Characterization of Oct4‐GFP spermatogonial stem cell line and its application in the reprogramming studies

Abstract

Establishment of mouse spermatogonial stem cell (SSC) culture systems offers a useful stem cell model for studies of proliferation and self-renewal of mammalian germline stem cells. In addition, spontaneous development of pluripotent stem cells from cultured SSCs emphasizes their possible applications in regenerative medicine as a substitute for embryonic stem cells (ESCs). These pluripotent stem cells termed multipotent germline stem cells (mGSCs) or germline-derived pluripotent stem cells (gPSCs) exhibit almost identical properties in terms of morphology and gene expression patterns to mouse ESCs (mESCs). In this study, to help understand mechanisms underlying reprogramming of SSCs into pluripotent stem cells, we established a culture system of SSCs derived from mice harboring green fluorescence protein (GFP) transgene whose expression is modulated by Oct4 regulatory sequences. Our results indicated that GFP intensity faithfully reflected cellular states upon reprogramming of SSCs or treatment with a selective extracellular signal-regulated kinase (ERK) inhibitor PD0325901. Moreover, in contrast to mESCs, regulation of Nanog expression did not appear to couple to the Oct4 level in SSCs. Further analysis of Oct4-GFP SSCs demonstrated that a posttranscriptional control of pluripotency marker genes such as Oct4 and Sox2 might play an important role as an inhibitory mechanism preventing the acquisition of pluripotency.