title: Increased proportion of donor primordial germ cells in chimeric gonads by sterilisation of recipient embryos using busulfan sustained-release emulsion in chickens.
abstract: The aim of the present study was to improve the efficiency of endogenous primordial germ cell (PGC) depletion and to increase the ratio of donor PGCs in the gonads of recipient chicken embryos. A sustained-release emulsion was prepared by emulsifying equal amounts of Ca(2+)- and Mg(2+)-free phosphate-buffered saline containing 10% busulfan solubilised in N,N-dimethylformamide and sesame oil, using a filter. Then, 75 microg per 50 microL busulfan sustained-release emulsion was injected into the yolk. To determine the depletion and repopulation of PGCs in the gonads after 6 days incubation, whole-mount immunostaining was performed. The busulfan sustained-release emulsion significantly reduced the number of endogenous PGCs compared with control (P < 0.05). Moreover, the busulfan sustained-release emulsion significantly depleted endogenous PGCs compared with other previously reported busulfan delivery systems (P < 0.05), but with less variation, suggesting that the sustained-release emulsion delivered a consistent amount of busulfan to the developing chicken embryos. The PGC transfer study showed that the proportion of donor PGCs in the gonads of busulfan sustained-release emulsion-treated embryos after 6 days incubation increased 28-fold compared with control. In conclusion, the results demonstrate that exogenous PGCs are capable of migrating and settling in gonads from which endogenous PGCs have been removed using a busulfan sustained-release emulsion.
authors.affiliation: Faculty of Agriculture, Shinshu University, Minamiminowa, Nagano, Japan.
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Nakamura
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Yamamoto
authors.forename: Fumitake
authors.initials: F
authors.lastname: Usui
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Atsumi
authors.forename: Yohei
authors.initials: Y
authors.lastname: Ito
authors.forename: Tamao
authors.initials: T
authors.lastname: Ono
authors.forename: Kumiko
authors.initials: K
authors.lastname: Takeda
authors.forename: Keijiro
authors.initials: K
authors.lastname: Nirasawa
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kagami
authors.forename: Takahiro
authors.initials: T
authors.lastname: Tagami
mesh_terms: D000477:Alkylating Agents / Q000008:administration & dosage / Q000494:pharmacology*; D000818:Animals; D002066:Busulfan / Q000008:administration & dosage / Q000494:pharmacology*; D002465:Cell Movement / Q000187:drug effects; D017690:Cell Transplantation / Q000379:methods*; D002642:Chick Embryo / Q000166:cytology / Q000187:drug effects*; D003692:Delayed-Action Preparations; D047108:Embryonic Development / Q000187:drug effects; D004655:Emulsions; D005854:Germ Cells / Q000166:cytology / Q000187:drug effects*; D007267:Injections; D013245:Sterilization, Reproductive; D018183:Transplantation Chimera
chemical_list: D000477:Alkylating Agents; D003692:Delayed-Action Preparations; D004655:Emulsions; D002066:Busulfan