Development of mouse spermatogonial purification system and its application for fertility restoration in mice

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We attempted to restore fertility to infertile mouse by increasing the donor cell colonization level. For this purpose, we used two approaches. First, we used immature mice for recipients. We have regularly used mature adult mice for fertility restoration by spermatogonial transplantation. In the present experiment, we used 5-10 day old pups as recipients. This was based on the observation that there are no tight junction between Sertoli cells at this stage. We therefore hypothesized that the absence of tight junction, thereby increasing the the colonization process of transplanted stem cells.We found that colonization efficiency in the pup testis is 10-fold higher than in the adult testis. Furthermore, 80 to 100% of the recipiet males became fertile upon mating with females. Surprisingly, the recipients sired offspring at age equivalent to wild type mice. As a second approach to increase donor cell colonization, we generated monoclonal antibodies to immature testis. The testes contain enriched population of undifferentiated spermatogonia, including stem cells. We immunized these testis cells in rats, and obtained several clones. One of the clones were able to enrich stem cells by 5-fold after spermatogonial transplantation. We are now trying to characterize the molecular structure of the antigen.In summary, we have established an efficient system for fertility restoration by spermatogonial transplantation in mice. We also found a novel antigen on spermatogonial stem cells. These results have important implications for developing technique for fertility restoration in humans.