Culture and Differentiation of Goat Spermatogonial Stem Cells

Abstract: The objective of this study was to develop a practical in vitro culture system of goat spermatogonial stem cells (SSCs) to facilitate further studies on transgenesis. 12 testis from 2-month-old bucks (GuanZhong milk goat) were decapsulated and dissociated enzymatically by one step to recover seminiferous tubule cells. 1×106 cells/ml with a viability 82.7% was cultured in 25 cm2 flasks containing DMEM/F12 supplemented with 15% fetal bovine serum (FBS) for first 4 week， less FBS after each medium exchange thereafter with 10% FBS as final concentration at 37 ℃ in a humidified atmosphere with 5% CO₂. During the first two week of culture， the number of SSCs declined then increased， pairs and chains of spermatogonia appeared ，in the third and fourth week， bird-nest-like and mountain-like colonies were observed with positive alkaline phosphatase (AKP) staining. During the second month of culture， more and larger colonies were found， there were anchored and free elongate-spermatid-like cells around colonies， selected the colonies and cultured them confluent monolayer of Sertoli cells， spermatid were formed around. For the first time to our knowledge， goat spermatogonial stem cells have been cultured and differentiated into spermatids in vitro， this culture system would provide targeted cells for tansgenesis， greatly accelerate the research on goat spermatogenesis and utilization of excellent male gene resources.

CSTR: 32200.14.cjcb.2005.02.0026