title: Investigations of motility and fertilization potential in thawed cryopreserved mouse sperm from cold-stored epididymides.
abstract: Cold transport of epididymides from genetically modified mice is an efficient alternative to the shipment of live animals between research facilities. Mouse sperm from epididymides cold-stored for short periods can maintain viability. We previously reported that cold storage of mouse epididymides in Lifor® perfusion medium prolonged sperm motility and fertilization potential and that the sperm efficiently fertilized oocytes when reduced glutathione was added to the fertilization medium. Cryopreservation usually results in decreased sperm viability; an optimized protocol for cold storage of epididymides plus sperm cryopreservation has yet to be established. Here, we examined the motility and fertilization potential of cryopreserved, thawed (frozen-thawed) sperm from previously cold-stored mouse epididymides. We also examined the protective effect of sphingosine-1-phosphate (S1P) on sperm viability when S1P was added to the preservation medium during cold storage. We assessed viability of frozen-thawed sperm from mouse epididymides that had been cold-transported domestically or internationally and investigated whether embryos fertilized in vitro with these sperm developed normally when implanted in pseudo-pregnant mice. Our results indicate that frozen-thawed sperm from epididymides cold-stored for up to 48 h maintained high fertilization potential. Fertilization potential was reduced after cold storage for 72 h, but not if S1P was included in the cold storage medium. Live pups were born normally to recipients after in vitro fertilization using frozen-thawed sperm from cold-transported epididymides. In summary, we demonstrate an improved protocol for cold-storage of epididymides that can facilitate transport of genetically engineered-mice and preserve sperm viability after cryopreservation.
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Takeo
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
authors.forename: Kiyoko
authors.initials: K
authors.lastname: Fukumoto
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
authors.forename: Tomoko
authors.initials: T
authors.lastname: Kondo
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
authors.forename: Yukie
authors.initials: Y
authors.lastname: Haruguchi
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
authors.forename: Yumi
authors.initials: Y
authors.lastname: Takeshita
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
authors.forename: Yuko
authors.initials: Y
authors.lastname: Nakamuta
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
authors.forename: Shuuji
authors.initials: S
authors.lastname: Tsuchiyama
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
authors.forename: Hidetaka
authors.initials: H
authors.lastname: Yoshimoto
authors.affiliation: Animal Laboratory for Medical Research, Asahikawa Medical University, 2-1-1-1 Midorigaoka-Higashi, Asahikawa 078-8510, Japan.
authors.forename: Norihiko
authors.initials: N
authors.lastname: Shimizu
authors.affiliation: Mouse Biology Program, University of California, Davis, CA 95618, USA.
authors.forename: Ming-Wen
authors.initials: MW
authors.lastname: Li
authors.affiliation: Mouse Biology Program, University of California, Davis, CA 95618, USA.
authors.forename: Kristy
authors.initials: K
authors.lastname: Kinchen
authors.affiliation: Mouse Biology Program, University of California, Davis, CA 95618, USA.
authors.forename: Jadine
authors.initials: J
authors.lastname: Vallelunga
authors.affiliation: Mouse Biology Program, University of California, Davis, CA 95618, USA.
authors.forename: K C Kent
authors.initials: KC
authors.lastname: Lloyd
authors.affiliation: Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan. Electronic address: nakagata@kumamoto-u.ac.jp.
authors.forename: Naomi
authors.initials: N
authors.lastname: Nakagata
mesh_terms: D000818:Animals; D015925:Cryopreservation / Q000379:methods*; D010064:Embryo Implantation / Q000187:drug effects / Q000502:physiology; D004822:Epididymis / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D005260:Female; D005306:Fertilization / Q000187:drug effects / Q000502:physiology; D005307:Fertilization in Vitro / Q000379:methods; D005615:Freezing; D005978:Glutathione / Q000494:pharmacology*; D008246:Lysophospholipids / Q000494:pharmacology*; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D019188:Organ Preservation Solutions / Q000737:chemistry; D011247:Pregnancy; D013081:Sperm Motility / Q000187:drug effects* / Q000502:physiology; D013094:Spermatozoa / Q000187:drug effects* / Q000502:physiology; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: D008246:Lysophospholipids; D019188:Organ Preservation Solutions; C060506:sphingosine 1-phosphate; D005978:Glutathione; D013110:Sphingosine
keywords: Cold storage; Cold transport; Epididymides; In vitro fertilization; Motility; Sperm; Sphingosine-1-phosphate