Although chicken spermatogonial stem cells (SSCs) have received considerable attention in recent years, only a few studies so far have focused on their derivation and characterization in vitro. Identification of specific molecular biomarkers and differentiation capacity of chicken SSCs would not only help us to understand cell and molecular biology of these cells, but also can contribute to their applications in biotechnology. In this regard, we found that colony-forming cells (SSCs) in newborn chicken testicular cell cultures were positive for alkaline phosphatase activity and also expressed specific markers including DAZL,STRA-8,CVH,PLZF SPRY-1,GFRα1,GDNF,POU5F1,NANOG,GPR125,THY-1, c-KIT,and BCL6B, at mRNA level. Moreover, these cells expressed POU5F1 and GPR125 proteins as reliable intracellular and cell surface markers, respectively; whereas they were negative for SSEA-1. Furthermore, we showed that newborn chicken colony-forming cells had spermatogenesis potential and thus could be produced sperm-like cells in a three-dimensional matrix in vitro. In conclusion, this study reports novel insights into the molecular signature of newborn chicken SSCs in comparison with mammalian SSCs and for the first time we report a successful protocol for in vitro spermatogenesis and thus production of sperm-like cells from newborn chicken testicular cell cultures.

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