ABSTRACT: This study explores the role of porcine spermatogonia germ cells and testicular fibroblast in testis development and spermatogenesis through xenotransplantation. Previous research demonstrated the importance of Sertoli cells in maintaining testicular structure and function during testis development. However, the contribution of spermatogonia to these processes remains unclear. H&E staining revealed that FO tissues, no testicular structures were formed; instead, heterogeneous tissues with simple tubular formations were observed. Conversely, in GF tissues, structures resembling testicular tissue developed, displaying various stages of testis development including elongated spermatid. Immunofluorescence staining showed seminiferous tubules expressing PGP 9.5, VASA, and ACR2, confirming the presence of spermatogenesis. Interestingly, FO tissues were dominantly expressed ESR1, indicating bipotent gonadal property. RNA-seq analysis further revealed that GF tissue showed spermatogenesis-linked GOBPs with gene expression of mature Sertoli cells, including NR5A1, AR, and DMRT1. However, FO tissues displayed early tissue formation processes with expression of BMP4, WNT5A, ESR1, TGFBR3 and ACVR, generally inhibited in normal testis. These findings highlight the critical role of germ cells in formation of testicular structures through xenotransplantation and suggest the potential for spermatogenesis within these structures. However, further research is needed to verify the integrity and fertilizing capacity of the sperm, as these aspects have not been confirmed in this study.
