Serum- and feeder-free culture of mouse germline stem cells.
Spermatogonial stem cells (SSCs) undergo self-renewal divisions to support spermatogenesis. Although several in vitro SSC culture systems have been developed, these systems include serum or fibroblast feeders, which complicate SSC self-renewal analyses. Here, we developed a serum- and feeder-free culture system for long-term propagation of SSCs. In addition to the SSC self-renewal factors, including glial cell line-derived neurotrophic factor, supplementation with fetuin and lipid-associated molecules was required to drive SSC proliferation in vitro. Cultured cells proliferated for at least 6 mo at a rate comparable to that of serum-supplemented cultured cells. However, germline potential was reduced under serum- and feeder-free conditions, as indicated by a lower SSC frequency after germ cell transplantation. Nevertheless, the cultured cells completed spermatogenesis and produced offspring following spermatogonial transplantation into seminiferous tubules of infertile mice. This culture system provides a basic platform for understanding the regulation of SSC fate commitment in vitro and for improving SSC culture medium.
Kanatsu-Shinohara
Mito
M
Department of Molecular Genetics, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
Inoue
Kimiko
K
Ogonuki
Narumi
N
Morimoto
Hiroko
H
Ogura
Atsuo
A
Shinohara
Takashi
T
D000818:Animals; D002448:Cell Adhesion; D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation; D003470:Culture Media / Q000737:chemistry*; D008055:Lipids / Q000494:pharmacology; D008297:Male; D051379:Mice; D013093:Spermatogonia / Q000166:cytology*; D013234:Stem Cells / Q000166:cytology* / Q000502:physiology*; D000509:alpha-Fetoproteins / Q000378:metabolism
D003470:Culture Media; D008055:Lipids; D000509:alpha-Fetoproteins