Production of functional sperm from the red seabream (Pagrus major) ovotestis on a xenogeneic zebrafish Sertoli cell line in culture.

In vitro culture of germ cells has the potential to accelerate gametogenesis because it is independent of the body's endocrine control. However, the need to determine culture conditions for each species has limited the application of this method for many species. Here, we report the establishment of a culture system for germ cells from the ovotestes of red seabream to differentiate into functional sperm, as well as the use of a zebrafish Sertoli cell line as feeder cells for seabream germ cells. We first determined the culture conditions for red seabream germ cells based on a zebrafish spermatogonia culture system. Spermatogenesis was successfully induced by replacing cyprinid-derived body fluid components with red seabream serum and by using seabream collagen-coated dishes. Functional sperm were obtained from spermatogonia-enriched ovotestis dissociates after 28 days of culture. Furthermore, when red seabream germ cells were cultured on the zebrafish Sertoli cell line, they adhered to the feeder cells and differentiated into functional sperm. Given the phylogenetic distance between seabream and zebrafish, these results suggest that germ cell culture may be widely applied to teleost fish through modification of body fluid factors and the use of xenogeneic zebrafish Sertoli cells as feeder cells.