Abstract
BackgroundRegenerative medicine potentially offers the opportunity for curing male infertility. Native extracellular matrix (ECM) creates a reconstruction platform to replace the organs. In this study, we aimed to evaluate the efficiency of the testis decellularized scaffold as a proper niche for stem cell differentiation toward testis-specific cell lineages.MethodsRats’ testes were decellularized by freeze-thaw cycle followed by immersion in deionized distilled water for 2 h, perfused with 1% Triton X-100 through ductus deferens for 4 h, 1% SDS for 48 h and 1% DNase for 2 h. The decellularized samples were prepared for further in vitro and in vivo analyses.ResultHistochemical and immunohistochemistry studies revealed that ECM components such as Glycosaminoglycans (GAGs), neutral carbohydrate, elastic fibers, collagen I & IV, laminin, and fibronectin were well preserved, and the cells were completely removed after decellularization. Scanning electron microscopy (SEM) showed that 3D ultrastructure of the testis remained intact. In vivo and in vitro studies point out that decellularized scaffold was non-toxic and performed a good platform for cell division. In vivo implant of the scaffolds with or without mesenchymal stem cells (MSCs) showed that appropriate positions for transplantation were the mesentery and liver and the scaffolds could induce donor-loaded MSCs or host migrating cells to differentiate to the cells with phenotype of the sertoli- and leydig-like cells. The scaffolds also provide a good niche for migrating DAZL-positive cells; however, they could not differentiate into post meiotic-cell lineages.ConclusionThe decellularized testis can be considered as a promising vehicle to support cell transplantation and may provide an appropriate niche for testicular cell differentiation.
Highlights
Regenerative medicine potentially offers the opportunity for curing male infertility
We found that the 3D ultrastructure of the testis remained with minimal damage after decellularization which is consistent with previous studies [34, 35]
We did not use any growth factor supplementation or co-culture system and we found that DAZL-positive cells migrated from the host to both cell-loaded or cell-free decellularized testis
Summary
Regenerative medicine potentially offers the opportunity for curing male infertility. The proliferation and self-renewal of the spermatogonial stem cells were improved by culturing in 3D agarose hydrogel [4]. 3D collagen/alginate hydrogel improved the embryonic stem cell-derived primordial germ cell differentiation [8]. 3D culture systems can be considered as a biomimicry of the testis microenvironment and improve the spermatogonial stem cell differentiation condition. The extracellular matrix and basement membrane constitute the main niche for germ cell as well as sertoli and interstitial cell that support their functions [9]. Decellularized tissues with naïve microarchitecture can be considered as an appropriate niche for germ cell differentiation and has been reported to be used for adult testicular tissue culture [2]. The decellularized tissues degrade slowly after implantation and are generally replaced by the host cell-derived ECM [15]
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