The Effect of Silk Nanofibrous Scaffold and Co-Culture with Sertoli Cells on Spermatogonial Stem Cell Proliferation

Abstract

BACKGROUND AND OBJECTIVE: In vitro proliferation and maintenance of spermatogonial stem cells is one of the treatment options for infertile men. Creating a suitable microenvironment similar to the natural conditions of reproduction of these cells leads to the improvement of culture and spermatogenesis. Therefore, the aim of the present study was to use a 3D silk nanofibrous electrospun scaffold and co-culture with Sertoli cells in order to increase the proliferation of spermatogonial stem cells. METHODS: In this experimental study, spermatogonial stem cells were isolated from the testes of 50 2-4-day-old mice (5 samples each time) using two-step mechanical and enzymatic digestion within two weeks. After placing 2×104 cells on the surface of the silk scaffold, they were exposed to culture medium. The viability of these cells was assessed using MTT assay and their binding was evaluated by SEM microscopy. The studied variables, including Stra8, DAZL and Piwill2 specific genes were analyzed by real time PCR and immunocytochemistry. FINDINGS: The viability of cultured cells in the presence of scaffold and Sertoli (91±6), (90±2) was higher than the control group (85±2), (83±8). Real time PCR results confirmed a significant increase in the expression of specific markers of spermatogonial stem cells such as Stra8, Piwill2 and DAZL in cells cultured respectively on scaffold (1.47±6) (1.28±5) (1.43±2) compared to the 2D culture group (1.17±5) (1.05±3) (1.13±7) (p<0.05). CONCLUSION: The results showed that silk scaffold in the presence of Sertoli cell co-culture can increase in vitro proliferation of spermatogonial stem cells and can have a positive effect on the viability and proliferation of these cells.

Keywords

• scaffold
• silk
• sertoli
• proliferation
• spermatogonia.