Propagation of adult SSCs: from mouse to human

Abstract

Adult spermatogonial stem cells (SSCs) represent a distinctive source of stem cells in mammals for several reasons. First, by giving rise to spermatogenesis, SSCs are responsible for the propagation of a father's genetic material. As such, autologous SSCs have been considered for treatment of infertility and other purposes, including correction of inherited disorders. Second, adult spermatogonia can spontaneously produce embryonic-like stem cells in vitro, which could be used as an alternative for therapeutic, diagnostic, or drug discovery strategies for humans. Therefore, an increasing urgency is driving efforts to understand the biology of SSCs and improve techniques to manipulate them in vitro as a prerequisite to achieve the aforementioned goals. The characterization of adult SSCs also requires reproducible methods to isolate and maintain them in long-term culture. Herein, we describe recent major advances and challenges in propagation of adult SSCs from mice and humans during the past few years, including the use of unique cell surface markers and defined cultured conditions.

Figure 1

In vitro propagation of adult SSCs. SSCs are derived from the adult testis using somatic feeders and media containing diverse growth factors such as GDNF and FGF-2. In the case of human SSCs, a preenrichment sorting step (FACS or MACS) using previously identified surface marker, is critical for the successful expansion of SSCs. Mouse cultures established in such way can be maintained for over 1 year. Long-term expansion of SSCs in vitro is confirmed by analyzing the expression of molecular markers of spermatogonia. Furthermore, the number of stem cells expanded in the culture must be validated and quantified by an in vivo functional assay consisting of transplantation of SSCs into busulfan-treated recipient mouse testis. The fluorescent image corresponds to seminiferous tubules repopulated with donor GFP-positive cells. Potential clinical applications of SSCs include restoration of male fertility and/or in vitro correction of mutated alleles prior to transplant. Furthermore, in vitro SSCs can spontaneously reprogram to embryonic-like stem cells and could be used for regenerative therapy.