Yu Il-Jeoung
Department of Theriogenology and Reproductive Biotechnologies, College of Veterinary Medicine and Bio-safety Research Institute, Chonbuk National University, Jeonju, Republic of Korea.
Cryo Letters. 2014 Mar-Apr;35(2):101-7.
The study is to develop a glycerol-free extender using glucose for canine sperm cryopreservation,.
Tris [hydroxymethyl] aminomethane (TRIS) extender. Canine sperm were cooled to 4 degree C in TRIS containing 44.4 mM glucose for 100 min, and then cooled at 4 degree C in TRIS with glucose concentrations of 0 mM, 44.4 mM, 100 mM, 200 mM, or 300 mM for 30 min followed by cryopreservation. After thawing at 37 degree C for 25 sec, sperm motility, viability, and morphological abnormalities were evaluated. In addition, 300 mM glucose-TRIS was compared to TRIS extenders with a final concentration of 5 % glycerol. Sperm phosphatidylserine (PS) translocation after freezing and thawing was assayed by flow cytometry using Annexin V-FITC apoptosis detection kit.
Progressive motility and viability (42% and 41%, respectively) were significantly higher in the 300mM group than the other groups with lower concentrations of glucose (P < 0.05). PS translocation index was significantly lower in 300mM glucose-TRIS than those in extenders with glycerol (85 vs 93, P < 0.05).
These results indicate that cryopreservation of canine sperm using glycerol-free 300 mM glucose-TRIS is feasible and yields more motile sperm with lower PS translocation compared with extenders containing glycerol.
本研究旨在开发一种使用葡萄糖的无甘油稀释液用于犬精子冷冻保存。
采用三羟甲基氨基甲烷（TRIS）稀释液。将犬精子在含有44.4 mM葡萄糖的TRIS中冷却至4℃ 100分钟，然后在葡萄糖浓度为0 mM、44.4 mM、100 mM、200 mM或300 mM的TRIS中于4℃冷却30分钟，随后进行冷冻保存。在37℃解冻25秒后，评估精子活力、存活率和形态异常情况。此外，将300 mM葡萄糖 - TRIS与最终浓度为5%甘油的TRIS稀释液进行比较。使用膜联蛋白V - FITC凋亡检测试剂盒通过流式细胞术检测冻融后精子磷脂酰丝氨酸（PS）的易位情况。
300 mM组的前向运动性和存活率（分别为42%和41%）显著高于其他葡萄糖浓度较低的组（P < 0.05）。300 mM葡萄糖 - TRIS中的PS易位指数显著低于含甘油的稀释液（85对93，P < 0.05）。
这些结果表明，与含甘油的稀释液相比，使用无甘油的300 mM葡萄糖 - TRIS冷冻保存犬精子是可行的，且能产生更多活力精子，PS易位率更低。
