Enrichment and characterization of Thy1-positive male germline stem cells (mGSCs) from dairy goat (Capra hircus) testis using magnetic microbeads

摘要

In mammalian testis, male germline stem cells (mGSCs) are originated from primordial germ cells and developed into spermatocyte and spermatid. In our previous studies, we had isolated a pluripotent mGSCs from goat testes and tested their pluripotency and differentiation potential in vitro and in vivo, which revealed that the isolated and cultured dairy goat mGSCs maintained the characteristics of mGSCs. However, Thy1, a marker of mGSCs, was not examined in detail. In this study, the dairy goat mGSCs were purified by differential plating followed by magnetic-activated cell sorting (MACS) using Thy1 antibody. The quantitative reverse transcription polymerase chain reaction and immunofluorescence analyses revealed that the transcription and expression of Thy1, CD49f, Plzf, Oct4, Gfra1, and Vasa were higher in Thy1-positive cells when compared with Thy1-negative cells. The detection results of culturing dairy goat mGSCs indicated that the Thy1-positive cells maintained the characteristics of mGSCs, grew relatively faster than Thy1-negative cells, and the percentage of alkaline phosphatase-positive cells and colonies were significantly higher in Thy1-positive mGSCs than Thy1-negative cells. Collectively, these results indicate that THY1 is a marker of undifferentiated spermatogonia in goat testes, the technique of magnetic-activated cell sorting using Thy1 antibody could be an efficient method to enrich mGSCs in goat