Co-culture of mouse spermatogonial stem cells with sertoli cell as a feeder layer, stimulates the proliferation and spermatogonial stemness profile

Abstract

Abstract Objectives Sertoli cells effect the fate map of spermatogonial stem cells (SSCs) to self-renew via providing the special microenvironments. Maintenance of proliferation and self-renewal activity of SSCs may be usable as a therapeutic strategy, leads to increase the recovery of male fertility. This research was aimed to evaluate the effect of mouse sertoli cells on spermatogonia stem cells proliferation and the expression pattern of stemness markers. Methods Spermatogonia stem cells were collected from neonatal mouse testis using a two-step mechanical and enzymatic digestion. SSCs were cultured in three groups: The first group or co-culture group consists of spermatogonia and sertoli cells that were cultured together. The control group, only spermatogonial cells and the group no. 3 included spermatogonial cells in the presence of GDNF. The colony formation of mentioned groups, was monitored during one month in culture. Identification of the colonies, was confirmed using PLZF and Oct4 immunostaining. Spermatogonial stemness genes includes; Stra8, mvh and piwill2 were analyzed by RT-PCR. Results In the co-culture group, cells proliferated rapidly and many colonies were appeared whereas they were rarely formed in the control groups. Colonies were exhibited alkaline phosphatesase activity and were immunopositive to Oct4 and PLZF, strongly. The gene expression of srta8, mvh and piwill2, in SSCs that were cultivated with sertoli cells, were greater significantly than other control groups. Conclusion It is concluded that co-culture of SSCs with sertoli cells prepares conditions which leads to efficient proliferation and maintenance of stemness condition of SSCs, that is usable as a therapeutic approach for treatment of male fertility.