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As a valuable resource for cell therapy, spermatogonial stem cells (SSCs) have raised hopes for the treatment of male infertility. Various 3D methods have been developed to produce cellular aggregates and mimic the organization and function of the testis. In order to improve in vitro spermatogenesis, the present study was performed with the aim of forming testicular organoid (TO) from mouse testicular cells in porous three-dimensional macro scaffolds obtained from fibroin silk-alginate-laminin. The silk cocoons were placed in a solution of sodium carbonate to remove sericin. The purified silk fibers were then dissolved in lithium bromide solution and dialyzed. After characterizing the purified silk fibroin, 8% silk fibrin, 8% alginate, 8% fibroin silk -8% alginate (1:1), and 8% fibroin silk -8% alginate (1:1) -5% laminate solutions were dehydrated. The morphology, porosity, FTIR, swelling, degradability, non-toxicity, and in vivo biocompatibility of scaffolds for SSCs were examined. Then, the differentiation of mouse testicular cells on porous structures was investigated using Real Time-PCR, ICC, flow cytometry techniques, and H&E staining. The functionality of Leydig and Sertoli cells was determined by their ability for hormone secretion. Results showed scaffolds obtained from fibroin silk-alginate-laminin could increase the hydrophilicity and subsequently enhance the swelling properties, proper surface properties, and good biocompatibility than other groups. Furthermore, inoculation of neonatal mouse testicular cells onto composite structures resulted in the generation of multicellular TOs in which the differentiation of SSCs to post-meiotic cells was confirmed. Hormonal analysis of composite scaffolds revealed the functionality of TOs in the secretion of testosterone and inhibin B. In general, our study suggested that a porous composite scaffold can be used for designing a functional bioartificial testis in vitro that would offer new fertility restoration options.
20941804008
