title: Conditions for Long-Term Culture of Cattle Undifferentiated Spermatogonia.
abstract: Continual and robust spermatogenesis relies on the actions of an undifferentiated spermatogonial population that contains stem cells. A remarkable feature of spermatogonial stem cells (SSCs) is the capacity to regenerate spermatogenesis following isolation from a donor testis and transplantation into a permissive recipient testis. This capacity has enormous potential as a tool for enhancing the reproductive capacity of livestock, which can improve production efficiency. Because SSCs are a rare subset of the undifferentiated spermatogonial population, a period of in vitro amplification in number following isolation from donor testicular tissue is essential. Here, we describe methodology for isolation of a cell fraction from prepubertal bull testes that is enriched for undifferentiated spermatogonia and long-term maintenance of the cells in both the feeder cell coculture and the feeder-free format. To achieve this method, we derived bovine fetal fibroblasts (BFF) to serve as feeders for optimizing medium conditions that promote maintenance of bovine undifferentiated spermatogonia for at least 2 mo. In addition, we devised a feeder-free system with BFF-conditioned medium that sustained bovine undifferentiated spermatogonia for at least 1 mo in vitro. The methodologies described could be optimized to provide platforms for exponential expansion of bovine SSCs that will provide the numbers needed for transplantation into recipient testes.
authors.affiliation: Center for Reproductive Biology, School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Pullman, Washington.
authors.forename: Melissa J
authors.initials: MJ
authors.lastname: Oatley
authors.affiliation: Center for Reproductive Biology, School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Pullman, Washington.
authors.forename: Amy V
authors.initials: AV
authors.lastname: Kaucher
authors.affiliation: Center for Reproductive Biology, School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Pullman, Washington.
authors.forename: Qi-En
authors.initials: QE
authors.lastname: Yang
authors.affiliation: Center for Reproductive Biology, School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Pullman, Washington.
authors.forename: Muhammad Salman
authors.initials: MS
authors.lastname: Waqas
authors.affiliation: Center for Reproductive Biology, School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Pullman, Washington joatley@vetmed.wsu.edu.
authors.forename: Jon M
authors.initials: JM
authors.lastname: Oatley
mesh_terms: D000818:Animals; D002417:Cattle; D018929:Cell Culture Techniques / Q000379:methods*; D002478:Cells, Cultured; D003470:Culture Media; D008297:Male; D013091:Spermatogenesis / Q000502:physiology*; D013093:Spermatogonia / Q000166:cytology*; D013234:Stem Cells / Q000166:cytology*; D013737:Testis / Q000166:cytology*
chemical_list: D003470:Culture Media
keywords: camel; cows; culture; llama; ruminants; sheep; spermatogonia; spermatogonial stem cells