Proliferation of human spermatogonial stem cells on optimized PCL/Gelatin nanofibrous scaffolds.

Abstract

Improvement of culture system and increasing the proliferation of spermatogonia stem cells under in vitro condition are the essential treatment options for infertility before autologous transplantation. Therefore, the present study aimed to evaluate the proliferation of human spermatogonia stem cells on the electrospun polycaprolactone/gelatin nanocomposite. Therefore, for this purpose, nanofiber porous scaffolds were prepared using the electrospinning method and their structures were then confirmed by SEM. After performing swelling, biodegradability and cell adhesion tests, human spermatogonia stem cells were cultured on scaffolds. In addition, both cell viability and proliferation were assessed using immunocytochemistry, flow cytometry and real-time PCR techniques in culturing during a 3-week period. SEM images indicated the presence of fibres with suitable diameters and arrangement as well as a sufficient porosity in nanocomposite scaffolds, showing good biocompatibility and biodegradability. The results show a significant increase in the number of spermatogonia stem cells in the cultured group on scaffold compared with the control group (p≤0.05). As well, the results show that the expressions of integrin ɑ6 and β1 and Plzf genes estimated using real-time PCR in nanofiber scaffolds were significantly higher than those of the control group (p≤0.05). However, the expression of c-Kit gene in the 3D group showed a significant decrease compared with the 2D group. Flow cytometry analysis also showed that the number of Plzf-positive cells was significantly higher in nanofiber porous scaffolds compared with the control group (p≤0.05). Additionally, immunocytochemistry findings confirmed the presence of human spermatogonia stem cell colonies. In general, it seems that the designed nanocomposite scaffold could provide a suitable capacity for self-renewal of human spermatogonia stem cells, which can have a good application potential in research and reconstructive medicine related to the field of male infertility.