Development of a long-term culture system for functional Sertoli cells.

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It has not been possible to culture functional Sertoli cells for long-term. We have developed a new system for culturing mouse Sertoli cells that provide microenvironment for spermatogonial stem cells. In this culture system, Sertoli cells could maintain spermatogonial stem cell activity for more than five months when they were supplemented with epidermal growth factor and fibroblast growth factor 2. Spermatogonial stem cells migrated under the Sertoli cell feeder layer and formed cobblestone colonies. Cultured spermatogonial stem cells could produce functional sperm upon transplantation into the seminiferous tubules of infertile mouse testes. Because cobblestone colony formation was suppressed by inhibiting CXCR4 or RET, these molecules may be involved in migration of spermatogonial stem cells to germline niche. Taken together, our novel culture system will provide a new system for functional assay of spermatogonial stem cells.