Effect of sugar molecules on the cryopreservation of mouse spermatogonial stem cells.
OBJECTIVE
To study the influence of sugars and establish a serum-free freezing method for the cryopreservation of spermatogonial stem cells (SSCs).


METHODS
Animal study.


METHODS
University laboratory.


METHODS
C57BL/6-TgEGFP, C57BL/6 mice.


METHODS
Germ cells enriched from testis cells were frozen using standard freezing medium containing sugars, including monosaccharides, disaccharides, and trisaccharides at 50, 100, and 200 mM, respectively. To study the feasibility of establishing a serum-free freezing method, fetal bovine serum was substituted with knockout serum replacement.


METHODS
Freeze-thawed germ cells were evaluated for recovery rate, proliferation capacity, and stem cell activity after transplantation to recipient testes.


RESULTS
Supplementation of freezing medium with 200 mM disaccharide is an effective method for cryopreservation of SSCs. Trehalose is the most effective cryoprotectant among all the sugars tested and only lactose was comparable to trehalose. Our proliferation and transplantation data show that serum-free freezing can be achieved in freezing medium supplemented with 200 mM trehalose, 10% knockout serum replacement, and 10% dimethyl sulfoxide (DMSO) for cryopreservation of SSCs.


CONCLUSIONS
These findings raise the possibility of effectively banking frozen SSCs from various species, including humans, in a traditional serum-free medium for germ cell research and male infertility treatments.
Department of Animal Science and Technology, Chung-Ang University, Ansung, Gyeonggi-Do, South Korea.
Yong-An
YA
Lee
Department of Animal Science and Technology, Chung-Ang University, Ansung, Gyeonggi-Do, South Korea.
Yong-Hee
YH
Kim
Department of Animal Science and Technology, Chung-Ang University, Ansung, Gyeonggi-Do, South Korea.
Seung-Jung
SJ
Ha
Department of Animal Science and Technology, Chung-Ang University, Ansung, Gyeonggi-Do, South Korea.
Bang-Jin
BJ
Kim
Department of Animal Science and Technology, Chung-Ang University, Ansung, Gyeonggi-Do, South Korea.
Ki-Jung
KJ
Kim
Department of Animal Science and Technology, Chung-Ang University, Ansung, Gyeonggi-Do, South Korea.
Mi-Seon
MS
Jung
Department of Animal Science and Technology, Chung-Ang University, Ansung, Gyeonggi-Do, South Korea.
Byung-Gak
BG
Kim
Department of Animal Science and Technology, Chung-Ang University, Ansung, Gyeonggi-Do, South Korea. Electronic address: byryu@cau.ac.kr.
Buom-Yong
BY
Ryu
D053687:Adult Stem Cells / Q000737:chemistry / Q000166:cytology* / Q000187:drug effects / Q000637:transplantation*; D000818:Animals; D002241:Carbohydrates / Q000737:chemistry* / Q000494:pharmacology*; D049109:Cell Proliferation; D002478:Cells, Cultured; D015925:Cryopreservation; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D012662:Semen Preservation / Q000379:methods*; D013094:Spermatozoa / Q000737:chemistry / Q000166:cytology / Q000187:drug effects / Q000637:transplantation; D013737:Testis / Q000166:cytology* / Q000601:surgery*
D002241:Carbohydrates
Cryopreservation; serum-free cryopreservation; spermatogonial stem cell; sugars