| Abstrakt: |
Introduction: Decellularizing testis tissue and recellularizing with spermatogonial stem cells (SSCs) seems to be a promising approach to restore fertility in prepubertal boys who undergoes cytotoxic therapies. Methods: Testis samples were obtained from brain death donors. Testis tissue decellularization was performed by adding 1% SDS and confirmed by histological analysis. The MTT assay was performed for biocompatibility analyses. SSCs were derived from male mice and were seeded onto the decellularized testicular matrix (DTM) scaffold. The recellularized DTM scaffold was cultured in a static cultivation system for 1 week, then transferred in a dynamic mini-perfusion bioreactor for 2 weeks. The expression of Id4, Plzf, Gfra-1, Prm1, Sycp3, Abp, Ki67, Bax, and Bcl2 genes were assessed in cellular suspension of 0 day and recellularized DTM after 1 week static and 2 weeks dynamic cultivations. Results: DNA qualification indicated that approximately 99% of the DNA components were removed from DTMs. Hematoxylin Eosin, Masson's Trichrome, and DAPI staining confirmed the effective recellularization. Dynamic cultivation of recellularized DTMs at the flow rate of 10 ml/h provided optimum conditions. The expression of SSCs-specific genes of Id4, Plzf, and Gfra-1 and post-meiosis genes of Scp3, prm1, and Abp was insignificantly higher in the dynamic cultivation of recellularized DTMs than in the cellular suspension of day 0. Ki67 expression was shown no difference between dynamic cultivation and day 0. An insignificant lower expression of the Bax and higher expression of Bcl2 genes was detected in the dynamic cultivation of recellularized DTMs compared to day 0. Conclusion: Our results indicated that SSCs can successfully be attached to the DTMs and effectively proliferate in the mini-perfusion bioreactor. [ABSTRACT FROM AUTHOR] |
