Isolation And Characterization Of Human Spermatogonial Stem Cell-like Cells Derived Under Feeder-free And Xeno-free System

As the key carriers for sexual reproducing creatures to transmit their genetic information through generations,germ cells play an irreplaceable role in reproduction.Spermatogenesis is a multi-step process,during which spermatogonial stem cells(SSCs)are essential for maintaining fertility via continuously producing male gametes.Male factor contributes to half of infertility incidence all over the world,which increases continuously over the last several years.Therefore,it will be of great benefit to induce germline cells in vitro for the fundamental research of reproductive development but also for the treatment of male infertility.In this study,I charicterized PLZF+SSC-like cells(SSCLCs)that were derived under the feeder-free and xeno-free culture condition from human pluripotent stem cells(PSCs).I found that these SSCLCs showed similar functional properties as in vivo isolated SSCs to migrate to the basement membrane.In addition,NANOS3 played an essential role in SSCLCs formation.Interestingly,I found that these germline cells were co-stained with multiple surface antigens.I further developed a method to isolate and purify these SSCLCs by sorting Ep-CAM-/TNAP+population after removing residual PSCs by TRA-1-81 exclusion.Taken together,my work provides experimental methods to isolate and characterize these PSC-derived SSCLCs both in vitro and in vivo.