activities of superoxide dismutase(SOD), catalase(CAT), glutathione peroxidase(GSH-PX)and glutathione reductase(GR)were investigated by sperm normal quality test and enzyme linked immunosorbent assay(ELISA). The results showed that: 1)In 5. 0 mg/mL glutathione group, the TM, PM and MI were higher than that in other groups, MDA level was lower than that in other groups, and addition of glutathione significantly improveed the activity of CAT and GSH-PX(P<0. 05). In 35 and 50 mg/mL trehalose groups, TM, PM, AI and MI were higher than that in other groups, and 35. 0 mg/mL trehalose significantly reduceed MDA content(P<0. 05); 2)The combination of glutathione and trehalose(5. 0+35. 0 mg/mL)significantly increaseed TM, PM, AI, and MI(P<0. 05), significantly reduceed MDA content(P<0. 05), and improveed the activity of SOD and GSH-PX. In conclusion, the combined addition of 5. 0 mg/mL glutathione with 35. 0 mg/mL trehalose protected the sperm from oxidative stress in a certain extent by changing the enzyme activity, thus improving the semen quality after frozen-thawed.