title: Optimization of ethylene glycol concentrations, freezing rates and holding times in liquid nitrogen vapor for cryopreservation of rhesus macaque (Macaca mulatta) sperm.
abstract: Ethylene glycol (EG) has been speculated to be the most appropriate penetrating cryoprotectant for cryopreservation of rhesus macaque sperm due to its higher permeability coefficient. The present study aimed to determine the optimal EG concentration, freezing rate and holding time in liquid nitrogen (LN(2)) vapor for rhesus sperm cryopreservation. Among six tested EG concentrations (0, 0.18, 0.35, 0.7, 1.4 and 2.1 M), 0.7 M EG showed the most effective cryoprotection (P<0.05). Sperm frozen with 0.7 M EG at -183°C/min showed higher post-thaw motility than sperm frozen at -10, -67 or -435°C/min (P<0.05). Sperm frozen in LN(2) vapor at -183°C/min with 0.7 M EG and a holding time of 10 min showed higher post-thaw motility compared with a holding time of 5 or 15 min (P<0.05). The function of sperm cryopreserved at the optimized EG concentration, freezing rate and holding time was further evaluated by in vitro fertilization. Of the 36 oocytes collected from gonadotropin-stimulated rhesus macaques, 61.1% were fertilized, and 61.1, 44.4 and 36.1% of the oocytes developed to 2 cells, morulae and blastocysts, respectively. Our findings provide an alternative penetrating cryoprotectant and optimal protocol for genetic preservation purposes in this important species.
authors.affiliation: Laboratory of Moleculac Genetics of Aging and Tumor, Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming 650224, PR China.
authors.forename: Shihua
authors.initials: S
authors.lastname: Yang
authors.forename: Shuhuang
authors.initials: S
authors.lastname: Ping
authors.forename: Wei
authors.initials: W
authors.lastname: Si
authors.forename: Xiechao
authors.initials: X
authors.lastname: He
authors.forename: Xinyi
authors.initials: X
authors.lastname: Wang
authors.forename: Yongqing
authors.initials: Y
authors.lastname: Lu
authors.forename: Shaohui
authors.initials: S
authors.lastname: Ji
authors.forename: Yuyu
authors.initials: Y
authors.lastname: Niu
authors.forename: Weizhi
authors.initials: W
authors.lastname: Ji
mesh_terms: D000177:Acrosome / Q000502:physiology; D000818:Animals; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D003451:Cryoprotective Agents; D019855:Ethylene Glycol; D005260:Female; D005307:Fertilization in Vitro / Q000662:veterinary; D008253:Macaca mulatta; D008297:Male; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility / Q000502:physiology; D013094:Spermatozoa
chemical_list: D003451:Cryoprotective Agents; D019855:Ethylene Glycol